Mutation of HIV-1 Genomes in a Clinical Population Treated with the Mutagenic Nucleoside KP1461

The deoxycytidine analog KP1212, and its prodrug KP1461, are prototypes of a new class of antiretroviral drugs designed to increase viral mutation rates, with the goal of eventually causing the collapse of the viral population. Here we present an extensive analysis of viral sequences from HIV-1 infected volunteers from the first “mechanism validation” phase II clinical trial of a mutagenic base analog in which individuals previously treated with antiviral drugs received 1600 mg of KP1461 twice per day for 124 days. Plasma viral loads were not reduced, and overall levels of viral mutation were not increased during this short-term study, however, the mutation spectrum of HIV was altered. A large number (N = 105 per sample) of sequences were analyzed, each derived from individual HIV-1 RNA templates, after 0, 56 and 124 days of therapy from 10 treated and 10 untreated control individuals (>7.1 million base pairs of unique viral templates were sequenced). We found that private mutations, those not found in more than one viral sequence and likely to have occurred in the most recent rounds of replication, increased in treated individuals relative to controls after 56 (p = 0.038) and 124 (p = 0.002) days of drug treatment. The spectrum of mutations observed in the treated group showed an excess of A to G and G to A mutations (p = 0.01), and to a lesser extent T to C and C to T mutations (p = 0.09), as predicted by the mechanism of action of the drug. These results validate the proposed mechanism of action in humans and should spur development of this novel antiretroviral approach.Koronis Pharmaceutical

Specificity of private mutational change over time in the treated (panel A) and control (panel B) groups.

<p>Open bars are from day 0, shaded bars are from day 56 and black bars are from day 125. The y-axis shows the total number of private mutations in the two groups of 10 subjects.</p

Differences in private site mutations observed in treated and control groups.

1<p>Mean number of private site mutations over all participants at time 0.</p>2<p>Difference (treated minus control) after subtracting the difference at baseline.</p>3<p>All p-values are two-sided and based on permutations tests with 1000 repetitions.</p>4<p>Difference (treated minus control) at day 124 minus difference at day 56.</p

Genetic diversity of 1.0 kb segments of HIV-1 <i>gag</i> genes through time in individuals treated and untreated with KP1461.

<p>Average population diversity of samplings of 105 1.0 kb <i>gag</i> gene segments through time is shown for treated (panel A) and untreated (panel B) groups.</p

Demographics of treated and control groups.

<p>*At Day 0 of treatment.</p

Chemical structures of KP1212 and its prodrug, KP1461, administered in the Phase IIa trial.

<p>Chemical structures of KP1212 and its prodrug, KP1461, administered in the Phase IIa trial.</p

Private mutations detected in HIV-1 <i>gag</i> genes in treated and control groups.

<p>The average number of private mutations (found in only one of 105 sequences in each sample) per kilobase of sequence are shown for treated (Panel A) and control (Panel C) subjects at three time points and in Phase I trial participants (Clay et al, manuscript in preparation). In panels A and C, open bars are from day 0 (t1), shaded bars are from day 56 (t2) and black bars are from day 125 (t3). Patient 1 underwent 2 courses of treatment, pt1 and pt1.2, with a 9-day gap in between. The trajectory lines drawn on top of each 3-bar data set (for each individual) track the accumulation or loss of mutations over the study. In Panel A, 8 of 11 comparisons revealed an accumulation of mutations, whereas in control participants depicted in Panel C, only 2 0f 10 had an accumulation of mutations over the study. Panel B shows the number of private mutations per subject at each of the three time points. Data from treated subjects are shown with diamond symbols and controls with circles. Phase I trial participants sampled at day 0 (Panel B) are also shown with triangle symbols. Median levels for each group are shown with a horizontal line. Panel D shows the overall change in the number of private mutations over the 125-day sampling period in the treated group and over comparable intervals in the untreated control subjects. Wilcoxon ranked sum tests were used to assess differences in total private site counts within subjects and between treated and untreated groups. In panel B, p values for treated vs. phase I subjects  = 0.894; phase I subjects vs. phase 2 controls  = 0.549.</p