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

    <schemaVersion>X0206</schemaVersion>

    <documentType>3</documentType>

    <periodOfReport>2018-09-25</periodOfReport>

    <noSecuritiesOwned>0</noSecuritiesOwned>

    <issuer>
        <issuerCik>0001724344</issuerCik>
        <issuerName>Entasis Therapeutics Holdings Inc.</issuerName>
        <issuerTradingSymbol>ETTX</issuerTradingSymbol>
    </issuer>

    <reportingOwner>
        <reportingOwnerId>
            <rptOwnerCik>0001388325</rptOwnerCik>
            <rptOwnerName>Novo Holdings A/S</rptOwnerName>
        </reportingOwnerId>
        <reportingOwnerAddress>
            <rptOwnerStreet1>TUBORG HAVNEVEJ 19</rptOwnerStreet1>
            <rptOwnerStreet2></rptOwnerStreet2>
            <rptOwnerCity>HELLERUP</rptOwnerCity>
            <rptOwnerState>G7</rptOwnerState>
            <rptOwnerZipCode>2900</rptOwnerZipCode>
            <rptOwnerStateDescription>DENMARK</rptOwnerStateDescription>
        </reportingOwnerAddress>
        <reportingOwnerRelationship>
            <isDirector>0</isDirector>
            <isOfficer>0</isOfficer>
            <isTenPercentOwner>1</isTenPercentOwner>
            <isOther>0</isOther>
        </reportingOwnerRelationship>
    </reportingOwner>

    <derivativeTable>
        <derivativeHolding>
            <securityTitle>
                <value>Series B Preferred Stock</value>
            </securityTitle>
            <conversionOrExercisePrice>
                <footnoteId id="F1"/>
            </conversionOrExercisePrice>
            <exerciseDate>
                <footnoteId id="F1"/>
            </exerciseDate>
            <expirationDate>
                <footnoteId id="F1"/>
            </expirationDate>
            <underlyingSecurity>
                <underlyingSecurityTitle>
                    <value>Common Stock</value>
                </underlyingSecurityTitle>
                <underlyingSecurityShares>
                    <value>7000000</value>
                </underlyingSecurityShares>
            </underlyingSecurity>
            <ownershipNature>
                <directOrIndirectOwnership>
                    <value>D</value>
                </directOrIndirectOwnership>
            </ownershipNature>
        </derivativeHolding>
        <derivativeHolding>
            <securityTitle>
                <value>Series B-1 Preferred Stock</value>
            </securityTitle>
            <conversionOrExercisePrice>
                <footnoteId id="F1"/>
            </conversionOrExercisePrice>
            <exerciseDate>
                <footnoteId id="F1"/>
            </exerciseDate>
            <expirationDate>
                <footnoteId id="F1"/>
            </expirationDate>
            <underlyingSecurity>
                <underlyingSecurityTitle>
                    <value>Common Stock</value>
                </underlyingSecurityTitle>
                <underlyingSecurityShares>
                    <value>14237288</value>
                </underlyingSecurityShares>
            </underlyingSecurity>
            <ownershipNature>
                <directOrIndirectOwnership>
                    <value>D</value>
                </directOrIndirectOwnership>
            </ownershipNature>
        </derivativeHolding>
    </derivativeTable>

    <footnotes>
        <footnote id="F1">Series B and B-1 redeemable convertible preferred stock (collectively, the &quot;Preferred Stock&quot;) has no expiration date and is convertible at any time, at the holder's election. Represents the number of shares of preferred stock held. Upon the closing of the Issuer's initial public offering, the Preferred Stock will automatically convert into shares of the Issuer's Common Stock based on the Series B Preferred Stock Conversion Price, Series B-1 Tranche A Preferred Stock Conversion Price and Series B-1 Tranche B Preferred Stock Conversion Price (each as defined in the Issuer's Amended and Restated Certificate of Incorporation).</footnote>
    </footnotes>

    <remarks>Novo Holdings A/S is a Danish limited liability company.  The board of directors of Novo Holdings A/S (the &quot;Novo Board&quot;), which is currently comprised of Lars Rebien Sorensen, Steen Riisgaard, Francis Cuss, Jean-Luc Butel, Jeppe Christiansen and Vivian Monges, has shared investment and voting control over the securities of the Issuer held by Novo Holdings A/S (the &quot;Shares&quot;) and may exercise such control only with the support of a majority of the Novo Board.  As such, no individual member of the Novo Board is deemed to hold any beneficial ownership or reportable pecuniary interest in the Shares.</remarks>

    <ownerSignature>
        <signatureName>/s/ Peter Haahr, Chief Financial Officer of Novo Holdings A/S</signatureName>
        <signatureDate>2018-09-25</signatureDate>
    </ownerSignature>
</ownershipDocument>
