Different kinetics of viral replication and DNA integration in the main HIV-1 cellular reservoirs in the presence and absence of integrase inhibitors

Abstract

To compare the kinetics of integration, p24 production and equilibrium of the different HIV-DNA forms in human primary cells in the presence/absence of integrase-inhibitors (INIs) in vitro. Monocyte-derived-macrophages (MDMs), CD4⁺ T-cells and peripheral blood mononuclear cells (PBMCs) were infected with HIV-1 in the presence/absence of raltegravir and dolutegravir. HIV-DNA levels and p24 production were measured by qPCR and ELISA assays, respectively. In the absence of INIs, levels of HIV-DNA forms were initially very low, with an increase in the integration process starting at 3 dpi. HIV-DNA increased more slowly in MDMs than it did in CD4⁺ T-cells and PMBCs peaking at 21 dpi with a mean of 1580 (±890) and 615 (±37) copies/10³ cells for proviral and unintegrated HIV-DNA, and 455,972 (±213,255) pg/mL of p24 at the same time point. In CD4⁺ T-cells the proviral HIV-DNA increased together with unintegrated HIV-DNA peaking at 7 dpi (583 ± 261 and 338 ± 254 copies/10³ cells) when the p24 was 218,000 (±75,600) pg/mL. A similar trend was observed in PBMCs (494 ± 361 and 350 ± 123 copies/10³ cells for proviral and unintegrated HIV-DNA, and p24 production of 149,400 ± 131,800 pg/mL). Both INIs inhibited viral replication and integration in all the cell types that were tested, especially starting at 3 dpi. However, a small but measurable amount of HIV-DNA (<5 copies/10³ cells) was still observed in treated-MDMs up to 30 dpi. In conclusion, our study showed differences in HIV-DNA kinetic integration between CD4⁺ T-cells and MDMs, which could explain the divergent kinetics of viral-replication. Both INIs inhibited HIV-1 integration and replication with no difference found between CD4⁺ T-cells and MDMs. However, residual HIV-DNA remained detectable up to 30 dpi in INI-treated MDMs although complete inhibition of HIV replication was achieved. The clinical significance of this minor DNA persistence deserves further investigation considering the role of macrophages as reservoirs.

Keywords: 2-LTR; CD4(+) T-cells; HIV-1 reservoir; Integrase inhibitors; Macrophages; Proviral HIV-DNA.