Meredith R. Clark

Despite advances in oral and injectable HIV prevention options and oral prophylaxis for sexually transmitted infections (STIs) of bacterial origin, there remains a critical need for effective on-demand topical (vaginal and rectal) products for pre- and post-exposure prophylaxis (PrEP and PEP). To fill this gap, we have developed single and first-in-kind multi-active topical inserts for bacterial STIs and HIV/STIs prevention. We have formulated two different inserts, one containing doxycycline (DOX) at 10, 50, and 100 mg doses for bacterial STI prevention, and a multipurpose prevention product (TED insert) that combines DOX (10 mg) with the antiretrovirals tenofovir alafenamide (TAF; 20 mg) and elvitegravir (EVG; 16 mg) to target both bacterial STIs and HIV. Inserts were manufactured using a simple and cost-effective process. Drug loading was within 95-105% of the labeled amount, confirming a robust manufacturing process. In vitro, the inserts disintegrated within 10 min with > 95% drug release within 60 min. The dissolution behavior of DOX inserts showed surface erosion but was affected by medium volume and drug amount. The inserts met key physicochemical targets: hardness (5-8 kg), friability (< 1%), moisture content (< 2%), and osmolality (< 550 mOsm/kg). Based on 6-month storage stability, DOX inserts maintained their physicochemical properties, indicating a shelf life of > 2 years. Preliminary 1-month stability of TED inserts under accelerated conditions showed preservation of their physicochemical properties. These data describe the formulation development and in vitro characterization of topical inserts containing DOX alone or in combination with antiretrovirals (TAF and EVG). Both inserts offer a novel, on-demand topical STI prevention option that supports flexible PrEP/PEP use by both women and men.

Purpose: Despite advances in oral and injectable HIV prevention options and oral prophylaxis for sexually transmitted infections (STIs) of bacterial origin, there remains a critical need for effective on-demand topical (vaginal/rectal) products for pre- and post-exposure prophylaxis (PrEP and PEP). To fill this gap, we have developed single and first-in-kind multi-active topical inserts for bacterial STIs and HIV/STIs prevention. Methods: We have formulated two different inserts, one containing doxycycline (DOX) at 10, 50, and 100mg doses for bacterial STI prevention, and a multipurpose prevention product (TED insert) that combines DOX (10mg) with the antiretrovirals tenofovir alafenamide (TAF; 20mg) and elvitegravir (EVG; 16mg) to target both bacterial STIs and HIV. Results: Inserts were manufactured through a simple, cost-effective process. Drug loading was within 95-105% of the labeled amount, confirming a robust manufacturing process. In vitro, they disintegrated within 10min with >95% drug release within 60min. The dissolution behavior of DOX inserts showed surface erosion but was affected by medium volume and drug amount. The inserts met key physicochemical targets: hardness (5-8kg), friability (<1%), moisture content (<2%), and osmolality (2years. Preliminary 1-month stability of TED inserts under accelerated conditions showed preservation of their physicochemical properties. Conclusion: This study represents the first formulation development report on topical inserts containing DOX alone or in combination with antiretrovirals. Both inserts offer a novel, on-demand topical STI prevention option that supports flexible PrEP/PEP use by both women and men.

On-demand topical inserts for HIV prevention may be a good option for people who have a low frequency of sexual activity. We recently demonstrated in a preclinical macaque model that rectal application of inserts containing tenofovir alafenamide fumarate (TAF) and elvitegravir (EVG) conferred protection against SHIV infection (93% efficacy) when applied as pre-exposure prophylaxis 4 hours before SHIV exposure. Here, we show that the same inserts provide significant postexposure protection (82.9% efficacy) when applied 4 hours after SHIV exposure. Our findings further support the ongoing clinical development of TAF/EVG inserts for on-demand HIV prevention.

Surface-enhanced Raman spectroscopy (SERS) has exceptional analytical sensitivity and selectivity. However, SERS irreproducibility presents an obstacle when using it for precise quantitative measurements. In this study, colloidal nanoparticles evaporated to dryness are used as a SERS active surface for the detection of the HIV drug emtricitabine (FTC; trade name Emtriva). Despite the irreproducibility of the SERS resulting from the stochastic process of evaporation, using a SERS scanning instrument, the SERS enhancement factors of spatially resolved spectra have a well-defined distribution of signals for a given analyte concentration. This distribution follows a power law function ranging from weak (very abundant signals) to very strong (sparse signals). By definition, a power law distribution cannot have a true mean. Hence a cumulative distribution function was used to model the concentration of emtricitabine, and calibration curves were constructed. For stochastically generated quantitative data sets, the precision of this approach is superior to methods utilizing signal averaging and improves analytical sensitivity.

