

<!--XBRL Document Created with Wdesk from Workiva-->
<!-- -->
<link:linkbase xsi:schemaLocation="http://www.xbrl.org/2003/linkbase http://www.xbrl.org/2003/xbrl-linkbase-2003-12-31.xsd" xmlns:link="http://www.xbrl.org/2003/linkbase" xmlns:xbrldt="http://xbrl.org/2005/xbrldt" xmlns:xlink="http://www.w3.org/1999/xlink" xmlns:xsi="http://www.w3.org/2001/XMLSchema-instance">
  <link:roleRef roleURI="http://www.autolus.com/role/AccruedExpensesAndOtherLiabilities" xlink:href="autl-20190630.xsd#AccruedExpensesAndOtherLiabilities" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/AccruedExpensesAndOtherLiabilitiesDetails" xlink:href="autl-20190630.xsd#AccruedExpensesAndOtherLiabilitiesDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/AccruedExpensesAndOtherLiabilitiesNarrativesDetails" xlink:href="autl-20190630.xsd#AccruedExpensesAndOtherLiabilitiesNarrativesDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/AccruedExpensesAndOtherLiabilitiesTables" xlink:href="autl-20190630.xsd#AccruedExpensesAndOtherLiabilitiesTables" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/CommitmentsAndContingencies" xlink:href="autl-20190630.xsd#CommitmentsAndContingencies" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/CommitmentsAndContingenciesFutureLeasePaymentsDetails" xlink:href="autl-20190630.xsd#CommitmentsAndContingenciesFutureLeasePaymentsDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/CommitmentsAndContingenciesFutureMinimumLeasePaymentsBeforeAdoptionDetails" xlink:href="autl-20190630.xsd#CommitmentsAndContingenciesFutureMinimumLeasePaymentsBeforeAdoptionDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/CommitmentsAndContingenciesLeaseBalanceSheetLocationDetails" xlink:href="autl-20190630.xsd#CommitmentsAndContingenciesLeaseBalanceSheetLocationDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/CommitmentsAndContingenciesLeaseCostDetails" xlink:href="autl-20190630.xsd#CommitmentsAndContingenciesLeaseCostDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/CommitmentsAndContingenciesNarrativesDetails" xlink:href="autl-20190630.xsd#CommitmentsAndContingenciesNarrativesDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/CommitmentsAndContingenciesOtherInformationDetails" xlink:href="autl-20190630.xsd#CommitmentsAndContingenciesOtherInformationDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/CommitmentsAndContingenciesTables" xlink:href="autl-20190630.xsd#CommitmentsAndContingenciesTables" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/CondensedConsolidatedBalanceSheetsUnaudited" xlink:href="autl-20190630.xsd#CondensedConsolidatedBalanceSheetsUnaudited" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/CondensedConsolidatedBalanceSheetsUnauditedParentheticals" xlink:href="autl-20190630.xsd#CondensedConsolidatedBalanceSheetsUnauditedParentheticals" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/CondensedConsolidatedStatementsOfCashFlowsUnaudited" xlink:href="autl-20190630.xsd#CondensedConsolidatedStatementsOfCashFlowsUnaudited" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/CondensedConsolidatedStatementsOfOperationsAndComprehensiveLossUnaudited" xlink:href="autl-20190630.xsd#CondensedConsolidatedStatementsOfOperationsAndComprehensiveLossUnaudited" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/CondensedConsolidatedStatementsOfShareholdersEquityUnaudited" xlink:href="autl-20190630.xsd#CondensedConsolidatedStatementsOfShareholdersEquityUnaudited" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/DocumentAndEntityInformation" xlink:href="autl-20190630.xsd#DocumentAndEntityInformation" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/EmployeeBenefitPlans" xlink:href="autl-20190630.xsd#EmployeeBenefitPlans" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/EmployeeBenefitPlansDetails" xlink:href="autl-20190630.xsd#EmployeeBenefitPlansDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/IncomeTaxes" xlink:href="autl-20190630.xsd#IncomeTaxes" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/IncomeTaxesDetails" xlink:href="autl-20190630.xsd#IncomeTaxesDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/LicenseAgreements" xlink:href="autl-20190630.xsd#LicenseAgreements" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/LicenseAgreementsDetails" xlink:href="autl-20190630.xsd#LicenseAgreementsDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/NatureOfBusiness" xlink:href="autl-20190630.xsd#NatureOfBusiness" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/NatureOfBusinessDetails" xlink:href="autl-20190630.xsd#NatureOfBusinessDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/PrepaidExpensesAndOtherCurrentAssets" xlink:href="autl-20190630.xsd#PrepaidExpensesAndOtherCurrentAssets" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/PrepaidExpensesAndOtherCurrentAssetsDetails" xlink:href="autl-20190630.xsd#PrepaidExpensesAndOtherCurrentAssetsDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/PrepaidExpensesAndOtherCurrentAssetsTables" xlink:href="autl-20190630.xsd#PrepaidExpensesAndOtherCurrentAssetsTables" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/PropertyAndEquipmentNet" xlink:href="autl-20190630.xsd#PropertyAndEquipmentNet" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/PropertyAndEquipmentNetDetails" xlink:href="autl-20190630.xsd#PropertyAndEquipmentNetDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/PropertyAndEquipmentNetNarrativesDetails" xlink:href="autl-20190630.xsd#PropertyAndEquipmentNetNarrativesDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/PropertyAndEquipmentNetTables" xlink:href="autl-20190630.xsd#PropertyAndEquipmentNetTables" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/RelatedPartyTransactions" xlink:href="autl-20190630.xsd#RelatedPartyTransactions" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/RelatedPartyTransactionsDetails" xlink:href="autl-20190630.xsd#RelatedPartyTransactionsDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareBasedCompensation" xlink:href="autl-20190630.xsd#ShareBasedCompensation" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareBasedCompensation2018PlanAnd2017PlanDetails" xlink:href="autl-20190630.xsd#ShareBasedCompensation2018PlanAnd2017PlanDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareBasedCompensationOptionsValuationAssumptionsDetails" xlink:href="autl-20190630.xsd#ShareBasedCompensationOptionsValuationAssumptionsDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareBasedCompensationRestrictedAndOrdinarySharesNarrativesDetails" xlink:href="autl-20190630.xsd#ShareBasedCompensationRestrictedAndOrdinarySharesNarrativesDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareBasedCompensationRestrictedOrdinarySharesRollforwardDetails" xlink:href="autl-20190630.xsd#ShareBasedCompensationRestrictedOrdinarySharesRollforwardDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareBasedCompensationShareBasedCompensationAllocationDetails" xlink:href="autl-20190630.xsd#ShareBasedCompensationShareBasedCompensationAllocationDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareBasedCompensationShareOptionsNarrativesDetails" xlink:href="autl-20190630.xsd#ShareBasedCompensationShareOptionsNarrativesDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareBasedCompensationShareOptionsRollforwardDetails" xlink:href="autl-20190630.xsd#ShareBasedCompensationShareOptionsRollforwardDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareBasedCompensationTables" xlink:href="autl-20190630.xsd#ShareBasedCompensationTables" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareholdersEquity" xlink:href="autl-20190630.xsd#ShareholdersEquity" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareholdersEquityDetails" xlink:href="autl-20190630.xsd#ShareholdersEquityDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareholdersEquityInitialPublicOfferingAndImpactOfCorporateReorganizationDetails" xlink:href="autl-20190630.xsd#ShareholdersEquityInitialPublicOfferingAndImpactOfCorporateReorganizationDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareholdersEquityOrdinarySharesDetails" xlink:href="autl-20190630.xsd#ShareholdersEquityOrdinarySharesDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/ShareholdersEquityTables" xlink:href="autl-20190630.xsd#ShareholdersEquityTables" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SubsequentEvent" xlink:href="autl-20190630.xsd#SubsequentEvent" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SummaryOfSignificantAccountingPolicies" xlink:href="autl-20190630.xsd#SummaryOfSignificantAccountingPolicies" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesAntidilutiveSharesDetails" xlink:href="autl-20190630.xsd#SummaryOfSignificantAccountingPoliciesAntidilutiveSharesDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesComprehensiveLossDetails" xlink:href="autl-20190630.xsd#SummaryOfSignificantAccountingPoliciesComprehensiveLossDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesForeignCurrencyRemeasurementAndTranslationDetails" xlink:href="autl-20190630.xsd#SummaryOfSignificantAccountingPoliciesForeignCurrencyRemeasurementAndTranslationDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesIntangibleAssetsSubjectToAmortizationDetails" xlink:href="autl-20190630.xsd#SummaryOfSignificantAccountingPoliciesIntangibleAssetsSubjectToAmortizationDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesPatentCostsDetails" xlink:href="autl-20190630.xsd#SummaryOfSignificantAccountingPoliciesPatentCostsDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesPolicies" xlink:href="autl-20190630.xsd#SummaryOfSignificantAccountingPoliciesPolicies" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesPropertyPlantAndEquipmentDetails" xlink:href="autl-20190630.xsd#SummaryOfSignificantAccountingPoliciesPropertyPlantAndEquipmentDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesResearchAndDevelopmentDetails" xlink:href="autl-20190630.xsd#SummaryOfSignificantAccountingPoliciesResearchAndDevelopmentDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesRestrictedCashDetails" xlink:href="autl-20190630.xsd#SummaryOfSignificantAccountingPoliciesRestrictedCashDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesSegmentInformationDetails" xlink:href="autl-20190630.xsd#SummaryOfSignificantAccountingPoliciesSegmentInformationDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesShareBasedCompensationDetails" xlink:href="autl-20190630.xsd#SummaryOfSignificantAccountingPoliciesShareBasedCompensationDetails" xlink:type="simple" />
  <link:roleRef roleURI="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesTables" xlink:href="autl-20190630.xsd#SummaryOfSignificantAccountingPoliciesTables" xlink:type="simple" />
  <link:arcroleRef arcroleURI="http://xbrl.org/int/dim/arcrole/all" xlink:href="http://www.xbrl.org/2005/xbrldt-2005.xsd#all" xlink:type="simple" />
  <link:arcroleRef arcroleURI="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:href="http://www.xbrl.org/2005/xbrldt-2005.xsd#dimension-default" xlink:type="simple" />
  <link:arcroleRef arcroleURI="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:href="http://www.xbrl.org/2005/xbrldt-2005.xsd#dimension-domain" xlink:type="simple" />
  <link:arcroleRef arcroleURI="http://xbrl.org/int/dim/arcrole/domain-member" xlink:href="http://www.xbrl.org/2005/xbrldt-2005.xsd#domain-member" xlink:type="simple" />
  <link:arcroleRef arcroleURI="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:href="http://www.xbrl.org/2005/xbrldt-2005.xsd#hypercube-dimension" xlink:type="simple" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/AccruedExpensesAndOtherLiabilities" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/AccruedExpensesAndOtherLiabilitiesDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/AccruedExpensesAndOtherLiabilitiesNarrativesDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/AccruedExpensesAndOtherLiabilitiesTables" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/CommitmentsAndContingencies" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/CommitmentsAndContingenciesFutureLeasePaymentsDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/CommitmentsAndContingenciesFutureMinimumLeasePaymentsBeforeAdoptionDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/CommitmentsAndContingenciesLeaseBalanceSheetLocationDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/CommitmentsAndContingenciesLeaseCostDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LesseeLeaseDescriptionLineItems" xlink:label="loc_us-gaap_LesseeLeaseDescriptionLineItems_6AAA0F60F02683C3D6E354A6DEF3F1F7" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LesseeLeaseDescriptionTable" xlink:label="loc_us-gaap_LesseeLeaseDescriptionTable_03B0C7D3609DB66B90D554A6DEF2711A" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_LesseeLeaseDescriptionLineItems_6AAA0F60F02683C3D6E354A6DEF3F1F7" xlink:to="loc_us-gaap_LesseeLeaseDescriptionTable_03B0C7D3609DB66B90D554A6DEF2711A" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_IncomeStatementLocationAxis" xlink:label="loc_us-gaap_IncomeStatementLocationAxis_110C1E7B19D062BB680554A6DEF281C0" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_LesseeLeaseDescriptionTable_03B0C7D3609DB66B90D554A6DEF2711A" xlink:to="loc_us-gaap_IncomeStatementLocationAxis_110C1E7B19D062BB680554A6DEF281C0" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_IncomeStatementLocationDomain" xlink:label="loc_us-gaap_IncomeStatementLocationDomain_B83F9AAFADFC4E517E1254A6DEF2AF91_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_IncomeStatementLocationAxis_110C1E7B19D062BB680554A6DEF281C0" xlink:to="loc_us-gaap_IncomeStatementLocationDomain_B83F9AAFADFC4E517E1254A6DEF2AF91_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_IncomeStatementLocationDomain" xlink:label="loc_us-gaap_IncomeStatementLocationDomain_B83F9AAFADFC4E517E1254A6DEF2AF91" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_IncomeStatementLocationAxis_110C1E7B19D062BB680554A6DEF281C0" xlink:to="loc_us-gaap_IncomeStatementLocationDomain_B83F9AAFADFC4E517E1254A6DEF2AF91" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ResearchAndDevelopmentExpenseMember" xlink:label="loc_us-gaap_ResearchAndDevelopmentExpenseMember_7CFF17135FB81AB0415D54A6DEF3E135" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncomeStatementLocationDomain_B83F9AAFADFC4E517E1254A6DEF2AF91" xlink:to="loc_us-gaap_ResearchAndDevelopmentExpenseMember_7CFF17135FB81AB0415D54A6DEF3E135" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_GeneralAndAdministrativeExpenseMember" xlink:label="loc_us-gaap_GeneralAndAdministrativeExpenseMember_65D59D0D8D8B263B7E8754A6DEF37F4E" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncomeStatementLocationDomain_B83F9AAFADFC4E517E1254A6DEF2AF91" xlink:to="loc_us-gaap_GeneralAndAdministrativeExpenseMember_65D59D0D8D8B263B7E8754A6DEF37F4E" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LeaseCostAbstract" xlink:label="loc_us-gaap_LeaseCostAbstract_5CC3FE1F60253E74CEFF54A6DEF3695B" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LesseeLeaseDescriptionLineItems_6AAA0F60F02683C3D6E354A6DEF3F1F7" xlink:to="loc_us-gaap_LeaseCostAbstract_5CC3FE1F60253E74CEFF54A6DEF3695B" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_OperatingLeaseCost" xlink:label="loc_us-gaap_OperatingLeaseCost_BE637794F90BC4B5567254A6DEF43231" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LeaseCostAbstract_5CC3FE1F60253E74CEFF54A6DEF3695B" xlink:to="loc_us-gaap_OperatingLeaseCost_BE637794F90BC4B5567254A6DEF43231" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_VariableLeaseCost" xlink:label="loc_us-gaap_VariableLeaseCost_0C2550B0300269361DAE54A6DEF5BB4B" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LeaseCostAbstract_5CC3FE1F60253E74CEFF54A6DEF3695B" xlink:to="loc_us-gaap_VariableLeaseCost_0C2550B0300269361DAE54A6DEF5BB4B" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShortTermLeaseCost" xlink:label="loc_us-gaap_ShortTermLeaseCost_6886E15F892E7CCC2C4954A6DEF653E9" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LeaseCostAbstract_5CC3FE1F60253E74CEFF54A6DEF3695B" xlink:to="loc_us-gaap_ShortTermLeaseCost_6886E15F892E7CCC2C4954A6DEF653E9" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LeaseCost" xlink:label="loc_us-gaap_LeaseCost_81F858900E80C913D8D754A6DEF63DD0" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LeaseCostAbstract_5CC3FE1F60253E74CEFF54A6DEF3695B" xlink:to="loc_us-gaap_LeaseCost_81