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. 1996 Jun;40(6):1491–1497. doi: 10.1128/aac.40.6.1491

Human serum alpha 1 acid glycoprotein reduces uptake, intracellular concentration, and antiviral activity of A-80987, an inhibitor of the human immunodeficiency virus type 1 protease.

J A Bilello 1, P A Bilello 1, K Stellrecht 1, J Leonard 1, D W Norbeck 1, D J Kempf 1, T Robins 1, G L Drusano 1
PMCID: PMC163355  PMID: 8726025

Abstract

The therapeutic utility of a human immunodeficiency virus type 1 (HIV-1) protease inhibitor may depend on its intracellular concentration, which is a property of its uptake, metabolism, and/or efflux. Previous studies in our laboratory indicated that the addition of alpha 1 acid glycoprotein (alpha 1 AGP) to the medium markedly increased the amount of the drug required to limit infection in vitro. In this study, physiologically relevant concentrations of alpha 1 AGP and a radiolabeled inhibitor, A-80987, were used to determine both the uptake and activity of the agent in HIV-1-infected human peripheral blood mononuclear cells and cell lines. Both the uptake and efflux of 14C-labeled A-80987 were rapid (t1/2, < 5 min). Uptake of the drug was linearly dependent on the concentration but insensitive to the metabolic inhibitors KF, sodium cyanide, or CCCP (carbonyl cyanide m-chlorophenyl hydrazone). The amount of A-80987 which entered the cells was inversely proportional to the concentration of alpha 1 AGP (r2, 0.99) and directly proportional to the amount of extracellular non-protein-bound drug (r2, 0.99). Most importantly, the antiviral activity of the drug in HIV-1-infected peripheral blood mononuclear cells and MT-2 cells was directly related to the amount of intracellular A-80987. This study demonstrates that A-80987 binds to alpha 1 AGP, resulting in a free fraction below 10%. Cellular uptake of A-80987 is proportionally decreased in the presence of alpha 1 AGP, which results in less-than-expected antiviral activity. Importantly, we demonstrate for the first time that the inhibition of HIV protease is highly correlated with the amount of intracellular inhibitor.

