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. 1994 Jan;68(1):233–239. doi: 10.1128/jvi.68.1.233-239.1994

Generation and characterization of a human immunodeficiency virus type 1 (HIV-1) mutant resistant to an HIV-1 protease inhibitor.

M A el-Farrash 1, M J Kuroda 1, T Kitazaki 1, T Masuda 1, K Kato 1, M Hatanaka 1, S Harada 1
PMCID: PMC236282  PMID: 8254733

Abstract

A synthetic peptide, RPI 312, that specifically inhibits the protease of the human immunodeficiency virus type 1 (HIV-1) showed a potent inhibition on virus production, maturation, and infectivity. Treatment with this agent prevented the cleavage of Gag protein at the site between p17 and p24 in HIV-1 chronically infected MOLT-4 cells as well as in the released virus. Passage of HIV-1 in the presence of gradually increasing concentrations of this protease inhibitor resulted in emergence of a variant that could evade the drug effects. In the resistant variant the maturation of Gag proteins appeared normal, but its infectivity was reduced compared with that of the parent virus. The nucleotides coding the amino acids at and around the cleavage site between Gag proteins p17 and p24 were not changed. One point mutation (A-->G) at site 2082 of the pol gene that resulted in one amino acid change at site 84 of the protease from isoleucine to valine (I-84-->V) could be detected in the resistant variant. An HIV-1 infectious DNA clone with the I-84-->V mutation also showed reduced sensitivity to this protease inhibitor. The findings that the resistant variant had lower infectivity and was still affected by higher doses of the drug support the speculation that resistance to protease inhibitors may not be as problematic as other drug resistance.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Adachi A., Gendelman H. E., Koenig S., Folks T., Willey R., Rabson A., Martin M. A. Production of acquired immunodeficiency syndrome-associated retrovirus in human and nonhuman cells transfected with an infectious molecular clone. J Virol. 1986 Aug;59(2):284–291. doi: 10.1128/jvi.59.2.284-291.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Gao Q., Gu Z. X., Parniak M. A., Li X. G., Wainberg M. A. In vitro selection of variants of human immunodeficiency virus type 1 resistant to 3'-azido-3'-deoxythymidine and 2',3'-dideoxyinosine. J Virol. 1992 Jan;66(1):12–19. doi: 10.1128/jvi.66.1.12-19.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Gingeras T. R., Prodanovich P., Latimer T., Guatelli J. C., Richman D. D., Barringer K. J. Use of self-sustained sequence replication amplification reaction to analyze and detect mutations in zidovudine-resistant human immunodeficiency virus. J Infect Dis. 1991 Dec;164(6):1066–1074. doi: 10.1093/infdis/164.6.1066. [DOI] [PubMed] [Google Scholar]
  4. Graham F. L., van der Eb A. J. A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology. 1973 Apr;52(2):456–467. doi: 10.1016/0042-6822(73)90341-3. [DOI] [PubMed] [Google Scholar]
  5. Gyotoku J., el-Farrash M. A., Fujimoto S., Germeraad W. T., Watanabe Y., Teshigawara K., Harada S., Katsura Y. Inhibition of human immunodeficiency virus replication in a human T cell line by antisense RNA expressed in the cell. Virus Genes. 1991 Jul;5(3):189–202. doi: 10.1007/BF00568969. [DOI] [PubMed] [Google Scholar]
  6. Harada S., Koyanagi Y., Yamamoto N. Infection of HTLV-III/LAV in HTLV-I-carrying cells MT-2 and MT-4 and application in a plaque assay. Science. 1985 Aug 9;229(4713):563–566. doi: 10.1126/science.2992081. [DOI] [PubMed] [Google Scholar]
  7. Hattori T., Sagawa K., Matsushita S., Koito A., Suto H., Matsuoka M., Yokoyama M., Takatsuki K. Characterization of three monoclonal antibodies (VAK3-5) that identify p24, core protein of human immunodeficiency virus, and its precursors. Jpn J Cancer Res. 1987 Mar;78(3):235–241. [PubMed] [Google Scholar]
  8. Kohl N. E., Emini E. A., Schleif W. A., Davis L. J., Heimbach J. C., Dixon R. A., Scolnick E. M., Sigal I. S. Active human immunodeficiency virus protease is required for viral infectivity. Proc Natl Acad Sci U S A. 1988 Jul;85(13):4686–4690. doi: 10.1073/pnas.85.13.4686. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Koyanagi Y., Harada S., Yamamoto N. Establishment of a high production system for AIDS retroviruses with a human T-leukemic cell line Molt-4. Cancer Lett. 1986 Mar;30(3):299–310. doi: 10.1016/0304-3835(86)90054-6. [DOI] [PubMed] [Google Scholar]
  10. Larder B. A., Kemp S. D., Purifoy D. J. Infectious potential of human immunodeficiency virus type 1 reverse transcriptase mutants with altered inhibitor sensitivity. Proc Natl Acad Sci U S A. 1989 Jul;86(13):4803–4807. doi: 10.1073/pnas.86.13.4803. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Masuda T., Matsushita S., Kuroda M. J., Kannagi M., Takatsuki K., Harada S. Generation of neutralization-resistant HIV-1 in vitro due to amino acid interchanges of third hypervariable env region. J Immunol. 1990 Nov 15;145(10):3240–3246. [PubMed] [Google Scholar]
