SAMMA, a mandelic acid condensation polymer, inhibits dendritic cell-mediated HIV transmission.

SAMMA, a mandelic acid condensation polymer, exhibits a broad antimicrobial activity against several sexually transmitted pathogens including human immunodeficiency virus (HIV). Here we demonstrated that SAMMA suppressed HIV transmission by dendritic cells (DCs), one of the first target cells for primary infection. The greatest inhibitory effect was achieved when SAMMA was present during the co-culture with target cells. The inhibitory effect of SAMMA on DC-mediated HIV transmission was not due to cytotoxicity. Analysis of the level of DC-associated HIV p24 antigen revealed that SAMMA prevented HIV internalization by DCs when the virus was pre-incubated with the compound. In contrast, pre-incubation of DCs with SAMMA followed by wash-off did not affect the amount of cell-associated HIV p24 antigen. In addition, SAMMA blocked HIV glycoprotein-mediated cell-cell fusion. This study suggests that SAMMA prevents HIV infection through multiple mechanisms.

SAMMA induces premature human acrosomal loss by Ca2+ signaling dysregulation.

SAMMA is licensed for development as a contraceptive microbicide. Understanding mechanisms of its biological activity is prerequisite to designing more active second generation products. This study examined Ca(2+) involvement in SAMMA-induced premature acrosomal loss (SAL) in noncapacitated human spermatozoa. SAMMA causes acrosomal loss (AL) in a dose-dependent manner (ED(50) = 0.25 microg/mL). SAL requires extracellular Ca(2+) (ED(50) = 85 microM). SAL is inhibited by verapamil (nonspecific voltage-dependent Ca(2+) channel blocker; IC(50) = 0.4 microM), diphenylhydantoin and NiCl(2) (T-type [Ca(v)3.x] channel blockers; IC(50) 210 microM and 75 microM, respectively). Verapamil blockade of L-type (Ca(v)1.x) channels is use-dependent; activated channels are more sensitive to inhibition. However, verapamil inhibition of SAL does not increase after repeated SAMMA stimulation. SAL is unaffected by 10 microM nifedipine (selective L-type channel blocker). This contrasts to 40% inhibition (P < .001) of AL induced by 1 microM thapsigargin (Ca(2+)-ATPase inhibitor; releases intracellular Ca(2+) stores, promotes capacitative Ca(2+) entry). SAL is unaffected by 1 microM BAPTA-AM (intracellular Ca(2+) chelator), and 50 microM 2-APB (blocks InsP3 receptors and store-operated channels). This contrasts with thapsigargin-induced AL, inhibited nearly 65% by BAPTA-AM (P < .005) and 91% by 2-APB (P, .001). The results suggest that SAL is mediated by Ca(2+) entry through channels pharmacologically similar to the T-type (Ca(v)3.2) class. This process appears distinct from that caused by physiological stimuli such as progesterone or zona pellucida-derived proteins. SAMMA's contraceptive activity may be caused by induction of premature AL through dysregulation of Ca(2+) signaling.

A pilot clinical trial comparing an acid-buffering formulation (ACIDFORM gel) with metronidazole gel for the treatment of symptomatic bacterial vaginosis.

AIM: To compare the effectiveness of an acid-buffering formulation gel (ACIDFORM) with metronidazole gel in the treatment of symptomatic bacterial vaginosis (BV). METHODS: After a confirmed diagnosis of BV according to the criteria established by Nugent and Amsel, 30 nonpregnant women were enrolled in a randomized, double-blind clinical study. The women were randomly assigned to receive either 5 g ACIDFORM gel (n = 13) or 10% metronidazole gel (n = 17) intravaginally once daily for five consecutive days. Participants were evaluated in two follow-up visits (7-12 days and 28-35 days after treatment). Therapeutic success was defined as the presence of less than three of Amsel's criteria. If three or more criteria were present at first or second follow-up visit, the woman was excluded from the study and treated orally with metronidazole. Nugent scores were recorded at each visit but these were not used to define cure. RESULTS: At the first follow-up visit, 15 (88%) of the women in the metronidazole group were cured compared with only three (23%) in the ACIDFORM group (P < 0.001). The remaining 12 women (10 of the ACIDFORM group and two of the metronidazole group) were considered as failure and were treated orally with metronidazole. At the second follow-up visit, two of the ACIDFORM-treated women and six of the metronidazole-treated women presented recurrent BV. Four women in the ACIDFORM group and one in the metronidazole group reported occasional burning and itching during product use. CONCLUSION: ACIDFORM gel was significantly less effective than high-dose metronidazole gel for the treatment of symptomatic BV.

