Immunogenic evaluation of recombinant proteins expressed in Pichia pastoris based on Plasmodium vivax antigens from different parasite stages

Abstract

Malaria is a serious public health problem worldwide that is recognized as a priority for research and vaccine development. This infectious disease is caused by the parasite Plasmodium. P. vivax is the specie of higher distribution around the world and higher prevalence in the Americas. Despite the intense research efforts in this area, so far, there is no effective vaccine against malaria infection. The complexity of the life cycle and intense antigenic diversity of the parasite infer that it is unlikely that an effective vaccine formulation will be achieved using a single target antigen. In this context, combination of immunodominant regions of antigens from different stages of life cycle of the parasite in a single vaccine formulation has a better prognosis for the induction of protective immune responses. Multistage multi-antigen strategies will aim at improving the anti-parasitic activity and reduce the significant selection of parasites resistant to human immune action. Thus, we propose to evaluate the immunogenic properties in mice of a vaccine formulation composed of pre-erythrocytic and asexual erythrocytic stages of the parasite. Two recombinant proteins, PvAMA-1 and PvCs-All, will be obtained by the yeast expression system Pichia pastoris. They will be purified by affinity chromatography followed by ion exchange chromatography. CS-All chimeric construct is a novel of Circumsporozoite Protein (CS) that merges epitopes mapped in the repeats of central region of three P. vivax strain variants, VK210, VK247 and P. vivax like, permitting coverage of the allelic variability of this immunodominant pre-erythrocytic stage antigen. It is our hypothesis that CS-All is capable of inducing a significant immune response which can be enhanced by the combination with an Apical Membrane Antigen (AMA-1), that has been previously characterized by our group. Immunizations will be performed in the same vaccine formulation with the recombinant proteins inoculated individually or in combination. We will evaluate the specific antibodies by ELISA and by indirect immunofluorescence to confirm the recognition of the native protein. Cellular immune responses will be determined by T cell proliferation and cytokine production. (AU)