This study evaluated heterogeneity in the HCV NS3 protease sequences isolated from HCV from a group of Brazilian patients with chronic hepatitis C whose responses to PEG-IFN and RBV treatment (SVR, REL and NR) were being monitored. We assessed genetic variability and the presence of protease inhibitor resistance mutations in the NS3 protease in the isolates before, during, and after treatment in patients with viral loads > 500 IU/ml. We investigated specific protease inhibitor resistance mutations, namely, V36A/L/M/G, Q41R, F43C/S, T54S/A, V55A, Q80R/K, S138T, R155K/T/G/I/Q/M/S/L and A156S/T/V/I. Of the 68 patients analyzed 3 (4.4%), had at least one of the mutations known to confer telaprevir and boceprevir resistance (V36L, T54S and V55A). These resistance mutations have been described previously [17–22]. In Brazil, knowledge is lacking about the temporal changes that occur in PI resistance mutations during the course of IFN/RBV treatment. One report  described the presence of V36L and T54S in HCVs in a group of Brazilian patients chronically infected with HCV. Recent studies have also highlighted the presence of naturally ocurring mutations in the NS3 protease in patients. For example, a study of four Spanish patients identified V36A, T54A, V55A, R155S, and A156T/V mutations in the NS3 protease . In addition, a Japanese study identified T54S, Q80K, I153V and D168E NS3 protease mutations among 261 patients (13.4% had a single mutation, while 2.3% had double mutations) . NS3 protease mutations (V36L, T54S, V55A/I, and Q80K/L) were also observed in 29% of genotype 1a patients in an Italian population .
In the present study, V36L, T54S and V55A mutations were identified in Brazilian patients treated with PEG-IFN and RBV. To assess the maintenance of protease mutation variants, this analysis was performed before, during and after treatment completion, thus allowing us to determine the fate of the genetic variants over an 18 months period. Negative selective pressure was observed acting on the NS3 protease region analyzed here. In addition, the observed mutations may confer resistance to telaprevir and boceprevir, making it possible to predict that patients harboring viruses with such mutations may not benefit from a future treatment with these drugs; therefore, the null responder patients should be engaged in the new Protease Therapy Protocol [10, 12, 16, 29]. These two drugs (Boceprevir and Telaprevir), already approved by FDA and by ANVISA in Brazil , are now components of licensed therapies. Thus, mapping NS3 protease resistance mutations to protease inhibitors may be an important tool to direct anti-HCV treatment.