Vaccine trials
A vaccine is developed and tested along a pathway from antigen discovery through clinical phases to licensure and surveillance. Its efficacy trial is sized from the incidence of the disease in the population studied, its safety data include reactogenicity collected from the participants themselves, its immune response can change the results of diagnostic tests, its efficacy is sometimes inferred from that immune response, and its trials need approvals, manufacturing capacity and site infrastructure in place before they can run.
The development pathway
An HIV vaccine is developed in stages: an exploratory stage, preclinical studies, clinical development, approval and licensure, and post-marketing surveillance. The exploratory stage is basic laboratory research by academic or government researchers, who discover natural or synthetic antigens that might help prevent HIV or modify the effects of the virus. The antigens may include virus-like particles, made in the laboratory for preventive vaccines, or other substances derived from HIV, and the stage runs two to four years.
Preclinical studies are conducted by biopharmaceutical companies. They use tissue and cell culture systems to assess the candidate's safety and potential toxicities and, crucially, its immunogenicity, which is its ability to induce an immune response. Extensive animal testing in challenge studies, with non-human primates and other animals, usually sheds some light on the cellular responses that might be expected in human beings, though predicting protection from these models has so far been particularly difficult with candidate HIV vaccines. Preclinical studies may also test for a safe starting dose and a safe way to administer the vaccine, and injections, including needle-free biojectors, and infusions have been tested. The stage runs one to two years. Many candidate vaccines fail at this stage because they do not induce the desired immune response, and successful candidates proceed to clinical studies.
Clinical development does not involve vaccinating participants and then intentionally exposing them to HIV. It starts when the sponsor, typically a private company, submits an investigational new drug application to the regulatory authorities of the countries where the vaccine may be marketed. The application includes a description of the vaccine manufacturing and testing processes, a summary of the laboratory reports and the clinical study proposal. The study is also subject to ethical review, and it must have both ethical and regulatory approvals before it starts. Participants are given information about the experimental vaccine, the risks and benefits of taking part and their rights and responsibilities before their consent is sought and throughout the trial. Like other drugs, a vaccine then goes through clinical trials in phases I to IV, which may be blinded or open-label.
Phase I enrols a small number of adults, 20 to 80, at low risk of HIV acquisition. It assesses safety in humans, including tolerability and dose ranges, and determines immunisation regimens. Participants are followed up for adverse effects and reactogenicity, and blood samples are drawn to estimate the type and extent of the preliminary immune response to HIV that the vaccine elicits. These responses may or may not protect against HIV, and larger trials are needed to determine this. Promising results lead to the next phase.
Phase II involves up to several hundred participants and may include some at higher risk of HIV acquisition. It collects more safety data, a more detailed assessment of immune responses and dosing data, determines the best immunisation schedule and investigates the best method of delivery. Phase IIb studies place more emphasis on estimating efficacy, and phase II mostly uses a randomised, double-blind, placebo-controlled, parallel-group design. Successful candidates move on to larger trials.
After a successful phase III trial proves efficacy, the sponsor submits a biologics license application to the regulatory authorities, which inspect the vaccine's manufacturing facility and approve its product labelling following usability testing. As with other drugs and vaccines, a licensed HIV vaccine would be monitored by an adverse event reporting system and database into which healthcare providers and consumers can report a suspected side effect, by continuous inspection of its manufacturing facilities by the regulatory authorities, by their review or conduct of batch tests to ensure the vaccine is consistently safe for public use, unadulterated and efficacious, and by phase IV trials. The licence holder might elect to conduct phase IV studies once the vaccine is approved and on the market, to continue testing its safety, efficacy and other potential issues.
In HIV vaccine development, what is most important is not which phase a trial is in but that studies are designed and carried out in a way that supports sound and ethical science, which is ultimately what progress toward an approved HIV vaccine needs.
Sample size and incidence
A phase III HIV vaccine efficacy trial is large, involving thousands of high-risk participants, and is usually a public-private partnership. Incidence data from the regions where the vaccine will be tested are gathered while the study is designed, to inform the sample size calculation and the duration of follow-up. With an HIV incidence of 1.5% per year in a population, for example, around 5,000 volunteers would be needed to power the study adequately. The higher the incidence, the fewer the participants needed, the shorter the follow-up, or both.
