Human immunodeficiency virus type-1 episomal cDNA in semen

BACKGROUND: Episomal 2-long terminal repeat (LTR) HIV-1 cDNA, a by-product of HIV-1 infection, is used in clinical trials as a marker for ongoing viral replication. It would be useful to employ 2-LTR cDNA to monitor cryptic HIV-1 infection in the genital tract of men on antiretroviral therapy (ART) to predict the evolution of sexually transmissible drug-resistant HIV-1, but studies thus far have failed to detect this marker in semen. The objectives of this study were: 1) to use a technique that maximizes DNA recovery from HIV-1 infected white blood cells in semen to determine if episomal 2-LTR cDNA is detectable in semen of ART-naïve men with other evidence of genital tract HIV-1 infection, and 2) to compare levels of HIV-1 2-LTR cDNA, RNA, and proviral DNA in semen from HIV-1+ men on ART. RESULTS: Using a somatic cell DNA extraction technique, 2-LTR cDNA was detected by PCR/ELISA in 4 out of 8 semen samples from ART-naïve men selected for other signs of seminal HIV-1 infection (positive controls). Southern blot and DNA sequencing confirmed that the amplified sequences were HIV-1 2-LTR cDNA; copy numbers ranged from 55 to 504 copies/sample. Two semen samples from a cohort of 22 HIV-1-infected men on dual nucleoside therapy, one with and one without detectable seminal HIV-1 RNA, were 2-LTR cDNA positive (336 and 8,560 copies/sample). Following addition of indinavir to the therapy regimen, no semen samples from 21 men with controlled peripheral and seminal viral loads were 2-LTR cDNA positive at 1 and 6 month time points, despite the persistence of HIV-1 proviral DNA+ semen cells and seminal cytomegalovirus (CMV) shedding in some cases. However, one individual who failed indinavir therapy and later developed distinct protease inhibitor (PI) drug resistance mutations in semen, maintained elevated levels of HIV-1 RNA and 2-LTR cDNA in semen. CONCLUSION: 2-LTR HIV-1 cDNA is detectable in semen of HIV-1-infected men. Two men on ART had 2-LTR HIV-1 cDNA in semen, suggesting that this marker may prove to be useful to monitor HIV-1 infection in the genital tract of men on ART to predict the evolution of drug resistance mutations in semen

Human immunodeficiency virus type-1 episomal cDNA in semen

Abstract Background Episomal 2-long terminal repeat (LTR) HIV-1 cDNA, a by-product of HIV-1 infection, is used in clinical trials as a marker for ongoing viral replication. It would be useful to employ 2-LTR cDNA to monitor cryptic HIV-1 infection in the genital tract of men on antiretroviral therapy (ART) to predict the evolution of sexually transmissible drug-resistant HIV-1, but studies thus far have failed to detect this marker in semen. The objectives of this study were: 1) to use a technique that maximizes DNA recovery from HIV-1 infected white blood cells in semen to determine if episomal 2-LTR cDNA is detectable in semen of ART-naïve men with other evidence of genital tract HIV-1 infection, and 2) to compare levels of HIV-1 2-LTR cDNA, RNA, and proviral DNA in semen from HIV-1+ men on ART. Results Using a somatic cell DNA extraction technique, 2-LTR cDNA was detected by PCR/ELISA in 4 out of 8 semen samples from ART-naïve men selected for other signs of seminal HIV-1 infection (positive controls). Southern blot and DNA sequencing confirmed that the amplified sequences were HIV-1 2-LTR cDNA; copy numbers ranged from 55 to 504 copies/sample. Two semen samples from a cohort of 22 HIV-1-infected men on dual nucleoside therapy, one with and one without detectable seminal HIV-1 RNA, were 2-LTR cDNA positive (336 and 8,560 copies/sample). Following addition of indinavir to the therapy regimen, no semen samples from 21 men with controlled peripheral and seminal viral loads were 2-LTR cDNA positive at 1 and 6 month time points, despite the persistence of HIV-1 proviral DNA+ semen cells and seminal cytomegalovirus (CMV) shedding in some cases. However, one individual who failed indinavir therapy and later developed distinct protease inhibitor (PI) drug resistance mutations in semen, maintained elevated levels of HIV-1 RNA and 2-LTR cDNA in semen. Conclusion 2-LTR HIV-1 cDNA is detectable in semen of HIV-1-infected men. Two men on ART had 2-LTR HIV-1 cDNA in semen, suggesting that this marker may prove to be useful to monitor HIV-1 infection in the genital tract of men on ART to predict the evolution of drug resistance mutations in semen.</p

