PCR products were directly sequenced and multiple alignment of nucleotides and deduced Ipilimumab order amino acid sequences was inferred using Clustal_W, version 1.74 (Conway Institute UCD, Dublin, Ireland). We retrieved 501 available sequences (476 genotype 1 and 25 genotype 4) from the GenBank database as a control group. These sequences were chosen from HCV-monoinfected patients only and to concern exclusively the

NS3 protease domain. Phylogenetic criteria were used to exclude very closely related sequences (that is, cases of clonal sequences from the same patient and time-point) from the data set. Fisher’s exact test was used to compare the frequencies of mutations at positions 36, 54, 155, 156 and 170, which are known to confer resistance to HCV PIs selleck chemicals [3,5], in the sequences obtained from HIV/HCV-coinfected individuals and the GenBank control group. Patients’ characteristics were compared according to the presence of HCV PI resistance mutations using a Fisher’s exact test for qualitative variables and a Wilcoxon–Mann–Whitney test for quantitative variables.

Distributions are described as medians [with 25th and 75th percentiles, and interquartile range (IQR)]. All statistical tests were two-sided. Statistical analyses were performed using sas 9.1 (SAS Institute Inc., Cary, NC, USA). At the time of HCV protease analysis, the median age of the HIV/HCV-coinfected patients was 47 years (IQR 45–49 years). Eighty patients were male. One hundred and ten patients had received antiretroviral therapy for at least 6 months. The median HIV load was 40 HIV-1 RNA copies/mL (range 20–560 800 copies/mL) and the median CD4 cell count was 474 cells/μL (range 3–1671 cells/μL). Eighty-two patients had never been treated for their chronic hepatitis C, whereas 38 were relapsers or nonresponders to previous anti-HCV treatment. Of 76 sequences from HIV/HCV genotype 1-coinfected patients, six (7.9%) showed amino acid substitutions associated with HCV PI resistance.

Three patients showed a mutation at position N-acetylglucosamine-1-phosphate transferase 36 known to confer low-level resistance to HCV PIs: V36L in one patient and V36M in the other two. Three patients carried mutations conferring intermediate or high levels of resistance to HCV PIs: R155K and T54S in one and two patients, respectively. In 31 (6.5%) of 476 HCV genotype 1 sequences retrieved from the GenBank database, HCV PI resistance mutations were found. Amino acid mutations detected in the sequences were as follows: V36L in six sequences, V36M in six, T54S in 11, R155K in five, V170A in one, and T54S+R155K in two. The proportion of patients with HCV PI resistance mutations was not significantly different between HIV/HCV-coinfected and HCV-monoinfected patients (P=0.6).