The trial is randomised, double-blind, placebo-controlled and parallel-group, and normal saline or another inactive substance may be used as placebo. Follow-up usually runs two to three years, with regular HIV testing and risk-reduction counselling, and the study emphasises to participants that the vaccine is experimental and not yet proven effective. Vaccine efficacy is evaluated by comparing the rate of HIV infection in the vaccine arm with that in the placebo arm, and any difference is analysed further to see whether it is due to chance or attributable to the vaccine. The trial also tests whether the vaccine delays progression to AIDS in those who become infected, judged by viral load or CD4 count, and whether it causes the production of antibodies to HIV or other immune responses. It also assesses safety in a large group of people, for rare adverse effects.
Effective or partially effective non-vaccine prevention programmes, with agents such as oral pre-exposure prophylaxis, are projected to decrease the incidence of HIV-1, the most common and infectious type, and current and future efficacy trials have to consider this in the sample size calculation and other design matters. A lower incidence means larger cohorts are needed to power a study to demonstrate efficacy while also assessing safety, so uptake of the prevention programme has to be monitored. Depending on uptake or other events, an ongoing study may need design adjustments or modified endpoints.
The efficacy trial design assumes that most participants will be exposed to HIV during follow-up, through unprotected sex or needle sharing. Whether and how well a vaccine works should ideally be answered by a well-designed, well-planned, well-executed and well-controlled efficacy trial with a statistically significant result, but with HIV vaccines more than one phase III trial of the same candidate may be needed. Key beneficial effects of an HIV vaccine may be realised not in vaccinees directly but at the population level, through indirect effects such as reduced infectiousness of infected vaccinees, and typical efficacy trial endpoints do not capture them.
Vaccine trials usually enrol more participants than non-vaccine drug trials, because vaccines are generally tested more thoroughly and approval bodies scrutinise them more intensely. The time and cost of this testing deter manufacturers from investing in vaccine development, an investment perceived as risky, and even more so for HIV. HIV vaccine development is scientifically and operationally challenging, complex and lengthy, and its success requires many different candidate vaccines to be studied simultaneously in different populations around the world.
Follow-up and correlates analysis
Sufficient antibodies take time to develop before the full protective picture can be elicited and evaluated, and this adds to the length of participant follow-up. In a population at high risk of acquiring HIV, where more events could occur in a shorter time, follow-up could be three years, and it could be shorter if the sample size is very large. Correlates analyses should be planned prospectively in efficacy trial designs. The timing and frequency of collecting specimens such as serum, plasma, blood cells and mucosal cells after vaccination and after infection must be considered, together with their handling and storage. All HIV infections that occur during prevention trials should be characterised by subtyping and sequencing, and the effect on viral load of any antiretrovirals started should be factored in. All this need for data should be weighed against the operational costs and logistics of collecting it: participant study visits, risky or invasive sampling, and sample processing.
Vaccine efficacy from the cases in each arm
In December 2020 the EMA recommended a conditional marketing authorisation for Comirnaty, developed by BioNTech and Pfizer, to prevent COVID-19 in people from 16 years of age. Its trial involved around 44,000 people, half given the vaccine and half a dummy injection, without knowing which they received. Efficacy was calculated in over 36,000 people from 16 years of age, including people over 75, who had no sign of previous infection. Of the vaccinated people, 8 of 18,198 got COVID-19 symptoms, against 162 of 18,325 given the dummy injection, a 95% reduction in symptomatic cases that the EMA reported as 95% efficacy. The trial also showed around 95% efficacy in participants at risk of severe COVID-19, including those with asthma, chronic lung disease, diabetes, high blood pressure or a body-mass index of 30 kg/m² or more, and the high efficacy was maintained across genders and racial and ethnic groups. The most common side effects were usually mild or moderate and got better within a few days: pain and swelling at the injection site, tiredness, headache, muscle and joint pain, chills and fever. The vaccine's safety and effectiveness were to be monitored through the EU pharmacovigilance system and additional studies by the company and European authorities.
The statistical analysis plan
Statistical analysis plans submitted to approvers should be clear from the beginning about the analysis of overall efficacy and subgroup results, the timing of unblinding for analysis purposes, and the modified intention-to-treat analysis. Where results are discordant, attention should be paid to why.
