Background:  Recently it has been shown that V38A and Q40H+L45M correlated with a gain and with a loss of CD4 count, respectively, in HIV-infected patients receiving enfuvirtide (T-20). The aim of this study is to investigate in a large cohort of T-20-treated patients the long-term association of such mutations with the viro-immunological parameters, and whether selected gp41 mutational clusters correlate with viro-immunological outcome

Methods:  We analyzed 195 sequences of HIV-1 gp41 and clinical follow-up from 77 T-20-treated patients at baseline and monthly until week 48. The association of mutations with T-20 treatment and between mutations at baseline and during treatment was assessed by χ² test. Association of mutations with viremia CD4 count (copies/µL) was assessed by Mann-Whitney test. Covariation analysis was based on binomial correlation coefficient (φ) and hierarchical clustering.

Results:  We found that 9 mutations (A30T, L54M, E119Q, S129D/G, N126K, N140I, D239H, T268A) are positively associated with T-20 treatment and correlate with known T-20 resistance mutations. In particular, strong correlation (φ >0.30; p <0.01) is observed for N140I with V38A and for D239H with Q40H and L45M. Cluster analysis reveals the existence of 4 clusters of mutations:  V38A with N140I, S129G and A30T; N43D with S138A; G36V with N126K; and Q40H, L45M with the L54M, E119Q, S129D, T286A, and D239H. Co-presence of N140I with V38A is associated (p <0.05) with a CD4 increase from baseline (40 copies/µL) of 2-fold (210 copies/µL) at week 24 and 4.7-fold (249 copies/µL) at week 48 compared with V38A alone, without significant changes in viral load (from 4.9 log at baseline to 4.5 log at week 24 and to 4.8 log at week 48, p = ns). In contrast, the presence of D329H duplicates CD4 loss from baseline (124 copies/µL) to week 48 (35 copies/µL) given by Q40H + L45M (p = 0.05), without significant changes in viral load. Moreover, specific polymorphisms at bseline, correlated (p <0.05) with the on treatment development of T-20 resistance mutations, were identified. In particular, baseline presence of P213Q and V321I correlated with development of V38A, while baseline presence of R236Q correlated with development of N43D

Conclusions:  Gp41-mutational patterns under T-20 pressure are more complex than currently known, suggesting that an ordered network of mutations, regulated by natural polymorphisms present before T-20 treatment, modulates positively and negatively the HIV ability to damage the immune system. Their knowledge is important for a correct use of T-20 and for innovative therapeutic strategies