Short communication: dynamic constraints on the second phase compartment of HIV-infected cells

Abstract

The cells responsible for the second phase decay of HIV-1 viremia following the initiation of antiretroviral therapy have yet to be identified. A dynamic model that considers where drugs act in the virus life cycle places constraints on candidate cell types. In this regard, the rapid drop in viremia in patients starting regimens containing the integrase inhibitor raltegravir is of particular interest. We show here that the time delay between reverse transcription and integration is short in differentiated macrophages, making these cells poor candidates for the second phase compartment under the assumptions of standard models of viral dynamics.

FIG. 1.

(A) A model of HIV-1 dynamics reflecting the action of RT inhibitors and integrase inhibitors. The model is based on the work of Sedaghat et al. and envisions first (T) and second (M) phase compartments, each consisting of uninfected cells (T_u, M_u), early stage infected cells (T₁, M₁), and late stage infected cells (T₂, M₂). For this analysis, we consider early stage infected cells to be cells in which the virus has completed reverse transcription but not integration. The conversion of uninfected cells to early stage infected cells occurs at a rate that depends on the concentrations of uninfected cells and free virus and the rate constants β_T and β_M and is blocked by RT inhibitors. Late stage cells are cells in which integration has occurred. The conversion of early to late stage infected cells is governed by the rate constants k_T and k_M and is blocked by integrase inhibitors. Each cell population decays at a characteristic rate (δ). The accumulation of a large number of M₁ cells can in principle explain the more rapid decay observed in patients on RAL. (B) Decay dynamics are predicted by the model shown in (A) in patients on RT inhibitor-based (red lines) or integrase inhibitor-based (green lines) regimens. First phase decay (solid lines) and second phase decay (dotted lines) are shown for each regimen. In patients on RT inhibitor-based regimens, there is a shoulder representing the time it takes for M₁ cells to convert to M₂ cells. The length of this shoulder is given by 1/(δ_M1 + k_M). Note that this shoulder can be obscured by first phase viremia. The time required for viremia to fall below the limit of detection for each regimen is shown with a colored arrow. (C) Experimental measurements of the time required for completion of the fusion, reverse transcription, and integrase reactions in primary CD4⁺ T lymphoblasts and MDM. CD4⁺ T lymphoblasts were generated by culturing peripheral blood mononuclear cells (PBMCs) from multiple healthy blood donors for 72 h in the presence of phytohemagglutinin, followed by negative selection to isolate CD4⁺ cells. To prepare MDM, monocytes isolated from PBMCs by positive selection with CD14⁺ microbeads (Miltenyi Biotec) were cultured in the presence of 1 ng/ml M-CSF (R&D Systems) for 7 days. Cells were infected with GFP-expressing HIV-1 pseudoviruses capable of a single round of viral replication as previously described.^,, Maximally inhibitory concentrations of the antiretroviral drugs enfuvirtide, EFV, and RAL were added at various times after infection to block fusion, reverse transcription, and integration, respectively. For MDM infections, the virus was pseudotyped vesicular stomatitis virus G protein instead of an X4 HIV-1 envelope, and thus enfurvitide could not be used. The progressive loss of inhibition with later addition of drug allowed calculation of the weighted average time for completion of each process.