Assessing suitability of next-generation viral outgrowth assays as proxies for classic QVOA to measure HIV-1 latent reservoir size

J Infect Dis. 2020 Mar 9;jiaa089. doi: 10.1093/infdis/jiaa089. Online ahead of print.

Abstract

Background: Evaluations of HIV curative interventions require reliable and efficient quantification of replication-competent latent reservoirs (LR). The "classic" quantitative viral outgrowth assay (QVOA) has been regarded as "gold standard," although prohibitively resource- and labor-intensive. We compared six "next-gen" VOA employing PCR or ultrasensitive p24 to assess their suitability as scalable proxies for QVOA.

Methods: Next-gen VOA were compared to classic QVOA using single leukapheresis-derived samples from five ART-suppressed HIV+ participants and one HIV- control; each lab tested blinded batches of three frozen and one fresh sample. Markov chain Monte Carlo methods estimated extra-Poisson variation at aliquot, batch, and lab levels. Models also estimated the effect of testing frozen versus fresh samples.

Results: Next-gen VOA had similar estimates of variation to QVOA. Assays with ultrasensitive readout reported higher IUPM than classic QVOA. Within-batch testing had 2.5-fold extra-Poisson variation (95%CI 2.1,3.5) for next-gen assays. Between-lab variation increased extra-Poisson variation to 3.4-fold (95% CI 2.6,5.4). Frozen storage did not substantially alter IUPM (-18%(-52%,+39%)).

Conclusions: The data offer cautious support for use of next-gen VOA as proxies for more laborious QVOA, while providing greater sensitivities and dynamic ranges. Measurement of LR in eradication strategies would benefit from high throughput and scalable assays.

Keywords: HIV cure; HIV reservoir; IUPM; Inducible HIV RNA; assay comparison; latency; leukapheresis; quantitative viral out growth assay (QVOA).