Evaluation of adjuvants for a pulmonary vaccine against tuberculosis

Tuberculosis (TB) is a highly infectious disease transmitted by Mycobacterium tuberculosis and causing infection in 1/3 of the world population. The current vaccine against TB, the Bacille of Calmette and Guérin (BCG), has failed to control the disease. This has led to the search for new vaccines that could efficiently boost BCG-induced immune responses and increase protective immunity. Pulmonary vaccination appears a promising route for vaccination as the lungs are the natural site of infection of M. tuberculosis. This thesis work aimed to evalutate immunogenicity and safety, after pulmonary delivery, of three potential mucosal adjuvants and a biodegradable copolymer in combination with antigen 85A (Ag85A), an immunodominant antigen of M. tuberculosis. For this purpose, the following mucosal adjuvants were selected: an oligonucleotide containing CpG motifs (CpG), the monophosphoryl lipid A of Salmonella Minnesota (MPLA) and the B subunit of heat-labile enterotoxin of Escherichia coli (LTB). Poloxamer 407 (P407) copolymer was chosen for its gelation properties. P407 can be administered in liquid frorm and acts as sustained release depot at body temperature. In a first time, we evaluated the optimal delivery site within the lungs of Ag85A. We further analyzed immune responses after pulmonary administration of Ag85A combined to CpG oligonucleotide and P407 as well as the safety in the lungs of CpG, P407 and the combination CpG-P407. Results showed that targeting the deep lungs induced most intense immune responses than targeting the upper airways. Thereby, the deep lungs were chosen as preferential site for vaccine delivery. The combination of CpG with P407 induced the highest Th-1 and Th-17 immune responses. Moroever, P407 showed to increase the residence time of the antigen within the deep lungs, suggesting the mechanism of P407 to enhance immune responses. However, P407 produced a reversible alteration of the alveolo-capillary barrier. We compared, in a second time, the abilities of CpG adjuvant to MPLA and LTB to induce immune responses and protective efficacy to Ag85A after delivery to the lungs of mice. We also compared the safety of CpG, MPLA and LTB after pulmonary administration. Both CpG and MPLA showed a Th-1 immune response. In addition, MPLA stimulates Th-17 immmune response. By contrast, LTB adjuvant promoted a Th-2 biased immune response. CpG did not induce inflammation in the lungs while LTB and MPLA showed a transient inflammation with neutrophil influx one day after administration. CpG, MPLA and LTB did not appear strong enough adjuvants to induce protection against a challenge with virulent M. tuberculosis H37Rv. In conclusion, the pulmonary delivery of CpG and MPLA provided good immune responses with Th-1 and Th-17 orientations and acceptable safety profile. However the vaccination protocol needs further optimization to induce good protection. Combining CpG and MPLA might be promising as CpG could enhance adjuvanticity of MPLA without increasing its toxicity.