F858900E80C913D8D754A6DEF63DD0" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/CommitmentsAndContingenciesNarrativesDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LossContingenciesLineItems" xlink:label="loc_us-gaap_LossContingenciesLineItems_C43A68C25CA86BA7AC8A54A5BFE2E984" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LossContingenciesTable" xlink:label="loc_us-gaap_LossContingenciesTable_6DE10A04251958D0F82C54A5BFE20D3C" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_LossContingenciesLineItems_C43A68C25CA86BA7AC8A54A5BFE2E984" xlink:to="loc_us-gaap_LossContingenciesTable_6DE10A04251958D0F82C54A5BFE20D3C" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LossContingenciesByNatureOfContingencyAxis" xlink:label="loc_us-gaap_LossContingenciesByNatureOfContingencyAxis_4ADF71183C187C56506A54A5BFE2B1C7" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_LossContingenciesTable_6DE10A04251958D0F82C54A5BFE20D3C" xlink:to="loc_us-gaap_LossContingenciesByNatureOfContingencyAxis_4ADF71183C187C56506A54A5BFE2B1C7" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LossContingencyNatureDomain" xlink:label="loc_us-gaap_LossContingencyNatureDomain_88F1A22017447DC4AF5C54A5BFE29506_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_LossContingenciesByNatureOfContingencyAxis_4ADF71183C187C56506A54A5BFE2B1C7" xlink:to="loc_us-gaap_LossContingencyNatureDomain_88F1A22017447DC4AF5C54A5BFE29506_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LossContingencyNatureDomain" xlink:label="loc_us-gaap_LossContingencyNatureDomain_88F1A22017447DC4AF5C54A5BFE29506" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_LossContingenciesByNatureOfContingencyAxis_4ADF71183C187C56506A54A5BFE2B1C7" xlink:to="loc_us-gaap_LossContingencyNatureDomain_88F1A22017447DC4AF5C54A5BFE29506" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LicensingAgreementsMember" xlink:label="loc_us-gaap_LicensingAgreementsMember_718126C659E41656538454A5BFE24CC0" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LossContingencyNatureDomain_88F1A22017447DC4AF5C54A5BFE29506" xlink:to="loc_us-gaap_LicensingAgreementsMember_718126C659E41656538454A5BFE24CC0" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentByTypeAxis" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentByTypeAxis_6C7362BC3C2D8ABCBFC754A5BFE2CF66" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_LossContingenciesTable_6DE10A04251958D0F82C54A5BFE20D3C" xlink:to="loc_us-gaap_PropertyPlantAndEquipmentByTypeAxis_6C7362BC3C2D8ABCBFC754A5BFE2CF66" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentTypeDomain" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_FD56D7189AD09AE2036B54A5BFE2B76F_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentByTypeAxis_6C7362BC3C2D8ABCBFC754A5BFE2CF66" xlink:to="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_FD56D7189AD09AE2036B54A5BFE2B76F_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentTypeDomain" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_FD56D7189AD09AE2036B54A5BFE2B76F" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentByTypeAxis_6C7362BC3C2D8ABCBFC754A5BFE2CF66" xlink:to="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_FD56D7189AD09AE2036B54A5BFE2B76F" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_BuildingMember" xlink:label="loc_us-gaap_BuildingMember_C0FAC47D621BEB6F82EB54A5BFE2BD84" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_FD56D7189AD09AE2036B54A5BFE2B76F" xlink:to="loc_us-gaap_BuildingMember_C0FAC47D621BEB6F82EB54A5BFE2BD84" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_AdditionalSpaceMember" xlink:label="loc_autl_AdditionalSpaceMember_A392E748B88BD1F49FB054A5BFE2CF2D" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_FD56D7189AD09AE2036B54A5BFE2B76F" xlink:to="loc_autl_AdditionalSpaceMember_A392E748B88BD1F49FB054A5BFE2CF2D" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_RealEstateAndAccumulatedDepreciationDescriptionOfPropertyAxis" xlink:label="loc_srt_RealEstateAndAccumulatedDepreciationDescriptionOfPropertyAxis_CC99A008C4154934E72054A5BFE26F0A" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_LossContingenciesTable_6DE10A04251958D0F82C54A5BFE20D3C" xlink:to="loc_srt_RealEstateAndAccumulatedDepreciationDescriptionOfPropertyAxis_CC99A008C4154934E72054A5BFE26F0A" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_RealEstateAndAccumulatedDepreciationNameOfPropertyDomain" xlink:label="loc_srt_RealEstateAndAccumulatedDepreciationNameOfPropertyDomain_DFE845C74592E55C225054A5BFE25BBA_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_srt_RealEstateAndAccumulatedDepreciationDescriptionOfPropertyAxis_CC99A008C4154934E72054A5BFE26F0A" xlink:to="loc_srt_RealEstateAndAccumulatedDepreciationNameOfPropertyDomain_DFE845C74592E55C225054A5BFE25BBA_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_RealEstateAndAccumulatedDepreciationNameOfPropertyDomain" xlink:label="loc_srt_RealEstateAndAccumulatedDepreciationNameOfPropertyDomain_DFE845C74592E55C225054A5BFE25BBA" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_srt_RealEstateAndAccumulatedDepreciationDescriptionOfPropertyAxis_CC99A008C4154934E72054A5BFE26F0A" xlink:to="loc_srt_RealEstateAndAccumulatedDepreciationNameOfPropertyDomain_DFE845C74592E55C225054A5BFE25BBA" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_ImperialForestHouseLimitedMember" xlink:label="loc_autl_ImperialForestHouseLimitedMember_FD8C77463BE3F133AC3654A5BFE22B41" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_srt_RealEstateAndAccumulatedDepreciationNameOfPropertyDomain_DFE845C74592E55C225054A5BFE25BBA" xlink:to="loc_autl_ImperialForestHouseLimitedMember_FD8C77463BE3F133AC3654A5BFE22B41" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_TherapyCatapultManufacturingCenterinStevenageUnitedKingdomMember" xlink:label="loc_autl_TherapyCatapultManufacturingCenterinStevenageUnitedKingdomMember_F98B23CDAADF823983F954A5BFE2DB63" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_srt_RealEstateAndAccumulatedDepreciationNameOfPropertyDomain_DFE845C74592E55C225054A5BFE25BBA" xlink:to="loc_autl_TherapyCatapultManufacturingCenterinStevenageUnitedKingdomMember_F98B23CDAADF823983F954A5BFE2DB63" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_LaboratorySpaceInWhiteCityLondonMember" xlink:label="loc_autl_LaboratorySpaceInWhiteCityLondonMember_AB6F54AE27176AB23EE654A5BFE22414" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_srt_RealEstateAndAccumulatedDepreciationNameOfPropertyDomain_DFE845C74592E55C225054A5BFE25BBA" xlink:to="loc_autl_LaboratorySpaceInWhiteCityLondonMember_AB6F54AE27176AB23EE654A5BFE22414" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_ManufacturingSpaceInEnfieldUnitedKingdomMember" xlink:label="loc_autl_ManufacturingSpaceInEnfieldUnitedKingdomMember_A2B84717F743F74E5E1C54A5BFE291D5" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_srt_RealEstateAndAccumulatedDepreciationNameOfPropertyDomain_DFE845C74592E55C225054A5BFE25BBA" xlink:to="loc_autl_ManufacturingSpaceInEnfieldUnitedKingdomMember_A2B84717F743F74E5E1C54A5BFE291D5" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_OfficeSpaceInRockvilleMarylandMember" xlink:label="loc_autl_OfficeSpaceInRockvilleMarylandMember_2914BDF8E7A42AAA3C3B54A5BFE2E9C3" xlink:type="locator" />
    <link:definitionArc order="5" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_srt_RealEstateAndAccumulatedDepreciationNameOfPropertyDomain_DFE845C74592E55C225054A5BFE25BBA" xlink:to="loc_autl_OfficeSpaceInRockvilleMarylandMember_2914BDF8E7A42AAA3C3B54A5BFE2E9C3" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_OfficeAndManufacturingSpaceinRockvilleMarylandMember" xlink:label="loc_autl_OfficeAndManufacturingSpaceinRockvilleMarylandMember_640679EF8B364031430A54A5BFE28DB7" xlink:type="locator" />
    <link:definitionArc order="6" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_srt_RealEstateAndAccumulatedDepreciationNameOfPropertyDomain_DFE845C74592E55C225054A5BFE25BBA" xlink:to="loc_autl_OfficeAndManufacturingSpaceinRockvilleMarylandMember_640679EF8B364031430A54A5BFE28DB7" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LossContingencyAccrualAtCarryingValue" xlink:label="loc_us-gaap_LossContingencyAccrualAtCarryingValue_DDE0EC445452EE3CF47854A5BFE2A2AB" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LossContingenciesLineItems_C43A68C25CA86BA7AC8A54A5BFE2E984" xlink:to="loc_us-gaap_LossContingencyAccrualAtCarryingValue_DDE0EC445452EE3CF47854A5BFE2A2AB" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LesseeOperatingLeaseTermOfContract" xlink:label="loc_us-gaap_LesseeOperatingLeaseTermOfContract_78642069881721A5DD1854A5BFE2FD43" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LossContingenciesLineItems_C43A68C25CA86BA7AC8A54A5BFE2E984" xlink:to="loc_us-gaap_LesseeOperatingLeaseTermOfContract_78642069881721A5DD1854A5BFE2FD43" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_OperatingLeaseRentFreePeriod" xlink:label="loc_autl_OperatingLeaseRentFreePeriod_0009DCD19CAFFE38BAC754A5BFE29452" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LossContingenciesLineItems_C43A68C25CA86BA7AC8A54A5BFE2E984" xlink:to="loc_autl_OperatingLeaseRentFreePeriod_0009DCD19CAFFE38BAC754A5BFE29452" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LeaseholdImprovementsGross" xlink:label="loc_us-gaap_LeaseholdImprovementsGross_A2D188D990145635904D54A5BFE2FF36" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LossContingenciesLineItems_C43A68C25CA86BA7AC8A54A5BFE2E984" xlink:to="loc_us-gaap_LeaseholdImprovementsGross_A2D188D990145635904D54A5BFE2FF36" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ConstructionInProgressGross" xlink:label="loc_us-gaap_ConstructionInProgressGross_A08D4FD3E10293B2F4EA54A5BFE2C125" xlink:type="locator" />
    <link:definitionArc order="5" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LossContingenciesLineItems_C43A68C25CA86BA7AC8A54A5BFE2E984" xlink:to="loc_us-gaap_ConstructionInProgressGross_A08D4FD3E10293B2F4EA54A5BFE2C125" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_NumberOfUnitsInRealEstateProperty" xlink:label="loc_us-gaap_NumberOfUnitsInRealEstateProperty_43DA8AB14A163C2E14B154A5BFE202BE" xlink:type="locator" />
    <link:definitionArc order="6" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LossContingenciesLineItems_C43A68C25CA86BA7AC8A54A5BFE2E984" xlink:to="loc_us-gaap_NumberOfUnitsInRealEstateProperty_43DA8AB14A163C2E14B154A5BFE202BE" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_LesseeOperatingLeaseLeaseNotYetCommencedUndiscountedAmount" xlink:label="loc_autl_LesseeOperatingLeaseLeaseNotYetCommencedUndiscountedAmount_AA5CE9C9DC4F08B7F21354A5BFE2E7CF" xlink:type="locator" />
    <link:definitionArc order="7" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LossContingenciesLineItems_C43A68C25CA86BA7AC8A54A5BFE2E984" xlink:to="loc_autl_LesseeOperatingLeaseLeaseNotYetCommencedUndiscountedAmount_AA5CE9C9DC4F08B7F21354A5BFE2E7CF" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LeaseAndRentalExpense" xlink:label="loc_us-gaap_LeaseAndRentalExpense_3F245326D3BAC093084754A5BFE2657A" xlink:type="locator" />
    <link:definitionArc order="8" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LossContingenciesLineItems_C43A68C25CA86BA7AC8A54A5BFE2E984" xlink:to="loc_us-gaap_LeaseAndRentalExpense_3F245326D3BAC093084754A5BFE2657A" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/CommitmentsAndContingenciesOtherInformationDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/CommitmentsAndContingenciesTables" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/CondensedConsolidatedBalanceSheetsUnaudited" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementLineItems" xlink:label="loc_us-gaap_StatementLineItems_50CDC3667EADAD77DA6343AEDCD314C7" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementTable" xlink:label="loc_us-gaap_StatementTable_DFC9E4DFC3E99614B0E243AEDCD1F1C0" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_StatementLineItems_50CDC3667EADAD77DA6343AEDCD314C7" xlink:to="loc_us-gaap_StatementTable_DFC9E4DFC3E99614B0E243AEDCD1F1C0" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementClassOfStockAxis" xlink:label="loc_us-gaap_StatementClassOfStockAxis_C72D6F804FD0C74669A643AEDCD1C13A" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_StatementTable_DFC9E4DFC3E99614B0E243AEDCD1F1C0" xlink:to="loc_us-gaap_StatementClassOfStockAxis_C72D6F804FD0C74669A643AEDCD1C13A" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_8246678EF863F29CCDB543AEDCD14939_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_StatementClassOfStockAxis_C72D6F804FD0C74669A643AEDCD1C13A" xlink:to="loc_us-gaap_ClassOfStockDomain_8246678EF863F29CCDB543AEDCD14939_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_8246678EF863F29CCDB543AEDCD14939" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_StatementClassOfStockAxis_C72D6F804FD0C74669A643AEDCD1C13A" xlink:to="loc_us-gaap_ClassOfStockDomain_8246678EF863F29CCDB543AEDCD14939" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_OrdinarySharesMember" xlink:label="loc_autl_OrdinarySharesMember_87E761371E33E9426D4543AEDCD2823F" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_8246678EF863F29CCDB543AEDCD14939" xlink:to="loc_autl_OrdinarySharesMember_87E761371E33E9426D4543AEDCD2823F" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassASharesMember" xlink:label="loc_autl_DeferredClassASharesMember_5B9F8DEE3283349D35A443AEDCD215EF" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_8246678EF863F29CCDB543AEDCD14939" xlink:to="loc_autl_DeferredClassASharesMember_5B9F8DEE3283349D35A443AEDCD215EF" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassBSharesMember" xlink:label="loc_autl_DeferredClassBSharesMember_68FF82104D04E0F26ADD43AEDCD238D0" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_8246678EF863F29CCDB543AEDCD14939" xlink:to="loc_autl_DeferredClassBSharesMember_68FF82104D04E0F26ADD43AEDCD238D0" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassCSharesMember" xlink:label="loc_autl_DeferredClassCSharesMember_4414A4FD78D276C1F4B643AEDCD385F2" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_8246678EF863F29CCDB543AEDCD14939" xlink:to="loc_autl_DeferredClassCSharesMember_4414A4FD78D276C1F4B643AEDCD385F2" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AssetsAbstract" xlink:label="loc_us-gaap_AssetsAbstract_A515F0DD1AA0C9BCC25E43AEDCD3BE1B" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_StatementLineItems_50CDC3667EADAD77DA6343AEDCD314C7" xlink:to="loc_us-gaap_AssetsAbstract_A515F0DD1AA0C9BCC25E43AEDCD3BE1B" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AssetsCurrentAbstract" xlink:label="loc_us-gaap_AssetsCurrentAbstract_2033AE73ABE1EBE93C1643AEDCD3269B" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_AssetsAbstract_A515F0DD1AA0C9BCC25E43AEDCD3BE1B" xlink:to="loc_us-gaap_AssetsCurrentAbstract_2033AE73ABE1EBE93C1643AEDCD3269B" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_Cash" xlink:label="loc_us-gaap_Cash_F08A832FDEE0C650F55943AEDCD9936C" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_AssetsCurrentAbstract_2033AE73ABE1EBE93C1643AEDCD3269B" xlink:to="loc_us-gaap_Cash_F08A832FDEE0C650F55943AEDCD9936C" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RestrictedCashCurrent" xlink:label="loc_us-gaap_RestrictedCashCurrent_B4DF7ED4FEB5E1D1032C43AEDCD91D3F" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_AssetsCurrentAbstract_2033AE73ABE1EBE93C1643AEDCD3269B" xlink:to="loc_us-gaap_RestrictedCashCurrent_B4DF7ED4FEB5E1D1032C43AEDCD91D3F" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PrepaidExpenseAndOtherAssetsCurrent" xlink:label="loc_us-gaap_PrepaidExpenseAndOtherAssetsCurrent_D81147057BE22F1DE7FE43AEDCD939DB" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_AssetsCurrentAbstract_2033AE73ABE1EBE93C1643AEDCD3269B" xlink:to="loc_us-gaap_PrepaidExpenseAndOtherAssetsCurrent_D81147057BE22F1DE7FE43AEDCD939DB" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AssetsCurrent" xlink:label="loc_us-gaap_AssetsCurrent_BE592B6D96A88523F8EE43AEDCDA55FB" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_AssetsCurrentAbstract_2033AE73ABE1EBE93C1643AEDCD3269B" xlink:to="loc_us-gaap_AssetsCurrent_BE592B6D96A88523F8EE43AEDCDA55FB" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AssetsNoncurrentAbstract" xlink:label="loc_us-gaap_AssetsNoncurrentAbstract_7C875E7D3F3C105DF90C43AEDCDA6850" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