• 1.
  

  a) T b) F c) T d) F e) F

It is a common misconception that microbicides are only being developed for vaginal use in women. Microbicides are being developed for both vaginal and rectal use; the latter is particularly focused on men who have sex with men but also for women. The rectal formulations of tenofovir gel being developed for rectal use are different from the vaginal gel – rectal formulations have a lower osmolality. The CAPRISA 004 trial has provided proof-of-concept that tenofovir gel can prevent HIV infection. As there is no correlate for HIV prevention, the only way to demonstrate that a microbicide is effective in preventing HIV is to conduct large trials where the incidence of HIV infection in microbicide users is compared with HIV incidence in placebo users. Although it is widely thought that there is likely to be a drug level that correlates with HIV prevention, this has not been empirically demonstrated yet. One analysis to date has suggested 1000 ng/ml as a potential cut-off, but this needs to be prospectively assessed. There is no correlation between clinical trial outcomes with outcomes in monkeys (e.g. PRO 2000 ^® was found to be highly effective in monkeys but not found to be protective in humans). The correlations seem to be better for antiretroviral microbicides, but insufficient data are available to make this determination at this time.

• 2.
  

  a) F b) T c) T d) F e) T

Antiretroviral drugs that inhibit viral integrase have been considered for microbicide development, but no candidates with this class of ARVs have been developed yet. An example of a candidate microbicide with this mechanism of action is Buffergel ^® , which acts by enhancing the vagina’s naturally low pH, has and been tested as a candidate microbicide. Several polyanions (e.g. PRO 2000 ^® and Carraguard ^® ) were developed as candidate microbicides and tested for efficacy against HIV infection. No candidate microbicides have been developed targeting DC-sign. Viral replication inhibitors, such as tenofovir, have been developed and tested against HIV.

• 3.
  

  a) F b) F c) T d) T e) T

The use of tenofovir as a first-line treatment drug is a good reason not to develop it for prevention because of the potential problem of cross-resistance. The decision to develop tenofovir as a gel preceded the point at which it became the preferred drug in first-line AIDS treatment. Tenofovir is a small molecule that is readily absorbed and has an excellent safety profile: it is one of the safest antiretroviral drugs and hence its selection as the preferred drug in first-line AIDS treatment regimens. A microbicide requires a long half-life as it is difficult to take a prophylactic drug more than once a day. The high levels of efficacy of tenofovir in preventing SIV infection in monkeys provided animal data that made the case for prioritising tenofovir. In several studies of tenofovir gel in monkeys, tenofovir gel was found to have high efficacy, thereby making a promising candidate for development as a topical microbicide.

• 4.
  

  a) T b) F c) T d) F e) T

The presence of ulcerative sexually transmitted infections (e.g. HSV-2) results in breaks in the genital mucosa, allowing access for HIV to submucosal target cells to facilitate infection. In addition, these ulcerative sexually transmitted infections (STI) are also very inflammatory, and this inflammation is associated with infiltration into the ulcers of high concentrations of target cells for HIV infection. The presence of aerobic Lactobacillus species in the vaginal flora is associated with protection against HIV infection because Lactobacillus species, by their production of lactic acid and other factors, lower the pH of the vaginal milieu. Human immunodeficiency virus infection is less efficient under conditions of low vaginal pH compared with more neutral pH. The higher the viral load of the transmitting partner, the higher the concentration of HIV will be shed in semen, and the higher the risk for transmission to his female partner. Men with viral loads less than 1500 RNA copies/ml in their plasma will not transmit to their female partners, making the use of antiretroviral therapy to reduce viral loads in HIV-infected partners an important intervention strategy. Cervical mucous has been shown to be relatively efficient at trapping HIV particles and preventing contact of HIV with the epithelial barrier. Although it is plausible that cervical mucous may retard HIV infection, there is currently little science to support this conclusion, and HIV infection does happen even in women with cervical mucous present. Bacterial vaginosis is a condition whereby non-commensal anaerobic bacteria colonise the genital tract, outcompeting natural Lactobacillus species. Bacterial vaginosis is associated with increased pH in the genital tract, which favours more efficient HIV infection. In addition, bacterial vaginosis induces genital tract inflammation that recruits target cells for HIV infection to this site.