HIV/AIDS remains a global public health issue, and products available for the prevention of HIV infections are limited, especially those for short-acting, on-demand, user-controlled applications. Topical inserts are products that can be applied vaginally or rectally and have been explored as drug delivery systems. To fill the gap in the HIV prevention product pipeline, CONRAD has developed a topical insert containing tenofovir alafenamide fumarate (TAF) and elvitegravir (EVG), two potent and synergistic antiretrovirals, as a simple, low-cost, and discreet option that can be self-administered vaginally and/or rectally, before and after coitus. In this review, we have described the development path of the TAF/EVG insert up to its current point in clinical testing, highlighting findings from acceptability, preclinical safety, pharmacokinetics, and efficacy evaluations and early clinical studies. In summary, the TAF/EVG inserts are stable, easy to manufacture, low-cost, acceptable, and show highly promising preclinical and clinical results for on-demand topical pre- or post-exposure HIV prevention.

Vaginal inserts that can be used on demand before or after sex may be a desirable human immunodeficiency virus (HIV) prevention option for women. We recently showed that inserts containing tenofovir alafenamide fumarate (TAF, 20 mg) and elvitegravir (EVG, 16 mg) were highly protective against repeated simian/human immunodeficiency virus (SHIV) vaginal exposures when administered to macaques 4 hours before or after virus exposure (93% and 100%, respectively). Here, we show in the same macaque model that insert application 8 hours or 24 hours after exposure maintains high efficacy (94.4% and 77.2%, respectively). These data extend the protective window by TAF/EVG inserts and inform their clinical development for on-demand prophylaxis in women.

Surface-enhanced Raman spectroscopy (SERS) is an ultrasensitive spectroscopic technique that generates signal-enhanced fingerprint vibrational spectra of small molecules. However, without rigorous control of SERS substrate active sites, geometry, surface area, or surface functionality, SERS is notoriously irreproducible, complicating the consistent quantitative analysis of small molecules. While evaporatively prepared samples yield significant SERS enhancement resulting in lower detection limits, the distribution of these enhancements along the SERS surface is inherently stochastic. Acquiring spatially resolved SERS spectra of these dried surfaces, we have shown that this enhancement is governed by a power law as a function of analyte concentration. Consequently, by definition, there is no true mean of SERS enhancement, requiring an alternative approach to achieve reproducible quantitative results. In this study, we introduce a new method of analysis of SERS data using a cumulative distribution function (CDF). The antiviral drug tenofovir (TFV) in an aqueous matrix was quantified down to a clinically relevant concentration of 25 ng/mL using hydroxylamine-reduced silver colloids evaporated to dryness. The data presented in this study provide a rationale for the benefits of combining a novel statistical approach using CDFs with simple and inexpensive experimental techniques to increase the precision, accuracy, and analytical sensitivity of aqueous TFV quantification by SERS. TFV calibration curves generated using CDF analysis showed higher analytical sensitivity (in the form of a normalized calibration curve average slope increase of 0.25) compared to traditional SERS intensity calculations. A second aliquot of nanoparticles and analyte dried on the SERS surface followed by CDF analysis showed further analytical sensitivity with a normalized calibration curve slope increase of 0.23 and decreased variation among replicates represented by an average standard deviation decrease of 0.02 with a second aliquot. The quantitative analysis of SERS data using CDFs presented here shows promise to be a reproducible method for quantitative analysis of SERS data, a significant step toward implementing SERS as an analytical method in clinical and industrial settings.

