Background:  The diketo analogue S-1360 is the first HIV-1 integrase strand transfer inhibitor (INSTI) that has entered clinical development.

Methods:  We have examined the development of HIV-1 resistance to S-1360 by selecting HIV-1(IIIB) in the presence of increasing concentrations of the compound. The IIIB/S-1360^res strains selected after 30, 50, and 70 passages in the presence of S-1360 were evaluated genotypically by sequencing analysis and phenotypically using the MT-4/MTT system.

Results:  Multiple mutations, nine in total, emerged in the catalytic domain of integrase as a result of the selection process. They included T66I and L74M, that have both been previously associated with resistance against the diketo acid L-708,906. The progressive accumulation of mutations coincided with an increasing level of resistance of the selected  strains towards S-1360. After 30, 50, and 70 passages in the presence of S-1360, IIIB/S-1360^res displayed a 3.8, 7.9, and >63-fold reduction in susceptibility to S-1360, respectively. Phenotypic cross-resistance to L-708,906 was observed for the strain selected after 70 passages. In contrast, IIIB/S-1360^res remained fully susceptible to V-165, an integrase binding inhibitor (INBI) belonging to the class of pyranodipyrimidines. All selected strains exhibited wild type sensitivity to inhibitors of viral entry, reverse transcriptase or protease. Recombination of the mutant integrase genes into wild type background by integrase-Chimeric Virus Technology entirely reproduced the observed resistance profile of all in vitro selected strains, confirming that the mutations found in integrase are indeed fully responsible for the resistance phenotype of IIIB/S-1360^res.

Conclusions:  Selection of resistance to the clinical drug candidate S-1360 in cell culture, is associated with the emergence of up to nine mutations, some of which have not previously been linked to resistance to integrase inhibitors. The accumulation of 9 mutations in a conserved protein suggests a complex mechanism of resistance development and may point to adaptation at the enzymatic level.

Keywords: resistance; integrase; S-1360