As filed with the Securities and Exchange Commission on April 1, 2024

Registration No. 333-

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UNITED STATES

SECURITIES AND EXCHANGE COMMISSION

Washington, D.C. 20549

FORM S-8

REGISTRATION STATEMENT

UNDER

THE SECURITIES ACT OF 1933

Xilio Therapeutics, Inc.

(Exact Name of Registrant as Specified in Its Charter)

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2021 Stock Incentive Plan

2021 Employee Stock Purchase Plan

(Full Title of the Plan)

René Russo

President and Chief Executive Officer

Xilio Therapeutics, Inc.

828 Winter Street, Suite 300

Waltham, Massachusetts 02451

(Name and Address of Agent for Service)

(857) 524-2466

(Telephone Number, Including Area Code, of Agent for Service)

Copy to:

Christopher Frankenfield

Chief Operating Officer

Xilio Therapeutics, Inc.

828 Winter Street, Suite 300

Waltham, Massachusetts 02451

(857) 524-2466

Indicate by check mark whether the registrant is a large accelerated filer, an accelerated filer, a non-accelerated filer, a smaller reporting company, or an emerging growth company. See the definitions of “large accelerated filer,” “accelerated filer,” “smaller reporting company,” and “emerging growth company” in Rule 12b-2 of the Exchange Act.

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If an emerging growth company, indicate by check mark if the registrant has elected not to use the extended transition period for complying with any new or revised financial accounting standards provided pursuant to Section 7(a)(2)(B) of the Securities Act. ☐

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Statement of Incorporation by Reference

This Registration Statement on Form S 8, relating to the 2021 Stock Incentive Plan (the “2021 Plan”) and the 2021 Employee Stock Purchase Plan (the “2021 ESPP”) of Xilio Therapeutics, Inc. (the “Registrant”), is being filed for the purpose of registering additional securities of the same class as other securities for which a Registration Statement on Form S-8 has previously been filed and is effective. Accordingly, this Registration Statement incorporates by reference the contents of (i) the Registration Statement on Form S 8 (File No. 333-260460) filed with the Securities and Exchange Commission (“SEC”) on October 25, 2021 by the Registrant relating to the 2021 Plan and the 2021 ESPP, (ii) the Registration Statement on Form S 8 (File No. 333-263178) filed with the SEC on March 1, 2022 by the Registrant relating to the 2021 Plan and the 2021 ESPP, and (iii) the Registration Statement on Form S-8 (File No. 333-270240) filed with the SEC on March 2, 2023 by the Registrant relating to the 2021 Plan and the 2021 ESPP, except in each case, for “Item 8. Exhibits” with respect to which the exhibit index set forth below is incorporated herein by reference.

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Item 8.Exhibits.

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The following exhibits are incorporated herein by reference:

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SIGNATURES

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Pursuant to the requirements of the Securities Act of 1933, as amended, the registrant certifies that it has reasonable grounds to believe that it meets all of the requirements for filing on Form S-8 and has duly caused this registration statement to be signed on its behalf by the undersigned, thereunto duly authorized, in the City of Waltham, Commonwealth of Massachusetts on this 1st day of April, 2024.

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POWER OF ATTORNEY AND SIGNATURES

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We, the undersigned officers and directors of Xilio Therapeutics, Inc., hereby severally constitute and appoint René Russo and Chris Frankenfield, and each of them singly (with full power to each of them to act alone), our true and lawful attorneys with full power of substitution and resubstitution in each of them for him and in his name, place and stead, and in any and all capacities, to sign for us and in our names in the capacities indicated below, the registration statement on Form S-8 filed herewith and any and all subsequent amendments to said registration statement, and generally to do all such things in our names and on our behalf in our capacities as officers and directors to enable Xilio Therapeutics, Inc. to comply with the provisions of the Securities Act of 1933, as amended, and all requirements of the Securities and Exchange Commission, hereby ratifying and confirming our signatures as they may be signed by our said attorneys, or any of them, to said registration statement and any and all amendments thereto.

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Pursuant to the requirements of the Securities Act of 1933, as amended, this registration statement has been signed by the following persons in the capacities and on the dates indicated.

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Exhibit 5.1

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April 1, 2024

Xilio Therapeutics, Inc.

828 Winter Street, Suite 300

Waltham, Massachusetts 02451

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Xilio Therapeutics, Inc. 2021 Employee Stock Purchase Plan

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Ladies and Gentlemen:

We have assisted in the preparation of a Registration Statement on Form S-8 (the “Registration Statement”) to be filed with the Securities and Exchange Commission (the “Commission”) under the Securities Act of 1933, as amended (the “Securities Act”), relating to an aggregate of 1,656,795 shares of common stock, $0.0001 par value per share (the “Shares”), of Xilio Therapeutics, Inc., a Delaware corporation (the “Company”), of which (i) 1,380,663 shares are issuable under the Company’s 2021 Stock Incentive Plan (the “SIP”) and (ii) 276,132 shares are issuable under the Company’s 2021 Employee Stock Purchase Plan (the “ESPP,” and, together with the SIP, the “Plans”).

We have examined the Certificate of Incorporation and By-Laws of the Company, each as amended and restated to date, and originals, or copies certified to our satisfaction, of all pertinent records of the meetings of the board of directors and stockholders of the Company, the Registration Statement, the Plans and such other documents relating to the Company as we have deemed material for the purposes of this opinion.

In our examination of the foregoing documents, we have assumed the genuineness of all signatures, the authenticity of all documents submitted to us as originals, the conformity to original documents of all documents submitted to us as certified, photostatic or other copies, the authenticity of the originals of any such documents and the legal competence of all signatories to such documents.

We assume that the appropriate action will be taken, prior to the offer and sale of the Shares in accordance with the Plans, to register and qualify the Shares for sale under all applicable state securities or “blue sky” laws.

We express no opinion herein as to the laws of any state or jurisdiction other than the General Corporation Law of the State of Delaware.

It is understood that this opinion is to be used only in connection with the offer and sale of the Shares while the Registration Statement is in effect.

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April 1, 2024

Page 2

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Please note that we are opining only as to the matters expressly set forth herein, and no opinion should be inferred as to any other matters.

Based on the foregoing, we are of the opinion that the Shares have been duly authorized for issuance and, when the Shares are issued and paid for in accordance with the terms and conditions of the Plans, the Shares will be validly issued, fully paid and nonassessable.

We hereby consent to the filing of this opinion with the Commission in connection with the Registration Statement in accordance with the requirements of Item 601(b)(5) of Regulation S-K under the Securities Act. In giving such consent, we do not hereby admit that we are in the category of persons whose consent is required under Section 7 of the Securities Act or the rules and regulations of the Commission.

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Exhibit 23.1

Consent of Independent Registered Public Accounting Firm

We consent to the incorporation by reference in the Registration Statement (Form S-8) pertaining to the 2021 Stock Incentive Plan and the 2021 Employee Stock Purchase Plan of Xilio Therapeutics, Inc. of our report dated April 1, 2024, with respect to the consolidated financial statements of Xilio Therapeutics, Inc. included in its Annual Report (Form 10-K) for the year ended December 31, 2023, filed with the Securities and Exchange Commission.

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Exhibit 107

Calculation of Filing Fee Tables

Table 1—Newly Registered Securities

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