title: Impact of maraviroc-resistant and low-CCR5-adapted mutations induced by in vitro passage on sensitivity to anti-envelope neutralizing antibodies.
abstract: The aim of this study was to generate maraviroc (MVC)-resistant viruses in vitro using a human immunodeficiency virus type 1 subtype B clinical isolate (HIV-1KP-5) to understand the mechanism(s) of resistance to MVC. To select HIV-1 variants resistant to MVC in vitro, we exposed high-chemokine (C-C motif) receptor 5 (CCR5)-expressing PM1/CCR5 cells to HIV-1KP-5 followed by serial passage in the presence of MVC. We also passaged HIV-1KP-5 in PM1 cells, which were low CCR5 expressing to determine low-CCR5-adapted substitutions and compared the Env sequences of the MVC-selected variants. Following 48 passages with MVC (10 µM), HIV-1KP-5 acquired a resistant phenotype [maximal per cent inhibition (MPI) 24%], whilst the low-CCR5-adapted variant had low sensitivity to MVC (IC50 ~200 nM), but not reduction of the MPI. The common substitutions observed in both the MVC-selected and low-CCR5-adapted variants were selected from the quasi-species, in V1, V3 and V5. After 14 passages, the MVC-selected variants harboured substitutions around the CCR5 N-terminal-binding site and V3 (V200I, T297I, K305R and M434I). The low-CCR5-adapted infectious clone became sensitive to anti-CD4bs and CD4i mAbs, but not to anti-V3 mAb and autologous plasma IgGs. Conversely, the MVC-selected clone became highly sensitive to the anti-envelope (Env) mAbs tested and the autologous plasma IgGs. These findings suggest that the four MVC-resistant mutations required for entry using MVC-bound CCR5 result in a conformational change of Env that is associated with a phenotype sensitive to anti-Env neutralizing antibodies.
authors.affiliation: Center for AIDS Research, Kumamoto University, 2-2-1 Honjo, Chuo-ku, Kumamoto 860-0811, Japan.|AIDS Research Centre, National Institute of Infectious Diseases, 1-23-1 Toyama, Shinjuku-ku, Tokyo 162-8640, Japan.
authors.forename: Kazuhisa
authors.initials: K
authors.lastname: Yoshimura
authors.affiliation: Center for AIDS Research, Kumamoto University, 2-2-1 Honjo, Chuo-ku, Kumamoto 860-0811, Japan.|AIDS Research Centre, National Institute of Infectious Diseases, 1-23-1 Toyama, Shinjuku-ku, Tokyo 162-8640, Japan.
authors.forename: Shigeyoshi
authors.initials: S
authors.lastname: Harada
authors.affiliation: Center for AIDS Research, Kumamoto University, 2-2-1 Honjo, Chuo-ku, Kumamoto 860-0811, Japan.|AIDS Research Centre, National Institute of Infectious Diseases, 1-23-1 Toyama, Shinjuku-ku, Tokyo 162-8640, Japan.
authors.forename: Samatchaya
authors.initials: S
authors.lastname: Boonchawalit
authors.affiliation: Center for AIDS Research, Kumamoto University, 2-2-1 Honjo, Chuo-ku, Kumamoto 860-0811, Japan.
authors.forename: Yoko
authors.initials: Y
authors.lastname: Kawanami
authors.affiliation: Center for AIDS Research, Kumamoto University, 2-2-1 Honjo, Chuo-ku, Kumamoto 860-0811, Japan.
authors.forename: Shuzo
authors.initials: S
authors.lastname: Matsushita
mesh_terms: D019380:Anti-HIV Agents / Q000494:pharmacology*; D057134:Antibodies, Neutralizing / Q000276:immunology*; D003510:Cyclohexanes / Q000494:pharmacology*; D024882:Drug Resistance, Viral; D015483:HIV Antibodies / Q000276:immunology*; D015497:HIV-1 / Q000187:drug effects / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D006801:Humans; D000077592:Maraviroc; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000235:genetics / Q000276:immunology; D020125:Mutation, Missense; D017422:Sequence Analysis, DNA; D012692:Serial Passage; D014230:Triazoles / Q000494:pharmacology*; D054299:env Gene Products, Human Immunodeficiency Virus / Q000235:genetics / Q000276:immunology*
chemical_list: D019380:Anti-HIV Agents; D057134:Antibodies, Neutralizing; D003510:Cyclohexanes; D015483:HIV Antibodies; D050505:Mutant Proteins; D014230:Triazoles; D054299:env Gene Products, Human Immunodeficiency Virus; D000077592:Maraviroc