High Resolution Imaging of Epithelial Injury in the Sheep Cervicovaginal Tract: A Promising Model for Testing Safety of Candidate Microbicides

Self Cite

Vaginal microbicides may play an important role in protecting women from HIV infection. A strong synergy between HSV and HIV has been observed, and epidemiological studies demonstrate that HSV infection increases the risk of HIV acquisition. Incorporation of the antiretroviral tenofovir (TFV) along with the antiherpetic acyclovir (ACV) into combination intravaginal rings (IVRs) for sustained mucosal delivery of both compounds could lead to increased microbicide product adherence and efficacy compared with conventional vaginal formulations. A novel, dual-protection "pod IVR" platform developed in-house and delivering ACV and TFV was evaluated in rabbit and sheep models. The devices were safe and exhibited sustained release of both drugs independently and at controlled rates over the 28-day studies. Daily release rates were estimated based on residual drug content of the used devices: rabbits, 343 ؎ 335 g day ؊1 (ACV) and 321 ؎ 207 g day ؊1 (TFV); sheep, 174 ؎ 14 g day ؊1 (ACV) and 185 ؎ 34 g day ؊1 (TFV). Mean drug levels in sheep vaginal samples were as follows: secretions, 5.25 ؎ 7.31 g ml ؊1 (ACV) and 20.6 ؎ 16.2 g ml ؊1 (TFV); cervicovaginal lavage fluid, 118 ؎ 113 ng ml ؊1 (ACV) and 191 ؎ 125 ng ml ؊1 (TFV); tissue, 173 ng g ؊1 (ACV) and 93 ng g ؊1 (TFV). An in vitro-in vivo correlation was established for both drugs and will allow the development of future formulations delivering target levels for prophylaxis and therapy. These data suggest that the IVR based on the pod design has potential in the prevention of transmission of HIV-1 and other sexually transmitted pathogens. Significant progress has been achieved in providing increased access to HIV/AIDS services in several low-and middleincome countries (44). Despite these encouraging findings, women in the developing world remain disproportionately at risk of HIV infection. Seventy-six percent of new HIV infections in sub-Saharan Africa occur in women aged 15 to 24 years (42). The proportion is as high as 90% in South Africa (38). In the absence of a preventative vaccine, effective prophylactic biomedical technologies are urgently required. Campaigns aimed at encouraging monogamy and condom use have had limited success in areas where marriage has been identified as the major risk factor for HIV acquisition in women (3,15,16,21,25).Human immunodeficiency virus type 1 (HIV-1) and herpes simplex virus type 2 (HSV-2), the serotype most commonly associated with genital herpes, are responsible for two intersecting epidemics, where the disease caused by one virus facilitates the transmission of and pathogenesis by the other. A strong synergy between HSV and HIV has been observed, and epidemiological studies demonstrate that HSV infection increases the risk of HIV-1 acquisition (33,34,43). A meta-analysis found that prevalent HSV-2 infection is associated with a threefold-increased risk of HIV acquisition among both men and women. These results suggest that, in areas of high HSV-2 prevalence, a substantial proportion of HIV infection is linked to HSV-2 infection (...

Section: Discussionmentioning

confidence: 99%

“…The sheep vaginal toxicity model has been described as a good model based on gross and microscopic anatomical similarities between the sheep and human vaginal tract (41). Like the human, the sheep vaginal epithelium is stratified squamous tissue, but it is thinner, providing a more sensitive model of toxicity.…”

Section: Discussionmentioning

confidence: 99%

Simultaneous Delivery of Tenofovir and Acyclovir via an Intravaginal Ring

Moss

Malone

Smith

et al. 2012

Self Cite

“…Similar to humans, the sheep vaginal epithelium is stratified squamous and thus is pertinent for toxicological irritation studies, such as the one described herein. The vaginal microflora species and vaginal fluid pH of sheep and women are generally different, although large variation in both variables exists in both populations (29,31,57). Another limitation of sheep is their seasonal breeding and shorter estrous cycle (approximately 17 days) compared to women, a difference which may potentially alter drug pharmacokinetics via changes in vaginal epithelium thickness and permeability (33,39).…”

Section: Fig 7 Proximal and Distal Tfv Vaginal Tissue Concentrations mentioning

confidence: 99%

“…The modest increase in both leukocytic infiltration and vaginal irritation potential of sheep receiving placebo and TFV IVRs warrants further evaluation to determine any ring and/or TFV effect on long-term safety and prophylactic efficacy. Though sheep have been used for the systemic safety evaluation of microbicide IVRs (37) and to evaluate the effects of candidate vaginal microbicides on the epithelium (56,57), they are yet to be considered a validated model for the testing of vaginal microbicide products. Nonetheless, this is the first time, to the best of the authors' knowledge, in which the vaginal irritation scoring methodology developed by Eckstein et al (9) has been applied to sheep.…”

