Structured observations reveal slow HIV-1 CTL escape.

The existence of viral variants that escape from the selection pressures imposed by cytotoxic T-lymphocytes (CTLs) in HIV-1 infection is well documented, but it is unclear when they arise, with reported measures of the time to escape in individuals ranging from days to years. A study of participants enrolled in the SPARTAC (Short Pulse Anti-Retroviral Therapy at HIV Seroconversion) clinical trial allowed direct observation of the evolution of CTL escape variants in 125 adults with primary HIV-1 infection observed for up to three years. Patient HLA-type, longitudinal CD8+ T-cell responses measured by IFN-γ ELISpot and longitudinal HIV-1 gag, pol, and nef sequence data were used to study the timing and prevalence of CTL escape in the participants whilst untreated. Results showed that sequence variation within CTL epitopes at the first time point (within six months of the estimated date of seroconversion) was consistent with most mutations being transmitted in the infecting viral strain rather than with escape arising within the first few weeks of infection. Escape arose throughout the first three years of infection, but slowly and steadily. Approximately one third of patients did not drive any new escape in an HLA-restricted epitope in just under two years. Patients driving several escape mutations during these two years were rare and the median and modal numbers of new escape events in each patient were one and zero respectively. Survival analysis of time to escape found that possession of a protective HLA type significantly reduced time to first escape in a patient (p = 0.01), and epitopes escaped faster in the face of a measurable CD8+ ELISpot response (p = 0.001). However, even in an HLA matched host who mounted a measurable, specific, CD8+ response the average time before the targeted epitope evolved an escape mutation was longer than two years.

Toxicogenomics screening of small molecules using high-density, nanocapillary real-time PCR.

Compounds which induce toxicity through similar mechanisms lead to characteristic gene expression patterns. The concept that structurally similar compounds may have similar biological profiles, the so-called generalized neighborhood behavior, is less obvious to be demonstrated. We screened 625 compounds from a fully combinatorial library for their gene expression profiles in vitro over a selected toxicity panel of 56 genes. We used the novel nanocapillary, quantitative real-time PCR OpenArray technology that is coupling outstanding analytical performance with the medium-throughput ideal for such a sample-per-feature ratio. Applying a hybrid clustering on the gene expression data, correlation was analyzed between molecular scaffold and biological fingerprint. Structurally highly dissimilar, but similarly hepatotoxic compounds show similar fingerprint on our toxicity panel, however compounds of the same scaffold and of unknown biological effect do not always share similar fingerprints. Out of 12 different scaffolds, 4 families show non-correlating, uniform distribution among clusters whilst 8 families show neighborhood behavior of varying strength. Structurally not similar compounds may have highly similar biological activity, on the other hand, compounds of the same scaffold family do not all share the same biological effects based on toxicology related gene expression fingerprint.