• 5.
  

  a) T b) T c) F d) T e) T

The genital tract of adolescent females, during puberty, has a more prominent cervical ectropion, with the transformation zone being more exposed in the vaginal vault and accessible for HIV infection. As the transformation zone has been shown to be particularly vulnerable to HIV infection, caused partly by the concentration of inflammatory molecules and susceptible target cells for HIV infection found here, and by the fact that single layer columnar epithelial cells are easier for HIV to traverse than the multilayered squamous epithelial cells. Progesterone has been shown to reduce inflammatory pathways and fluctuations in female hormones during adolescence may result in suppression of both the innate and adaptive immune system. Adolescent females have a greater prevalence of anaerobic bacteria than older females. Normal commensal Lactobacillus species colonise the genital tract with onset of sexual activity and parity.

• 6.
  

  a) T b) F c) T d) T e) T

Repeated exposure to N-9 and cellulose sulfate during sexual intercourse was associated with mucosal inflammation and disruption of the cervical squamous epithelium. Both N-9 and cellulose sulfate were shown to increase inflammatory signals in genital tissue, activating nuclear factor kappa-light-chain-enhancer of activated B cell activity. This resulted in recruitment of high concentration of activated target cells for HIV infection to the genital mucosa to increase the efficiency of HIV infection. Early microbicides (N-9) were shown to alter vaginal flora, and Lactobacillus species colonisation of the genital mucosa was compromised in women using these microbicide products compared with those using the placebo. N-9 was shown to disrupt the phospholipid membrane of epithelial cells, causing non-specific damage to the vaginal epithelium. The resultant damage to the vaginal epithelium made it an easier barrier for HIV to traverse as well as making submucosal target cells accessible for HIV infection.

• 7.
  

  a) T b) T c) T d) F e) F

Given that inflammation, caused by STIs, is an important risk factor for HIV infection, management of genital tract inflammation in combination with topical microbicide agents would improve the protective effect of a microbicide. As Lactobacillus species are normal commensals of the female genital tract that may be out competed by anaerobic organisms, combination strategies, including probiotic Lactobacilli with an anti-retroviral microbicide, would improve the protective effect of a microbicide. Douching agents have been implicated in increased susceptibility to HIV infection, so would not be considered to be a suitable combination treatment to improve the efficacy of topical microbicides. Estrogen containing compounds have not been shown to reduce the rates of BV.

• 8.
  

  a) T b) T c) T d) F e) T

An advantage to all topical microbicide formulations is that they are controlled by the woman seeking protecting from human immunodeficiency virus (HIV). The goal of a topically applied compound to prevent HIV infection is to have high concentrations at the site of application, but limited systemic absorption. Gels, in addition to serving as microbicides, also provide women with lubrication during intercourse. Drugs that are not stable in aqueous solution cannot be formulated as a microbicide gel and are often more stable as films or tablets. Because of the limited systemic absorption of topically applied gels, there is a decreased chance that women will experience the systemic side-effects that may be associated with an oral dosage form.

• 9.
  

  a) F b) T c) F d) F e) F

Tenofovir acts as a nucleoside analogue that terminates HIV transcription from RNA to DNA before integration into the host cell genome. Dapivirine is a non-nucleoside reverse transcriptase inhibitor that allosterically blocks the reverse transcriptase enzyme and thereby prevents the complete transcription of HIV RNA to DNA. RC-101 is a retrocyclin 1 peptide analogue that works as an entry inhibitor preventing the binding of HIV to both CXCR4 and CCR5 co-receptors located on CD4+ cells. Nonoxynol-9 is a spermicidal polymer with bio-adhesive properties only. Maraviroc prevents HIV from binding to the cell surface co-receptor CCR5.

• 10.
  

  a) T b) T c) T d) T e) T

Some pharmaceutically active compounds are not stable in an aqueous formulation, such as a gel or ring, and would be more stably formulated in a film or solid tablet. Regardless of the dosage form, it is important that all mucosal surfaces that may be exposed to HIV be protected. Both pH and osmolality must be compatible with the vagina in order to avoid irritation, which could increase the risk of HIV acquisition. Films and tablets must have reliable disintegration over a predictable amount of time in order to ensure that women will be protected at all exposed mucosal surfaces regardless of the amount of vaginal moisture.

