Preclinical safety and tolerability of a repeatedly administered human leishmaniasis DNA vaccine

Riede, O; Seifert, K; Oswald, D; Endmann, A; Hock, C; Winkler, A; Salguero, F J; Schroff, M; Croft, S L; Juhls, C

LEISHDNAVAX is an equimass mixture of five MIDGE-Th1 vectors, dissolved in sterile PBS. Each vector encodes one Leishmania antigen: KMP11, CPA, CPB, P74 or TSA, respectively. Their DNA sequences and expression cassettes of MIDGE-Th1 vectors have been described previously. 22 MIDGE-Th1 vectors were synthesized in a simple and standardized procedure. 18 Forty Wistar rats (20 per sex, Janvier Labs, Saint-Berthevin, France), 8 weeks of age, were allocated to single-or four-dose study groups with 10 animals each (five per sex). Animals received needle injections of 120 μg LEISHDNAVAX per injection (~5.3 × 10 13 MIDGE-Th1 vector molecules, injection volume 30 μl) i.d. at the dorsal base of tail, either once or four times in weekly intervals. Animals injected with a single dose were killed 24 h after administration. Four-dose groups were either killed 24 h, 14 days or 60 days after the fourth injection. Clinical signs were recorded before, 30 min and 4 h after treatment, thereafter once daily. Twenty-four hours after injection, a modified Irwin test 39 was performed on the single-dose group and on a non-treated control group to assess acute neurological toxicity of the vaccine. Throughout the study, mortality was evaluated once daily and body weight once weekly. At killing, animals were anesthetized deeply by isoflurane inhalation and blood was collected from the plexus ophthalmicus. Gross pathology was performed and samples of the following tissues were taken: blood, thigh bone marrow, brain, heart, kidneys, liver (Lobus quadratus), lymph nodes (Lnn. axillares, inguinales, mesenteriales), lung, thigh muscle, ovaries, skin of injection site, spleen and testes. To avoid cross-contamination with vector DNA single-use scalpels were applied for each sample. All other equipment was thoroughly decontaminated after each dissection. Samples were immediately frozen in liquid nitrogen and stored at − 80°C until processing.