Each polymorphism was introduced into a functional LAI envelope clone by site-directed mutagenesis.5 Only the M426P substitution resulted in a 3-fold increase in viral susceptibility to temsavir compared with the wild-type LAI virus (Supplemental Digital Content Table 4, http://links.lww.com/QAI/B94). There was no correlation between substitutions at gp120 positions S375, M426, M434, and M475 and the number of subjects qualifying for resistance testing through week 48, regardless of whether the substitutions were linked to a 3-fold or 3-fold reduction in viral susceptibility to temsavir (or a previous attachment inhibitor, BMS-488043) in this study or previous in vitro studies (Fig. an evaluable phenotype using PhenoSense Flurbiprofen Entry (which tests for viral susceptibility to temsavir) and 13/29 exhibited 3-fold increase in temsavir IC50 from BL. gp120 population sequencing was successful in 11/13 subjects and 7 had emergent substitutions in gp120 associated with reduced temsavir susceptibility (S375, M426, or M434). However, 5/13 fostemsavir-treated subjects achieved subsequent suppression to 50 copies/mL before the week 48 database lock, regardless of key gp120 substitutions. Conclusions: Response rates remained similar across study arms regardless of BL nucleoside reverse transcriptase inhibitor, nonnucleoside reverse transcriptase inhibitor, and protease inhibitor resistance-associated mutations. Emergent changes in viral susceptibility occurred more frequently with fostemsavir compared with ATV/r. However, the full impact of temsavir IC50 changes and emergent HIV-1 gp120 substitutions, and thus appropriate clinical cutoffs, requires further study. Fostemsavir is being evaluated in a phase 3 trial in heavily treatment-experienced subjects. were as previously described.5 In most cases, uncloned purified polymerase chain reaction products were used for population sequencing of gp160 using a library of envelope-specific primers (Supplemental Digital Content Table S2, http://links.lww.com/QAI/B94). HIV-1 sequences were aligned to the HIV-1 subtype B consensus sequence available in the Los Alamos National Laboratories HIV sequence database (http://www.hiv.lanl.gov) using the AlignX software in the Vector NTI Advance package (version 11.5; Invitrogen, Carlsbad, CA). BL sequences were deposited in GenBank with the accession numbers “type”:”entrez-nucleotide-range”,”attrs”:”text”:”MF990378 to MF990556″,”start_term”:”MF990378″,”end_term”:”MF990556″,”start_term_id”:”1246707825″,”end_term_id”:”1246708003″MF990378 to MF990556. Amino acid positions were numbered Rabbit Polyclonal to C1S per the HIV-1 HXB2 strain sequence. Amino acid substitutions in gp120 were visualized using GeneDoc software11 in difference display mode, and Flurbiprofen specific changes at gp120 positions S375, M434, M426, Flurbiprofen and M475 were assessed. If the nucleotide sequence had more than one possible base, all possibilities were expanded within the codon and amino acids were assigned as previously described.4 In addition, changes at positions L116 and A204, previously linked to reduced in vitro viral susceptibility to temsavir,5 were assessed. In the case of a novel polymorphism, the mutations were introduced into clinical isolates or the wild-type HIV-1 LAI strain by site-directed mutagenesis, and viral susceptibility to temsavir was assessed using a cellCcell fusion assay as described previously.5,12 All data were reported as FC in temsavir half-maximal effective concentration (EC50), normalized to an internal control.5 RESULTS BL Characteristics and Genotypic Resistance Profile A total of 581 subjects were screened, 254 were randomly assigned to treatment Flurbiprofen groups in the study, and 251 received treatment (200 subjects received fostemsavir).8 BL characteristics were generally well balanced between treatment arms.8 Most subjects had HIV-1 subtype B (65.7%) or C (19.9%); the remainder had HIV-1 subtype A (0.4%), A1 (4.0%), BF (0.4%), complex (6.8%), F1 (2.4%), or G (0.4%). The median BL viral load was 4.85 log10 copies/mL (43% 100,000 copies/mL), median CD4+ T-cell count was 229.5 cells/L (38% 200 cells/L), and median BL temsavir IC50 was 0.67 nM (range: 0.05C161 nM). One subject with a BL temsavir IC50 of 161 nM, which was higher than the IC50 Flurbiprofen cutoff of 100 nM specified in the entry criteria, was randomized to the study but achieved the primary efficacy endpoint (HIV-1 RNA 50 copies/mL at week 24) and remained on study through week 48. BL genotypic resistance to PIs and NRTIs was described previously8 and is expanded in Supplemental Digital Content Table S3, http://links.lww.com/QAI/B94. Overall, 49% of subjects had 1 major PI, NRTI, or NNRTI mutation, and across all treatment arms, 22%C34% had 1 NRTI and NNRTI mutation at BL. The most common BL mutations (other than minor PI substitutions) were M184V (22%C40% of samples across all treatment arms), K103N (20%C39%), and thymidine analogue mutations (8%C16%). In line with study-entry criteria, no subject had virus with integrase resistance-associated mutations (RAMs) at BL. Three subjects had virus with the K70E substitution; however, this was not associated with reduced.