• 11.
  

  a) F b) F c) T d) T e) T

It is often difficult to identify the population at risk in prevention trials. If trials are being conducted in general populations, they typically have to recruit a large number of study participants who may never be at risk of the disease of interest. In treatment trials, however, specific patient groups can be targeted much more easily.

• 12.
  

  a) T b) F c) F d) F e) F

The incidence of HIV determines the number of events that drives the power of the trial. All of the other factors will be determined by the trial once undertaken.

• 13.
  

  a) F b) T c) T d) F e) F

Adherence data is used to help estimate the product efficacy. A microbicide trial measures effectiveness, which will be reduced as adherence to the product falls. The trial result can only therefore be compared to other ones if adherence data is known.

• 14.
  

  a) F b) F c) F d) T e) T

The role of the IDMC is to review unblinded data regularly to ensure study participants are not being put at risk by being on the trial.

• 15.
  

  a) F b) F c) F d) T e) F

Answers a) and b) are inclusion criteria for microbicide trials and are therefore incorrect. Answer c) was an inclusion criterion for the CAPRISA 004 study. The investigators of this review believe it should be part of any trial going forward. Answer d) is the correct answer, as a woman who is planning to have a pregnancy over the trial duration would meet the exclusion criteria and should not be enrolled.

• 16.
  

  a) F b) F c) T d) T e) T

Tenofovir an antiretroviral, and Savvy, a surfactant, are not believed to have contraceptive effects. Nonoxynol-9 is a spermicide that has been used as a contraceptive method for many years. In addition, cellulose sulfate has been shown to have contraceptive properties.

• 17.
  

  a) F b) T c) F d) F e) F

The correct answer is depot medroxyprogesterone acetate, as it is the only non-barrier form of contraception listed. The female condom, male condom, diaphragm, and cervical cap are all barrier forms of contraception, which were not considered an eligible contraceptive method in CAPRISA 004.

• 18.
  

  a) F b) F c) T d) F e) F

The only correct answer is the Population Council Carraguard trial, which was the only clinical trial that withdrew a study participant from the trial if she became pregnant. The other trials halted the study product after a positive pregnancy test and re-instituted the product after a confirmed negative pregnancy test. This approach helped to limit the time-off study product because of pregnancy.

• 19.
  

  a) T b) F c) T d) T e) T

Female control is a key issue within HIV prevention and the future agenda for research in women. Historically, HIV prevention has given men a greater advantage with prevention techniques that they can control (i.e. condoms and male circumcision). Women need to be given HIV prevention techniques that will allow them to control their health issues and empower them to take responsibility without being dependent on their partners. Although poverty alleviation and infrastructure are important issues in the HIV pandemic, they are one part of a broader social, political and structural change that needs to occur in order to fight the HIV pandemic. This needs to include men and women, and is not specific to the female research agenda. It is critical to advance the roll-out and use of available female barrier methods. Currently, female condoms remain poorly rolled out in health facilities in South Africa, and women remain poorly educated and misinformed about how female condoms function, and how to use them. Men remain largely ignorant of female condoms, and greater awareness campaigns need to occur. This would attenuate and enhance the protective effects that microbicides would have for protecting women against HIV infection when a successful candidate microbicide is found and rolled out. Male and female dynamics and relationships are key factors in HIV prevention. Couples counselling and a joint sense of responsibility for protecting one another against HIV infection is an essential factor for HIV research.

• 20.
  

  a) T b) F c) T d) F e) F

Current epidemiological research suggests the women aged 24 years and younger are particularly at risk in the Sub-Saharan epidemic, with high rates of HIV infection. This is a particularly important group in which HIV prevention efforts need to be focused. Although an at-risk group and an important focus of HIV prevention efforts, the Sub-Saharan epidemic remains a heterosexually driven epidemic. Therefore, current focus needs to remain particularly aware of the heterosexual risk factors; men who have sex with men populations need to remain part of prevention as well. Pregnant women remain an essential part of HIV prevention efforts in Sub-Saharan Africa. This is because prevalence remains high in this population and also because it is an essential point of entry for antenatal services and prevention of mother-to-child transmission of HIV. As with men who have sex with men, injecting drug users are an important risk group; however, because of the heterosexually driven epidemic in Sub-Saharan Africa, the heterosexual epidemic needs to remain key point of HIV prevention efforts. While Hep B +ve women are important and the two infections may co-exist, Hep B positivity alone should not be the focus of efforts.