Section: Fig 7 Proximal and Distal Tfv Vaginal Tissue Concentrations mentioning

confidence: 99%

A 90-Day Tenofovir Reservoir Intravaginal Ring for Mucosal HIV Prophylaxis

Johnson

Clark

Albright

et al. 2012

117

A vaginal gel containing the antiretroviral tenofovir (TFV) recently demonstrated 39% protection against HIV infection in women. We designed and evaluated a novel reservoir TFV intravaginal ring (IVR) to potentially improve product effectiveness by providing a more controlled and sustained vaginal dose to maintain cervicovaginal concentrations. Polyurethane tubing of various hydrophilicities was filled with a high-density TFV/glycerol/water semisolid paste and then end-sealed to create IVRs. In vitro, TFV release increased with polyurethane hydrophilicity, with 35 weight percent water-swelling polyurethane IVRs achieving an approximately 10-mg/day release for 90 days with mechanical stiffness similar to that of the commercially available NuvaRing. This design was evaluated in two 90-day in vivo sheep studies for TFV pharmacokinetics and safety. Overall, TFV vaginal tissue, vaginal fluid, and plasma levels were relatively time independent over the 90-day duration at approximately 10 4 ng/g, 10 6 ng/g, and 10 1 ng/ml, respectively, near or exceeding the highest observed concentrations in a TFV 1% gel control group. TFV vaginal fluid concentrations were approximately 1,000-fold greater than levels shown to provide significant protection in women using the TFV 1% gel. There were no toxicological findings following placebo and TFV IVR treatment for 28 or 90 days, although slight to moderate increases in inflammatory infiltrates in the vaginal epithelia were observed in these animals compared to naïve animals. In summary, the controlled release of TFV from this reservoir IVR provided elevated sheep vaginal concentrations for 90 days to merit its further evaluation as an HIV prophylactic. R ecent progress in antiretroviral HIV prevention research has advanced the field from concept toward medical practice (46). The CAPRISA 004 study demonstrated that a vaginal gel containing the reverse transcriptase inhibitor tenofovir (TFV) was partially effective in preventing HIV transmission in women (1), with significant protection observed in women who maintained preventative TFV concentrations of at least 1,000 ng/ml in vaginal fluid (23). However, the overall effectiveness (39%) was likely reduced by poor user adherence to the inconvenient before-andafter-sex dosing regimen. The correlation of adherence and TFV vaginal fluid concentrations to protection was a key finding (23,24), indicating the need for vaginal drug delivery systems that attain and maintain elevated user adherence and vaginal drug concentrations. More recently, the VOICE trial tested the same TFV 1% gel formulation as CAPRISA 004 but with a once-daily dosage regimen and failed to show any effectiveness in women. Here, as well, low adherence may have contributed to the gel's inability to prevent HIV transmission (54). As a result, we (6, 21) and others (4,35,36,44,49) aim to develop TFV drug delivery systems to provide sustained protective vaginal tissue concentrations and potentially increase user adherence.The micromolar anti-HIV activity of TFV motivated sel...

“…The sheep has a number of characteristics that make it suitable for use in preclinical evaluations of microbicides. In particular, the cervix and vagina are anatomically and histologically similar to those of women (9), and since the dimensions of the lower female reproductive tract are also similar in the two species, intravaginal products intended for human use, including medical devices, such as intravaginal rings, can be evaluated in sheep without modification. In addition, sheep are widely available, relatively inexpensive, and have a temperament that makes them easy to handle and examine under laboratory conditions; therefore, they can be used in studies conducted in compliance with the principles of good laboratory practice (GLP).…”

mentioning

confidence: 99%

The Sheep as a Model of Preclinical Safety and Pharmacokinetic Evaluations of Candidate Microbicides

Holt

Cameron²,

Dias³

et al. 2015

dWhen developing novel microbicide products for the prevention of HIV infection, the preclinical safety program must evaluate not only the active pharmaceutical ingredient but also the product itself. To that end, we applied several relatively standard toxicology study methodologies to female sheep, incorporating an assessment of the pharmacokinetics, safety, tolerability, and local toxicity of a dapivirine-containing human vaginal ring formulation (Dapivirine Vaginal Ring-004). We performed a 3-month general toxicology study, a preliminary pharmacokinetic study using drug-loaded vaginal gel, and a detailed assessment of the kinetics of dapivirine delivery to plasma, vaginal, and rectal fluid and rectal, vaginal, and cervical tissue over 28 days of exposure and 3 and 7 days after removal of the ring. The findings of the general toxicology study supported the existing data from both preclinical and clinical studies in that there were no signs of toxicity related to dapivirine. In addition, the presence of the physical dapivirine ring did not alter local or systemic toxicity or the pharmacokinetics of dapivirine. Pharmacokinetic studies indicated that the dapivirine ring produced significant vaginal tissue levels of dapivirine. However, no dapivirine was detected in cervical tissue samples using the methods described here. Plasma and vaginal fluid levels were lower than those in previous clinical studies, while there were detectable dapivirine levels in the rectal tissue and fluid. All tissue and fluid levels tailed off rapidly to undetectable levels following removal of the ring. The sheep represents a very useful model for the assessment of the safety and pharmacokinetics of microbicide drug delivery devices, such as the vaginal ring. With an estimated 2.3 million new human immunodeficiency virus type 1 (HIV-1) infections in 2012 (1) and a total number of people living with HIV estimated at 35.3 million in the same year, the need for effective prevention strategies remains critical. Although barrier prevention strategies, such as male and female condoms, are effective, and behavioral programs teaching the benefits of monogamy and abstinence have shown some success (2-4), they have not significantly impacted the epidemic. One area of key concern is sub-Saharan Africa, where 57% of HIV-infected people are women (1). In order to provide prevention strategies to women in this high-risk population, locally (vaginally or rectally) applied products containing potent antiretroviral drugs are being developed by many different groups. These microbicides represent one of the most promising prophylactic strategies currently being developed in terms of being simple to self-administer, relatively easy to produce, and by virtue of the lower drug levels required compared to those with systemic approaches, the potential to be within reach of the health system and nongovernmental organization (NGO) budgets in the developing world. A key advantage to the topical microbicide approach is also that the same targeted active ingredient...