_AssetsAbstract_A515F0DD1AA0C9BCC25E43AEDCD3BE1B" xlink:to="loc_us-gaap_AssetsNoncurrentAbstract_7C875E7D3F3C105DF90C43AEDCDA6850" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentNet" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentNet_A773537869F21910A32A43AEDCDADE15" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_AssetsNoncurrentAbstract_7C875E7D3F3C105DF90C43AEDCDA6850" xlink:to="loc_us-gaap_PropertyPlantAndEquipmentNet_A773537869F21910A32A43AEDCDADE15" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_OperatingLeaseRightOfUseAsset" xlink:label="loc_us-gaap_OperatingLeaseRightOfUseAsset_6C0503A02CAEA6E2D62743AEDCDBF877" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_AssetsNoncurrentAbstract_7C875E7D3F3C105DF90C43AEDCDA6850" xlink:to="loc_us-gaap_OperatingLeaseRightOfUseAsset_6C0503A02CAEA6E2D62743AEDCDBF877" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_DepositsAssetsNoncurrent" xlink:label="loc_us-gaap_DepositsAssetsNoncurrent_111D8831F2A62CE7C77043AEDCDB4B93" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_AssetsNoncurrentAbstract_7C875E7D3F3C105DF90C43AEDCDA6850" xlink:to="loc_us-gaap_DepositsAssetsNoncurrent_111D8831F2A62CE7C77043AEDCDB4B93" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_Assets" xlink:label="loc_us-gaap_Assets_EE7C6F2526C2326988F343AEDCDB8E1B" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_AssetsAbstract_A515F0DD1AA0C9BCC25E43AEDCD3BE1B" xlink:to="loc_us-gaap_Assets_EE7C6F2526C2326988F343AEDCDB8E1B" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LiabilitiesAndStockholdersEquityAbstract" xlink:label="loc_us-gaap_LiabilitiesAndStockholdersEquityAbstract_2F8DE51281C804E8F4E543AEDCDBCBC3" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_StatementLineItems_50CDC3667EADAD77DA6343AEDCD314C7" xlink:to="loc_us-gaap_LiabilitiesAndStockholdersEquityAbstract_2F8DE51281C804E8F4E543AEDCDBCBC3" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LiabilitiesCurrentAbstract" xlink:label="loc_us-gaap_LiabilitiesCurrentAbstract_51177DEA16D9D67000E043AEDCDBACDD" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LiabilitiesAndStockholdersEquityAbstract_2F8DE51281C804E8F4E543AEDCDBCBC3" xlink:to="loc_us-gaap_LiabilitiesCurrentAbstract_51177DEA16D9D67000E043AEDCDBACDD" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AccountsPayableCurrent" xlink:label="loc_us-gaap_AccountsPayableCurrent_81345F9E2DB44254A5DD43AEDCDC2499" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LiabilitiesCurrentAbstract_51177DEA16D9D67000E043AEDCDBACDD" xlink:to="loc_us-gaap_AccountsPayableCurrent_81345F9E2DB44254A5DD43AEDCDC2499" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AccruedLiabilitiesCurrent" xlink:label="loc_us-gaap_AccruedLiabilitiesCurrent_5D3020A49DD4BA6B9B3943AEDCDC2DC3" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LiabilitiesCurrentAbstract_51177DEA16D9D67000E043AEDCDBACDD" xlink:to="loc_us-gaap_AccruedLiabilitiesCurrent_5D3020A49DD4BA6B9B3943AEDCDC2DC3" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_OperatingLeaseLiabilityCurrent" xlink:label="loc_us-gaap_OperatingLeaseLiabilityCurrent_44A2945DEE2BE33AA1FD43AEDCDCC03F" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LiabilitiesCurrentAbstract_51177DEA16D9D67000E043AEDCDBACDD" xlink:to="loc_us-gaap_OperatingLeaseLiabilityCurrent_44A2945DEE2BE33AA1FD43AEDCDCC03F" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LiabilitiesCurrent" xlink:label="loc_us-gaap_LiabilitiesCurrent_F09859F541BED62A9DF943AEDCDD8183" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LiabilitiesCurrentAbstract_51177DEA16D9D67000E043AEDCDBACDD" xlink:to="loc_us-gaap_LiabilitiesCurrent_F09859F541BED62A9DF943AEDCDD8183" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LiabilitiesNoncurrentAbstract" xlink:label="loc_us-gaap_LiabilitiesNoncurrentAbstract_7C4378B428E80258BCEE43AEDCDD2B06" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LiabilitiesAndStockholdersEquityAbstract_2F8DE51281C804E8F4E543AEDCDBCBC3" xlink:to="loc_us-gaap_LiabilitiesNoncurrentAbstract_7C4378B428E80258BCEE43AEDCDD2B06" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_OperatingLeaseLiabilityNoncurrent" xlink:label="loc_us-gaap_OperatingLeaseLiabilityNoncurrent_5E8DFDB40913FBA705E543AEDCDD1FCE" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LiabilitiesNoncurrentAbstract_7C4378B428E80258BCEE43AEDCDD2B06" xlink:to="loc_us-gaap_OperatingLeaseLiabilityNoncurrent_5E8DFDB40913FBA705E543AEDCDD1FCE" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_IncentiveFromLessor" xlink:label="loc_us-gaap_IncentiveFromLessor_55FA9C2DE4E47B68C26D43AEDCDD0FCC" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LiabilitiesNoncurrentAbstract_7C4378B428E80258BCEE43AEDCDD2B06" xlink:to="loc_us-gaap_IncentiveFromLessor_55FA9C2DE4E47B68C26D43AEDCDD0FCC" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LongTermDebtNoncurrent" xlink:label="loc_us-gaap_LongTermDebtNoncurrent_AB067EA8E692B25F934843AEDCDDC275" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LiabilitiesNoncurrentAbstract_7C4378B428E80258BCEE43AEDCDD2B06" xlink:to="loc_us-gaap_LongTermDebtNoncurrent_AB067EA8E692B25F934843AEDCDDC275" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_Liabilities" xlink:label="loc_us-gaap_Liabilities_3AA75C6535D8BC1A53D143AEDCDE3E7E" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LiabilitiesAndStockholdersEquityAbstract_2F8DE51281C804E8F4E543AEDCDBCBC3" xlink:to="loc_us-gaap_Liabilities_3AA75C6535D8BC1A53D143AEDCDE3E7E" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StockholdersEquityAbstract" xlink:label="loc_us-gaap_StockholdersEquityAbstract_EA1EAF6EEAD3C4F44E8543AEDCDEB277" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LiabilitiesAndStockholdersEquityAbstract_2F8DE51281C804E8F4E543AEDCDBCBC3" xlink:to="loc_us-gaap_StockholdersEquityAbstract_EA1EAF6EEAD3C4F44E8543AEDCDEB277" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_CommonStockValue" xlink:label="loc_us-gaap_CommonStockValue_CE1CAA9058242569D2FE43AEDCDE7795" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_StockholdersEquityAbstract_EA1EAF6EEAD3C4F44E8543AEDCDEB277" xlink:to="loc_us-gaap_CommonStockValue_CE1CAA9058242569D2FE43AEDCDE7795" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AdditionalPaidInCapital" xlink:label="loc_us-gaap_AdditionalPaidInCapital_78207289E84E70F06AE343AEDCDE6EB7" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_StockholdersEquityAbstract_EA1EAF6EEAD3C4F44E8543AEDCDEB277" xlink:to="loc_us-gaap_AdditionalPaidInCapital_78207289E84E70F06AE343AEDCDE6EB7" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AccumulatedOtherComprehensiveIncomeLossNetOfTax" xlink:label="loc_us-gaap_AccumulatedOtherComprehensiveIncomeLossNetOfTax_EB6B6EC7242E658FC24143AEDCDED142" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_StockholdersEquityAbstract_EA1EAF6EEAD3C4F44E8543AEDCDEB277" xlink:to="loc_us-gaap_AccumulatedOtherComprehensiveIncomeLossNetOfTax_EB6B6EC7242E658FC24143AEDCDED142" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RetainedEarningsAccumulatedDeficit" xlink:label="loc_us-gaap_RetainedEarningsAccumulatedDeficit_A1F9DAF339829A96098243AEDCDF7E77" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_StockholdersEquityAbstract_EA1EAF6EEAD3C4F44E8543AEDCDEB277" xlink:to="loc_us-gaap_RetainedEarningsAccumulatedDeficit_A1F9DAF339829A96098243AEDCDF7E77" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StockholdersEquity" xlink:label="loc_us-gaap_StockholdersEquity_EFEB7F1699DEEE2A90E243AEDCDF287D" xlink:type="locator" />
    <link:definitionArc order="5" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_StockholdersEquityAbstract_EA1EAF6EEAD3C4F44E8543AEDCDEB277" xlink:to="loc_us-gaap_StockholdersEquity_EFEB7F1699DEEE2A90E243AEDCDF287D" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LiabilitiesAndStockholdersEquity" xlink:label="loc_us-gaap_LiabilitiesAndStockholdersEquity_F0D0F15DCCD0AFD90FB343AEDCDFEE2C" xlink:type="locator" />
    <link:definitionArc order="5" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_LiabilitiesAndStockholdersEquityAbstract_2F8DE51281C804E8F4E543AEDCDBCBC3" xlink:to="loc_us-gaap_LiabilitiesAndStockholdersEquity_F0D0F15DCCD0AFD90FB343AEDCDFEE2C" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/CondensedConsolidatedBalanceSheetsUnauditedParentheticals" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementLineItems" xlink:label="loc_us-gaap_StatementLineItems_0AC8FDA0E2DB85AC28DD43AEDCB8FD65" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementTable" xlink:label="loc_us-gaap_StatementTable_212E25AE4B902F01816043AEDCB69565" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_StatementLineItems_0AC8FDA0E2DB85AC28DD43AEDCB8FD65" xlink:to="loc_us-gaap_StatementTable_212E25AE4B902F01816043AEDCB69565" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementClassOfStockAxis" xlink:label="loc_us-gaap_StatementClassOfStockAxis_3EA0693513235A26763343AEDCB6667E" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_StatementTable_212E25AE4B902F01816043AEDCB69565" xlink:to="loc_us-gaap_StatementClassOfStockAxis_3EA0693513235A26763343AEDCB6667E" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_BC49ACE255DFDE41980443AEDCB7918D_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_StatementClassOfStockAxis_3EA0693513235A26763343AEDCB6667E" xlink:to="loc_us-gaap_ClassOfStockDomain_BC49ACE255DFDE41980443AEDCB7918D_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_BC49ACE255DFDE41980443AEDCB7918D" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_StatementClassOfStockAxis_3EA0693513235A26763343AEDCB6667E" xlink:to="loc_us-gaap_ClassOfStockDomain_BC49ACE255DFDE41980443AEDCB7918D" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_OrdinarySharesMember" xlink:label="loc_autl_OrdinarySharesMember_D2AF974467E7FA604BF543AEDCB76CD2" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_BC49ACE255DFDE41980443AEDCB7918D" xlink:to="loc_autl_OrdinarySharesMember_D2AF974467E7FA604BF543AEDCB76CD2" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassASharesMember" xlink:label="loc_autl_DeferredClassASharesMember_0DB379C5BE43B3A2D49343AEDCB7E665" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_BC49ACE255DFDE41980443AEDCB7918D" xlink:to="loc_autl_DeferredClassASharesMember_0DB379C5BE43B3A2D49343AEDCB7E665" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassBSharesMember" xlink:label="loc_autl_DeferredClassBSharesMember_31CFD0A0DE192F1D287943AEDCB788FB" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_BC49ACE255DFDE41980443AEDCB7918D" xlink:to="loc_autl_DeferredClassBSharesMember_31CFD0A0DE192F1D287943AEDCB788FB" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassCSharesMember" xlink:label="loc_autl_DeferredClassCSharesMember_650CB39F23C8BB63DA4243AEDCB83FE2" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_BC49ACE255DFDE41980443AEDCB7918D" xlink:to="loc_autl_DeferredClassCSharesMember_650CB39F23C8BB63DA4243AEDCB83FE2" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StockholdersEquityAbstract" xlink:label="loc_us-gaap_StockholdersEquityAbstract_E69823F4A567CE8374E443AEDCB84C49" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_StatementLineItems_0AC8FDA0E2DB85AC28DD43AEDCB8FD65" xlink:to="loc_us-gaap_StockholdersEquityAbstract_E69823F4A567CE8374E443AEDCB84C49" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_CommonStockParOrStatedValuePerShare" xlink:label="loc_us-gaap_CommonStockParOrStatedValuePerShare_72B05DA97B2164A9EA7243AEDCB8F284" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_StockholdersEquityAbstract_E69823F4A567CE8374E443AEDCB84C49" xlink:to="loc_us-gaap_CommonStockParOrStatedValuePerShare_72B05DA97B2164A9EA7243AEDCB8F284" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_CommonStockSharesAuthorized" xlink:label="loc_us-gaap_CommonStockSharesAuthorized_7AA4117DD04F9072F36D43AEDCB90B4B" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_StockholdersEquityAbstract_E69823F4A567CE8374E443AEDCB84C49" xlink:to="loc_us-gaap_CommonStockSharesAuthorized_7AA4117DD04F9072F36D43AEDCB90B4B" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_CommonStockSharesIssued" xlink:label="loc_us-gaap_CommonStockSharesIssued_F71444F1C5405319320F43AEDCB98B6B" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_StockholdersEquityAbstract_E69823F4A567CE8374E443AEDCB84C49" xlink:to="loc_us-gaap_CommonStockSharesIssued_F71444F1C5405319320F43AEDCB98B6B" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_CommonStockSharesOutstanding" xlink:label="loc_us-gaap_CommonStockSharesOutstanding_6A6200A72361B27E7A9343AEDCB9AD1C" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_StockholdersEquityAbstract_E69823F4A567CE8374E443AEDCB84C49" xlink:to="loc_us-gaap_CommonStockSharesOutstanding_6A6200A72361B27E7A9343AEDCB9AD1C" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/CondensedConsolidatedStatementsOfCashFlowsUnaudited" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/CondensedConsolidatedStatementsOfOperationsAndComprehensiveLossUnaudited" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/CondensedConsolidatedStatementsOfShareholdersEquityUnaudited" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementLineItems" xlink:label="loc_us-gaap_StatementLineItems_D5DCCDCA80D3D6A11DAE6C7725DD20C6" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementTable" xlink:label="loc_us-gaap_StatementTable_6A4D6DF89056D053673E6C7725D3E209" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_StatementLineItems_D5DCCDCA80D3D6A11DAE6C7725DD20C6" xlink:to="loc_us-gaap_StatementTable_6A4D6DF89056D053673E6C7725D3E209" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementEquityComponentsAxis" xlink:label="loc_us-gaap_StatementEquityComponentsAxis_CA2B3152300EDD4BDA7F6C7725D41B48" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_StatementTable_6A4D6DF89056D053673E6C7725D3E209" xlink:to="loc_us-gaap_StatementEquityComponentsAxis_CA2B3152300EDD4BDA7F6C7725D41B48" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_EquityComponentDomain" xlink:label="loc_us-gaap_EquityComponentDomain_9A88B2C801C0605610006C7725D42EBB_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_StatementEquityComponentsAxis_CA2B3152300EDD4BDA7F6C7725D41B48" xlink:to="loc_us-gaap_EquityComponentDomain_9A88B2C801C0605610006C7725D42EBB_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_EquityComponentDomain" xlink:label="loc_us-gaap_EquityComponentDomain_9A88B2C801C0605610006C7725D42EBB" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_StatementEquityComponentsAxis_CA2B3152300EDD4BDA7F6C7725D41B48" xlink:to="loc_us-gaap_EquityComponentDomain_9A88B2C801C0605610006C7725D42EBB" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_CommonStockMember" xlink:label="loc_us-gaap_CommonStockMember_EF9262C61699F8E57E936C7725D49BF7" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_EquityComponentDomain_9A88B2C801C0605610006C7725D42EBB" xlink:to="loc_us-gaap_CommonStockMember_EF9262C61699F8E57E936C7725D49BF7" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AdditionalPaidInCapitalMember" xlink:label="loc_us-gaap_AdditionalPaidInCapitalMember_A4AA334C8CE3B21EEF0B6C7725D5631B" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_EquityComponentDomain_9A88B2C801C0605610006C7725D42EBB" xlink:to="loc_us-gaap_AdditionalPaidInCapitalMember_A4AA334C8CE3B21EEF0B6C7725D5631B" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AccumulatedOtherComprehensiveIncomeMember" xlink:label="loc_us-gaap_AccumulatedOtherComprehensiveIncomeMember_53990A1C5BB7A02E89326C7725D7B0ED" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_EquityComponentDomain_9A88B2C801C0605610006C7725D42EBB" xlink:to="loc_us-gaap_AccumulatedOtherComprehensiveIncomeMember_53990A1C5BB7A02E89326C7725D7B0ED" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RetainedEarningsMember" xlink:label="loc_us-gaap_RetainedEarningsMember_6911B5246EE1D07C84556C7725DC2E38" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_EquityComponentDomain_9A88B2C801C0605610006C7725D42EBB" xlink:to="loc_us-gaap_RetainedEarningsMember_6911B5246EE1D07C84556C7725DC2E38" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementClassOfStockAxis" xlink:label="loc_us-gaap_StatementClassOfStockAxis_1428307644F21AFA48AD6C7725DC6BF1" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_StatementTable_6A4D6DF89056D053673E6C7725D3E209" xlink:to="loc_us-gaap_StatementClassOfStockAxis_1428307644F21AFA48AD6C7725DC6BF1" