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Deep sequencing of HIV-1 envelope determines coreceptor usage and predicts virologic responses to antiretroviral treatment with CCR5 antagonist medication

University of British Columbia

Next-generation sequencing can be used to genotype an array of HIV variants within clinical specimens, in a process referred to as deep sequencing. When directed at the gene for HIV envelope, this approach can be used to generate a high-sensitivity overview of the viral tropism of the HIV quasispecies in an infected individual. Since HIV variants with tropism for the CXCR4 coreceptor are not susceptible to the CCR5 antagonist agent maraviroc, their detection is crucial in order to screen out patients unlikely to achieve viral load declines on maraviroc. There are several assays that test HIV tropism, but it has been unclear as to which are most useful in the clinical setting. The aim of this thesis is to compare next-generation sequencing using massively parallel pyrosequencing against several alternative tropism assays in a total of four large randomized clinical trials of maraviroc. The methodologies are refined and optimized in the early chapters and applied to clinical specimens from a total of 2864 HIV-infected individuals. The concordance of next-generation sequencing with phenotypic tropism assays reached 87%. Relative to both the original Trofile phenotypic assay and standard population-based sequencing, deep sequencing had higher sensitivity to detect minority non-R5 HIV. Where assays gave discordant results, deep sequencing tended to outperform the comparator assay and was able to better discriminate maraviroc responders from non-responders. Next-generation sequencing had excellent performance in populations of both treatment-experienced and treatment-naïve individuals. It was consistently able to determine coreceptor usage, and to predict which patients would respond to maraviroc. It could be performed using either HIV RNA from blood plasma or HIV DNA from peripheral blood mononuclear cells. Additionally, longitudinal deep sequencing was performed on samples taken prior to maraviroc administration and again at treatment failure. Phylogenetic analyses confirmed that the non-R5 variants present at time of maraviroc treatment failure were derived from variants detected by deep sequencing before treatment was initiated. In conclusion, next-generation sequencing was applied to thousands of samples from phase III clinical trials, and was a superior screening tool to those originally used during trial enrollment. This thesis demonstrates the clinical utility of next-generation sequencing.

Text

2014-05-21

Attribution 2.5 Canada

http://hdl.handle.net/2429/46764

Experimental Medicine

University of British Columbia

UBCV

http://creativecommons.org/licenses/by/2.5/ca/