3M-052 as an adjuvant for a PLGA microparticle-based Leishmania donovani recombinant protein vaccine.

It is believed that an effective vaccine against leishmaniasis will require a T helper type 1 (TH 1) immune response. In this study, we investigated the adjuvanticity of the Toll-like receptor (TLR) 7/8 agonist 3M-052 in combination with the Leishmania donovani 36-kDa nucleoside hydrolase recombinant protein antigen (NH36). NH36 and 3M-052 were encapsulated in separate batches of poly(lactic-co-glycolic acid) (PLGA) microparticles (MPs). The loading efficiency for NH36 was 83% and for 3M-052 was above 95%. In vitro stimulation of bone marrow-derived dendritic cells, measured by IL-12 secretion, demonstrated that 3M-052 (free or MP-formulated) had a concentration-dependent immunostimulatory effect with an optimum concentration of 2 µg/mL. In immunogenicity studies in BALB/c mice, MP-formulated NH36 and 3M-052 elicited the highest serum titers of TH 1-associated IgG2a and IgG2b antibodies and the highest frequency of IFNγ-producing splenocytes. No dose dependency was observed among MP/NH36/3M-052 groups over a dose range of 4-60 µg 3M-052 per injection. The ability of MP-formulated NH36 and 3M-052 to elicit a TH 1-biased immune response indicates the potential for PLGA MP-formulated 3M-052 to be used as an adjuvant for leishmaniasis vaccines. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 1587-1594, 2018.

Cysteine mutagenesis improves the production without abrogating antigenicity of a recombinant protein vaccine candidate for human chagas disease.

A therapeutic vaccine for human Chagas disease is under development by the Sabin Vaccine Institute Product Development Partnership. The aim of the vaccine is to significantly reduce the parasite burden of Trypanosoma cruzi in humans, either as a standalone product or in combination with conventional chemotherapy. Vaccination of mice with Tc24 formulated with monophosphoryl-lipid A (MPLA) adjuvant results in a Th1 skewed immune response with elevated IgG2a and IFNγ levels and a statistically significant decrease in parasitemia following T. cruzi challenge. Tc24 was therefore selected for scale-up and further evaluation. During scale up and downstream process development, significant protein aggregation was observed due to intermolecular disulfide bond formation. To prevent protein aggregation, cysteine codons were replaced with serine codons which resulted in the production of a non-aggregated and soluble recombinant protein, Tc24-C4. No changes to the secondary structure of the modified molecule were detected by circular dichroism. Immunization of mice with wild-type Tc24 or Tc24-C4, formulated with E6020 (TLR4 agonist analog to MPLA) emulsified in a squalene-oil-in-water emulsion, resulted in IgG2a and antigen specific IFNγ production levels from splenocytes that were not significantly different, indicating that eliminating putative intermolecular disulfide bonds had no significant impact on the immunogenicity of the molecule. In addition, vaccination with either formulated wild type Tc24 or Tc24-C4 antigen also significantly increased survival and reduced cardiac parasite burden in mice. Investigations are now underway to examine the efficacy of Tc24-C4 formulated with other adjuvants to reduce parasite burden and increase survival in pre-clinical studies.