Reactogenicity
Reactogenicity is the set of local or systemic signs and symptoms after vaccination, such as pain, swelling or redness at the injection site, fever or malaise. Collecting data on these specific adverse reactions is a study process that must be implemented well, usually through diary cards on which participants record their temperature, injection-site reactions such as swelling or redness, and other solicited symptoms. Because these data contribute to the safety endpoints of vaccine studies, participants need to be well trained in completing and returning the cards, with recall bias and their varying cognitive abilities in mind, so that the data collected are accurate and complete. The nature of each adverse event is defined in a standardised manner, such as injection site pain, and immunogenicity analyses include measurements of antibody levels and cell-mediated immunity.
Vaccine-induced seropositivity
Creating an antibody response is the goal of an HIV vaccine, but routine HIV tests usually detect antibodies to HIV in the blood rather than the virus itself, so a vaccine recipient who undergoes routine testing may get a reactive result. This is vaccine-induced seropositivity, and how often it is detected and how long it lasts vary greatly with the product's potency, durability and dosing and with the type of test used. For a participant it can sometimes lead to an incorrect diagnosis, causing stress and unnecessary complications with insurance, military service, blood or tissue donation, immigration and pregnancy, where a false positive antibody test could lead to unnecessary antiretroviral treatment of a baby.
Testing outside the study can also unblind a participant, when a "positive" result from routine testing outside the study is followed by HIV-negative results in the study, and if the participant then changes their risk behaviour, even unintentionally, the study data can be compromised. Education about vaccine-induced seropositivity therefore has to emphasise to all participants the importance of getting all HIV testing done through the study or research site until their vaccine-induced seropositivity is no longer detectable. A registry that verifies previous study participation and receipt of HIV vaccine product, so that further HIV testing can be done promptly, is indicated.
Efficacy inferred from the immune response
The clinical efficacy of the chikungunya vaccines Ixchiq, recommended by the EMA for adults from 18 years, and Vimkunya, recommended in January 2025 for adolescents from 12 years and adults, was inferred from a post-vaccination CHIKV-specific neutralising antibody titre threshold. For Vimkunya the threshold was selected as a surrogate marker and referred to as seroresponse.
The opinion on Ixchiq rested largely on a placebo-controlled study in which the immune response was evaluated in 362 participants, 266 given Ixchiq and 96 placebo. Twenty-eight days after vaccination, 98.9% of those given Ixchiq had antibody titres above the threshold, and titres above it persisted in 99.5% at 12 months and 97.1% at 24 months, with titres to be monitored for up to five years. The opinion on Vimkunya rested largely on two placebo-controlled studies, one in 3,258 people aged 12 to 64 and one in 413 older adults, with the immune response evaluated in 3,355 participants, 2,748 given Vimkunya and 607 placebo. In the first study the difference in seroresponse rates between vaccine and placebo was 46.1% on day 8, 96% on day 15, 96.6% on day 22 and 84% on day 183, and in the second it was 79.5%, 86.2% and 74.4% on days 15, 22 and 183. For each vaccine the EMA's human medicines committee requested a post-authorisation efficacy study to confirm its effectiveness in preventing chikungunya.
In September 2024 the EMA recommended extending the indication of the smallpox and mpox vaccine Imvanex, which contains a live, highly weakened form of modified vaccinia virus Ankara (MVA-BN), from adults to adolescents aged 12 to 17, on the interim results of a study that compared its ability to generate virus-specific antibodies in 315 adolescents and 211 adults. Because the immune response in adolescents was similar to that in adults, it was inferred that the vaccine will provide similar protection in adolescents to that expected in adults. The safety profile in adolescents was comparable to that in adults, with no additional risk identified, and the marketing authorisation holder was asked to submit the final results of the study by 30 May 2025 to further characterise safety in adolescents.
