<?xml version="1.0"?>
<ownershipDocument>

    <schemaVersion>X0206</schemaVersion>

    <documentType>3</documentType>

    <periodOfReport>2016-04-06</periodOfReport>

    <noSecuritiesOwned>0</noSecuritiesOwned>

    <issuer>
        <issuerCik>0001636282</issuerCik>
        <issuerName>Aeglea BioTherapeutics, Inc.</issuerName>
        <issuerTradingSymbol>AGLE</issuerTradingSymbol>
    </issuer>

    <reportingOwner>
        <reportingOwnerId>
            <rptOwnerCik>0001655962</rptOwnerCik>
            <rptOwnerName>York Charles N II</rptOwnerName>
        </reportingOwnerId>
        <reportingOwnerAddress>
            <rptOwnerStreet1>C/O AEGLEA BIOTHERAPEUTICS, INC.</rptOwnerStreet1>
            <rptOwnerStreet2>901 S. MOPAC EXPRESSWAY, SUITE 250</rptOwnerStreet2>
            <rptOwnerCity>AUSTIN</rptOwnerCity>
            <rptOwnerState>TX</rptOwnerState>
            <rptOwnerZipCode>78746</rptOwnerZipCode>
            <rptOwnerStateDescription></rptOwnerStateDescription>
        </reportingOwnerAddress>
        <reportingOwnerRelationship>
            <isDirector>0</isDirector>
            <isOfficer>1</isOfficer>
            <isTenPercentOwner>0</isTenPercentOwner>
            <isOther>0</isOther>
            <officerTitle>Chief Financial Officer and VP</officerTitle>
            <otherText></otherText>
        </reportingOwnerRelationship>
    </reportingOwner>

    <nonDerivativeTable>
        <nonDerivativeHolding>
            <securityTitle>
                <value>Common Stock</value>
            </securityTitle>
            <postTransactionAmounts>
                <sharesOwnedFollowingTransaction>
                    <value>7504</value>
                </sharesOwnedFollowingTransaction>
            </postTransactionAmounts>
            <ownershipNature>
                <directOrIndirectOwnership>
                    <value>D</value>
                </directOrIndirectOwnership>
            </ownershipNature>
        </nonDerivativeHolding>
    </nonDerivativeTable>