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_5708530C24C48EFAEFFB6C7725DC1C2D_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_StatementClassOfStockAxis_1428307644F21AFA48AD6C7725DC6BF1" xlink:to="loc_us-gaap_ClassOfStockDomain_5708530C24C48EFAEFFB6C7725DC1C2D_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_5708530C24C48EFAEFFB6C7725DC1C2D" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_StatementClassOfStockAxis_1428307644F21AFA48AD6C7725DC6BF1" xlink:to="loc_us-gaap_ClassOfStockDomain_5708530C24C48EFAEFFB6C7725DC1C2D" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_OrdinarySharesMember" xlink:label="loc_autl_OrdinarySharesMember_FDCEBAA14EEE9A92B1DE6C7725DD4A34" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_5708530C24C48EFAEFFB6C7725DC1C2D" xlink:to="loc_autl_OrdinarySharesMember_FDCEBAA14EEE9A92B1DE6C7725DD4A34" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassASharesMember" xlink:label="loc_autl_DeferredClassASharesMember_F47BFC08FFFD64C316D16C7725DDCC80" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_5708530C24C48EFAEFFB6C7725DC1C2D" xlink:to="loc_autl_DeferredClassASharesMember_F47BFC08FFFD64C316D16C7725DDCC80" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassBSharesMember" xlink:label="loc_autl_DeferredClassBSharesMember_A9E4EB5D52151AEDFA976C7725DD6C4D" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_5708530C24C48EFAEFFB6C7725DC1C2D" xlink:to="loc_autl_DeferredClassBSharesMember_A9E4EB5D52151AEDFA976C7725DD6C4D" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassCSharesMember" xlink:label="loc_autl_DeferredClassCSharesMember_8F57BC2D70A0E63EE39E6C7725DD2F27" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_5708530C24C48EFAEFFB6C7725DC1C2D" xlink:to="loc_autl_DeferredClassCSharesMember_8F57BC2D70A0E63EE39E6C7725DD2F27" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_IncreaseDecreaseInStockholdersEquityRollForward" xlink:label="loc_us-gaap_IncreaseDecreaseInStockholdersEquityRollForward_2CC715ACF4D82676D2B46C7725DEC99E" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_StatementLineItems_D5DCCDCA80D3D6A11DAE6C7725DD20C6" xlink:to="loc_us-gaap_IncreaseDecreaseInStockholdersEquityRollForward_2CC715ACF4D82676D2B46C7725DEC99E" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StockholdersEquity" xlink:label="loc_us-gaap_StockholdersEquity_4513A4947180F85ACB7D6C7725DE228B" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncreaseDecreaseInStockholdersEquityRollForward_2CC715ACF4D82676D2B46C7725DEC99E" xlink:to="loc_us-gaap_StockholdersEquity_4513A4947180F85ACB7D6C7725DE228B" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SharesOutstanding" xlink:label="loc_us-gaap_SharesOutstanding_80CCE41DAD052B0DDF6F6C7725DEB94A" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncreaseDecreaseInStockholdersEquityRollForward_2CC715ACF4D82676D2B46C7725DEC99E" xlink:to="loc_us-gaap_SharesOutstanding_80CCE41DAD052B0DDF6F6C7725DEB94A" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AdjustmentsToAdditionalPaidInCapitalSharebasedCompensationRequisiteServicePeriodRecognitionValue" xlink:label="loc_us-gaap_AdjustmentsToAdditionalPaidInCapitalSharebasedCompensationRequisiteServicePeriodRecognitionValue_9305318826B50A75D4AA6C7725DE5B4C" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncreaseDecreaseInStockholdersEquityRollForward_2CC715ACF4D82676D2B46C7725DEC99E" xlink:to="loc_us-gaap_AdjustmentsToAdditionalPaidInCapitalSharebasedCompensationRequisiteServicePeriodRecognitionValue_9305318826B50A75D4AA6C7725DE5B4C" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StockIssuedDuringPeriodSharesRestrictedStockAwardForfeited" xlink:label="loc_us-gaap_StockIssuedDuringPeriodSharesRestrictedStockAwardForfeited_C4AE0119B3437535D8046C7725DE2ADC" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncreaseDecreaseInStockholdersEquityRollForward_2CC715ACF4D82676D2B46C7725DEC99E" xlink:to="loc_us-gaap_StockIssuedDuringPeriodSharesRestrictedStockAwardForfeited_C4AE0119B3437535D8046C7725DE2ADC" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StockIssuedDuringPeriodValueStockOptionsExercised" xlink:label="loc_us-gaap_StockIssuedDuringPeriodValueStockOptionsExercised_E44804AC3F6B9667F8D86C7725DFB5C5" xlink:type="locator" />
    <link:definitionArc order="5" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncreaseDecreaseInStockholdersEquityRollForward_2CC715ACF4D82676D2B46C7725DEC99E" xlink:to="loc_us-gaap_StockIssuedDuringPeriodValueStockOptionsExercised_E44804AC3F6B9667F8D86C7725DFB5C5" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StockIssuedDuringPeriodSharesStockOptionsExercised" xlink:label="loc_us-gaap_StockIssuedDuringPeriodSharesStockOptionsExercised_AF1F7162850CFE68B5B46C7725DF8D35" xlink:type="locator" />
    <link:definitionArc order="6" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncreaseDecreaseInStockholdersEquityRollForward_2CC715ACF4D82676D2B46C7725DEC99E" xlink:to="loc_us-gaap_StockIssuedDuringPeriodSharesStockOptionsExercised_AF1F7162850CFE68B5B46C7725DF8D35" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StockIssuedDuringPeriodValueNewIssues" xlink:label="loc_us-gaap_StockIssuedDuringPeriodValueNewIssues_75E576C7CCE3670580766C7725DF9A2E" xlink:type="locator" />
    <link:definitionArc order="7" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncreaseDecreaseInStockholdersEquityRollForward_2CC715ACF4D82676D2B46C7725DEC99E" xlink:to="loc_us-gaap_StockIssuedDuringPeriodValueNewIssues_75E576C7CCE3670580766C7725DF9A2E" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StockIssuedDuringPeriodSharesNewIssues" xlink:label="loc_us-gaap_StockIssuedDuringPeriodSharesNewIssues_9FF75366BCED477129CF6C7725DF36F1" xlink:type="locator" />
    <link:definitionArc order="8" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncreaseDecreaseInStockholdersEquityRollForward_2CC715ACF4D82676D2B46C7725DEC99E" xlink:to="loc_us-gaap_StockIssuedDuringPeriodSharesNewIssues_9FF75366BCED477129CF6C7725DF36F1" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_OtherComprehensiveIncomeForeignCurrencyTransactionAndTranslationAdjustmentNetOfTaxPortionAttributableToParent" xlink:label="loc_us-gaap_OtherComprehensiveIncomeForeignCurrencyTransactionAndTranslationAdjustmentNetOfTaxPortionAttributableToParent_BF779F3B728EFAE128216C7725DF6BD2" xlink:type="locator" />
    <link:definitionArc order="9" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncreaseDecreaseInStockholdersEquityRollForward_2CC715ACF4D82676D2B46C7725DEC99E" xlink:to="loc_us-gaap_OtherComprehensiveIncomeForeignCurrencyTransactionAndTranslationAdjustmentNetOfTaxPortionAttributableToParent_BF779F3B728EFAE128216C7725DF6BD2" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_NetIncomeLoss" xlink:label="loc_us-gaap_NetIncomeLoss_C9526F593B5E4B5E05796C7725DF3627" xlink:type="locator" />
    <link:definitionArc order="10" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncreaseDecreaseInStockholdersEquityRollForward_2CC715ACF4D82676D2B46C7725DEC99E" xlink:to="loc_us-gaap_NetIncomeLoss_C9526F593B5E4B5E05796C7725DF3627" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/DocumentAndEntityInformation" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/EmployeeBenefitPlans" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/EmployeeBenefitPlansDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/IncomeTaxes" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/IncomeTaxesDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/LicenseAgreements" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/LicenseAgreementsDetails" xlink:type="extended">
    <link:loc xlink:href="autl-20190630.xsd#autl_ScheduleOfLicenseAgreementsLineItems" xlink:label="loc_autl_ScheduleOfLicenseAgreementsLineItems_D2888A1A78CCE9E317C043AEDB6DA86E" xlink:type="locator" />
    <link:loc xlink:href="autl-20190630.xsd#autl_ScheduleOfLicenseAgreementsTable" xlink:label="loc_autl_ScheduleOfLicenseAgreementsTable_DFFCB35C8FBA5975693C43AEDB6A0DD1" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_autl_ScheduleOfLicenseAgreementsLineItems_D2888A1A78CCE9E317C043AEDB6DA86E" xlink:to="loc_autl_ScheduleOfLicenseAgreementsTable_DFFCB35C8FBA5975693C43AEDB6A0DD1" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_CounterpartyNameAxis" xlink:label="loc_srt_CounterpartyNameAxis_83F857D5110E42C03E8043AEDB6ABC80" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_autl_ScheduleOfLicenseAgreementsTable_DFFCB35C8FBA5975693C43AEDB6A0DD1" xlink:to="loc_srt_CounterpartyNameAxis_83F857D5110E42C03E8043AEDB6ABC80" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_RepurchaseAgreementCounterpartyNameDomain" xlink:label="loc_srt_RepurchaseAgreementCounterpartyNameDomain_D5FB4DB91E9192ECAC4A43AEDB6A2955_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_srt_CounterpartyNameAxis_83F857D5110E42C03E8043AEDB6ABC80" xlink:to="loc_srt_RepurchaseAgreementCounterpartyNameDomain_D5FB4DB91E9192ECAC4A43AEDB6A2955_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_RepurchaseAgreementCounterpartyNameDomain" xlink:label="loc_srt_RepurchaseAgreementCounterpartyNameDomain_D5FB4DB91E9192ECAC4A43AEDB6A2955" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_srt_CounterpartyNameAxis_83F857D5110E42C03E8043AEDB6ABC80" xlink:to="loc_srt_RepurchaseAgreementCounterpartyNameDomain_D5FB4DB91E9192ECAC4A43AEDB6A2955" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_UCLBusinessplcMember" xlink:label="loc_autl_UCLBusinessplcMember_B3C73205CCC2B4CF5AAA43AEDB6ABFCD" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_srt_RepurchaseAgreementCounterpartyNameDomain_D5FB4DB91E9192ECAC4A43AEDB6A2955" xlink:to="loc_autl_UCLBusinessplcMember_B3C73205CCC2B4CF5AAA43AEDB6ABFCD" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_ConditionalPaymentsAxis" xlink:label="loc_autl_ConditionalPaymentsAxis_8420B09C1605FDF436E443AEDB6B42ED" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_autl_ScheduleOfLicenseAgreementsTable_DFFCB35C8FBA5975693C43AEDB6A0DD1" xlink:to="loc_autl_ConditionalPaymentsAxis_8420B09C1605FDF436E443AEDB6B42ED" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_ConditionalPaymentsDomain" xlink:label="loc_autl_ConditionalPaymentsDomain_365361C756BC9D3ABCB143AEDB6B89ED_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_autl_ConditionalPaymentsAxis_8420B09C1605FDF436E443AEDB6B42ED" xlink:to="loc_autl_ConditionalPaymentsDomain_365361C756BC9D3ABCB143AEDB6B89ED_default" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_ConditionalPaymentsDomain" xlink:label="loc_autl_ConditionalPaymentsDomain_365361C756BC9D3ABCB143AEDB6B89ED" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_autl_ConditionalPaymentsAxis_8420B09C1605FDF436E443AEDB6B42ED" xlink:to="loc_autl_ConditionalPaymentsDomain_365361C756BC9D3ABCB143AEDB6B89ED" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_RegulatoryApprovalMember" xlink:label="loc_autl_RegulatoryApprovalMember_84E2F386AE91941F323243AEDB6B573D" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_autl_ConditionalPaymentsDomain_365361C756BC9D3ABCB143AEDB6B89ED" xlink:to="loc_autl_RegulatoryApprovalMember_84E2F386AE91941F323243AEDB6B573D" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_ProductSalesMember" xlink:label="loc_autl_ProductSalesMember_ADAEEBCEDFAEAB16D9B643AEDB6C1288" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_autl_ConditionalPaymentsDomain_365361C756BC9D3ABCB143AEDB6B89ED" xlink:to="loc_autl_ProductSalesMember_ADAEEBCEDFAEAB16D9B643AEDB6C1288" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_NetSalesLevelsMember" xlink:label="loc_autl_NetSalesLevelsMember_D687332A479DBBED6E6F43AEDB6C1A3E" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_autl_ConditionalPaymentsDomain_365361C756BC9D3ABCB143AEDB6B89ED" xlink:to="loc_autl_NetSalesLevelsMember_D687332A479DBBED6E6F43AEDB6C1A3E" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_RangeAxis" xlink:label="loc_srt_RangeAxis_09C0BAABDB289CC6F48F43AEDB6C09AF" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_autl_ScheduleOfLicenseAgreementsTable_DFFCB35C8FBA5975693C43AEDB6A0DD1" xlink:to="loc_srt_RangeAxis_09C0BAABDB289CC6F48F43AEDB6C09AF" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_RangeMember" xlink:label="loc_srt_RangeMember_9FDDFA2334CCB826B55A43AEDB6C612D_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_srt_RangeAxis_09C0BAABDB289CC6F48F43AEDB6C09AF" xlink:to="loc_srt_RangeMember_9FDDFA2334CCB826B55A43AEDB6C612D_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_RangeMember" xlink:label="loc_srt_RangeMember_9FDDFA2334CCB826B55A43AEDB6C612D" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_srt_RangeAxis_09C0BAABDB289CC6F48F43AEDB6C09AF" xlink:to="loc_srt_RangeMember_9FDDFA2334CCB826B55A43AEDB6C612D" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_MinimumMember" xlink:label="loc_srt_MinimumMember_9FE7C3DD3FAE283A0EC343AEDB6C780F" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_srt_RangeMember_9FDDFA2334CCB826B55A43AEDB6C612D" xlink:to="loc_srt_MinimumMember_9FE7C3DD3FAE283A0EC343AEDB6C780F" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_MaximumMember" xlink:label="loc_srt_MaximumMember_CCCD7D021952D181DF6843AEDB6C0524" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_srt_RangeMember_9FDDFA2334CCB826B55A43AEDB6C612D" xlink:to="loc_srt_MaximumMember_CCCD7D021952D181DF6843AEDB6C0524" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StockIssuedDuringPeriodSharesNewIssues" xlink:label="loc_us-gaap_StockIssuedDuringPeriodSharesNewIssues_977E29393CDBEB98D27643AEDB6DF607" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_autl_ScheduleOfLicenseAgreementsLineItems_D2888A1A78CCE9E317C043AEDB6DA86E" xlink:to="loc_us-gaap_StockIssuedDuringPeriodSharesNewIssues_977E29393CDBEB98D27643AEDB6DF607" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_PaymentOfUpfrontFees" xlink:label="loc_autl_PaymentOfUpfrontFees_C074551217DAA48373B043AEDB6D5F34" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_autl_ScheduleOfLicenseAgreementsLineItems_D2888A1A78CCE9E317C043AEDB6DA86E" xlink:to="loc_autl_PaymentOfUpfrontFees_C074551217DAA48373B043AEDB6D5F34" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_AccruedLicenseFeesCurrent" xlink:label="loc_autl_AccruedLicenseFeesCurrent_81019077B333B02C3E4943AEDB6D77B7" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_autl_ScheduleOfLicenseAgreementsLineItems_D2888A1A78CCE9E317C043AEDB6DA86E" xlink:to="loc_autl_AccruedLicenseFeesCurrent_81019077B333B02C3E4943AEDB6D77B7" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_AnnualLicenseFeePayments" xlink:label="loc_autl_AnnualLicenseFeePayments_DFB0BD0C3330BEE9713543AEDB6D48D3" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_autl_ScheduleOfLicenseAgreementsLineItems_D2888A1A78CCE9E317C043AEDB6DA86E" xlink:to="loc_autl_AnnualLicenseFeePayments_DFB0BD0C3330BEE9713543AEDB6D48D3" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_ConditionalPaymentBasis" xlink:label="loc_autl_ConditionalPaymentBasis_730E84C0962A7BAC164B43AEDB6E48F9" xlink:type="locator" />
