Search for: All records

Background: Croton oligandrus Pierre & Hutch is a tropical tree that grows in West and Central Africa, used in ethnomedicine to treat cancer, diabetes, headaches, convulsions, urinary diseases, and inflammatory diseases. As other Croton species have been observed to possess chemical compounds that target HIV latency-reversal, we hypothesized that this species may have similar properties. Aim of the study: The identification of extracts and compounds of this species, which have HIV-1 latency-reversing activity in J-Lat T cell lines. Methods: The stem bark was obtained, air-dried, powdered, and extracted using dichloromethane. In vitro flow cytometry was used to monitor GFP expression, a marker of HIV latency reversal, following treatment of J-Lat T cells with extracts and compounds. Results: Four extracts were found to reverse HIV latency, the most active extract showing better activity (ie, latency reversal in 69.7 ± 7.1% [mean ± s.e.m.] of J-Lat 10.6 cells at 1 µg/mL) than control agents prostratin (46.2 ± 9.5% at 1.2 µg.mL) and the "Mukungulu" (Croton megalobotrys) extract (34.9 ± 24.2% at 1 µg/mL). Extracts reversed HIV latency through mechanisms over and above protein kinase C (PKC) activation and distinct from histone deacetylase (HDAC) inhibition. The most active extract also synergized with the control HDAC inhibitor romidepsin but did not synergize with other extracts. Isolated compounds (β-Stigmasterol and lupeol) had limited but consistent latency reversal on their own. Conclusion: The plant extracts and compounds reverse HIV latency through mechanisms additional to PKC activation and/or synergize with romidepsin in vitro. Extracts and compounds from this plant may enhance the activity of current HIV latency-reversing agents being assessed in HIV cure studies. 

A scalable approach for quantifying intact HIV-1 proviruses is critical for basic research and clinical trials directed at HIV-1 cure. The intact proviral DNA assay (IPDA) is a novel approach to characterizing the HIV-1 reservoir, focusing on the genetic integrity of individual proviruses independent of transcriptional status. It uses multiplex digital droplet PCR to distinguish and separately quantify intact proviruses, defined by a lack of overt fatal defects such as large deletions and APOBEC3G-mediated hypermutation, from the majority of proviruses that have such defects. This distinction is important because only intact proviruses cause viral rebound on ART interruption. To evaluate IPDA performance and provide benchmark data to support its implementation, we analyzed peripheral blood samples from 400 HIV-1 + adults on ART from several diverse cohorts, representing a robust sample of treated HIV-1 infection in the United States. We provide direct quantitative evidence that defective proviruses greatly outnumber intact proviruses (by >12.5 fold). However, intact proviruses are present at substantially higher frequencies (median, 54/10 6 CD4 + T cells) than proviruses detected by the quantitative viral outgrowth assay, which requires induction and in vitro growth (∼1/10 6 CD4 + T cells). IPDA amplicon signal issues resulting from sequence polymorphisms were observed in only 6.3% of individuals and were readily apparent and easily distinguished from low proviral frequency, an advantage of the IPDA over standard PCR assays which generate false-negative results in such situations. The large IPDA dataset provided here gives the clearest quantitative picture to date of HIV-1 proviral persistence on ART.