Results of a phase 1, randomized, placebo-controlled first-in-human trial of griffithsin formulated in a carrageenan vaginal gel

Abstract

HIV pre-exposure prophylaxis (PrEP) is dominated by clinical therapeutic antiretroviral (ARV) drugs. Griffithsin (GRFT) is a non-ARV lectin with potent anti-HIV activity. GRFT's preclinical safety, lack of systemic absorption after vaginal administration in animal studies, and lack of cross-resistance with existing ARV drugs prompted its development for topical HIV PrEP. We investigated safety, pharmacokinetics (PK), pharmacodynamics (PD), and immunogenicity of PC-6500 (0.1% GRFT in a carrageenan (CG) gel) in healthy women after vaginal administration. This randomized, placebo-controlled, parallel group, double-blind first-in-human phase 1 study enrolled healthy, HIV-negative, non-pregnant women aged 24-45 years. In the open label period, all participants (n = 7) received single dose of PC-6500. In the randomized period, participants (n = 13) were instructed to self-administer 14 doses of PC-6500 or its matching CG placebo (PC-535) once daily for 14 days. The primary outcomes were safety and PK after single dose, and then after 14 days of dosing. Exploratory outcomes were GRFT concentrations in cervicovaginal fluids, PD, inflammatory mediators and gene expression in ectocervical biopsies. This trial is registered with ClinicalTrials.gov, number NCT02875119. No significant adverse events were recorded in clinical or laboratory results or histopathological evaluations in cervicovaginal mucosa, and no anti-drug (GRFT) antibodies were detected in serum. No cervicovaginal proinflammatory responses and no changes in the ectocervical transcriptome were evident. Decreased levels of proinflammatory chemokines (CXCL8, CCL5 and CCL20) were observed. GRFT was not detected in plasma. GRFT and GRFT/CG in cervicovaginal lavage samples inhibited HIV and HPV, respectively, in vitro in a dose-dependent fashion. These data suggest GRFT formulated in a CG gel is a safe and promising on-demand multipurpose prevention technology product that warrants further investigation.

Conflict of interest statement

The authors have declared that no competing interests exist.

Figures

Fig 1. CONSORT flow diagram.

Fig 2. Clinical trial design.

Fig 3. GRFT concentrations in CVL from participants in the randomized phase.

GRFT concentrations 4h (or 6h) or 8h after single gel application and 24h after last gel application. 0h represents pre-gel dose BL. Each symbol represents an individual subject. Data below LLOQ are shown as 0. BL and 4h CVL samples from one subject in PC-6500 group were suspected to be switched at the time of collection. This has been adjusted for the presentation. Non-specific background was detected in 4h and 24h time points CVLs from a single subject in placebo group. These data were excluded.

Fig 4. Concentrations of mediators of inflammation in CVLs.

CVLs from participants in the randomized study (n = 10 PC-6500, n = 3 placebo) were collected pre-dose at BL and 24h post last gel application (D15) using saline. One subject (PC-6500 group) was excluded from the analysis as BL and 4h time-point CVLs were suspected to be switched at the time of collection. CVL samples with suspected blood (one post placebo gel, one post PC-6500 and one BL sample in PC-6500 group) were excluded from the analysis. Dotted lines represent lower limit of detection (LLD).

Fig 5. Ectocervical tissue transcriptome before and after repeated PC-6500 or placebo gel administration.

Ectocervical gene expression in biopsies collected at BL and 24h after 14 days repeated gel administration is shown as (A) MDS plot, (B) heatmap and (C, D) volcano plots. MDS plot was prepared through plot MDS function within edgeR (doi: 10.1093/bioinformatics/btp616). The heatmap was constructed by edgeR using top 500 genes based on LogCPM. Induction (red) and inhibition (blue) of expression (z-scores) are shown. The volcano plots of gene expression at BL and post gel administration according to the fold change and adjusted p values (q < 0.05) were prepared through in-house R code. The y-axis corresponds to the mean expression value of log10 (FDR; q), and the x-axis displays the log2 fold change (FC) value. Samples from 9 subjects in PC-6500 group and from n = 3 subjects in placebo group were analyzed. Due to poor RNA quality, sample from one subject in PC-6500 group was excluded from the analysis.

Fig 6. CVL activity against HIV-1 and HPV16 PsV.

(A) TMZ-bl cells were incubated with cell-free HIV-1ADA-M and different dilutions of CVLs collected at BL, 4 or 8h post single GRFT/CG gel administration and 24h after 14 day repeated gel administration (randomized study). The EC50 and 95% CI were calculated using a curve-fitting analysis with GraphPad Prism. The Spearman correlation analysis demonstrated that the higher the concentration of GRFT in CVL, the more potent the antiviral activity against HIV-1ADA-M in the MAGI assay. (B) HeLa cells were incubated with HPV16 PsV and different dilutions of CVLs collected at BL, 4 or 8h post single PC-6500 gel administration and 24h post last gel administration (randomized study). The EC50 and 95% CI were calculated using a curve-fitting analysis with GraphPad Prism. (C) Polarized cervical explant cultures were challenged with 500 TCID50 HIV-1BaL in the presence of CVLs collected at BL; 4, 6 or 8h post single and 24h after 14 day repeated gel administration (randomized study) applied on the epithelial surface for 2h (two explants per condition). Tissues were washed and cultured for 14d. Infection was monitored by HIV gag qRT-PCR using supernatants collected every 3-4d. 3TC (4μl of 500μM stock applied on the epithelium) control was included where feasible. The Spearman correlation analysis demonstrated that the higher the concentration of GRFT in CVLs, the more potent the antiviral activity against HIV-1BaL.