THIRD AMENDMENT TO THE SECOND AMENDED AND RESTATED STOCKHOLDERS AGREEMENT

EXHIBIT 4.11

THIRD AMENDMENT TO THE

SECOND AMENDED AND RESTATED STOCKHOLDERS’ AGREEMENT

THIS THIRD AMENDMENT (the “Amendment”) TO THE SECOND AMENDED AND RESTATED STOCKHOLDERS’ AGREEMENT dated as of August 4, 2004, as amended (the “Stockholders’ Agreement”), is entered into as of June 1, 2006 by and among Pharmasset, Inc., a Delaware corporation (the “Company”), and the undersigned holders of the Company’s capital stock (collectively, the “Stockholders”). Capitalized terms used but not otherwise defined in this Amendment shall have the meanings set forth in the Stockholders’ Agreement.

RECITALS

WHEREAS, the Board of Directors of the Company has approved an amendment to the Company’s Equity Incentive Plan, which increases the number of shares available for grants by 1,600,000 additional shares of the Company’s Common Stock from 3,675,522 to 5,275,522 shares;

WHEREAS, a same increase is being made concurrently to the number of shares of Common Stock set forth in clause (ii) of the definition of “Excluded Securities” in the Certificate of Incorporation;

WHEREAS, the Company and the Stockholders desire to amend the definition of “New Shares” set forth in Section 1.1 of the Stockholders’ Agreement;

NOW, THEREFORE, in consideration of the mutual promises and covenants hereinafter set forth, the receipt, sufficiency and adequacy of which are hereby acknowledged, the parties hereto agree to amend the Stockholders’ Agreement as follows:

Definition. (a) The definition of the term “New Shares” set forth in Section 1.1 of the Stockholders’ Agreement is hereby amended by deleting clause (iii) thereof and replacing it in its entirety with the following:

“(iii) up to 5,275,522 shares of Common Stock (appropriately adjusted to take account of any stock split, stock dividend, combination of shares, recapitalization or other similar event) issuable to officers, directors, employees and consultants of the Company or a subsidiary pursuant to the exercise of options granted or shares of Common Stock directly issued under the Equity Incentive Plan, subject to vesting, and such options and other rights to acquire Common Stock granted thereunder;”

Governing Law. This Amendment shall be governed by and construed under the laws of the State of Delaware (without giving effect to any conflicts or choice of laws provisions which would cause the application of the domestic substantive laws of any other jurisdiction).

Effective Date. This Amendment shall become effective as of the date first written above (the “Effective Date”) upon receipt by the Company of counterparts of this Amendment duly executed by (i) the Majority Investors and (ii) the Stockholders holding a majority of the outstanding shares of the Common Stock that are subject to this Agreement and the Preferred Stock (on an as-if-converted basis) owned by such Stockholders. Each Stockholder shall be bound by this Amendment whether or not such Stockholder shall have consented to this Amendment.

Miscellaneous. From and after the Effective Date of this Amendment, each reference in the Stockholders’ Agreement to “this Agreement”, “hereof”, “hereunder”, or words of like import in any and all agreements, instruments, documents, notes, certificates and other writings of any kind and nature shall be deemed to refer to the Stockholders’ Agreement as amended by this Amendment. This Amendment may be executed in any number of counterparts, each of which shall be an original, but all of which together constitute one instrument. The headings in this Amendment are for purposes of reference only and shall not limit or otherwise effect the meaning hereof.

[Signatures on Following Pages]

2

IN WITNESS WHEREOF, the parties hereto have executed this Third Amendment to the Second Amended and Restated Stockholders’ Agreement as of the date set forth in the first paragraph hereof.

Signature Page to Third Amendment to

Second Amended and Restated Stockholders’ Agreement

Signature Page to Third Amendment to

Second Amended and Restated Stockholders’ Agreement

Signature Page to Third Amendment to

Second Amended and Restated Stockholders’ Agreement

Signature Page to Third Amendment to

Second Amended and Restated Stockholders’ Agreement

Signature Page to Third Amendment to

Second Amended and Restated Stockholders’ Agreement

Signature Page to Third Amendment to

Second Amended and Restated Stockholders’ Agreement

Signature Page to Third Amendment to

Second Amended and Restated Stockholders’ Agreement