Condylomata acuminata (anogenital warts) contain accumulations of HIV-1 target cells that may provide portals for HIV transmission

Background: Condylomata acuminata [anogenital warts (AGW)] are prevalent in HIV-infected individuals and sexually active populations at risk for HIV acquisition, and have been associated with HIV transmission. We compared AGW to control tissue for abundance, types and location of HIV-target cells, and for susceptibility to HIV infection in vitro, to provide biological evidence that AGW facilitate HIV transmission. Methods: We used immunohistology to identify HIV-target cells in AGW and control skin. We also inoculated AGW and control tissue from HIV-negative men with HIV in vitro, and assessed infection by TZM-bl and p24 assays. Results: CD1a+ dendritic cells, CD4+ T cells and macrophages were significantly more abundant in the epidermis of AGW than control tissue. These HIV target cells also often appeared in large focal accumulations in the dermis of AGW. Two out of 8 AGW vs. 0 of 8 control tissues showed robust infection with HIV in vitro. Conclusions: Compared to normal skin, AGW contain significantly higher concentrations of HIV-target cells that may be susceptible to HIV infection. Condylomata may thus promote HIV transmission, especially in the setting of typical lesion vascularity and friability. Prevention or treatment of AGW may decrease the sexual transmission of HIV

Safety, acceptability, and pharmacokinetics of a monoclonal antibody-based vaginal multipurpose prevention film (MB66): A Phase I randomized trial.

BackgroundMB66 film is a multipurpose prevention technology (MPT) product with monoclonal antibodies (mAbs) against HIV-1 (VRC01-N) and HSV-1 and 2 (HSV8-N). The mAbs were produced by transient expression in Nicotiana benthamiana (N). We conducted a Phase I clinical trial to assess the safety, pharmacokinetics (PK), and ex vivo efficacy of single and repeated doses of MB66 when used intravaginally.Methods and findingsThe clinical trial enrolled healthy reproductive-aged, sexually abstinent women. In Segment A, 9 women received a single MB66 film which was inserted into the vaginal posterior fornix by a clinician. In Segment B, 29 women were randomly assigned to MB66 (Active) or Placebo film groups and were instructed to insert 1 film vaginally for 7 consecutive days. Visits and clinical sampling occurred predose and at various time points after single and repeated film doses. The primary endpoint was number of adverse events (AEs) Grade 2 or higher related to product use. Secondary endpoints included film dissolution rate, Nugent score (a Gram stain scoring system to diagnose bacterial vaginosis), vaginal pH, post-use survey results, cytokine concentrations in cervicovaginal lavage (CVL) specimens (assessed by Luminex assay), mAb concentrations in vaginal fluid collected from 4 sites (assessed by ELISA), and HIV and HSV neutralization activity of CVL samples ex vivo (assessed by TZM-bl and plaque reduction assay, respectively). The product was generally safe and well tolerated, with no serious AEs recorded in either segment. The AEs in this study were primarily genitourinary in nature with the most commonly reported AE being asymptomatic microscopic hematuria. There were no differences in vaginal pH or Nugent scores or significant increases in levels of proinflammatory cytokines for up to 7 days after film insertion in either segment or between Active and Placebo groups. Acceptability and willingness to use the product were judged to be high by post-use surveys. Concentrations of VRC01-N and HSV8-N in vaginal secretions were assessed over time to generate pharmacokinetic curves. Antibody levels peaked 1 hour postdosing with Active film (median: 35 μg/mL) and remained significantly elevated at 24 hours post first and seventh film (median: 1.8 μg/mL). Correcting for sample dilution (1:20), VRC01-N concentrations ranged from 36 to 700 μg/mL at the 24-hour time point, greater than 100-fold the IC50 for VRC01 (0.32 μg/mL); HSV8-N concentrations ranged from 80 to 601 μg/mL, well above the IC50 of 0.1 μg/m. CVL samples collected 24 hours after MB66 insertion significantly neutralized both HIV-1 and HSV-2 ex vivo. Study limitations include the small size of the study cohort, and the fact that no samples were collected between 24 hours and 7 days for pharmacokinetic evaluation.ConclusionsSingle and repeated intravaginal applications of MB66 film were safe, well tolerated, and acceptable. Concentrations and ex vivo bioactivity of both mAbs in vaginal secretions were significantly elevated and thus could provide protection for at least 24 hours postdose. However, further research is needed to evaluate the efficacy of MB66 film in women at risk for HIV and HSV infection. Additional antibodies could be added to this platform to provide protection against other sexually transmitted infections (STIs) and contraception.Trial registrationClinicalTrials.gov NCT02579083