Evolution of HIV-1 Provirus DNA load and PBMC DNA genotype after a 6 months CD4 guided-structured treatment interruption (STI)

I Pellegrin¹, P.Blanco², JF Viallard², MH Schrive¹, H Fleury¹, JF Moreau³, JL Pellegrin². Department of Virology¹, Internal Medicine Infectious Diseases², Immunology³, Bordeaux University Hospital (France).

Background: Reemergence of drug-susceptible HIV-1 and waning of drug-resistant HIV-1 in plasma after a STI offers the theoretical potential for a more durable response to salvage therapy. However, the persistence of low levels of drug-resistance virus in cellular reservoirs remains a concern. Material and Methods: Proviral DNA extracted from patients PBMC at W0 and W24 after a CD4+-driven STI was quantified by real time PCR and analysed for reverse transcriptase (RT) and protease (PR) genotypes (according to the IAS list 2002) by direct sequencing using the CEQ2000 sequencer. Results: Paired samples obtained at the time of STI (W0) and at W24 were available for 37 patients. At W0, median CD4+ count, plasma RNA and proviral DNA loads were 747 [637-935] cells/µl, 1.7 [1.7-3.4] log₁₀ copies/ml and 2.5 [2-3.1] log₁₀ copies/10⁶ PBMC, respectively. Discontinuation of therapy for 24 weeks was associated with a median decrease in the CD4+ count of 270 [158-341] cells/µl and an increase in the plasma HIV RNA level of 2.1 [1.2-2.9] log₁₀ copies/ml. At W24, median proviral DNA load was 3.4 [3-3.7] log₁₀ copies/10⁶ PBMC with a median increase of 0.7 [0.3-1.1] log₁₀ copies/10⁶ PBMC within 6 months. PBMC genotyping at W0 showed 70.3% (26/37) RT and PR with 1-4 NRTI- and 1-4 PI-related mutations. The evolution of genotypic resistance pattern in proviral DNA between W0 and W24 after STI was analysed. When considering sequences with detectable NRTI- or PI-related resistance mutations at baseline, we observed at W24 for RT sequences: 3 RT identical to baseline with 2 mutations, the loss of 1 to 4 mutations in 19 RT and detection of 1 additional mutation in 4 RT ; for PR sequences: 14 PR remained identical to baseline with 1 to 3 mutations, the loss of 1 to 2 mutations (including primary mutations) in 8 PR and 1 to 3 additional mutations in 4 PR. RT and PR wild-type patterns remained stable in  11 paired samples. Conclusion: Within 24W of STI, we observed: 1) viral replication replenishes the pool of latently infected cells. 2) a virus population containing less resistance mutations than those present before STI emerges in proviral DNA. 3) despite the repopulation of sensitive virus in plasma and the loss of some mutations in PBMC, resistant HIV-1 still persist in PBMC after STI. Decisions regarding salvage therapy must be made with the knowledge that all previously circulating wild-type and resistant virus can be archived in the reservoir and may emerge when conditions are favorable.