The population to vaccinate
In October 2018 the EMA's human medicines committee recommended granting marketing authorisation for Dengvaxia, a live attenuated tetravalent dengue vaccine, for the prevention of dengue caused by serotypes 1, 2, 3 and 4 in people aged 9 to 45 who live in an endemic area and have already had a dengue virus infection. Its benefits and safety were evaluated in 31 clinical studies conducted mostly in endemic areas of Latin America and Asia Pacific, which together included over 41,000 participants aged 9 months to 60 years who received at least one dose. For people aged 9 to 45, the data showed that the vaccine has positive effects in preventing symptomatic and severe dengue in those who have had a previous dengue infection and live in endemic areas, while people who have never had dengue have an increased risk of clinically severe dengue leading to hospitalisation when they are later infected. The committee therefore recommended limiting use to individuals with prior dengue infection for whom laboratory confirmation of that infection is available before vaccination. It did not recommend vaccinating people who live in non-endemic areas and travel to endemic ones, because there were no safety, immunogenicity or efficacy data to support it. The approved indication excludes the populations of the EU mainland and of territories outside tropical areas, where dengue is not endemic, while a number of EU territories, mainly overseas, lie in endemic areas and could benefit. Additional risk-minimisation measures, such as educational material for physicians and a guide for healthcare professionals, were to be put in place, and use of the vaccine should follow official recommendations from member states.
In October 2017 the EMA released a draft guideline on respiratory syncytial virus for six months of public consultation. It advises developers on vaccines and monoclonal antibodies to prevent RSV disease and on direct-acting antivirals to treat it. It focuses the assessment of safety and efficacy on the people most likely to develop RSV lower respiratory tract infection and severe disease, including newborns of 0 to 27 days, infants of 28 days to 11 months, toddlers of 12 to 23 months, older children likely to develop severe disease, and people over 65. It also addresses vaccinating pregnant women to prevent RSV disease in their babies once born, and gives guidance on study design, on assessing a vaccine's efficacy in different scenarios, and on selecting the recommended dose regimen for medicines.
Approvals and manufacturing
Institutional biosafety committees review research involving recombinant DNA, RNA, other potentially infectious material and transgenic animals, and recommend controls on the biohazards associated with the use of microbiological agents. HIV vaccines fall into that category, because they could consist of substances derived from HIV or from vector viruses such as canarypox, adenovirus or cytomegalovirus, so such a committee must review and approve an HIV vaccine study in addition to the usual regulatory and ethics permissions. These committees represent the local community's interests in public health and the environment. For similar reasons an HIV vaccine may need approval as a genetically modified organism product in some countries, and in South Africa the Department of Agriculture, Forestry and Fisheries must also approve the intentional introduction of GMOs into the environment.
It is vital that a test HIV vaccine for an efficacy trial has consistent batch-to-batch production with defined, reproducible specifications. Formulating a fully characterised vaccine, including its stability and immunogenicity data, takes time, and the capacity to produce it in large quantities over a given period is a further factor. Adenovirus vector vaccines are popular platforms because they satisfy all of these factors.
Site and regulatory capacity
Research sites with insufficient infrastructure often need time, which can take some years, to develop clinic, laboratory and human capacity. Development strategies have to factor this in if host countries and communities are to participate meaningfully in vaccine development, ensure the scientific and ethical conduct of vaccine studies, and function as equal partners with other stakeholders in a collaborative process. Regulatory expertise may also need strengthening to allow timely approvals of research, and regional regulatory harmonisation could hasten the process and enable a wide knowledge-sharing base. The WHO-supported African Vaccine Regulatory Forum is one such initiative, building regulatory capacity where the framework to approve vaccine studies is limited. WHO also supports the Developing Country Vaccine Regulatory Network in strengthening national regulatory authorities in low- and middle-income countries where vaccines are manufactured.
In Africa, the region hardest hit by the epidemic, HIV vaccine trials face unique community, ethical, political, regulatory and scientific challenges. These include national regulatory authorities that are weak or inexperienced in vaccine research, inadequately resourced institutions, undeveloped clinical and laboratory infrastructure, and substandard participant recruitment strategies that may exploit communities with high rates of illiteracy. No entity can single-handedly overcome the hurdles of HIV vaccine development, and there is an urgent ethical need for global support, political will and collaboration to find an HIV vaccine. This necessitates significant international cooperation over time, drawing on partners from health sectors, intergovernmental organisations, government, research institutions, industry and affected populations. Countries with scientific expertise and resources must assist countries that lack the infrastructure and the regulatory and ethical capacity to conduct trials.