A highly multiplexed droplet digital PCR assay to measure the intact HIV-1 proviral reservoir

Autor: Claire N. Levy, Sean M. Hughes, Pavitra Roychoudhury, Daniel B. Reeves, Chelsea Amstuz, Haiying Zhu, Meei-Li Huang, Yulun Wei, Marta E. Bull, Noah A.J. Cassidy, Jan McClure, Lisa M. Frenkel, Mars Stone, Sonia Bakkour, Elizabeth R. Wonderlich, Michael P. Busch, Steven G. Deeks, Joshua T. Schiffer, Robert W. Coombs, Dara A. Lehman, Keith R. Jerome, Florian Hladik
Jazyk: angličtina
Rok vydání: 2021
Předmět:
Zdroj: Cell Reports Medicine, Vol 2, Iss 4, Pp 100243- (2021)
Druh dokumentu: article
ISSN: 2666-3791
DOI: 10.1016/j.xcrm.2021.100243
Popis: Summary: Quantifying the replication-competent HIV reservoir is essential for evaluating curative strategies. Viral outgrowth assays (VOAs) underestimate the reservoir because they fail to induce all replication-competent proviruses. Single- or double-region HIV DNA assays overestimate it because they fail to exclude many defective proviruses. We designed two triplex droplet digital PCR assays, each with 2 unique targets and 1 in common, and normalize the results to PCR-based T cell counts. Both HIV assays are specific, sensitive, and reproducible. Together, they estimate the number of proviruses containing all five primer-probe regions. Our 5-target results are on average 12.1-fold higher than and correlate with paired quantitative VOA (Spearman's ρ = 0.48) but estimate a markedly smaller reservoir than previous DNA assays. In patients on antiretroviral therapy, decay rates in blood CD4+ T cells are faster for intact than for defective proviruses, and intact provirus frequencies are similar in mucosal and circulating T cells.
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