Development and characterization of bioadhesive vaginal films of sodium polystyrene sulfonate (PSS), a novel contraceptive antimicrobial agent.

PURPOSE: Polystyrene sulfonate (PSS) is a novel noncytotoxic antimicrobial contraceptive agent. A gel formulation of PSS was found safe for vaginal administration in phase I clinical trials. The purpose of the current study was to develop and evaluate novel bioadhesive vaginal film formulations of PSS. METHODS: PSS films were prepared by solvent evaporation and optimized for various physical, mechanical, and aesthetic properties. Further, films were evaluated for various biological activities and safety. RESULTS: Vaginal films containing 300 mg PSS per unit have been developed, using generally regarded as safe (GRAS) listed excipients. The films are colorless, transparent, thin, soft, and tough, dissolve rapidly in physiologic fluid to form a smooth, viscous and bioadhesive solution that could be retained in the vagina for prolonged intervals. Sperm function inhibition (hyaluronidase and cervical mucus penetration) and antimicrobial activities against human immunodeficiency virus (HIV) and herpes simplex virus (HSV) by PSS films were found comparable to PSS. Also, films did not inhibit normal vaginal microflora (Lactobacillus) and were noncytotoxic as indicated by negligible sperm immobilization and cytotoxicity to host cell assays. CONCLUSIONS: Rapidly dissolving bioadhesive vaginal film formulation of PSS with desired physical, mechanical, aesthetic, and biological properties is a suitable candidate vaginal microbicide for prevention of sexually transmitted disease (STDs) and is ready for toxicological and clinical evaluation.

Contraception by Ushercell (cellulose sulfate) in formulation: duration of effect and dose effectiveness.

This study evaluated contraception by formulated Ushercell, a uniquely high-molecular-weight form of cellulose sulfate, in the rabbit. Variables included (1) dose effectiveness, (2) duration of effectiveness, and (3) formulation excipients. Vaginally applied carboxymethyl-cellulose-based Ushercell gel is contraceptive. A 6% gel is active for at least 18 h; partial activity is observed for at least 24 h. With an application-insemination interval of 0.5 h, Ushercell as low as 0.1% is contraceptive. Contraception is incomplete with 2% Ushercell and an application-insemination interval of 24 h. Ushercell formulations containing a relatively high concentration of Carbopol are ineffective contraceptives, whether the gel is applied before insemination or is premixed with spermatozoa before insemination. Contraceptive activity is restored in Ushercell formulations with lower Carbopol content. This study shows that formulated Ushercell is an effective, long-lasting contraceptive and, hence, is bioavailable when vaginally applied. Activity is dependent on the type and relative concentration of formulation excipients. These data support a projected successful outcome of further clinical trials.

Candidate topical microbicides bind herpes simplex virus glycoprotein B and prevent viral entry and cell-to-cell spread.

Topical microbicides designed to prevent acquisition of sexually transmitted infections are urgently needed. Nonoxynol-9, the only commercially available spermicide, damages epithelium and may enhance human immunodeficiency virus transmission. The observation that herpes simplex virus (HSV) and human immunodeficiency virus bind heparan sulfate provided the rationale for the development of sulfated or sulfonated polymers as topical agents. Although several of the polymers have advanced to clinical trials, the spectrum and mechanism of anti-HSV activity and the effects on soluble mediators of inflammation have not been evaluated. The present studies address these gaps. The results indicate that PRO 2000, polystyrene sulfonate, cellulose sulfate, and polymethylenehydroquinone sulfonate inhibit HSV infection 10,000-fold and are active against clinical isolates, including an acyclovir-resistant variant. The compounds formed stable complexes with glycoprotein B and inhibit viral binding, entry, and cell-to-cell spread. The effects may be long lasting due to the high affinity and stability of the sulfated compound-virus complex, as evidenced by surface plasmon resonance studies. The candidate microbicides retained their antiviral activities in the presence of cervical secretions and over a broad pH range. There was little reduction in cell viability following repeated exposure of human endocervical cells to these compounds, although a reduction in secretory leukocyte protease inhibitor levels was observed. These studies support further development and rigorous evaluation of these candidate microbicides.

Development pharmaceutics of microbicide formulations. Part II: formulation, evaluation, and challenges.