    <link:definitionArc order="5" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_autl_ScheduleOfLicenseAgreementsLineItems_D2888A1A78CCE9E317C043AEDB6DA86E" xlink:to="loc_autl_ConditionalPaymentBasis_730E84C0962A7BAC164B43AEDB6E48F9" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/NatureOfBusiness" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/NatureOfBusinessDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockLineItems" xlink:label="loc_us-gaap_ClassOfStockLineItems_9BB672C289B5560D7CC95042E2F4E262" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfStockByClassTable" xlink:label="loc_us-gaap_ScheduleOfStockByClassTable_F529E1512380F059A5515042E2F328AF" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_ClassOfStockLineItems_9BB672C289B5560D7CC95042E2F4E262" xlink:to="loc_us-gaap_ScheduleOfStockByClassTable_F529E1512380F059A5515042E2F328AF" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementClassOfStockAxis" xlink:label="loc_us-gaap_StatementClassOfStockAxis_503AF1207BAF8992F8BA5042E2F4C9C9" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfStockByClassTable_F529E1512380F059A5515042E2F328AF" xlink:to="loc_us-gaap_StatementClassOfStockAxis_503AF1207BAF8992F8BA5042E2F4C9C9" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_E7FEEF6AEA6608959A385042E2F49C82_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_StatementClassOfStockAxis_503AF1207BAF8992F8BA5042E2F4C9C9" xlink:to="loc_us-gaap_ClassOfStockDomain_E7FEEF6AEA6608959A385042E2F49C82_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_E7FEEF6AEA6608959A385042E2F49C82" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_StatementClassOfStockAxis_503AF1207BAF8992F8BA5042E2F4C9C9" xlink:to="loc_us-gaap_ClassOfStockDomain_E7FEEF6AEA6608959A385042E2F49C82" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_OrdinarySharesMember" xlink:label="loc_autl_OrdinarySharesMember_E85F9BFBB964C8EF2BE85042E2F47BA0" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_E7FEEF6AEA6608959A385042E2F49C82" xlink:to="loc_autl_OrdinarySharesMember_E85F9BFBB964C8EF2BE85042E2F47BA0" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SubsidiarySaleOfStockAxis" xlink:label="loc_us-gaap_SubsidiarySaleOfStockAxis_B27354BCFB346613795E5042E2F4E66F" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfStockByClassTable_F529E1512380F059A5515042E2F328AF" xlink:to="loc_us-gaap_SubsidiarySaleOfStockAxis_B27354BCFB346613795E5042E2F4E66F" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SaleOfStockNameOfTransactionDomain" xlink:label="loc_us-gaap_SaleOfStockNameOfTransactionDomain_F42540E4CBC324C54F805042E2F4A964_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_SubsidiarySaleOfStockAxis_B27354BCFB346613795E5042E2F4E66F" xlink:to="loc_us-gaap_SaleOfStockNameOfTransactionDomain_F42540E4CBC324C54F805042E2F4A964_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SaleOfStockNameOfTransactionDomain" xlink:label="loc_us-gaap_SaleOfStockNameOfTransactionDomain_F42540E4CBC324C54F805042E2F4A964" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_SubsidiarySaleOfStockAxis_B27354BCFB346613795E5042E2F4E66F" xlink:to="loc_us-gaap_SaleOfStockNameOfTransactionDomain_F42540E4CBC324C54F805042E2F4A964" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_IPOMember" xlink:label="loc_us-gaap_IPOMember_EFC7C07F354B45F3C1525042E2F4796A" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_SaleOfStockNameOfTransactionDomain_F42540E4CBC324C54F805042E2F4A964" xlink:to="loc_us-gaap_IPOMember_EFC7C07F354B45F3C1525042E2F4796A" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_OverAllotmentOptionMember" xlink:label="loc_us-gaap_OverAllotmentOptionMember_ED29F0A97BE2C106DC4F5042E2F4613E" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_SaleOfStockNameOfTransactionDomain_F42540E4CBC324C54F805042E2F4A964" xlink:to="loc_us-gaap_OverAllotmentOptionMember_ED29F0A97BE2C106DC4F5042E2F4613E" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_PublicStockOfferingMember" xlink:label="loc_autl_PublicStockOfferingMember_7E2613BE96E741DA76885042E2F4ADE6" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_SaleOfStockNameOfTransactionDomain_F42540E4CBC324C54F805042E2F4A964" xlink:to="loc_autl_PublicStockOfferingMember_7E2613BE96E741DA76885042E2F4ADE6" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SaleOfStockNumberOfSharesIssuedInTransaction" xlink:label="loc_us-gaap_SaleOfStockNumberOfSharesIssuedInTransaction_7AA3FA216B4826E423A75042E2F45D18" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_9BB672C289B5560D7CC95042E2F4E262" xlink:to="loc_us-gaap_SaleOfStockNumberOfSharesIssuedInTransaction_7AA3FA216B4826E423A75042E2F45D18" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SaleOfStockPricePerShare" xlink:label="loc_us-gaap_SaleOfStockPricePerShare_EB7FC91BC9A4F3A487D85042E2F41EAC" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_9BB672C289B5560D7CC95042E2F4E262" xlink:to="loc_us-gaap_SaleOfStockPricePerShare_EB7FC91BC9A4F3A487D85042E2F41EAC" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SaleOfStockConsiderationReceivedOnTransaction" xlink:label="loc_us-gaap_SaleOfStockConsiderationReceivedOnTransaction_75A68595FD7A4779D5325042E2F4462F" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_9BB672C289B5560D7CC95042E2F4E262" xlink:to="loc_us-gaap_SaleOfStockConsiderationReceivedOnTransaction_75A68595FD7A4779D5325042E2F4462F" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_NetIncomeLoss" xlink:label="loc_us-gaap_NetIncomeLoss_D65FCC2874D492045F025042E2F48AED" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_9BB672C289B5560D7CC95042E2F4E262" xlink:to="loc_us-gaap_NetIncomeLoss_D65FCC2874D492045F025042E2F48AED" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RetainedEarningsAccumulatedDeficit" xlink:label="loc_us-gaap_RetainedEarningsAccumulatedDeficit_B3FCCED7C72793553FC45042E2F4A395" xlink:type="locator" />
    <link:definitionArc order="5" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_9BB672C289B5560D7CC95042E2F4E262" xlink:to="loc_us-gaap_RetainedEarningsAccumulatedDeficit_B3FCCED7C72793553FC45042E2F4A395" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_Cash" xlink:label="loc_us-gaap_Cash_589A1D4266DE0C52479C5042E2F4A260" xlink:type="locator" />
    <link:definitionArc order="6" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_9BB672C289B5560D7CC95042E2F4E262" xlink:to="loc_us-gaap_Cash_589A1D4266DE0C52479C5042E2F4A260" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/PrepaidExpensesAndOtherCurrentAssets" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/PrepaidExpensesAndOtherCurrentAssetsDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/PrepaidExpensesAndOtherCurrentAssetsTables" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/PropertyAndEquipmentNet" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/PropertyAndEquipmentNetDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentLineItems" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentLineItems_B4E7761B4458CAAE3DB043AEDBDD577F" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfPropertyPlantAndEquipmentTable" xlink:label="loc_us-gaap_ScheduleOfPropertyPlantAndEquipmentTable_2DAC8CAB79BCBCC5ED5443AEDBDBE2C6" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentLineItems_B4E7761B4458CAAE3DB043AEDBDD577F" xlink:to="loc_us-gaap_ScheduleOfPropertyPlantAndEquipmentTable_2DAC8CAB79BCBCC5ED5443AEDBDBE2C6" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentByTypeAxis" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentByTypeAxis_CC57DB8E00D72E41F49743AEDBDBEB6D" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfPropertyPlantAndEquipmentTable_2DAC8CAB79BCBCC5ED5443AEDBDBE2C6" xlink:to="loc_us-gaap_PropertyPlantAndEquipmentByTypeAxis_CC57DB8E00D72E41F49743AEDBDBEB6D" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentTypeDomain" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_708D9BEFE96340F162EE43AEDBDBB321_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentByTypeAxis_CC57DB8E00D72E41F49743AEDBDBEB6D" xlink:to="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_708D9BEFE96340F162EE43AEDBDBB321_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentTypeDomain" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_708D9BEFE96340F162EE43AEDBDBB321" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentByTypeAxis_CC57DB8E00D72E41F49743AEDBDBEB6D" xlink:to="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_708D9BEFE96340F162EE43AEDBDBB321" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_EquipmentMember" xlink:label="loc_us-gaap_EquipmentMember_7062DF0E8792F689A40843AEDBDC5F87" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_708D9BEFE96340F162EE43AEDBDBB321" xlink:to="loc_us-gaap_EquipmentMember_7062DF0E8792F689A40843AEDBDC5F87" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_OfficeEquipmentMember" xlink:label="loc_us-gaap_OfficeEquipmentMember_B3C40F19FFB64C02A52943AEDBDC9245" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_708D9BEFE96340F162EE43AEDBDBB321" xlink:to="loc_us-gaap_OfficeEquipmentMember_B3C40F19FFB64C02A52943AEDBDC9245" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_FurnitureAndFixturesMember" xlink:label="loc_us-gaap_FurnitureAndFixturesMember_AB623FAE15E5D407491143AEDBDC2E90" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_708D9BEFE96340F162EE43AEDBDBB321" xlink:to="loc_us-gaap_FurnitureAndFixturesMember_AB623FAE15E5D407491143AEDBDC2E90" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_LeaseholdImprovementsMember" xlink:label="loc_us-gaap_LeaseholdImprovementsMember_495A9E25A0425C76CA7543AEDBDC71AE" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_708D9BEFE96340F162EE43AEDBDBB321" xlink:to="loc_us-gaap_LeaseholdImprovementsMember_495A9E25A0425C76CA7543AEDBDC71AE" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AssetUnderConstructionMember" xlink:label="loc_us-gaap_AssetUnderConstructionMember_976935A1C5FC7930467243AEDBDDFDBD" xlink:type="locator" />
    <link:definitionArc order="5" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_708D9BEFE96340F162EE43AEDBDBB321" xlink:to="loc_us-gaap_AssetUnderConstructionMember_976935A1C5FC7930467243AEDBDDFDBD" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentGross" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentGross_80904ED1968C3E2ADDB843AEDBDDC566" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentLineItems_B4E7761B4458CAAE3DB043AEDBDD577F" xlink:to="loc_us-gaap_PropertyPlantAndEquipmentGross_80904ED1968C3E2ADDB843AEDBDDC566" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AccumulatedDepreciationDepletionAndAmortizationPropertyPlantAndEquipment" xlink:label="loc_us-gaap_AccumulatedDepreciationDepletionAndAmortizationPropertyPlantAndEquipment_D66752918FC64473A9E143AEDBDED14B" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentLineItems_B4E7761B4458CAAE3DB043AEDBDD577F" xlink:to="loc_us-gaap_AccumulatedDepreciationDepletionAndAmortizationPropertyPlantAndEquipment_D66752918FC64473A9E143AEDBDED14B" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentNet" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentNet_427AB4DEC4EAAB71CF4243AEDBDEB8E0" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentLineItems_B4E7761B4458CAAE3DB043AEDBDD577F" xlink:to="loc_us-gaap_PropertyPlantAndEquipmentNet_427AB4DEC4EAAB71CF4243AEDBDEB8E0" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/PropertyAndEquipmentNetNarrativesDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/PropertyAndEquipmentNetTables" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/RelatedPartyTransactions" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/RelatedPartyTransactionsDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RelatedPartyTransactionLineItems" xlink:label="loc_us-gaap_RelatedPartyTransactionLineItems_6D1922C881BE1782ED287216205C5989" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfRelatedPartyTransactionsByRelatedPartyTable" xlink:label="loc_us-gaap_ScheduleOfRelatedPartyTransactionsByRelatedPartyTable_3D9B9B54E9A1F8040F837216205AD3E8" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_RelatedPartyTransactionLineItems_6D1922C881BE1782ED287216205C5989" xlink:to="loc_us-gaap_ScheduleOfRelatedPartyTransactionsByRelatedPartyTable_3D9B9B54E9A1F8040F837216205AD3E8" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RelatedPartyTransactionsByRelatedPartyAxis" xlink:label="loc_us-gaap_RelatedPartyTransactionsByRelatedPartyAxis_FDDB00753DDB9BE4E2327216205ACFAE" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfRelatedPartyTransactionsByRelatedPartyTable_3D9B9B54E9A1F8040F837216205AD3E8" xlink:to="loc_us-gaap_RelatedPartyTransactionsByRelatedPartyAxis_FDDB00753DDB9BE4E2327216205ACFAE" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RelatedPartyDomain" xlink:label="loc_us-gaap_RelatedPartyDomain_2814BEE45120F17D0E9F7216205A7476_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_RelatedPartyTransactionsByRelatedPartyAxis_FDDB00753DDB9BE4E2327216205ACFAE" xlink:to="loc_us-gaap_RelatedPartyDomain_2814BEE45120F17D0E9F7216205A7476_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RelatedPartyDomain" xlink:label="loc_us-gaap_RelatedPartyDomain_2814BEE45120F17D0E9F7216205A7476" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_RelatedPartyTransactionsByRelatedPartyAxis_FDDB00753DDB9BE4E2327216205ACFAE" xlink:to="loc_us-gaap_RelatedPartyDomain_2814BEE45120F17D0E9F7216205A7476" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_AffiliatedEntityMember" xlink:label="loc_srt_AffiliatedEntityMember_17C43A72DB69DB1A19587216205B0CA5" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_RelatedPartyDomain_2814BEE45120F17D0E9F7216205A7476" xlink:to="loc_srt_AffiliatedEntityMember_17C43A72DB69DB1A19587216205B0CA5" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_CounterpartyNameAxis" xlink:label="loc_srt_CounterpartyNameAxis_F572E79C0801A3656AF07216205B7B50" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfRelatedPartyTransactionsByRelatedPartyTable_3D9B9B54E9A1F8040F837216205AD3E8" xlink:to="loc_srt_CounterpartyNameAxis_F572E79C0801A3656AF07216205B7B50" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_RepurchaseAgreementCounterpartyNameDomain" xlink:label="loc_srt_RepurchaseAgreementCounterpartyNameDomain_608A6F3E9FBAA20327C57216205B0382_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_srt_CounterpartyNameAxis_F572E79C0801A3656AF07216205B7B50" xlink:to="loc_srt_RepurchaseAgreementCounterpartyNameDomain_608A6F3E9FBAA20327C57216205B0382_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_RepurchaseAgreementCounterpartyNameDomain" xlink:label="loc_srt_RepurchaseAgreementCounterpartyNameDomain_608A6F3E9FBAA20327C57216205B0382" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_srt_CounterpartyNameAxis_F572E79C0801A3656AF07216205B7B50" xlink:to="loc_srt_RepurchaseAgreementCounterpartyNameDomain_608A6F3E9FBAA20327C57216205B0382" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_UniversityCollegeLondonandRelatedEntitiesMember" xlink:label="loc_autl_UniversityCollegeLondonandRelatedEntitiesMember_80C17106BE82673B21317216205C62D6" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_srt_RepurchaseAgreementCounterpartyNameDomain_608A6F3E9FBAA20327C57216205B0382" xlink:to="loc_autl_UniversityCollegeLondonandRelatedEntitiesMember_80C17106BE82673B21317216205C62D6" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RelatedPartyTransactionExpensesFromTransactionsWithRelatedParty" xlink:label="loc_us-gaap_RelatedPartyTransactionExpensesFromTransactionsWithRelatedParty_930D22F599B368F083E77216205C025D" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_RelatedPartyTransactionLineItems_6D1922C881BE1782ED287216205C5989" xlink:to="loc_us-gaap_RelatedPartyTransactionExpensesFromTransactionsWithRelatedParty_930D22F599B368F083E77216205C025D" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareBasedCompensation" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareBasedCompensation2018PlanAnd2017PlanDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_B48DDEC7FABFC75DF49643AEDBA52225" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" xlink:label="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_456C095715034CCB3EF743AEDBA24215" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_B48DDEC7FABFC75DF49643AEDBA52225" xlink:to="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_456C095715034CCB3EF743AEDBA24215" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AwardTypeAxis" xlink:label="loc_us-gaap_AwardTypeAxis_E4C2594D1AA7F219466843AEDBA25B31" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_456C095715034CCB3EF743AEDBA24215" xlink:to="loc_us-gaap_AwardTypeAxis_E4C2594D1AA7F219466843AEDBA25B31" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_72CEA08630D4F6FC55C943AEDBA30EFA_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_AwardTypeAxis_E4C2594D1AA7F219466843AEDBA25B31" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_72CEA08630D4F6FC55C943AEDBA30EFA_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_72CEA08630D4F6FC55C943AEDBA30EFA" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_AwardTypeAxis_E4C2594D1AA7F219466843AEDBA25B31" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_72CEA08630D4F6FC55C943AEDBA30EFA" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_PerformanceBasedMember" xlink:label="loc_autl_PerformanceBasedMember_32DCCD4FD79B552144D943AEDBA3DD1E" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_72CEA08630D4F6FC55C943AEDBA30EFA" xlink:to="loc_autl_PerformanceBasedMember_32DCCD4FD79B552144D943AEDBA3DD1E" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_EmployeeStockOptionMember" xlink:label="loc_us-gaap_EmployeeStockOptionMember_1F9352A6BF9C04E5E92743AEDBA33915" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_72CEA08630D4F6FC55C943AEDBA30EFA" xlink:to="loc_us-gaap_EmployeeStockOptionMember_1F9352A6BF9C04E5E92743AEDBA33915" