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Covalent inhibitors for eradication of drug-resistant HIV-1 reverse transcriptase: From design to protein crystallography

  1. William L. Jorgensenc,1
  1. aDepartment of Pharmacology, Yale University School of Medicine, New Haven, CT 06520-8066;
  2. bDepartment of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, CT 06520-8066;
  3. cDepartment of Chemistry, Yale University, New Haven, CT 06520-8107
  1. Contributed by William L. Jorgensen, July 26, 2017 (sent for review June 26, 2017; reviewed by Maurizio Botta and Dennis C. Liotta)


HIV-1 reverse transcriptase (RT) has been the prime target for anti-HIV chemotherapy; however, its rapid mutation often generates drug resistance. Prominent variant strains of HIV-1 that lead to treatment failure with nonnucleoside RT inhibitors (NNRTIs) bear the Tyr181Cys mutation in RT. Based on our previous discovery and crystallography for potent noncovalent NNRTIs, new compounds were designed with incorporation of chemical warheads intended to modify covalently Cys181. Here we report on the success of the strategy, including biochemical, biophysical, and cellular evidence of the desired irreversible covalent inhibition. The new compounds completely eliminate the activity of Cys181-bearing RT, and it may be possible to dose them less frequently than noncovalent inhibitors.


Development of resistance remains a major challenge for drugs to treat HIV-1 infections, including those targeting the essential viral polymerase, HIV-1 reverse transcriptase (RT). Resistance associated with the Tyr181Cys mutation in HIV-1 RT has been a key roadblock in the discovery of nonnucleoside RT inhibitors (NNRTIs). It is the principal point mutation that arises from treatment of HIV-infected patients with nevirapine, the first-in-class drug still widely used, especially in developing countries. We report covalent inhibitors of Tyr181Cys RT (CRTIs) that can completely knock out activity of the resistant mutant and of the particularly challenging Lys103Asn/Tyr181Cys variant. Conclusive evidence for the covalent modification of Cys181 is provided from enzyme inhibition kinetics, mass spectrometry, protein crystallography, and antiviral activity in infected human T-cell assays. The CRTIs are also shown to be selective for Cys181 and have lower cytotoxicity than the approved NNRTI drugs efavirenz and rilpivirine.


  • ?1To whom correspondence may be addressed. Email: karen.anderson{at}yale.edu or william.jorgensen{at}yale.edu.
  • Author contributions: K.S.A. and W.L.J. designed research; A.H.C., W.-G.L., K.A.S., J.A.C., S.N.K., Z.O.P., A.B.B., and W.L.J. performed research; A.H.C., W.-G.L., K.A.S., J.A.C., S.N.K., Z.O.P., A.B.B., K.S.A., and W.L.J. analyzed data; and A.H.C., K.S.A., and W.L.J. wrote the paper.

  • Reviewers: M.B., University of Siena; and D.C.L., Emory University.

  • The authors declare no conflict of interest.

  • Data deposition: The atomic coordinates and structure factors have been deposited in the Protein Data Bank, www.wwpdb.org (PDB ID codes 5VQQ, 5VQR, 5VQS, 5VQT, 5VQU, 5VQV, 5VQW, 5VQX, 5VQY, and 5VQZ).

  • This article contains supporting information online at www.danielhellerman.com/lookup/suppl/doi:10.1073/pnas.1711463114/-/DCSupplemental.

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