In recent years, AIDS and sexually transmitted diseases (STDs) have become a burgeoning problem and are spreading at an alarming rate. Microbicides are being developed as a new therapeutic category for prevention of transmission of sexually transmitted infections (STIs) and HIV. Many of the microbicide formulations (MF) may fail to elicit a protective response either because of a lack of efficacy or inadequate formulation. Manufacturing a stable, efficacious, safe, and optimal product is the main objective of formulation development programs. Preformulation parameters (PP), as discussed in Part I of this series, influence formulation development significantly and should be considered carefully before designing a formulation strategy. Initially, based on PP and market research, a target product profile (TPP) is generated, which defines product attributes that can be normally classified as "essential" and "desirable." A complex and dynamic process begins thereafter that takes into consideration myriad factors starting from selection of delivery system, selection of excipients, compatibility study, prototype composition, selection of process and optimization, stability testing, scale up, manufacturing under good manufacturing practices (GMP), and packaging development. Prototype formulations are evaluated for several performance characteristics (e.g., dispersion behavior, bioadhesion, retention, spreading, rheology). These compositions are also subjected to biologic evaluation by various in vitro and in vivo models. Such a well-planned, well-coordinated, and well-implemented formulation development program not only accelerates overall development but also minimizes failures in subsequent clinical development studies. The objective of this review is to highlight the importance of formulation science, outline the steps involved in this process, and explore how these can be exploited for achieving optimal MF.

Women's preferences for vaginal antimicrobial contraceptives. V: attitudes of Brazilian women to the insertion of vaginal products.

The rapid spread of HIV/AIDS in the female population increases the urgency of developing new formulations that offer protection from this disease as well as other sexually transmitted infections. In many cultures, women do not readily accept touching their genitals or inserting products into their vaginas. Information on this subject was collected during a study involving 635 women in Brazil to determine the preferred attributes of vaginal products. Seventy-six percent would use an idealized contraceptive method that offered dual protection even though it could only be inserted with a finger and 96% would use this method if it could only be placed with an applicator. Qualitative analyses of responses to open questions suggest that the majority of Brazilian women studied did not like to touch their vagina with their finger or to insert devices. Although the introduction of safe and effective vaginal microbicides into many cultural settings can be successful, it should be accompanied by significant efforts to educate women about their bodies.

Development pharmaceutics of microbicide formulations. Part I: preformulation considerations and challenges.

Microbicides, the compounds and formulations that can prevent transmission of sexually transmitted diseases (STDs)/HIV are being pursued actively as a promising AIDS intervention. The drug development chain for a topical microbicide differs significantly from that of any systemic or topical compound/formulation regarding to time line, cost, activities, and milestones. This is in part because of the lack of standard in vitro models to assess efficacy, and complex ethical issues in clinical trials of microbicides. Several factors, including changes in the physiology of the cervix and vagina with age and menstrual cycle, intercourse, as well as leakage of the formulation from the vagina may affect their design, development, and performance. Selection and development of optimal microbicide delivery systems (gel/cream, pessary, film, tablet, foam, etc.), their inactive ingredients, manufacturing details, and packaging system are dependent on the properties of active drug, or their preformulation parameters (PP). The PP of the active drug substance needs to be evaluated in initial stages of drug discovery and development so that the most suitable delivery system can be selected. Some PP of microbicide agents include physical state, organoleptic properties (color, odor, appearance, taste, etc.), molecular weight, aqueous solubility, hygroscopicity, acidity/alkalinity, permeability and absorption characteristics, stability in solid/solution state, and inherent bioadhesiveness. Thus, a well-coordinated, planned, and implemented preformulation program can help in not only accelerating microbicide formulation development, but also to minimize unforeseen failures in subsequent stages of the development. The objective of this review is to highlight the significance of PP, suggesting a systematic preformulation program.

Assemblies for in vitro measurement of bioadhesive strength and retention characteristics in simulated vaginal environment.

