Rebecca Pellett Madan

A randomized trial to assess anti-HIV activity in female genital tract secretions and soluble mucosal immunity following application of 1% tenofovir gel.

Background Preclinical and early phase clinical microbicide studies have not consistently predicted the outcome of efficacy trials. To address this gap, candidate biomarkers of microbicide pharmacodynamics and safety were evaluated in a double-blind, placebo-controlled trial of tenofovir gel, the first microbicide to demonstrate significant protection against HIV acquisition. Methods 30 women were randomized to apply a single daily dose of tenofovir or placebo gel for 14 consecutive days. Anti-HIV activity was measured in cervicovaginal lavage (CVL) on Days 0, 3, 7, 14 and 21 by luciferase assay as a surrogate marker of pharmacodynamics. Endogenous activity against E. coli and HSV-2 and concentrations of immune mediators were quantified in CVL as candidate biomarkers of safety. Tenofovir levels were measured in CVL and blood. Results A significant increase in anti-HIV activity was detected in CVL from women who applied tenofovir gel compared to their endogenous anti-HIV activity in genital tract secretions on Day 0 and compared to activity in CVL from women in the placebo group. The activity correlated significantly with CVL concentration of tenofovir (r = 0.6, p<0.001) and fit a sigmoid Emax pharmacodynamic model. Anti-HIV activity in CVL from women who applied tenofovir persisted when virus was introduced in semen, whereas endogenous anti-HIV activity decreased. Tenofovir did not trigger an inflammatory response or induce sustained loss in endogenous antimicrobial activity or immune mediators. Conclusions Tenofovir gel had no deleterious impact on soluble mucosal immunity. The increased anti-HIV activity in CVL, which persisted in the presence of semen and correlated with tenofovir concentration, is consistent with the efficacy observed in a recent clinical trial. These results promote quantified CVL anti-HIV activity as a surrogate of tissue pharmacodynamics and as a potential biomarker of adherence to product. This simple, feasible and inexpensive bioassay may promote the development of models more predictive of microbicide efficacy. Trial Registration ClinicalTrials.gov NCT00594373

Postcoital Bioavailability and Antiviral Activity of 0.5% PRO 2000 Gel: Implications for Future Microbicide Clinical Trials

Background The pharmacokinetics and pharmacodynamics of vaginal microbicides are typically assessed among sexually abstinent women. However, the physical act of sex may modulate gel distribution, and preclinical studies demonstrate seminal plasma interferes with the antiviral activity of several microbicides. This study compared the biological activity and concentration of PRO 2000 in cervicovaginal lavage (CVL) collected in the absence or following coitus. Methods CVL samples were collected from ten heterosexual couples at baseline, after sex, after a single dose of 0.5% PRO 2000 gel and sex, and after gel application without sex. The impact of CVL on HIV-1 infection of TZM-bl cells and HSV-2 infection of CaSki cells was monitored by luciferase and plaque assay, respectively. PRO 2000 concentrations were measured by fluorescence. Results CVL collected after PRO 2000 application significantly inhibited HIV-1 and HSV-2 (p = 0.01). However, the antiviral activity was reduced following sex and no significant protective effect was observed in postcoital CVL obtained in the presence compared to the absence of PRO 2000 for HIV (p = 0.45) or HSV-2 (p = 0.56). Less PRO 2000 was recovered in postcoital CVL, which, in conjunction with interference by seminal plasma, may have contributed to lower antiviral activity. Conclusions Postcoital responses to PRO 2000 differ from precoital measures and the results obtained may provide insights into the clinical trial findings in which there was no significant protection against HIV-1 or HSV-2. Postcoital studies should be incorporated into clinical studies before embarking on large-scale efficacy trials.