• 21.
  

  a) F b) F c) F d) T e) T

Although the discovery of the HIV virus was a critical point in HIV awareness, it also defined the HIV research agenda for decades, delaying the HIV prevention response and funds in heterosexually driven HIV infection. Condoms were important in the fight against HIV, but were the dominant form of HIV prevention for decades while funding remained lacking in female-driven, HIV-prevention technologies. They also remain an HIV prevention technique controlled by men, and increased efforts into female controlled HIV prevention are urgently needed. Zena Stein’s article in the American Journal of Public Health was the critical point in which the research community recognised the need for female-driven HIV prevention methods. It marked an important milestone in the microbicide advocacy movement around which advocacy for increased funding and research into female-driven HIV prevention technologies began. The CAPRISA 004 trial marked a new sense of hope for the microbicide field after many failures. It renewed hope that a female-driven technology was possible.

• 22.
  

  a) T b) F c) T d) T e) T

A critical point for the future of research in microbicides and barrier methods for use by women is that biomedical research is needed to test products and understand the mechanisms with which the products work. The behavioural fields, however, remain important for understanding issues such as adherence, product use, barriers to use, and larger social issues affecting use of products and HIV prevention technologies. Future research and roll-out of HIV-prevention strategies need to address the larger social issues around empowering women, changing gender roles and norms, and addressing the empowerment of women and responsibility of men towards combating the HIV epidemic. It is critical to ensure that HIV prevention efforts use what is available to reduce the number of infections. This is important for ensuring that female infections do not increase while more efficacious prevention strategies can be developed. HIV remains a social and biological disease. Thus, any prevention efforts need to address more than just one issue. Multi-disciplinary teams from all areas of society (from social services, health, education and other areas) will need to collaborate in order to address the HIV pandemic. Future HIV research looks towards combination and structural interventions.

• 23.
  

  a) T b) F c) T d) F e) F

The dapivirine ring is an off-white flexible ring containing 25 mg of drug dispersed in a platinum-cured silicone matrix. Dapivirine is one of a new generation of non-nucleoside reverse transcriptase inhibitors (NNRTIs). These bind to the HIV reverse transcriptase enzyme preventing viral replication and therefore production of infectious virus. NNRTIs, such as dapivirine bind specifically to the HIV-1 but not HIV-2 reverse transcriptase. As a result, dapivirine ring is not intended to protect against HIV-2 or other sexually transmitted infections, including HSV-2. The vaginal ring developed by IPM is designed to provide women with long-acting protection against HIV infection for a month or longer. The convenience of a monthly product may help ensure the product is used more consistently, thus increasing a woman’s chance of being protected at any given time. All other preventive products currently in use (i.e. condoms) or under research (i.e. oral tablets, vaginal and rectal gels) are intended for use around the time of sex, or once-daily. The dapivirine ring was designed to be an HIV-prevention product for women, and will be evaluated in HIV-negative women in randomised-, placebo-controlled trials to determine its efficacy in preventing male-to-female transmission of HIV-1. Men have several effective prevention options available to them currently, such as male condoms and male circumcision. One advantage of the dapivirine silicone elastomer vaginal ring, however, is that the cost for monthly use is relatively low, compared with daily or coitally associated use of applicator-based gels.

• 24.
  

  a) F b) F c) T d) T e) T

Data from ring-acceptability studies have shown that adherence rates were over 80%, and women indicated they would be extremely likely to use a long-acting vaginal ring for protection against HIV. Male partner interviews have indicated that men did not feel the ring during intercourse nor did it affect their sexual pleasure. Acceptability and market research studies have indicated that women’s preferences will be different depending, for example, on age, marital status and cultural background. Acceptability of formulations has been seen to vary by country. For example, in a market research study of three different vaginal formulations (film, tablet, and soft-gel capsule), the film was preferred in Burkina Faso and Zambia; whereas the soft-gel capsule was preferred in Tanzania. As the ring is able to deliver drug for at least 1 month, the burden of user adherence is lower than for once daily products. Product-acceptability studies and the experience gained from marketed vaginal ring products have established a high level of acceptance and adherence from women using vaginal rings with similar physical characteristics. Optimal dosage forms will depend on a variety of factors, including the physico-chemical characteristics of the active pharmaceutical ingredient, the mechanism of action, and the duration of effect required.