The vaginal route of administration offers a promising option for local and systemic delivery of drugs. Conventional vaginal formulations are associated with limitations of poor retention, leakage, and messiness, thereby causing inconvenience to users. To overcome these limitations, formulations that adhere to the vaginal mucosa for a sufficient period of time need to be developed. Bioadhesion and retention are desirable characteristics of a vaginal formulation to achieve desired efficacy. These properties can be built in during formulation development by the use of bioadhesive polymers. In the present study, assemblies for in vitro measurement of bioadhesive strength and retention characteristics of vaginal formulations have been developed. A modified simulated vaginal fluid (SVFM) was used to simulate vaginal conditions for bioadhesion studies. Cellophane hydrated with SVFM and isolated sheep vaginal mucosa were used as model membranes. The bioadhesive potential of various polymers and their combinations was evaluated. Among the polymers evaluated, xanthan gum (XG), sodium alginate (SA), Polycarbophil (PC), and their combinations (XG + SA and XG + PC) were found to possess significant bioadhesive strength. In retention experiments, XG, SA, and combinations (XG + SA and XG + PC) were retained in isolated sheep vaginal tissue, while PC exhibited poor retention under experimental conditions. Based on the results of the study conducted, XG, SA, and combinations (XG + SA and XG + PC) have been proposed as potential candidates for developing bioadhesive vaginal drug delivery systems.

Use of mandelic acid condensation polymer (SAMMA), a new antimicrobial contraceptive agent, for vaginal prophylaxis.

OBJECTIVE: To assess the contraceptive properties, antimicrobial activity, and safety of mandelic acid condensation polymer (SAMMA). DESIGN: Experimental study of SAMMA's in vitro and in vivo properties. SETTING: Academic research laboratories. PATIENT(S): Healthy volunteers for semen donation in an academic research environment. INTERVENTION(S): Inhibition of sperm function indicators, conception, sexually transmitted infection-causing pathogens (including HIV), and lactobacilli was evaluated. Safety indicators were studied. MAIN OUTCOME MEASURE(S): Quantitation of SAMMA's effect on microbial infectivity or multiplication and on sperm function in vitro; evaluation of contraceptive efficacy in vivo; assessment of safety in vitro and in vivo. RESULT(S): Mandelic acid condensation polymer is not cytotoxic toward lactobacilli, microbial host cells, and spermatozoa. The compound inhibits hyaluronidase and acrosin, induces sperm acrosomal loss, and is contraceptive in the rabbit model. Mandelic acid condensation polymer prevents infectivity of HIV and herpesviruses 1 and 2 and, to a lesser extent, of Chlamydia trachomatis. It inhibits the multiplication of Neisseria gonorrhoeae. Mandelic acid condensation polymer is not mutagenic, has low acute oral toxicity, and is safe in the rabbit vaginal irritation assay. CONCLUSION(S): Mandelic acid condensation polymer inhibits sperm function, is contraceptive, has broad-spectrum antimicrobial activity, and is highly safe. Further development as a microbicide is warranted.

Two novel vaginal microbicides (polystyrene sulfonate and cellulose sulfate) inhibit Gardnerella vaginalis and anaerobes commonly associated with bacterial vaginosis.

This is the first report demonstrating the in vitro inhibitory activity of two novel microbicides (cellulose sulfate and polystyrene sulfonate) against bacterial vaginosis (BV)-associated bacteria. Vaginal application of these microbicides not only may reduce the risk of acquisition of human immunodeficiency virus and other sexually transmitted infection-causing organisms but may also decrease the incidence of BV.

Preclinical evaluation of sodium cellulose sulfate (Ushercell) as a contraceptive antimicrobial agent.

The spread of sexually transmitted infections (STIs) and limited methods for control of pregnancies presents high risks to the reproductive health of women. Methods controlled by women and directed toward disease prevention and contraception are needed. We report on preclinical studies of the biological properties of sodium cellulose sulfate (Ushercell) currently being developed for use as a topical contraceptive antimicrobial agent. Ushercell was evaluated with tests designed to identify its contraceptive and antimicrobial properties. Ushercell inhibits hyaluronidase (reversible; IC50 = 1.7 mg/mL), impairs sperm penetration of cervical mucus (approximately 70% inhibition at 1 mg/mL), and acts as a stimulus for acrosomal loss (IC50 = 52 ng/mL). It prevents conception in rabbits when added to spermatozoa (approximately 95% inhibition at 1 mg/mL) or when vaginally applied (complete contraception by 45 mg) before insemination. However, up to 50 mg/mL, Ushercell does not irreversibly immobilize spermatozoa, suggesting that Ushercell is not cytotoxic. Ushercell has a broad spectrum of antimicrobial activity in vitro. Inhibited microbes include human immunodeficiency viruses (different laboratory strains and clinical isolates; IC50 values range from 3 to 78 microg/mL), herpes viruses, HSV-1 (IC50 = 59 ng/mL) and HSV-2 (lC50 = 24 ng/mL), Neisseria gonorrhoeae (IC50 = 2 microg/mL), and Chlamydia trachomatis (IC50 = 78 microg/mL). In contrast, Ushercell does not inhibit growth of beneficial vaginal bacteria, Lactobacillus gasseri, at 5 mg/mL. These results suggest that the antimicrobial effects of Ushercell are selective, and not likely mediated by nonspecific cytotoxic mechanisms. These data provide the basis for further clinical development of Ushercell as a vaginal agent to prevent unplanned pregnancy and STIs.