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_VestingAxis" xlink:label="loc_us-gaap_VestingAxis_E6C09EE04C36A79E9F1943AEDBA4E4B0" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_456C095715034CCB3EF743AEDBA24215" xlink:to="loc_us-gaap_VestingAxis_E6C09EE04C36A79E9F1943AEDBA4E4B0" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_VestingDomain" xlink:label="loc_us-gaap_VestingDomain_03715FB9B2F95E9F448643AEDBA49BDE_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_VestingAxis_E6C09EE04C36A79E9F1943AEDBA4E4B0" xlink:to="loc_us-gaap_VestingDomain_03715FB9B2F95E9F448643AEDBA49BDE_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_VestingDomain" xlink:label="loc_us-gaap_VestingDomain_03715FB9B2F95E9F448643AEDBA49BDE" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_VestingAxis_E6C09EE04C36A79E9F1943AEDBA4E4B0" xlink:to="loc_us-gaap_VestingDomain_03715FB9B2F95E9F448643AEDBA49BDE" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationAwardTrancheOneMember" xlink:label="loc_us-gaap_ShareBasedCompensationAwardTrancheOneMember_D1E6EC5F60FF140AF06F43AEDBA5419D" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_VestingDomain_03715FB9B2F95E9F448643AEDBA49BDE" xlink:to="loc_us-gaap_ShareBasedCompensationAwardTrancheOneMember_D1E6EC5F60FF140AF06F43AEDBA5419D" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationAwardTrancheTwoMember" xlink:label="loc_us-gaap_ShareBasedCompensationAwardTrancheTwoMember_001B95B8B6C8B4654E0443AEDBA5B008" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_VestingDomain_03715FB9B2F95E9F448643AEDBA49BDE" xlink:to="loc_us-gaap_ShareBasedCompensationAwardTrancheTwoMember_001B95B8B6C8B4654E0443AEDBA5B008" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardAwardVestingPeriod1" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardAwardVestingPeriod1_0AF51DEEDF987DACB14943AEDBA51377" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_B48DDEC7FABFC75DF49643AEDBA52225" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardAwardVestingPeriod1_0AF51DEEDF987DACB14943AEDBA51377" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SharebasedCompensationArrangementBySharebasedPaymentAwardAwardVestingRightsPercentage" xlink:label="loc_us-gaap_SharebasedCompensationArrangementBySharebasedPaymentAwardAwardVestingRightsPercentage_82B21D767499299BE13543AEDBA62A41" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_B48DDEC7FABFC75DF49643AEDBA52225" xlink:to="loc_us-gaap_SharebasedCompensationArrangementBySharebasedPaymentAwardAwardVestingRightsPercentage_82B21D767499299BE13543AEDBA62A41" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SharebasedCompensationArrangementBySharebasedPaymentAwardExpirationPeriod" xlink:label="loc_us-gaap_SharebasedCompensationArrangementBySharebasedPaymentAwardExpirationPeriod_9DD27BABDD2B12B21DA943AEDBA6C83D" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_B48DDEC7FABFC75DF49643AEDBA52225" xlink:to="loc_us-gaap_SharebasedCompensationArrangementBySharebasedPaymentAwardExpirationPeriod_9DD27BABDD2B12B21DA943AEDBA6C83D" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareBasedCompensationOptionsValuationAssumptionsDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_7DE52F6FDBEFEB3F628543AEDB9B8127" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" xlink:label="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_C9C212C7F62987059FEC43AEDB993934" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_7DE52F6FDBEFEB3F628543AEDB9B8127" xlink:to="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_C9C212C7F62987059FEC43AEDB993934" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AwardTypeAxis" xlink:label="loc_us-gaap_AwardTypeAxis_32D7399C68E4A429125D43AEDB9AD300" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_C9C212C7F62987059FEC43AEDB993934" xlink:to="loc_us-gaap_AwardTypeAxis_32D7399C68E4A429125D43AEDB9AD300" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_6CA63DD4A9CD1BA8EEA743AEDB9AF9EF_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_AwardTypeAxis_32D7399C68E4A429125D43AEDB9AD300" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_6CA63DD4A9CD1BA8EEA743AEDB9AF9EF_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_6CA63DD4A9CD1BA8EEA743AEDB9AF9EF" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_AwardTypeAxis_32D7399C68E4A429125D43AEDB9AD300" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_6CA63DD4A9CD1BA8EEA743AEDB9AF9EF" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_EmployeeStockOptionMember" xlink:label="loc_us-gaap_EmployeeStockOptionMember_31CCA0DA5692096B71EB43AEDB9A053E" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_6CA63DD4A9CD1BA8EEA743AEDB9AF9EF" xlink:to="loc_us-gaap_EmployeeStockOptionMember_31CCA0DA5692096B71EB43AEDB9A053E" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_RangeAxis" xlink:label="loc_srt_RangeAxis_4085854F32AAA96E294D43AEDB9B06BF" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_C9C212C7F62987059FEC43AEDB993934" xlink:to="loc_srt_RangeAxis_4085854F32AAA96E294D43AEDB9B06BF" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_RangeMember" xlink:label="loc_srt_RangeMember_E07EB10CAE76317DE9D443AEDB9B976D_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_srt_RangeAxis_4085854F32AAA96E294D43AEDB9B06BF" xlink:to="loc_srt_RangeMember_E07EB10CAE76317DE9D443AEDB9B976D_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_RangeMember" xlink:label="loc_srt_RangeMember_E07EB10CAE76317DE9D443AEDB9B976D" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_srt_RangeAxis_4085854F32AAA96E294D43AEDB9B06BF" xlink:to="loc_srt_RangeMember_E07EB10CAE76317DE9D443AEDB9B976D" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_MinimumMember" xlink:label="loc_srt_MinimumMember_5B520628AB9563D1C9CB43AEDB9BBE3E" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_srt_RangeMember_E07EB10CAE76317DE9D443AEDB9B976D" xlink:to="loc_srt_MinimumMember_5B520628AB9563D1C9CB43AEDB9BBE3E" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/srt/2019/elts/srt-2019-01-31.xsd#srt_MaximumMember" xlink:label="loc_srt_MaximumMember_6FE3EC11A2B46454309243AEDB9B1A56" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_srt_RangeMember_E07EB10CAE76317DE9D443AEDB9B976D" xlink:to="loc_srt_MaximumMember_6FE3EC11A2B46454309243AEDB9B1A56" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsAndMethodologyAbstract" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsAndMethodologyAbstract_D9C71A7E3DF8BCB5158443AEDB9C001B" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_7DE52F6FDBEFEB3F628543AEDB9B8127" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsAndMethodologyAbstract_D9C71A7E3DF8BCB5158443AEDB9C001B" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SharebasedCompensationArrangementBySharebasedPaymentAwardFairValueAssumptionsExpectedTerm1" xlink:label="loc_us-gaap_SharebasedCompensationArrangementBySharebasedPaymentAwardFairValueAssumptionsExpectedTerm1_08DDE72E276F839F57D243AEDB9C69CA" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsAndMethodologyAbstract_D9C71A7E3DF8BCB5158443AEDB9C001B" xlink:to="loc_us-gaap_SharebasedCompensationArrangementBySharebasedPaymentAwardFairValueAssumptionsExpectedTerm1_08DDE72E276F839F57D243AEDB9C69CA" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsRiskFreeInterestRateMinimum" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsRiskFreeInterestRateMinimum_19B1AD1CECE1C3013A0943AEDB9CBE77" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsAndMethodologyAbstract_D9C71A7E3DF8BCB5158443AEDB9C001B" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsRiskFreeInterestRateMinimum_19B1AD1CECE1C3013A0943AEDB9CBE77" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsRiskFreeInterestRateMaximum" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsRiskFreeInterestRateMaximum_4DC5398D96BAF5424E3C43AEDB9C8FA0" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsAndMethodologyAbstract_D9C71A7E3DF8BCB5158443AEDB9C001B" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsRiskFreeInterestRateMaximum_4DC5398D96BAF5424E3C43AEDB9C8FA0" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsExpectedVolatilityRateMinimum" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsExpectedVolatilityRateMinimum_420AAB1E340A7D343DFF43AEDB9CB8CC" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsAndMethodologyAbstract_D9C71A7E3DF8BCB5158443AEDB9C001B" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsExpectedVolatilityRateMinimum_420AAB1E340A7D343DFF43AEDB9CB8CC" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsExpectedVolatilityRateMaximum" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsExpectedVolatilityRateMaximum_F496D81C960411F09F2F43AEDB9DAAD7" xlink:type="locator" />
    <link:definitionArc order="5" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsAndMethodologyAbstract_D9C71A7E3DF8BCB5158443AEDB9C001B" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsExpectedVolatilityRateMaximum_F496D81C960411F09F2F43AEDB9DAAD7" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsExpectedDividendRate" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsExpectedDividendRate_307A1F522B35BD627FC343AEDB9D559D" xlink:type="locator" />
    <link:definitionArc order="6" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsAndMethodologyAbstract_D9C71A7E3DF8BCB5158443AEDB9C001B" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardFairValueAssumptionsExpectedDividendRate_307A1F522B35BD627FC343AEDB9D559D" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareBasedCompensationRestrictedAndOrdinarySharesNarrativesDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_383E5984CC33D04B385C43AEDB7B8DDE" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" xlink:label="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_B67286F47A20268F908A43AEDB7A1597" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_383E5984CC33D04B385C43AEDB7B8DDE" xlink:to="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_B67286F47A20268F908A43AEDB7A1597" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AwardTypeAxis" xlink:label="loc_us-gaap_AwardTypeAxis_9ABD139E2B39E2ABC6FD43AEDB7A55B5" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_B67286F47A20268F908A43AEDB7A1597" xlink:to="loc_us-gaap_AwardTypeAxis_9ABD139E2B39E2ABC6FD43AEDB7A55B5" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_1176CFAE5697DA6C10B143AEDB7A5E4F_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_AwardTypeAxis_9ABD139E2B39E2ABC6FD43AEDB7A55B5" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_1176CFAE5697DA6C10B143AEDB7A5E4F_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_1176CFAE5697DA6C10B143AEDB7A5E4F" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_AwardTypeAxis_9ABD139E2B39E2ABC6FD43AEDB7A55B5" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_1176CFAE5697DA6C10B143AEDB7A5E4F" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RestrictedStockMember" xlink:label="loc_us-gaap_RestrictedStockMember_C426D108ED7CEBEDBD7C43AEDB7BAF20" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_1176CFAE5697DA6C10B143AEDB7A5E4F" xlink:to="loc_us-gaap_RestrictedStockMember_C426D108ED7CEBEDBD7C43AEDB7BAF20" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognizedShareBasedAwardsOtherThanOptions" xlink:label="loc_us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognizedShareBasedAwardsOtherThanOptions_D3C3D1EDF47CCFE605FD43AEDB7B2D82" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_383E5984CC33D04B385C43AEDB7B8DDE" xlink:to="loc_us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognizedShareBasedAwardsOtherThanOptions_D3C3D1EDF47CCFE605FD43AEDB7B2D82" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareBasedCompensationRestrictedOrdinarySharesRollforwardDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_FA7E251CD0B78968618743AEDB81B235" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" xlink:label="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_DEF50F21EDA5133E93D943AEDB80B3FB" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_FA7E251CD0B78968618743AEDB81B235" xlink:to="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_DEF50F21EDA5133E93D943AEDB80B3FB" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AwardTypeAxis" xlink:label="loc_us-gaap_AwardTypeAxis_84382FA6887FA7CCC94243AEDB804257" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_DEF50F21EDA5133E93D943AEDB80B3FB" xlink:to="loc_us-gaap_AwardTypeAxis_84382FA6887FA7CCC94243AEDB804257" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_2A0B6C47D95E7524D99543AEDB8068AC_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_AwardTypeAxis_84382FA6887FA7CCC94243AEDB804257" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_2A0B6C47D95E7524D99543AEDB8068AC_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_2A0B6C47D95E7524D99543AEDB8068AC" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_AwardTypeAxis_84382FA6887FA7CCC94243AEDB804257" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_2A0B6C47D95E7524D99543AEDB8068AC" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RestrictedStockMember" xlink:label="loc_us-gaap_RestrictedStockMember_943C977EC4DF5ED47DEA43AEDB80D365" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_2A0B6C47D95E7524D99543AEDB8068AC" xlink:to="loc_us-gaap_RestrictedStockMember_943C977EC4DF5ED47DEA43AEDB80D365" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward_6C6187F85E0F96466BE343AEDB816F6B" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_FA7E251CD0B78968618743AEDB81B235" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward_6C6187F85E0F96466BE343AEDB816F6B" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedNumber" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedNumber_B51B0875AD4F3FCB084F43AEDB81AE60" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward_6C6187F85E0F96466BE343AEDB816F6B" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedNumber_B51B0875AD4F3FCB084F43AEDB81AE60" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsGrantsInPeriod" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsGrantsInPeriod_2B9198F1BD08AF7FA52443AEDB8240D3" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward_6C6187F85E0F96466BE343AEDB816F6B" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsGrantsInPeriod_2B9198F1BD08AF7FA52443AEDB8240D3" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsVestedInPeriod" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsVestedInPeriod_ECC1E1A4D28752CA4BCB43AEDB82CEC4" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward_6C6187F85E0F96466BE343AEDB816F6B" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsVestedInPeriod_ECC1E1A4D28752CA4BCB43AEDB82CEC4" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsForfeitedInPeriod" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsForfeitedInPeriod_DB36DDAD10EA64ED5C0E43AEDB822979" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedRollForward_6C6187F85E0F96466BE343AEDB816F6B" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsForfeitedInPeriod_DB36DDAD10EA64ED5C0E43AEDB822979" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward_A33EF3A4F396AFC9C30F43AEDB82944A" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_FA7E251CD0B78968618743AEDB81B235" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward_A33EF3A4F396AFC9C30F43AEDB82944A" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValue" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValue_ED4BF2EC2EB624788A8C43AEDB8290CC" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward_A33EF3A4F396AFC9C30F43AEDB82944A" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValue_ED4BF2EC2EB624788A8C43AEDB8290CC" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsGrantsInPeriodWeightedAverageGrantDateFairValue" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsGrantsInPeriodWeightedAverageGrantDateFairValue_30A15ED105631ECB89EC43AEDB83EFE8" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward_A33EF3A4F396AFC9C30F43AEDB82944A" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsGrantsInPeriodWeightedAverageGrantDateFairValue_30A15ED105631ECB89EC43AEDB83EFE8" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsVestedInPeriodWeightedAverageGrantDateFairValue" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsVestedInPeriodWeightedAverageGrantDateFairValue_14130FC471D40394943D43AEDB831210" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward_A33EF3A4F396AFC9C30F43AEDB82944A" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsVestedInPeriodWeightedAverageGrantDateFairValue_14130FC471D40394943D43AEDB831210" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsForfeituresWeightedAverageGrantDateFairValue" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsForfeituresWeightedAverageGrantDateFairValue_667481275400A254580843AEDB83F6CF" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsNonvestedWeightedAverageGrantDateFairValueRollForward_A33EF3A4F396AFC9C30F43AEDB82944A" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardEquityInstrumentsOtherThanOptionsForfeituresWeightedAverageGrantDateFairValue_667481275400A254580843AEDB83F6CF" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareBasedCompensationShareBasedCompensationAllocationDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_D4237AA905D33084FC305042E25129CC" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" xlink:label="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_6C999139D34508D6E7D95042E24E9382" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_D4237AA905D33084FC305042E25129CC" xlink:to="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_6C999139D34508D6E7D95042E24E9382" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_IncomeStatementLocationAxis" xlink:label="loc_us-gaap_IncomeStatementLocationAxis_D0472C31FA0BEE372A7C5042E24E455E" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_6C999139D34508D6E7D95042E24E9382" xlink:to="loc_us-gaap_IncomeStatementLocationAxis_D0472C31FA0BEE372A7C5042E24E455E" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_IncomeStatementLocationDomain" xlink:label="loc_us-gaap_IncomeStatementLocationDomain_5FC705540AC845782B0B5042E24E46F2_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_IncomeStatementLocationAxis_D0472C31FA0BEE372A7C5042E24E455E" xlink:to="loc_us-gaap_IncomeStatementLocationDomain_5FC705540AC845782B0B5042E24E46F2_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_IncomeStatementLocationDomain" xlink:label="loc_us-gaap_IncomeStatementLocationDomain_5FC705540AC845782B0B5042E24E46F2" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_IncomeStatementLocationAxis_D0472C31FA0BEE372A7C5042E24E455E" xlink:to="loc_us-gaap_IncomeStatementLocationDomain_5FC705540AC845782B0B5042E24E46F2" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ResearchAndDevelopmentExpenseMember" xlink:label="loc_us-gaap_ResearchAndDevelopmentExpenseMember_927A4878CFE7017A4A2E5042E24ECBE8" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncomeStatementLocationDomain_5FC705540AC845782B0B5042E24E46F2" xlink:to="loc_us-gaap_ResearchAndDevelopmentExpenseMember_927A4878CFE7017A4A2E5042E24ECBE8" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_GeneralAndAdministrativeExpenseMember" xlink:label="loc_us-gaap_GeneralAndAdministrativeExpenseMember_F2A2EDF28B1290F5A05D5042E25139AA" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_IncomeStatementLocationDomain_5FC705540AC845782B0B5042E24E46F2" xlink:to="loc_us-gaap_GeneralAndAdministrativeExpenseMember_F2A2EDF28B1290F5A05D5042E25139AA" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AllocatedShareBasedCompensationExpense" xlink:label="loc_us-gaap_AllocatedShareBasedCompensationExpense_5F717F31CA93BB8F9B745042E2517457" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_D4237AA905D33084FC305042E25129CC" xlink:to="loc_us-gaap_AllocatedShareBasedCompensationExpense_5F717F31CA93BB8F9B745042E2517457" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareBasedCompensationShareOptionsNarrativesDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_E980AB5C5F732DBF6D7343AEDB89120D" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable" xlink:label="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_C45BC4456B4ABD0AB76043AEDB88701C" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_E980AB5C5F732DBF6D7343AEDB89120D" xlink:to="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_C45BC4456B4ABD0AB76043AEDB88701C" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AwardTypeAxis" xlink:label="loc_us-gaap_AwardTypeAxis_C9DED7851D9BB8638CB943AEDB882DFC" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfShareBasedCompensationArrangementsByShareBasedPaymentAwardTable_C45BC4456B4ABD0AB76043AEDB88701C" xlink:to="loc_us-gaap_AwardTypeAxis_C9DED7851D9BB8638CB943AEDB882DFC" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_D4A23C5A171F39217E2543AEDB8851A4_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_AwardTypeAxis_C9DED7851D9BB8638CB943AEDB882DFC" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_D4A23C5A171F39217E2543AEDB8851A4_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_D4A23C5A171F39217E2543AEDB8851A4" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_AwardTypeAxis_C9DED7851D9BB8638CB943AEDB882DFC" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_D4A23C5A171F39217E2543AEDB8851A4" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_EmployeeStockOptionMember" xlink:label="loc_us-gaap_EmployeeStockOptionMember_1CE6B2A95568E3F6C44E43AEDB881B77" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementsByShareBasedPaymentAwardAwardTypeAndPlanNameDomain_D4A23C5A171F39217E2543AEDB8851A4" xlink:to="loc_us-gaap_EmployeeStockOptionMember_1CE6B2A95568E3F6C44E43AEDB881B77" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardOptionsGrantsInPeriodWeightedAverageGrantDateFairValue" xlink:label="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardOptionsGrantsInPeriodWeightedAverageGrantDateFairValue_B12ED3E883F08DA4C4C443AEDB89629D" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_E980AB5C5F732DBF6D7343AEDB89120D" xlink:to="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardOptionsGrantsInPeriodWeightedAverageGrantDateFairValue_B12ED3E883F08DA4C4C443AEDB89629D" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognized" xlink:label="loc_us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognized_26B615B21B8C7294040B43AEDB89F184" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_E980AB5C5F732DBF6D7343AEDB89120D" xlink:to="loc_us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognized_26B615B21B8C7294040B43AEDB89F184" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognizedPeriodForRecognition1" xlink:label="loc_us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognizedPeriodForRecognition1_67A488485CD049C5988943AEDB89E732" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ShareBasedCompensationArrangementByShareBasedPaymentAwardLineItems_E980AB5C5F732DBF6D7343AEDB89120D" xlink:to="loc_us-gaap_EmployeeServiceShareBasedCompensationNonvestedAwardsTotalCompensationCostNotYetRecognizedPeriodForRecognition1_67A488485CD049C5988943AEDB89E732" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareBasedCompensationShareOptionsRollforwardDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareBasedCompensationTables" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareholdersEquity" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareholdersEquityDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockLineItems" xlink:label="loc_us-gaap_ClassOfStockLineItems_7E6938A908D453CD78E243AEDBC1149C" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfStockByClassTable" xlink:label="loc_us-gaap_ScheduleOfStockByClassTable_20302029B08107D545BD43AEDBBFD4C6" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_ClassOfStockLineItems_7E6938A908D453CD78E243AEDBC1149C" xlink:to="loc_us-gaap_ScheduleOfStockByClassTable_20302029B08107D545BD43AEDBBFD4C6" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementClassOfStockAxis" xlink:label="loc_us-gaap_StatementClassOfStockAxis_C7A8698494D3380F5A3C43AEDBC0336F" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfStockByClassTable_20302029B08107D545BD43AEDBBFD4C6" xlink:to="loc_us-gaap_StatementClassOfStockAxis_C7A8698494D3380F5A3C43AEDBC0336F" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_251ADEBA7F49FB3C3C0543AEDBC02527_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_StatementClassOfStockAxis_C7A8698494D3380F5A3C43AEDBC0336F" xlink:to="loc_us-gaap_ClassOfStockDomain_251ADEBA7F49FB3C3C0543AEDBC02527_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_251ADEBA7F49FB3C3C0543AEDBC02527" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_StatementClassOfStockAxis_C7A8698494D3380F5A3C43AEDBC0336F" xlink:to="loc_us-gaap_ClassOfStockDomain_251ADEBA7F49FB3C3C0543AEDBC02527" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_OrdinarySharesMember" xlink:label="loc_autl_OrdinarySharesMember_25683E2B83371F68FB1B43AEDBC08A60" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_251ADEBA7F49FB3C3C0543AEDBC02527" xlink:to="loc_autl_OrdinarySharesMember_25683E2B83371F68FB1B43AEDBC08A60" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassASharesMember" xlink:label="loc_autl_DeferredClassASharesMember_D302FEBE6CC27DED6BA243AEDBC1AC11" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_251ADEBA7F49FB3C3C0543AEDBC02527" xlink:to="loc_autl_DeferredClassASharesMember_D302FEBE6CC27DED6BA243AEDBC1AC11" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassBSharesMember" xlink:label="loc_autl_DeferredClassBSharesMember_B06C218F2E30115BEE1943AEDBC12DAE" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_251ADEBA7F49FB3C3C0543AEDBC02527" xlink:to="loc_autl_DeferredClassBSharesMember_B06C218F2E30115BEE1943AEDBC12DAE" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassCSharesMember" xlink:label="loc_autl_DeferredClassCSharesMember_19029F2A71536B9D9A3743AEDBC106AA" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_251ADEBA7F49FB3C3C0543AEDBC02527" xlink:to="loc_autl_DeferredClassCSharesMember_19029F2A71536B9D9A3743AEDBC106AA" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SharesOutstanding" xlink:label="loc_us-gaap_SharesOutstanding_D7DB287378B051593FB843AEDBC17C6B" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_7E6938A908D453CD78E243AEDBC1149C" xlink:to="loc_us-gaap_SharesOutstanding_D7DB287378B051593FB843AEDBC17C6B" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareholdersEquityInitialPublicOfferingAndImpactOfCorporateReorganizationDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockLineItems" xlink:label="loc_us-gaap_ClassOfStockLineItems_0792510301FD47FA539E5042E299F4EC" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfStockByClassTable" xlink:label="loc_us-gaap_ScheduleOfStockByClassTable_A0524D5B2EB169A5B5855042E296B29D" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_ClassOfStockLineItems_0792510301FD47FA539E5042E299F4EC" xlink:to="loc_us-gaap_ScheduleOfStockByClassTable_A0524D5B2EB169A5B5855042E296B29D" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SubsidiarySaleOfStockAxis" xlink:label="loc_us-gaap_SubsidiarySaleOfStockAxis_02681B2CF562AF672AD85042E296B638" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfStockByClassTable_A0524D5B2EB169A5B5855042E296B29D" xlink:to="loc_us-gaap_SubsidiarySaleOfStockAxis_02681B2CF562AF672AD85042E296B638" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SaleOfStockNameOfTransactionDomain" xlink:label="loc_us-gaap_SaleOfStockNameOfTransactionDomain_0DDB765394D42AB7731D5042E2973D3D_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_SubsidiarySaleOfStockAxis_02681B2CF562AF672AD85042E296B638" xlink:to="loc_us-gaap_SaleOfStockNameOfTransactionDomain_0DDB765394D42AB7731D5042E2973D3D_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SaleOfStockNameOfTransactionDomain" xlink:label="loc_us-gaap_SaleOfStockNameOfTransactionDomain_0DDB765394D42AB7731D5042E2973D3D" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_SubsidiarySaleOfStockAxis_02681B2CF562AF672AD85042E296B638" xlink:to="loc_us-gaap_SaleOfStockNameOfTransactionDomain_0DDB765394D42AB7731D5042E2973D3D" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_IPOMember" xlink:label="loc_us-gaap_IPOMember_D3676CEBD4877100DA885042E2978238" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_SaleOfStockNameOfTransactionDomain_0DDB765394D42AB7731D5042E2973D3D" xlink:to="loc_us-gaap_IPOMember_D3676CEBD4877100DA885042E2978238" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_PublicStockOfferingMember" xlink:label="loc_autl_PublicStockOfferingMember_03140E58056311D18E4D507E9C600613" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_SaleOfStockNameOfTransactionDomain_0DDB765394D42AB7731D5042E2973D3D" xlink:to="loc_autl_PublicStockOfferingMember_03140E58056311D18E4D507E9C600613" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_OverAllotmentOptionMember" xlink:label="loc_us-gaap_OverAllotmentOptionMember_2EDDE8236341392D3862507E9E1A44EE" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_SaleOfStockNameOfTransactionDomain_0DDB765394D42AB7731D5042E2973D3D" xlink:to="loc_us-gaap_OverAllotmentOptionMember_2EDDE8236341392D3862507E9E1A44EE" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementClassOfStockAxis" xlink:label="loc_us-gaap_StatementClassOfStockAxis_51F750F4D2013D749ED95042E29792BC" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfStockByClassTable_A0524D5B2EB169A5B5855042E296B29D" xlink:to="loc_us-gaap_StatementClassOfStockAxis_51F750F4D2013D749ED95042E29792BC" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_08E32C64A619992846C75042E2979A3C_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_StatementClassOfStockAxis_51F750F4D2013D749ED95042E29792BC" xlink:to="loc_us-gaap_ClassOfStockDomain_08E32C64A619992846C75042E2979A3C_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_08E32C64A619992846C75042E2979A3C" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_StatementClassOfStockAxis_51F750F4D2013D749ED95042E29792BC" xlink:to="loc_us-gaap_ClassOfStockDomain_08E32C64A619992846C75042E2979A3C" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_OrdinarySharesMember" xlink:label="loc_autl_OrdinarySharesMember_AC548DBD3CFCA77477425042E298A83F" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_08E32C64A619992846C75042E2979A3C" xlink:to="loc_autl_OrdinarySharesMember_AC548DBD3CFCA77477425042E298A83F" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassASharesMember" xlink:label="loc_autl_DeferredClassASharesMember_4AD17BB45E3940A608845042E298CA4F" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_08E32C64A619992846C75042E2979A3C" xlink:to="loc_autl_DeferredClassASharesMember_4AD17BB45E3940A608845042E298CA4F" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassBSharesMember" xlink:label="loc_autl_DeferredClassBSharesMember_F43F27DF10DBBE3AED565042E298B04A" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_08E32C64A619992846C75042E2979A3C" xlink:to="loc_autl_DeferredClassBSharesMember_F43F27DF10DBBE3AED565042E298B04A" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_DeferredClassCSharesMember" xlink:label="loc_autl_DeferredClassCSharesMember_9103AF7353DF464B15575042E2992B28" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_08E32C64A619992846C75042E2979A3C" xlink:to="loc_autl_DeferredClassCSharesMember_9103AF7353DF464B15575042E2992B28" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SaleOfStockNumberOfSharesIssuedInTransaction" xlink:label="loc_us-gaap_SaleOfStockNumberOfSharesIssuedInTransaction_A106A02870FC00096B335042E2994971" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_0792510301FD47FA539E5042E299F4EC" xlink:to="loc_us-gaap_SaleOfStockNumberOfSharesIssuedInTransaction_A106A02870FC00096B335042E2994971" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SaleOfStockPricePerShare" xlink:label="loc_us-gaap_SaleOfStockPricePerShare_50A0A07360FAA5C9078D5042E2993253" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_0792510301FD47FA539E5042E299F4EC" xlink:to="loc_us-gaap_SaleOfStockPricePerShare_50A0A07360FAA5C9078D5042E2993253" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ProceedsFromIssuanceOfCommonStock" xlink:label="loc_us-gaap_ProceedsFromIssuanceOfCommonStock_0BD391ED25836CC3505C5042E2990119" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_0792510301FD47FA539E5042E299F4EC" xlink:to="loc_us-gaap_ProceedsFromIssuanceOfCommonStock_0BD391ED25836CC3505C5042E2990119" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PaymentsOfStockIssuanceCosts" xlink:label="loc_us-gaap_PaymentsOfStockIssuanceCosts_9E6E0356469AF9BDFBC15042E2990CBC" xlink:type="locator" />
    <link:definitionArc order="4" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_0792510301FD47FA539E5042E299F4EC" xlink:to="loc_us-gaap_PaymentsOfStockIssuanceCosts_9E6E0356469AF9BDFBC15042E2990CBC" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_EquityConversionRatio" xlink:label="loc_autl_EquityConversionRatio_E099550310CD5AC7C8385042E299129C" xlink:type="locator" />