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Selected References

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  1. Bilello J. A., Bauer G., Dudley M. N., Cole G. A., Drusano G. L. Effect of 2',3'-didehydro-3'-deoxythymidine in an in vitro hollow-fiber pharmacodynamic model system correlates with results of dose-ranging clinical studies. Antimicrob Agents Chemother. 1994 Jun;38(6):1386–1391. doi: 10.1128/aac.38.6.1386. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bilello J. A., Bilello P. A., Kort J. J., Dudley M. N., Leonard J., Drusano G. L. Efficacy of constant infusion of A-77003, an inhibitor of the human immunodeficiency virus type 1 (HIV-1) protease, in limiting acute HIV-1 infection in vitro. Antimicrob Agents Chemother. 1995 Nov;39(11):2523–2527. doi: 10.1128/aac.39.11.2523. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bilello J. A., Bilello P. A., Prichard M., Robins T., Drusano G. L. Reduction of the in vitro activity of A77003, an inhibitor of human immunodeficiency virus protease, by human serum alpha 1 acid glycoprotein. J Infect Dis. 1995 Mar;171(3):546–551. doi: 10.1093/infdis/171.3.546. [DOI] [PubMed] [Google Scholar]
  4. Debouck C., Gorniak J. G., Strickler J. E., Meek T. D., Metcalf B. W., Rosenberg M. Human immunodeficiency virus protease expressed in Escherichia coli exhibits autoprocessing and specific maturation of the gag precursor. Proc Natl Acad Sci U S A. 1987 Dec;84(24):8903–8906. doi: 10.1073/pnas.84.24.8903. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Dudley M. N., Blaser J., Gilbert D., Zinner S. H. Significance of "extravascular" protein binding for antimicrobial pharmacodynamics in an in vitro capillary model of infection. Antimicrob Agents Chemother. 1990 Jan;34(1):98–101. doi: 10.1128/aac.34.1.98. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Erickson J., Neidhart D. J., VanDrie J., Kempf D. J., Wang X. C., Norbeck D. W., Plattner J. J., Rittenhouse J. W., Turon M., Wideburg N. Design, activity, and 2.8 A crystal structure of a C2 symmetric inhibitor complexed to HIV-1 protease. Science. 1990 Aug 3;249(4968):527–533. doi: 10.1126/science.2200122. [DOI] [PubMed] [Google Scholar]
  7. Göttlinger H. G., Sodroski J. G., Haseltine W. A. Role of capsid precursor processing and myristoylation in morphogenesis and infectivity of human immunodeficiency virus type 1. Proc Natl Acad Sci U S A. 1989 Aug;86(15):5781–5785. doi: 10.1073/pnas.86.15.5781. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Holodniy M., Katzenstein D. A., Sengupta S., Wang A. M., Casipit C., Schwartz D. H., Konrad M., Groves E., Merigan T. C. Detection and quantification of human immunodeficiency virus RNA in patient serum by use of the polymerase chain reaction. J Infect Dis. 1991 Apr;163(4):862–866. doi: 10.1093/infdis/163.4.862. [DOI] [PubMed] [Google Scholar]
  9. Huff J. R. HIV protease: a novel chemotherapeutic target for AIDS. J Med Chem. 1991 Aug;34(8):2305–2314. doi: 10.1021/jm00112a001. [DOI] [PubMed] [Google Scholar]
  10. Kageyama S., Anderson B. D., Hoesterey B. L., Hayashi H., Kiso Y., Flora K. P., Mitsuya H. Protein binding of human immunodeficiency virus protease inhibitor KNI-272 and alteration of its in vitro antiretroviral activity in the presence of high concentrations of proteins. Antimicrob Agents Chemother. 1994 May;38(5):1107–1111. doi: 10.1128/aac.38.5.1107. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kempf D. J., Marsh K. C., Fino L. C., Bryant P., Craig-Kennard A., Sham H. L., Zhao C., Vasavanonda S., Kohlbrenner W. E., Wideburg N. E. Design of orally bioavailable, symmetry-based inhibitors of HIV protease. Bioorg Med Chem. 1994 Sep;2(9):847–858. doi: 10.1016/s0968-0896(00)82036-2. [DOI] [PubMed] [Google Scholar]
  12. Kempf D. J., Marsh K. C., Paul D. A., Knigge M. F., Norbeck D. W., Kohlbrenner W. E., Codacovi L., Vasavanonda S., Bryant P., Wang X. C. Antiviral and pharmacokinetic properties of C2 symmetric inhibitors of the human immunodeficiency virus type 1 protease. Antimicrob Agents Chemother. 1991 Nov;35(11):2209–2214. doi: 10.1128/aac.35.11.2209. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. 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]