Efficacy and safety of a new vaginal contraceptive antimicrobial formulation containing high molecular weight poly(sodium 4-styrenesulfonate).

Host cell infection by sexually transmitted disease (STD)-causing microbes and fertilization by spermatozoa may have some mechanisms in common. If so, certain noncytotoxic agents could inhibit the functional activity of both organisms. High molecular mass poly(sodium 4-styrenesulfonate) (T-PSS) may be one of these compounds. T-PSS alone (1 mg/ml) or in a gel (2% or 5% T-PSS) completely prevented conception in the rabbit. Contraception was not due to sperm cytotoxicity or to an effect on sperm migration. However, T-PSS inhibited sperm hyaluronidase (IC(50) = 5.3 microg/ml) and acrosin (IC(50) = 0.3 microg/ml) and caused the loss of acrosomes from spermatozoa (85% maximal loss by 0.5 microg/ml). T-PSS (5% in gel) also reduced sperm penetration into bovine cervical mucus (73% inhibition by 1 mg gel/ml). T-PSS (5% in gel) inhibited human immunodeficiency virus (HIV; IC(50)= 16 microg gel/ml) and herpes simplex viruses (HSV-1 and HSV-2; IC(50) = 1.3 and 1.0 microg gel/ml, respectively). The drug showed high efficacy against a number of clinical isolates and laboratory strains. T-PSS (5% in gel) also inhibited Neisseria gonorrhea (IC(50) < 1.0 gel/ml) and Chlamydia trachomatis (IC(50) = 1.2 microg gel/ml) but had no effect on lactobacilli. These results imply that T-PSS is an effective functional inhibitor of both spermatozoa and certain STD-causing microbes. The noncytotoxic nature should make T-PSS safe for vaginal use. T-PSS was nonmutagenic in vitro and possessed an acute oral toxicity of >5 g/kg (rat). Gel with 10% T-PSS did not irritate the skin or penile mucosa (rabbit) and caused no dermal sensitization (guinea pig). Vaginal administration of the 5% T-PSS gel to the rabbit for 14 consecutive days caused no systemic toxicity and only mild (acceptable) vaginal irritation. T-PSS in gel form is worthy of clinical evaluation as a vaginal contraceptive HIV/STD preventative.

Prevençäo de doenças sexualmente transmissíveis em mulheres: associaçäo com variáveis sócio-econômicas e demográficas / Prevention of sexually transmitted diseases among women: association with socioeconomic and demographic variables

As doenças sexualmente transmissíveis (DST) têm sido debatidas no ambiente científico e nos meios de comunicaçäo de massa, em especial, por sua associaçäo a maior risco de infecçäo pelo Vírus da Imunodeficiência Humana (HIV). Estudou-se a adoçäo de comportamentos por mulheres para proteçäo das DST, tal como a associaçäo destes a variáveis sócio-econômicas e demográficas. Trata-se de estudo descritivo, com dados secundários de pesquisa feita em Campinas, Säo Paulo, na qual foram entrevistadas 635 mulheres selecionadas mediante a técnica de amostragem "bola de neve". Foram classificadas em: adolescentes e adultas de status sócio-econômico médio-alto ou baixo. Grande proporçäo delas näo se prevenia das DST, em particular, as de status baixo. Em todos os grupos, o condom masculino foi o método de prevençäo mais referido. Houve associaçäo negativa entre parceiro fixo e uso de condom, e a principal razäo para näo usá-lo foi "só ter um parceiro e confiar nele". Em meio às adolescentes, ocorreu associaçäo positiva entre escolaridade acima da oitava série e uso de condom, bem como negativa entre idade e uso desse método. Entre adultas o uso exclusivo de condom esteve, em geral, positivamente associado a status sócio-econômico.