• 25.
  

  a) F b) F c) F d) T e) F

Young women in low income countries are at disproportionally high risk of HIV infection compared to their male counterparts. Preventions options that are women controlled, and may be used discreetly without partner consent if needed, appear to be most suitable for young women. HIV prevention options for women are limited currently to abstinence from sexual activity and female condom use. It is globally recognised that young people, especially women in the 15 – 24 year age band are at highest risk for new infections and also represent the largest number of HIV cases amongst those infected in low income countries. HIV prevention for young women is therefore a public health priority. Incident HIV infections in young women in low income countries are drivers of generalised HIV epidemics. HIV prevention in young women is urgently required.

• 26.
  

  a) F b) T c) F d) F e) F

Although microbicides are not yet registered and available for use, the challenges related to maintaining clinical trial effectiveness in practice are complex and not limited to availability of products. It has been previously demonstrated that even highly effective interventions will only be marginally more effective than inferior interventions unless health systems achieve high performance at each interaction with patients. Although greater coverage will be achieved if healthcare workers promote effective HIV prevention interventions, this is not likely to influence the effectiveness of the product during real-life use. Cost of microbicides, when they become available, will certainly influence the wide-scale roll out of the product in countries limited by budget restrictions but the cost is unrelated to the intrinsic effectiveness of the product. Quality improvement approaches aim to strengthen heath systems and will be an important tool to facilitate programmatic scale up of new interventions but are not likely to influence the effectiveness of the product during real-life use.

• 27.
  

  a) T b) F c) F d) F e) F

This definition best describes the implementation sciences approach which strives to integrate evidence-based interventions into healthcare systems with the aim to improve health outcomes. Randomised trials are required to ascertain effectiveness of candidate agents and are conducted prior to implementation sciences research. Pre-clinical assessment of interventions involves in vitro and animal testing of compounds to ascertain early effectiveness, dosing and safety. Monitoring and evaluation of health systems is important to understand performance and to manage processes but is not synonymous with implementation sciences approach. Post marketing surveillance or phase IV trials ascertains post registration, long term safety of products and is not related to integrating successful biomedical intervention into health systems

• 28.
  

  a) F b) F c) T d) F e) T

Microbicides by virtue of their mode of use, safety profile and potential user profile are ideal for integration into existing healthcare services. A microbicide product will only be made available for implementation once effectiveness has been proven in clinical trials. Low income countries, particularly in Sub-Saharan Africa have a high incidence and prevalence of HIV infections and contribute the majority of cases worldwide. HIV prevention options such as microbicides are urgently needed and excluding HIV infection at the start and monitoring throughout prophylaxis will be challenging. The required regulatory approval processes (product licensing) would occur prior to implementation and roll out of microbicides. The unknown cost of initial production cost and future cost is regarded as potential barrier to implementation.

• 29.
  

  a) T b) T c) T d) T e) T

All of the above are significant challenges for prophylaxis researchers in any setting but particularly in low resource settings.

• 30.
  

  a) F b) T c) F d) F e) F

Although it is true that with topical microbicides systemic toxicities are limited, clinical monitoring for HIV, pregnancy and other safety markers will still be required. Sexually active women at high risk for HIV infection often utilize other healthcare services such as family planning clinics, STI and primary healthcare clinics for minor ailments or antenatal and postnatal care. These points of interaction with healthcare workers provide an ideal opportunity to engage HIV prevention services, including microbicides, to women. Microbicides may be formulated as gels, rings, films for either vaginal or rectal use. At present no microbicides are registered for clinical use as an HIV prevention agent. It is unclear at this stage of development and research how access to future microbicides will be controlled. There is a possibility that these agents may be prescription only.