    <derivativeTable>
        <derivativeHolding>
            <securityTitle>
                <value>Series A Preferred Stock</value>
            </securityTitle>
            <conversionOrExercisePrice>
                <value>0</value>
                <footnoteId id="F1"/>
            </conversionOrExercisePrice>
            <exerciseDate>
                <footnoteId id="F1"/>
            </exerciseDate>
            <expirationDate>
                <footnoteId id="F1"/>
            </expirationDate>
            <underlyingSecurity>
                <underlyingSecurityTitle>
                    <value>Common Stock</value>
                </underlyingSecurityTitle>
                <underlyingSecurityShares>
                    <value>9523</value>
                </underlyingSecurityShares>
            </underlyingSecurity>
            <ownershipNature>
                <directOrIndirectOwnership>
                    <value>I</value>
                </directOrIndirectOwnership>
                <natureOfOwnership>
                    <value>By IRA</value>
                </natureOfOwnership>
            </ownershipNature>
        </derivativeHolding>
        <derivativeHolding>
            <securityTitle>
                <value>Series B Preferred Stock</value>
            </securityTitle>
            <conversionOrExercisePrice>
                <value>0</value>
                <footnoteId id="F1"/>
            </conversionOrExercisePrice>
            <exerciseDate>
                <footnoteId id="F1"/>
            </exerciseDate>
            <expirationDate>
                <footnoteId id="F1"/>
            </expirationDate>
            <underlyingSecurity>
                <underlyingSecurityTitle>
                    <value>Common Stock</value>
                </underlyingSecurityTitle>
                <underlyingSecurityShares>
                    <value>2037</value>
                </underlyingSecurityShares>
            </underlyingSecurity>
            <ownershipNature>
                <directOrIndirectOwnership>
                    <value>I</value>
                </directOrIndirectOwnership>
                <natureOfOwnership>
                    <value>By IRA</value>
                </natureOfOwnership>
            </ownershipNature>
        </derivativeHolding>
        <derivativeHolding>
            <securityTitle>
                <value>Series B Preferred Stock</value>
            </securityTitle>
            <conversionOrExercisePrice>
                <value>0</value>
                <footnoteId id="F1"/>
            </conversionOrExercisePrice>
            <exerciseDate>
                <footnoteId id="F1"/>
            </exerciseDate>
            <expirationDate>
                <footnoteId id="F1"/>
            </expirationDate>
            <underlyingSecurity>
                <underlyingSecurityTitle>
                    <value>Common Stock</value>
                </underlyingSecurityTitle>
                <underlyingSecurityShares>
                    <value>1324</value>
                </underlyingSecurityShares>
            </underlyingSecurity>
            <ownershipNature>
                <directOrIndirectOwnership>
                    <value>D</value>
                </directOrIndirectOwnership>
            </ownershipNature>
        </derivativeHolding>
        <derivativeHolding>
            <securityTitle>
                <value>Employee Stock Option (right to buy)</value>
            </securityTitle>
            <conversionOrExercisePrice>
                <value>3.465</value>
            </conversionOrExercisePrice>
            <exerciseDate>
                <footnoteId id="F2"/>
            </exerciseDate>
            <expirationDate>
                <value>2025-03-31</value>
            </expirationDate>
            <underlyingSecurity>
                <underlyingSecurityTitle>
                    <value>Common Stock</value>
                </underlyingSecurityTitle>
                <underlyingSecurityShares>
                    <value>10183</value>
                </underlyingSecurityShares>
            </underlyingSecurity>
            <ownershipNature>
                <directOrIndirectOwnership>
                    <value>D</value>
                </directOrIndirectOwnership>
            </ownershipNature>
        </derivativeHolding>
        <derivativeHolding>
            <securityTitle>
                <value>Employee Stock Option (right to buy)</value>
            </securityTitle>
            <conversionOrExercisePrice>
                <value>3.465</value>
            </conversionOrExercisePrice>
            <exerciseDate>
                <footnoteId id="F3"/>
            </exerciseDate>
            <expirationDate>
                <value>2025-03-31</value>
            </expirationDate>
            <underlyingSecurity>
                <underlyingSecurityTitle>
                    <value>Common Stock</value>
                </underlyingSecurityTitle>
                <underlyingSecurityShares>
                    <value>22258</value>
                </underlyingSecurityShares>
            </underlyingSecurity>
            <ownershipNature>
                <directOrIndirectOwnership>
                    <value>D</value>
                </directOrIndirectOwnership>
            </ownershipNature>
        </derivativeHolding>
        <derivativeHolding>
            <securityTitle>
                <value>Employee Stock Option (right to buy)</value>
            </securityTitle>
            <conversionOrExercisePrice>
                <value>3.465</value>
            </conversionOrExercisePrice>
            <exerciseDate>
                <footnoteId id="F4"/>
            </exerciseDate>
            <expirationDate>
                <value>2025-03-31</value>
            </expirationDate>
            <underlyingSecurity>
                <underlyingSecurityTitle>
                    <value>Common Stock</value>
                </underlyingSecurityTitle>
                <underlyingSecurityShares>
                    <value>15027</value>
                </underlyingSecurityShares>
            </underlyingSecurity>
            <ownershipNature>
                <directOrIndirectOwnership>
                    <value>D</value>
                </directOrIndirectOwnership>
            </ownershipNature>
        </derivativeHolding>
    </derivativeTable>

    <footnotes>
        <footnote id="F1">Each share of Series A Preferred Stock and Series B Preferred Stock will be automatically converted into one (1) share of Common Stock, for no additional consideration, immediately prior to the consummation of the Issuer's initial public offering, and has no expiration date.</footnote>
        <footnote id="F2">The stock option vested and became exercisable as to 25% of the shares subject to the option on July 16, 2015, and thereafter vests as to 1/16th of the shares at the end of each three month period in equal installments, until such time as the option is 100% vested, subject to the continuing employment of the Reporting Person on each vesting date.</footnote>
        <footnote id="F3">The stock option vested and became exercisable as to 25% of the shares subject to the option on March 20, 2016, and thereafter vests as to 1/16th of the shares at the end of each three month period in equal installments, until such time as the option is 100% vested, subject to the continuing employment of the Reporting Person on each vesting date.</footnote>
        <footnote id="F4">The stock option vests and becomes exercisable as to 25% of the shares subject to the option on March 20, 2017, and thereafter vests as to 1/16th of the shares at the end of each three month period in equal installments, until such time as the option is 100% vested, subject to the continuing employment of the Reporting Person on each vesting date.</footnote>
    </footnotes>

    <ownerSignature>
        <signatureName>/s/ Charles N. York II</signatureName>
        <signatureDate>2016-04-06</signatureDate>
    </ownerSignature>
</ownershipDocument>