Female Genital Tract Secretions Inhibit Herpes Simplex Virus Infection: Correlation with Soluble Mucosal Immune Mediators and Impact of Hormonal Contraception

Citation Shust GF, Cho S, Kim M, Madan RP, Guzman EM, Pollack M, Epstein J, Cohen HW, Keller MJ, Herold BC. Female genital tract secretions inhibit herpes simplex virus infection: correlation with soluble mucosal immune mediators and impact of hormonal contraception. Am J Reprod Immunol 2010; 63: 110–119

Humoral and Cell-Mediated Immune Responses to Monovalent 2009 Influenza A/H1N1 and Seasonal Trivalent Influenza Vaccines in High-Risk Children

OBJECTIVE Humoral and cell-mediated immune responses to monovalent 2009 pandemic influenza A (H1N1/2009) and seasonal trivalent influenza (TIV) vaccines were evaluated in healthy children and children with asthma, sickle cell disease (SCD), systemic lupus erythematosus (SLE), and solid organ transplantation (SOT). STUDY DESIGN Blood was collected from 112 subjects at the time of H1N1/2009 vaccination and 46 ± 15 days later for hemagglutination inhibition titers and γ-interferon ELISPOT responses to H1N1/2009 vaccine and TIV; unvaccinated children also received TIV at enrollment. RESULTS A significant increase in the percentage of subjects with seroprotective hemagglutination inhibition titers to both vaccines was observed in all high-risk groups. Children with asthma and SCD were most likely to achieve seroprotective titers to H1N1/2009, whereas <50% of subjects with SOT and SLE had a seroprotective response. Subjects with SOT and SLE also had lower rates of seroprotection after TIV, and subjects with SLE had the lowest ELISPOT responses to both vaccines. Overall, 73% of healthy children exhibited protective responses to TIV; only 35% achieved seroprotection for H1N1/2009. CONCLUSIONS This evaluation of immune responses to H1N1/2009 in high-risk children suggests suboptimal responses for SOT and SLE subjects, but not for subjects with SCD or asthma. Higher antigen dose, additional dose regimens, or both for immunocompromised children warrant further investigation.

A hybrid strategy for the prevention of cytomegalovirus-related complications in pediatric liver transplantation recipients.

Background. This single center, retrospective study describes experience with a hybrid prevention strategy combining short-course antiviral prophylaxis and preemptive cytomegalovirus (CMV) polymerase chain reaction (PCR) monitoring. Methods. One hundred twenty-two pediatric liver transplantation recipients were followed up for a median of 2.3 years posttransplantation. Subjects received a minimum of 14 days of postoperative ganciclovir, followed by monthly CMV PCR monitoring. Results. Forty-three CMV seronegative recipients received seropositive grafts and were considered high risk for CMV; 79 subjects were routine risk. CMV was detected by PCR in the absence of symptoms in 34.4% of subjects and was more likely in high risk than in routine risk recipients (58.1% vs. 21.8%, P=0.0001). Twelve subjects (9.8%) developed CMV disease (8 high risk vs. 4 routine risk, P=0.03). Three subjects developed acute rejection in the 6 months after detection of CMV, but CMV was preceded by rejection in 13 subjects. There were no mortalities secondary to CMV. A total of 38.5% of subjects were spared antiviral medications beyond their initial postoperative prophylaxis. Conclusions. These results suggest that a hybrid preventative approach for CMV is a reasonable alternative to prolonged antiviral prophylaxis and may reduce unnecessary exposure to antiviral therapy. However, patients who receive intensified immunosuppression after acute rejection are at increased risk for CMV and may require extended prophylaxis and closer monitoring.

Lactobacillus proteins are associated with the bactericidal activity against E. coli of female genital tract secretions.

Background Female genital tract secretions are bactericidal for Escherichia (E.) coli ex vivo. However, the intersubject variability and molecules that contribute to this activity have not been defined. Methods The bactericidal activity and concentration of immune mediators in cervicovaginal lavage (CVL) collected from 99 healthy women were determined. Results CVL reduced the number of E. coli colonies by 68% [−26, 100] (median [range]). CVL were active against laboratory and clinical isolates of E. coli, but were inactive against Lactobacillus species. Bactericidal activity correlated with the concentration of protein recovered (p<0.001), but not with cytokines, chemokines or antimicrobial peptides. Four CVL with>90% inhibitory activity (active) and two with<30% activity were subjected to MS/MS proteomic analysis. 215 proteins were identified and six were found exclusively in active samples. Four of these corresponded to Lactobacillus crispatus or jensenii proteins. Moreover, culture supernatants from Lactobacillus jensenii were bactericidal for E. coli. Conclusion Both host and commensal microbiota proteins contribute to mucosal defense. Identification of these proteins will facilitate the development of strategies to maintain a healthy vaginal microbiome and prevent colonization with pathogenic bacteria such as E. coli that increase the risk for urinary tract infections, preterm labor and perinatal infection.

