title: In vitro selection and characterization of human immunodeficiency virus type 1 (HIV-1) isolates with reduced sensitivity to hydroxyethylamino sulfonamide inhibitors of HIV-1 aspartyl protease.
abstract: Human immunodeficiency virus type 1 (HIV-1) variants with reduced sensitivity to the hydroxyethylamino sulfonamide protease inhibitors VB-11,328 and VX-478 have been selected in vitro by two independent serial passage protocols with HIV-1 in CEM-SS and MT-4 cell lines. Virus populations with greater than 100-fold-increased resistance to both inhibitors compared with the parental virus have been obtained. DNA sequence analyses of the protease genes from VB-11,328- and VX-478-resistant variants reveal a sequential accumulation of point mutations, with similar resistance patterns occurring for the two inhibitors. The deduced amino acid substitutions in the resistant protease are Leu-10-->Phe, Met-46-->Ile, Ile-47-->Val, and Ile-50-->Val. This is the first observation in HIV protease resistance studies of an Ile-50-->Val mutation, a mutation that appears to arise uniquely against the sulfonamide inhibitor class. When the substitutions observed were introduced as single mutations into an HIV-1 infectious clone (HXB2), only the Ile-50-->Val mutant showed reduced sensitivity (two- to threefold) to VB-11,328 and VX-478. A triple protease mutant infectious clone carrying the mutations Met-46-->Ile, Ile-47-->Val, and Ile-50-->Val, however, showed much greater reduction in sensitivity (14- to 20-fold) to VB-11,328 and VX-478. The same mutations were studied in recombinant HIV protease. The mutant protease Ile-50-->Val displays a much lower affinity for the inhibitors than the parent enzyme (< or = 80-fold). The protease triply mutated at Met-46-->Ile, Ile-47-->Val, and Ile-50-->Val shows an even greater decrease in inhibitor binding (< or = 270-fold). The sulfonamide-resistant HIV protease variants remain sensitive to inhibitors from other chemical classes (Ro 31-8959 and L-735,524), suggesting possibilities for clinical use of HIV protease inhibitors in combination or serially.
authors.affiliation: Vertex Pharmaceuticals Incorporated, Cambridge, Massachusetts 02139-4211, USA.
authors.forename: J A
authors.initials: JA
authors.lastname: Partaledis
authors.forename: K
authors.initials: K
authors.lastname: Yamaguchi
authors.forename: M
authors.initials: M
authors.lastname: Tisdale
authors.forename: E E
authors.initials: EE
authors.lastname: Blair
authors.forename: C
authors.initials: C
authors.lastname: Falcione
authors.forename: B
authors.initials: B
authors.lastname: Maschera
authors.forename: R E
authors.initials: RE
authors.lastname: Myers
authors.forename: S
authors.initials: S
authors.lastname: Pazhanisamy
authors.forename: O
authors.initials: O
authors.lastname: Futer
authors.forename: A B
authors.initials: AB
authors.lastname: Cullinan
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D002219:Carbamates; D002460:Cell Line; D017931:DNA Primers; D005663:Furans; D016333:HIV Protease / Q000737:chemistry / Q000378:metabolism*; D017320:HIV Protease Inhibitors / Q000494:pharmacology*; D015497:HIV-1 / Q000187:drug effects* / Q000302:isolation & purification / Q000502:physiology; D006801:Humans; D007700:Kinetics; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000737:chemistry; D013329:Structure-Activity Relationship; D013449:Sulfonamides / Q000494:pharmacology*; D013601:T-Lymphocytes; D014779:Virus Replication / Q000187:drug effects
chemical_list: D002219:Carbamates; D017931:DNA Primers; D005663:Furans; D017320:HIV Protease Inhibitors; D011994:Recombinant Proteins; D013449:Sulfonamides; C095107:VB 11328; C095108:amprenavir; D016333:HIV Protease