Identification of a rare mutation at reverse transcriptase Lys65 (K65E) in HIV-1-infected patients failing on nucleos(t)ide reverse transcriptase inhibitors

The HIV reverse transcriptase (RT) mutation K65R confers resistance to nucleos(t)ide reverse transcriptase inhibitors (NRTIs). Here, analysing a large database, we report the selection of another rare K65E mutation in patients failing on NRTI-containing regimens. Clinical and virological characteristics of patients harbouring the K65E mutation were analysed using a large RT sequence database from treatment-experienced individuals. Structural analysis of the K65E RT mutant complexwasperformed bymeansof docking simulations. The replication capacitywas assessed using viruses harbouring the K65E mutation introduced by site-directed mutagenesis (SDM) in pNL 4-3. Overall, in 23530 sequences frompatients failing on antiretroviral therapy, the prevalence of substitutions at position K65 in RTwas 2.4%. In addition to K65R (n1/4395) and K65N (n1/49), another mutation, K65E, was found in 15 patients. In 11 out of 15 cases, tenofovir, abacavir, didanosine or stavudine were present at the time of K65E selection. The molecular recognition of RTcontaining K65E supports evidence for the role of this mutation in resistance to tenofovir. The SDM pNL4-3 K65E variant harboured a very low replicative capacity (5% versus wild-type). We investigated the role of a novel rare NRTI mutation located at position Lys65 of RT (K65E), found in drug-experienced patients failing on NRTIs. The low frequency of this mutation is probably related to the high impairment of replicativecapacity inducedbythismutation. This studyshouldhave significant clinical implications, as these findings warn clinicians that other minor substitutions at Lys65 (such as K65E) play a role in NRTI resistance.