Female Genital Tract Secretions and Semen Impact the Development of Microbicides for the Prevention of HIV and Other Sexually Transmitted Infections

Citation 
Herold BC, Mesquita PM, Madan RP, Keller MJ. Female genital tract secretions and semen impact the development of microbicides for the prevention of human immunodeficiency virus (HIV) and other sexually transmitted infections. Am J Reprod Immunol 2011; 65: 325–333

The changing face of post-transplant lymphoproliferative disease in the era of molecular EBV monitoring.

Kerkar N, Morotti RA, Madan RP, Shneider B, Herold BC, Dugan C, Miloh T, Karabicak I, Strauchen JA, Emre S. The changing face of post‐transplant lymphoproliferative disease in the era of molecular EBV monitoring.
Pediatr Transplantation 2010: 14:504–511.

Altered biomarkers of mucosal immunity and reduced vaginal Lactobacillus concentrations in sexually active female adolescents.

Background Genital secretions collected from adult women exhibit in vitro activity against herpes simplex virus (HSV) and Escherichia coli (E. coli), but prior studies have not investigated this endogenous antimicrobial activity or its mediators in adolescent females. Methodology/Principal Findings Anti-HSV and anti-E.coli activity were quantified from cervicovaginal lavage (CVL) specimens collected from 20 sexually active adolescent females (15–18 years). Soluble immune mediators that may influence this activity were measured in CVL, and concentrations of Lactobacillus jensenii and crispatus were quantified by PCR from vaginal swabs. Results for adolescents were compared to those obtained from 54 healthy, premenopausal adult women. Relative to specimens collected from adults, CVL collected from adolescent subjects had significantly reduced activity against E. coli and diminished concentrations of protein, IgG, and IgA but significantly increased anti-HSV activity and concentrations of interleukin (IL)-1α, IL-6 and IL-1 receptor antagonist. Vaginal swabs collected from adolescent subjects had comparable concentrations of L. crispatus but significantly reduced concentrations of L. jensenii, relative to adult swabs. Conclusions/Significance Biomarkers of genital mucosal innate immunity may differ substantially between sexually active adolescents and adult women. These findings warrant further study and may have significant implications for prevention of sexually transmitted infections in adolescent females.

Prioritizing prevention of HIV and sexually transmitted infections: first-generation vaginal microbicides.

Purpose of review As the HIV/AIDS pandemic continues unabated, novel control measures for the spread of HIV and other sexually transmitted infections are urgently needed. Topical microbicides are designed to prevent transmission of sexually transmitted infections when applied vaginally. The microbicides discussed in this review may provide a new opportunity for decreasing the spread of HIV and other sexually transmitted infections. Recent findings Epidemiological studies suggest a synergistic relationship between HIV and sexually transmitted infections, particularly between HIV and genital herpes infection. Compounds have been developed to block transmission of HIV-1 and herpes simplex virus, as well as Neisseria gonorrhoea and Chlamydia trachomatis. Several of these compounds have advanced to clinical trials as candidate microbicides. Candidate compounds fall into the following categories: detergents or surfactants that inactivate viral particles, anionic polymers that block attachment of virus to target cells, vaginal acid-buffering agents that maintain a protective vaginal pH, and antiretroviral drugs specific for HIV. Evaluation of the safety of topical microbicides remains problematic. Clinical experiences indicate that current models to assess safety in vitro and in vivo may be insufficient to assess the safety of vaginal microbicides. A critical direction of future studies is to identify which assay(s) provide surrogate laboratory markers of safety that correlate with clinical outcomes. Summary The spread of HIV, and its increasing burden of disease in women, necessitates the development of novel prophylactic strategies. Topical microbicides offer women an empowering preventative option but require vigorous testing for safety and effectiveness.