BACKGROUND The systemic toxicity of three candidate HIV-1 vaccines plasmid pSG2.HIVconsv DNA (D), ChAdV63.HIVconsv (C) and MVA.HIVconsv (M) expressing chimeric immunogen derived from the most conserved regions of the HIV-1 proteome was evaluated in two repeat-dose studies in the male and female BALB/c mice. METHODS In study UNO011, mice received three doses of 2×10(7) plaque-forming units of MVA.HIVconsv vaccine (MMM). In study UNO012, mice received 3 doses of 50μg of pSG2.HIVconsv DNA followed by a single dose of 5.95×10(9) virus particles of ChAdV63.HIVconsv vaccine (DDDC). Similarly constituted control groups received the vehicle alone (phosphate buffered saline) at the same volume-dose. All vaccines were administered by intramuscular needle injection into the right hind limb at 14-day intervals and animals were sacrificed 7 days after the last dose. Assessment of local and systemic toxicity was made. Induction of HIV-1-specific responses was confirmed. Parameters assessed included clinical condition, body weight, food consumption, ophthalmoscopy, haematology, blood chemistry, organ weight and macroscopic and microscopic pathology. RESULTS In both studies, treatment with the candidate vaccines elicited strong HIV-1-specific T-cell responses. The vaccine treatment was well-tolerated without any adverse systemic toxicological changes. The local toxicity findings observed in these studies were consistent with the predicted response to a vaccine/substance administration by intramuscular injection. CONCLUSIONS The three novel anti-HIV-1 vaccines were well tolerated when administered by intramuscular injection to BALB/c mice. These results supported an application for authorisation by the Medicines and Healthcare Products Regulatory Agency of the UK to test these vaccines for the first time in phase I clinical trials in healthy both uninfected subjects and HIV-1-infected patients stable on antiretroviral treatment.