Antiretroviral therapy can reduce HIV-1 to undetectable levels in peripheral blood, but the effectiveness of treatment in suppressing replication in lymphoid tissue reservoirs has not been determined. Here we show in lymph node samples obtained before and during 6 mo of treatment that the tissue concentrations of five of the most frequently used antiretroviral drugs are much lower than in peripheral blood. These lower concentrations correlated with continued virus replication measured by the slower decay or increases in the follicular dendritic cell network pool of virions and with detection of viral RNA in productively infected cells. The evidence of persistent replication associated with apparently suboptimal drug concentrations argues for development and evaluation of novel therapeutic strategies that will fully suppress viral replication in lymphatic tissues. These strategies could avert the long-term clinical consequences of chronic immune activation driven directly or indirectly by low-level viral replication to thereby improve immune reconstitution.drug levels | pharmacokinetics | FDC network C ombination antiviral therapy (ART) to suppress HIV-1 replication and reduce plasma viremia to below the limits of detection in peripheral blood (PB) has reduced mortality and dramatically improved quality of life for patients. However, immune reconstitution, measured by changes in the size of populations of CD4 T cells, is often incomplete, even after years of therapy (1-3). During apparently effective therapy, CD4 T-cell populations in PB mononuclear cells (PBMCs), lymph node (LN), and gut-associated lymphoid tissue (GALT) remain abnormally low and innate and adaptive immunity is not fully restored (4). Levels of T-cell activation and innate system activation are often higher than that observed in well-matched uninfected adults (5, 6). These persistent abnormalities may contribute to abnormal vaccine responses (7, 8), a higher than normal incidence of non-AIDS-related cancers (9, 10) and increased risk for clinical conditions associated with chronic inflammation (e.g., cardiac disease, clotting disorders, pulmonary hypertension, emphysema, and stroke) (11-18). Thus, improvements over current approaches to treatment of HIV infection that more fully restore normal immune function might significantly improve health and life expectancy.To that end, we explore here the hypothesis that antiretroviral drug (ARV) concentrations might be insufficient to fully suppress replication in the lymphoid tissue compartments, which are the principal sites where virus is produced, stored as complexes on the follicular dendritic cell network (FDCn) (19-21), and persists in latently infected cells during ART (19,20,22). This hypothesis builds first on the link between the size of the reservoir and the degree of inflammation, arguing that persistent virus production during ART could sustain immune activation (IA) and downstream pathological consequences (23, 24), and second on drug distribution studies in animal models of AIDS in which ...

Antiretroviral therapy (ART) suppresses HIV replication in most individuals but cannot eradicate latently infected cells established before ART was initiated. Thus, infection rebounds when treatment is interrupted by reactivation of virus production from this reservoir. Currently, one or a few latently infected resting memory CD4 T cells are thought be the principal source of recrudescent infection, but this estimate is based on peripheral blood rather than lymphoid tissues (LTs), the principal sites of virus production and persistence before initiating ART. We, therefore, examined lymph node (LN) and gut-associated lymphoid tissue (GALT) biopsies from fully suppressed subjects, interrupted therapy, monitored plasma viral load (pVL), and repeated biopsies on 12 individuals as soon as pVL became detectable. Isolated HIV RNA-positive (vRNA+) cells were detected by in situ hybridization in LTs obtained before interruption in several patients. After interruption, multiple foci of vRNA+ cells were detected in 6 of 12 individuals as soon as pVL was measureable and in some subjects, in more than one anatomic site. Minimal estimates of the number of rebounding/founder (R/F) variants were determined by single-gene amplification and sequencing of viral RNA or DNA from peripheral blood mononuclear cells and plasma obtained at or just before viral recrudescence. Sequence analysis revealed a large number of R/F viruses representing recrudescent viremia from multiple sources. Together, these findings are consistent with the origins of recrudescent infection by reactivation from many latently infected cells at multiple sites. The inferred large pool of cells and sites to rekindle recrudescent infection highlights the challenges in eradicating HIV.

There is intense interest in developing curative interventions for HIV. How such a cure will be quantified and defined is not known. We applied a series of measurements of HIV persistence to the study of an HIV-infected adult who has exhibited evidence of cure after allogeneic hematopoietic stem cell transplant from a homozygous CCR5Δ32 donor. Samples from blood, spinal fluid, lymph node, and gut were analyzed in multiple laboratories using different approaches. No HIV DNA or RNA was detected in peripheral blood mononuclear cells (PBMC), spinal fluid, lymph node, or terminal ileum, and no replication-competent virus could be cultured from PBMCs. However, HIV RNA was detected in plasma (2 laboratories) and HIV DNA was detected in the rectum (1 laboratory) at levels considerably lower than those expected in ART-suppressed patients. It was not possible to obtain sequence data from plasma or gut, while an X4 sequence from PBMC did not match the pre-transplant sequence. HIV antibody levels were readily detectable but declined over time; T cell responses were largely absent. The occasional, low-level PCR signals raise the possibility that some HIV nucleic acid might persist, although they could also be false positives. Since HIV levels in well-treated individuals are near the limits of detection of current assays, more sensitive assays need to be developed and validated. The absence of recrudescent HIV replication and waning HIV-specific immune responses five years after withdrawal of treatment provide proof of a clinical cure.

Vaccine responses vary by geographic location. We have previously described how HIV-associated inflammation leads to fibrosis of secondary lymph nodes (LNs) and T cell depletion. We hypothesized that other infections may cause LN inflammation and fibrosis, in a process similar to that seen in HIV infection, which may lead to T cell depletion and affect vaccine responses. We studied LNs of individuals from Kampala, Uganda, before and after yellow fever vaccination (YFV) and found fibrosis in LNs that was similar to that seen in HIV infection. We found blunted antibody responses to YFV that correlated to the amount of LN fibrosis and loss of T cells, including T follicular helper cells. These data suggest that LN fibrosis is not limited to HIV infection and may be associated with impaired immunologic responses to vaccines. This may have an impact on vaccine development, especially for infectious diseases prevalent in the developing world.

Tumor response to radiation is associated with improved tumor control and overall survival rate, and p21 expression is a marker of tumor radiosensitivity in patients with rectal cancer. Furthermore, a high proliferating cell nuclear antigen labeling index and a low microvessel count in the preradiation biopsy specimen may be prognostic indicators for tumor recurrence.