  14. Kort J. J., Bilello J. A., Bauer G., Drusano G. L. Preclinical evaluation of antiviral activity and toxicity of Abbott A77003, an inhibitor of the human immunodeficiency virus type 1 protease. Antimicrob Agents Chemother. 1993 Jan;37(1):115–119. doi: 10.1128/aac.37.1.115. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kremer J. M., Wilting J., Janssen L. H. Drug binding to human alpha-1-acid glycoprotein in health and disease. Pharmacol Rev. 1988 Mar;40(1):1–47. [PubMed] [Google Scholar]
  16. Kräusslich H. G., Wimmer E. Viral proteinases. Annu Rev Biochem. 1988;57:701–754. doi: 10.1146/annurev.bi.57.070188.003413. [DOI] [PubMed] [Google Scholar]
  17. Loeb D. D., Hutchison C. A., 3rd, Edgell M. H., Farmerie W. G., Swanstrom R. Mutational analysis of human immunodeficiency virus type 1 protease suggests functional homology with aspartic proteinases. J Virol. 1989 Jan;63(1):111–121. doi: 10.1128/jvi.63.1.111-121.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Louis J. M., Wondrak E. M., Copeland T. D., Smith C. A., Mora P. T., Oroszlan S. Chemical synthesis and expression of the HIV-1 protease gene in E. coli. Biochem Biophys Res Commun. 1989 Feb 28;159(1):87–94. doi: 10.1016/0006-291x(89)92408-x. [DOI] [PubMed] [Google Scholar]
  19. Miller M., Schneider J., Sathyanarayana B. K., Toth M. V., Marshall G. R., Clawson L., Selk L., Kent S. B., Wlodawer A. Structure of complex of synthetic HIV-1 protease with a substrate-based inhibitor at 2.3 A resolution. Science. 1989 Dec 1;246(4934):1149–1152. doi: 10.1126/science.2686029. [DOI] [PubMed] [Google Scholar]
  20. Mulder J., McKinney N., Christopherson C., Sninsky J., Greenfield L., Kwok S. Rapid and simple PCR assay for quantitation of human immunodeficiency virus type 1 RNA in plasma: application to acute retroviral infection. J Clin Microbiol. 1994 Feb;32(2):292–300. doi: 10.1128/jcm.32.2.292-300.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Navia M. A., Fitzgerald P. M., McKeever B. M., Leu C. T., Heimbach J. C., Herber W. K., Sigal I. S., Darke P. L., Springer J. P. Three-dimensional structure of aspartyl protease from human immunodeficiency virus HIV-1. Nature. 1989 Feb 16;337(6208):615–620. doi: 10.1038/337615a0. [DOI] [PubMed] [Google Scholar]
  22. Oie S., Jacobson M. A., Abrams D. I. Alpha 1-acid glycoprotein levels in AIDS patients before and after short-term treatment with zidovudine (ZDV) J Acquir Immune Defic Syndr. 1993 May;6(5):531–533. [PubMed] [Google Scholar]
  23. Ou C. Y., Kwok S., Mitchell S. W., Mack D. H., Sninsky J. J., Krebs J. W., Feorino P., Warfield D., Schochetman G. DNA amplification for direct detection of HIV-1 in DNA of peripheral blood mononuclear cells. Science. 1988 Jan 15;239(4837):295–297. doi: 10.1126/science.3336784. [DOI] [PubMed] [Google Scholar]
  24. Redington J., Ebert S. C., Craig W. A. Role of antimicrobial pharmacokinetics and pharmacodynamics in surgical prophylaxis. Rev Infect Dis. 1991 Sep-Oct;13 (Suppl 10):S790–S799. doi: 10.1093/clinids/13.supplement_10.s790. [DOI] [PubMed] [Google Scholar]
  25. Reedijk M., Boucher C. A., van Bommel T., Ho D. D., Tzeng T. B., Sereni D., Veyssier P., Jurriaans S., Granneman R., Hsu A. Safety, pharmacokinetics, and antiviral activity of A77003, a C2 symmetry-based human immunodeficiency virus protease inhibitor. Antimicrob Agents Chemother. 1995 Jul;39(7):1559–1564. doi: 10.1128/aac.39.7.1559. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Robins T., Plattner J. HIV protease inhibitors: their anti-HIV activity and potential role in treatment. J Acquir Immune Defic Syndr. 1993 Feb;6(2):162–170. [PubMed] [Google Scholar]
  27. Siegler E., Schaefer A., Bilello J. A. Interferon-induced alterations in membrane functions and the growth of Daudi lymphoma and Friend leukemia cells. Biosci Rep. 1982 Aug;2(8):605–608. doi: 10.1007/BF01314223. [DOI] [PubMed] [Google Scholar]
  28. Wlodawer A., Erickson J. W. Structure-based inhibitors of HIV-1 protease. Annu Rev Biochem. 1993;62:543–585. doi: 10.1146/annurev.bi.62.070193.002551. [DOI] [PubMed] [Google Scholar]
  29. Wong A. K., Hsia J. C. In vitro binding of propranolol and progesterone to native and desialylated human orosomucoid. Can J Biochem Cell Biol. 1983 Oct;61(10):1114–1116. doi: 10.1139/o83-142. [DOI] [PubMed] [Google Scholar]
  30. Zimmerman T. P., Mahony W. B., Prus K. L. 3'-azido-3'-deoxythymidine. An unusual nucleoside analogue that permeates the membrane of human erythrocytes and lymphocytes by nonfacilitated diffusion. J Biol Chem. 1987 Apr 25;262(12):5748–5754. [PubMed] [Google Scholar]

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