  12. McKeating J. A., Gow J., Goudsmit J., Pearl L. H., Mulder C., Weiss R. A. Characterization of HIV-1 neutralization escape mutants. AIDS. 1989 Dec;3(12):777–784. doi: 10.1097/00002030-198912000-00001. [DOI] [PubMed] [Google Scholar]
  13. McQuade T. J., Tomasselli A. G., Liu L., Karacostas V., Moss B., Sawyer T. K., Heinrikson R. L., Tarpley W. G. A synthetic HIV-1 protease inhibitor with antiviral activity arrests HIV-like particle maturation. Science. 1990 Jan 26;247(4941):454–456. doi: 10.1126/science.2405486. [DOI] [PubMed] [Google Scholar]
  14. Meek T. D., Lambert D. M., Dreyer G. B., Carr T. J., Tomaszek T. A., Jr, Moore M. L., Strickler J. E., Debouck C., Hyland L. J., Matthews T. J. Inhibition of HIV-1 protease in infected T-lymphocytes by synthetic peptide analogues. Nature. 1990 Jan 4;343(6253):90–92. doi: 10.1038/343090a0. [DOI] [PubMed] [Google Scholar]
  15. Mervis R. J., Ahmad N., Lillehoj E. P., Raum M. G., Salazar F. H., Chan H. W., Venkatesan S. The gag gene products of human immunodeficiency virus type 1: alignment within the gag open reading frame, identification of posttranslational modifications, and evidence for alternative gag precursors. J Virol. 1988 Nov;62(11):3993–4002. doi: 10.1128/jvi.62.11.3993-4002.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Mitsuya H., Yarchoan R., Broder S. Molecular targets for AIDS therapy. Science. 1990 Sep 28;249(4976):1533–1544. doi: 10.1126/science.1699273. [DOI] [PubMed] [Google Scholar]
  17. Nunberg J. H., Schleif W. A., Boots E. J., O'Brien J. A., Quintero J. C., Hoffman J. M., Emini E. A., Goldman M. E. Viral resistance to human immunodeficiency virus type 1-specific pyridinone reverse transcriptase inhibitors. J Virol. 1991 Sep;65(9):4887–4892. doi: 10.1128/jvi.65.9.4887-4892.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Otto M. J., Garber S., Winslow D. L., Reid C. D., Aldrich P., Jadhav P. K., Patterson C. E., Hodge C. N., Cheng Y. S. In vitro isolation and identification of human immunodeficiency virus (HIV) variants with reduced sensitivity to C-2 symmetrical inhibitors of HIV type 1 protease. Proc Natl Acad Sci U S A. 1993 Aug 15;90(16):7543–7547. doi: 10.1073/pnas.90.16.7543. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Overton H. A., McMillan D. J., Gridley S. J., Brenner J., Redshaw S., Mills J. S. Effect of two novel inhibitors of the human immunodeficiency virus protease on the maturation of the HIV gag and gag-pol polyproteins. Virology. 1990 Nov;179(1):508–511. doi: 10.1016/0042-6822(90)90326-m. [DOI] [PubMed] [Google Scholar]
  20. Pauwels R., Balzarini J., Baba M., Snoeck R., Schols D., Herdewijn P., Desmyter J., De Clercq E. Rapid and automated tetrazolium-based colorimetric assay for the detection of anti-HIV compounds. J Virol Methods. 1988 Aug;20(4):309–321. doi: 10.1016/0166-0934(88)90134-6. [DOI] [PubMed] [Google Scholar]
  21. Popovic M., Sarngadharan M. G., Read E., Gallo R. C. Detection, isolation, and continuous production of cytopathic retroviruses (HTLV-III) from patients with AIDS and pre-AIDS. Science. 1984 May 4;224(4648):497–500. doi: 10.1126/science.6200935. [DOI] [PubMed] [Google Scholar]
  22. Prasad V. R., Lowy I., de los Santos T., Chiang L., Goff S. P. Isolation and characterization of a dideoxyguanosine triphosphate-resistant mutant of human immunodeficiency virus reverse transcriptase. Proc Natl Acad Sci U S A. 1991 Dec 15;88(24):11363–11367. doi: 10.1073/pnas.88.24.11363. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Ratner L., Haseltine W., Patarca R., Livak K. J., Starcich B., Josephs S. F., Doran E. R., Rafalski J. A., Whitehorn E. A., Baumeister K. Complete nucleotide sequence of the AIDS virus, HTLV-III. Nature. 1985 Jan 24;313(6000):277–284. doi: 10.1038/313277a0. [DOI] [PubMed] [Google Scholar]
  24. Richman D. D., Guatelli J. C., Grimes J., Tsiatis A., Gingeras T. Detection of mutations associated with zidovudine resistance in human immunodeficiency virus by use of the polymerase chain reaction. J Infect Dis. 1991 Dec;164(6):1075–1081. doi: 10.1093/infdis/164.6.1075. [DOI] [PubMed] [Google Scholar]
  25. Richman D., Shih C. K., Lowy I., Rose J., Prodanovich P., Goff S., Griffin J. Human immunodeficiency virus type 1 mutants resistant to nonnucleoside inhibitors of reverse transcriptase arise in tissue culture. Proc Natl Acad Sci U S A. 1991 Dec 15;88(24):11241–11245. doi: 10.1073/pnas.88.24.11241. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Roberts N. A., Martin J. A., Kinchington D., Broadhurst A. V., Craig J. C., Duncan I. B., Galpin S. A., Handa B. K., Kay J., Kröhn A. Rational design of peptide-based HIV proteinase inhibitors. Science. 1990 Apr 20;248(4953):358–361. doi: 10.1126/science.2183354. [DOI] [PubMed] [Google Scholar]
  27. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Szelke M., Leckie B., Hallett A., Jones D. M., Sueiras J., Atrash B., Lever A. F. Potent new inhibitors of human renin. Nature. 1982 Oct 7;299(5883):555–557. doi: 10.1038/299555a0. [DOI] [PubMed] [Google Scholar]

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