While oral pre-exposure prophylaxis (PrEP) can substantially reduce HIV risk, there are important barriers to uptake and adherence. We explored preferences for long-acting injectable and implantable PrEP among women and girls in Eswatini, Kenya, and South Africa. We conducted an online quantitative survey and discrete choice experiment (DCE) among adolescent girls (15–17), young women (18–29), and adult women (30–49). Participants completed a survey about their demographics and behavior and a DCE with 5 attributes (format, insertion location, number of insertions, dual-protection, and palpability). We recruited 1236 respondents (Eswatini = 420; Kenya = 350; South Africa = 493) in May 2022. Most participants were sexually active (72%), nearly 29% of whom reported recently engaging in transactional sex. 46% had heard of oral PrEP, but of those, only 16% reported having ever used it. Product format and dual-protection were significant predictors of product choice. Relative to a 2-month injection, participants had 1.76 times the odds (95% CI 1.08–2.04) of choosing a 6-month injectable, and 1.70 the odds (95% CI 1.06–1.92) of choosing a 12-month removable implant. Compared to a single-indication product, respondents had 2.46 times the odds (95% CI 1.04–2.68) of preferring a product also protecting against pregnancy, and 2.81 the odds (95% CI 1.04–3.05) of choosing a product that also protected against STIs. Adolescent girls and women in these countries showed strong preferences for longer-acting PrEP product formats, as well as those offering dual-protection. Introduction of long-acting options could improve PrEP uptake and reduce HIV burdens in east and southern African settings.

This study introduces a convenient and ultra-sensitive method of detection and quantification of the antiviral drug, tenofovir (TFV), by surface-enhanced Raman spectroscopy (SERS). Novel spatially resolved instrumentation for spectral acquisition and subsequent statistical analysis for hot spot selection was developed for convenient quantification of TFV in an aqueous matrix. Methods of statistical analysis include the use of partial least squares (PLS) regression vector analysis and spectral ranking by quality indices computed using CHAOS theory. Hydroxylamine-reduced Ag colloidal nanoparticles evaporated to dryness on an aluminum well-plate were used as the SERS substrate. To our knowledge, quantification of TFV down to 25 ng/mL by SERS, comprising clinically relevant concentrations, has not been previously reported. Furthermore, in this work we propose a novel method of quantification of aqueous TFV standards by SERS using statistical treatment of data by PLS and CHAOS theory. Based on these data, we propose future studies to develop a method of TFV detection and quantification in biological samples, beneficial to clinicians for rapid assessment of drug adherence during the treatment and prevention of viral diseases.

Background: New multi-purpose prevention technology (MPT) products are needed to prevent human immunodeficiency virus (HIV) and herpes simplex virus type 2 (HSV2). In this study, we evaluated a fast-dissolve insert that may be used vaginally or rectally for prevention of infection.Objective: To describe the safety, acceptability, multi-compartment pharmacokinetics (PK), and in vitro modeled pharmacodynamics (PD) after a single vaginal dose of an insert containing tenofovir alafenamide (TAF) and elvitegravir (EVG) in healthy women.Methods: This was a Phase I, open-label, study. Women (n=16) applied one TAF (20mg)/EVG (16mg) vaginal insert and were randomized (1:1) to sample collection time groups for up to 7 days post dosing. Safety was assessed by treatment-emergent adverse events (TEAEs). EVG, TAF and tenofovir (TFV) concentrations were measured in plasma, vaginal fluid and tissue, and TFV-diphosphate (TFV-DP) concentration in vaginal tissue. PD was modeled in vitro by quantifying the change in inhibitory activity of vaginal fluid and vaginal tissue against HIV and HSV2 from baseline to after treatment. Acceptability data was collected by a quantitative survey at baseline and post treatment.Results: The TAF/EVG insert was safe, with all TEAEs graded as mild, and acceptable to participants. Systemic plasma exposure was low, consistent with topical delivery, while high mucosal levels were detected, with median TFV vaginal fluid concentrations exceeding 200,000 ng/mL and 1,000 ng/mL for up to 24 hours and 7 days post dosing, respectively. All participants had vaginal tissue EVG concentrations of > 1 ng/mg at 4 and 24 hours post dosing. The majority had tissue TFV-DP concentrations exceeding 1000 fmol/mg by 24 – 72 hours post dosing. Vaginal fluid inhibition of HIV-1 and HSV-2 in vitro significantly increased from baseline and was similarly high at 4 and 24 hours post dosing. Consistent with high tissue TFV-DP concentrations, p24 HIV antigen production from ectocervical tissues infected ex vivo with HIV-1 significantly decreased from baseline at 4 hours post dosing. HSV-2 production from tissue also decreased post treatment.Conclusions: A single dose of TAF/EVG inserts met PK benchmarks, with PK data supporting an extended window of high mucosal protection. PD modeling supports mucosal protection against both HIV-1 and HSV-2. The inserts were safe and highly acceptable.Clinical trial registration: ClinicalTrials.gov, identifier NCT03762772.