    <link:definitionArc order="5" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_0792510301FD47FA539E5042E299F4EC" xlink:to="loc_autl_EquityConversionRatio_E099550310CD5AC7C8385042E299129C" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SharesOutstanding" xlink:label="loc_us-gaap_SharesOutstanding_B4C17784499399053ACD5042E2997C90" xlink:type="locator" />
    <link:definitionArc order="6" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_0792510301FD47FA539E5042E299F4EC" xlink:to="loc_us-gaap_SharesOutstanding_B4C17784499399053ACD5042E2997C90" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_CommonStockValue" xlink:label="loc_us-gaap_CommonStockValue_677EDFD083C7B958404C5042E299B2DE" xlink:type="locator" />
    <link:definitionArc order="7" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_0792510301FD47FA539E5042E299F4EC" xlink:to="loc_us-gaap_CommonStockValue_677EDFD083C7B958404C5042E299B2DE" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_ShareCapitalRegistrationThreshold" xlink:label="loc_autl_ShareCapitalRegistrationThreshold_14FFBCFE2EB488E253745042E2990524" xlink:type="locator" />
    <link:definitionArc order="8" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_0792510301FD47FA539E5042E299F4EC" xlink:to="loc_autl_ShareCapitalRegistrationThreshold_14FFBCFE2EB488E253745042E2990524" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_SaleOfStockConsiderationReceivedOnTransaction" xlink:label="loc_us-gaap_SaleOfStockConsiderationReceivedOnTransaction_DF9EEBBB0A61985C1C15507E993C9C4B" xlink:type="locator" />
    <link:definitionArc order="9" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_0792510301FD47FA539E5042E299F4EC" xlink:to="loc_us-gaap_SaleOfStockConsiderationReceivedOnTransaction_DF9EEBBB0A61985C1C15507E993C9C4B" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareholdersEquityOrdinarySharesDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockLineItems" xlink:label="loc_us-gaap_ClassOfStockLineItems_979C2C1BA0F7E2DFC9B043AEDBBB2C8E" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfStockByClassTable" xlink:label="loc_us-gaap_ScheduleOfStockByClassTable_24B3FFDA547EE476C91043AEDBBAEED8" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_ClassOfStockLineItems_979C2C1BA0F7E2DFC9B043AEDBBB2C8E" xlink:to="loc_us-gaap_ScheduleOfStockByClassTable_24B3FFDA547EE476C91043AEDBBAEED8" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_StatementClassOfStockAxis" xlink:label="loc_us-gaap_StatementClassOfStockAxis_DD0E44444A783C794F8C43AEDBBAC019" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfStockByClassTable_24B3FFDA547EE476C91043AEDBBAEED8" xlink:to="loc_us-gaap_StatementClassOfStockAxis_DD0E44444A783C794F8C43AEDBBAC019" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_793305C2A6C7E317FAA043AEDBBB8465_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_StatementClassOfStockAxis_DD0E44444A783C794F8C43AEDBBAC019" xlink:to="loc_us-gaap_ClassOfStockDomain_793305C2A6C7E317FAA043AEDBBB8465_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ClassOfStockDomain" xlink:label="loc_us-gaap_ClassOfStockDomain_793305C2A6C7E317FAA043AEDBBB8465" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_StatementClassOfStockAxis_DD0E44444A783C794F8C43AEDBBAC019" xlink:to="loc_us-gaap_ClassOfStockDomain_793305C2A6C7E317FAA043AEDBBB8465" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_OrdinarySharesMember" xlink:label="loc_autl_OrdinarySharesMember_68778CC745F67D1D9B7B43AEDBBB1807" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockDomain_793305C2A6C7E317FAA043AEDBBB8465" xlink:to="loc_autl_OrdinarySharesMember_68778CC745F67D1D9B7B43AEDBBB1807" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_CommonStockSharesAuthorized" xlink:label="loc_us-gaap_CommonStockSharesAuthorized_638C0A4A49FEE117145C43AEDBBC0A64" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_979C2C1BA0F7E2DFC9B043AEDBBB2C8E" xlink:to="loc_us-gaap_CommonStockSharesAuthorized_638C0A4A49FEE117145C43AEDBBC0A64" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_CommonStockParOrStatedValuePerShare" xlink:label="loc_us-gaap_CommonStockParOrStatedValuePerShare_B60D7640D74B7CB02CBA43AEDBBC6691" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_ClassOfStockLineItems_979C2C1BA0F7E2DFC9B043AEDBBB2C8E" xlink:to="loc_us-gaap_CommonStockParOrStatedValuePerShare_B60D7640D74B7CB02CBA43AEDBBC6691" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/ShareholdersEquityTables" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/SubsequentEvent" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/SummaryOfSignificantAccountingPolicies" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesAntidilutiveSharesDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareLineItems" xlink:label="loc_us-gaap_AntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareLineItems_38DD63DDFEF5E43BD9EB43AEDBFE70B5" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfAntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareTable" xlink:label="loc_us-gaap_ScheduleOfAntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareTable_2B80DED6422B96C9C8A543AEDBFD56E6" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_AntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareLineItems_38DD63DDFEF5E43BD9EB43AEDBFE70B5" xlink:to="loc_us-gaap_ScheduleOfAntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareTable_2B80DED6422B96C9C8A543AEDBFD56E6" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareByAntidilutiveSecuritiesAxis" xlink:label="loc_us-gaap_AntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareByAntidilutiveSecuritiesAxis_18232F4C74FF7CC285A743AEDBFD8EBD" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfAntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareTable_2B80DED6422B96C9C8A543AEDBFD56E6" xlink:to="loc_us-gaap_AntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareByAntidilutiveSecuritiesAxis_18232F4C74FF7CC285A743AEDBFD8EBD" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AntidilutiveSecuritiesNameDomain" xlink:label="loc_us-gaap_AntidilutiveSecuritiesNameDomain_856D1ECC87F3F90A8A0A43AEDBFEB305_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_AntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareByAntidilutiveSecuritiesAxis_18232F4C74FF7CC285A743AEDBFD8EBD" xlink:to="loc_us-gaap_AntidilutiveSecuritiesNameDomain_856D1ECC87F3F90A8A0A43AEDBFEB305_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AntidilutiveSecuritiesNameDomain" xlink:label="loc_us-gaap_AntidilutiveSecuritiesNameDomain_856D1ECC87F3F90A8A0A43AEDBFEB305" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_AntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareByAntidilutiveSecuritiesAxis_18232F4C74FF7CC285A743AEDBFD8EBD" xlink:to="loc_us-gaap_AntidilutiveSecuritiesNameDomain_856D1ECC87F3F90A8A0A43AEDBFEB305" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RestrictedStockUnitsRSUMember" xlink:label="loc_us-gaap_RestrictedStockUnitsRSUMember_353A4AD8DE48862C6EE743AEDBFE0E80" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_AntidilutiveSecuritiesNameDomain_856D1ECC87F3F90A8A0A43AEDBFEB305" xlink:to="loc_us-gaap_RestrictedStockUnitsRSUMember_353A4AD8DE48862C6EE743AEDBFE0E80" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_EmployeeStockOptionMember" xlink:label="loc_us-gaap_EmployeeStockOptionMember_AFAC21AC4A48AA05268B43AEDBFE7958" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_AntidilutiveSecuritiesNameDomain_856D1ECC87F3F90A8A0A43AEDBFEB305" xlink:to="loc_us-gaap_EmployeeStockOptionMember_AFAC21AC4A48AA05268B43AEDBFE7958" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_AntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareAmount" xlink:label="loc_us-gaap_AntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareAmount_24D1C52B670EB05F684743AEDBFF4419" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_AntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareLineItems_38DD63DDFEF5E43BD9EB43AEDBFE70B5" xlink:to="loc_us-gaap_AntidilutiveSecuritiesExcludedFromComputationOfEarningsPerShareAmount_24D1C52B670EB05F684743AEDBFF4419" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesComprehensiveLossDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesForeignCurrencyRemeasurementAndTranslationDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesIntangibleAssetsSubjectToAmortizationDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesPatentCostsDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesPolicies" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesPropertyPlantAndEquipmentDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentLineItems" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentLineItems_ADE153F04EE353A1389C43AEDC2D5938" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfPropertyPlantAndEquipmentTable" xlink:label="loc_us-gaap_ScheduleOfPropertyPlantAndEquipmentTable_724901D01B0637DB260143AEDC267F39" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentLineItems_ADE153F04EE353A1389C43AEDC2D5938" xlink:to="loc_us-gaap_ScheduleOfPropertyPlantAndEquipmentTable_724901D01B0637DB260143AEDC267F39" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentByTypeAxis" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentByTypeAxis_4B45472E93ED40F83FD243AEDC26F6EB" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfPropertyPlantAndEquipmentTable_724901D01B0637DB260143AEDC267F39" xlink:to="loc_us-gaap_PropertyPlantAndEquipmentByTypeAxis_4B45472E93ED40F83FD243AEDC26F6EB" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentTypeDomain" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_DE054B06723B7F84F9A043AEDC2702CF_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentByTypeAxis_4B45472E93ED40F83FD243AEDC26F6EB" xlink:to="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_DE054B06723B7F84F9A043AEDC2702CF_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentTypeDomain" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_DE054B06723B7F84F9A043AEDC2702CF" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentByTypeAxis_4B45472E93ED40F83FD243AEDC26F6EB" xlink:to="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_DE054B06723B7F84F9A043AEDC2702CF" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_OfficeEquipmentMember" xlink:label="loc_us-gaap_OfficeEquipmentMember_4CC3F7D695CAE04022B243AEDC27E236" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_DE054B06723B7F84F9A043AEDC2702CF" xlink:to="loc_us-gaap_OfficeEquipmentMember_4CC3F7D695CAE04022B243AEDC27E236" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_EquipmentMember" xlink:label="loc_us-gaap_EquipmentMember_8B6FC6078FEDF9DC625743AEDC2D9568" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_DE054B06723B7F84F9A043AEDC2702CF" xlink:to="loc_us-gaap_EquipmentMember_8B6FC6078FEDF9DC625743AEDC2D9568" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_FurnitureAndFixturesMember" xlink:label="loc_us-gaap_FurnitureAndFixturesMember_E772F866BBDBB758EA7643AEDC2DAF9C" xlink:type="locator" />
    <link:definitionArc order="3" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentTypeDomain_DE054B06723B7F84F9A043AEDC2702CF" xlink:to="loc_us-gaap_FurnitureAndFixturesMember_E772F866BBDBB758EA7643AEDC2DAF9C" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_PropertyPlantAndEquipmentUsefulLife" xlink:label="loc_us-gaap_PropertyPlantAndEquipmentUsefulLife_CD538A3BF38F2C54F44643AEDC2EB592" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentLineItems_ADE153F04EE353A1389C43AEDC2D5938" xlink:to="loc_us-gaap_PropertyPlantAndEquipmentUsefulLife_CD538A3BF38F2C54F44643AEDC2EB592" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ImpairmentOfLongLivedAssetsHeldForUse" xlink:label="loc_us-gaap_ImpairmentOfLongLivedAssetsHeldForUse_0D31B2CCDEEC9BD1BB3043B588D71BFC" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_PropertyPlantAndEquipmentLineItems_ADE153F04EE353A1389C43AEDC2D5938" xlink:to="loc_us-gaap_ImpairmentOfLongLivedAssetsHeldForUse_0D31B2CCDEEC9BD1BB3043B588D71BFC" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesResearchAndDevelopmentDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesRestrictedCashDetails" xlink:type="extended">
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RestrictedCashAndCashEquivalentsItemsLineItems" xlink:label="loc_us-gaap_RestrictedCashAndCashEquivalentsItemsLineItems_4D21C5473EA802B7C7BA43AEDC335743" xlink:type="locator" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_ScheduleOfRestrictedCashAndCashEquivalentsTable" xlink:label="loc_us-gaap_ScheduleOfRestrictedCashAndCashEquivalentsTable_62A99F72F1DE9E481A1D43AEDC3281FA" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/all" xlink:from="loc_us-gaap_RestrictedCashAndCashEquivalentsItemsLineItems_4D21C5473EA802B7C7BA43AEDC335743" xlink:to="loc_us-gaap_ScheduleOfRestrictedCashAndCashEquivalentsTable_62A99F72F1DE9E481A1D43AEDC3281FA" xlink:type="arc" xbrldt:closed="true" xbrldt:contextElement="segment" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RestrictedCashAndCashEquivalentsCashAndCashEquivalentsAxis" xlink:label="loc_us-gaap_RestrictedCashAndCashEquivalentsCashAndCashEquivalentsAxis_962F131A533B87D3BC5443AEDC32C0C9" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/hypercube-dimension" xlink:from="loc_us-gaap_ScheduleOfRestrictedCashAndCashEquivalentsTable_62A99F72F1DE9E481A1D43AEDC3281FA" xlink:to="loc_us-gaap_RestrictedCashAndCashEquivalentsCashAndCashEquivalentsAxis_962F131A533B87D3BC5443AEDC32C0C9" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RestrictedCashAndCashEquivalentsCashAndCashEquivalentsMember" xlink:label="loc_us-gaap_RestrictedCashAndCashEquivalentsCashAndCashEquivalentsMember_1121BC8ACBB23BEF139943AEDC329CAC_default" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-default" xlink:from="loc_us-gaap_RestrictedCashAndCashEquivalentsCashAndCashEquivalentsAxis_962F131A533B87D3BC5443AEDC32C0C9" xlink:to="loc_us-gaap_RestrictedCashAndCashEquivalentsCashAndCashEquivalentsMember_1121BC8ACBB23BEF139943AEDC329CAC_default" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RestrictedCashAndCashEquivalentsCashAndCashEquivalentsMember" xlink:label="loc_us-gaap_RestrictedCashAndCashEquivalentsCashAndCashEquivalentsMember_1121BC8ACBB23BEF139943AEDC329CAC" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/dimension-domain" xlink:from="loc_us-gaap_RestrictedCashAndCashEquivalentsCashAndCashEquivalentsAxis_962F131A533B87D3BC5443AEDC32C0C9" xlink:to="loc_us-gaap_RestrictedCashAndCashEquivalentsCashAndCashEquivalentsMember_1121BC8ACBB23BEF139943AEDC329CAC" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_LetterofCreditCollateralMember" xlink:label="loc_autl_LetterofCreditCollateralMember_B1FE39CE32B64D387EAB43AEDC33CE83" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_RestrictedCashAndCashEquivalentsCashAndCashEquivalentsMember_1121BC8ACBB23BEF139943AEDC329CAC" xlink:to="loc_autl_LetterofCreditCollateralMember_B1FE39CE32B64D387EAB43AEDC33CE83" xlink:type="arc" />
    <link:loc xlink:href="autl-20190630.xsd#autl_SecurityDepositMember" xlink:label="loc_autl_SecurityDepositMember_AFDDEA436BFD945C5E3443AEDC330588" xlink:type="locator" />
    <link:definitionArc order="2" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_RestrictedCashAndCashEquivalentsCashAndCashEquivalentsMember_1121BC8ACBB23BEF139943AEDC329CAC" xlink:to="loc_autl_SecurityDepositMember_AFDDEA436BFD945C5E3443AEDC330588" xlink:type="arc" />
    <link:loc xlink:href="http://xbrl.fasb.org/us-gaap/2019/elts/us-gaap-2019-01-31.xsd#us-gaap_RestrictedCashCurrent" xlink:label="loc_us-gaap_RestrictedCashCurrent_66707B3E91297F30AA5C43AEDC335CA2" xlink:type="locator" />
    <link:definitionArc order="1" xlink:arcrole="http://xbrl.org/int/dim/arcrole/domain-member" xlink:from="loc_us-gaap_RestrictedCashAndCashEquivalentsItemsLineItems_4D21C5473EA802B7C7BA43AEDC335743" xlink:to="loc_us-gaap_RestrictedCashCurrent_66707B3E91297F30AA5C43AEDC335CA2" xlink:type="arc" />
  </link:definitionLink>
  <link:definitionLink xlink:role="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesSegmentInformationDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesShareBasedCompensationDetails" xlink:type="extended" />
  <link:definitionLink xlink:role="http://www.autolus.com/role/SummaryOfSignificantAccountingPoliciesTables" xlink:type="extended" />
</link:linkbase>





