Structural and Immunological Characterization of Recombinant 6-Cysteine Domains of the Plasmodium falciparum Sexual Stage Protein Pfs230

Abstract

Development of a Plasmodium falciparum (Pf) transmission blocking vaccine (TBV) has the potential to significantly impact malaria control. Antibodies elicited against sexual stage proteins in the human bloodstream are taken up with the blood meal of the mosquitoes and inactivate parasite development in the mosquito. In a phase 1 trial, a leading TBV identified as Pfs25-EPA/Alhydrogel appeared safe and immunogenic, however, the level of Pfs25-specific antibodies were likely too low for an effective vaccine. Pfs230, a 230-kDa sexual stage protein expressed in gametocytes is an alternative vaccine candidate. A unique 6-cysteine-rich domain structure within Pfs230 have thwarted its recombinant expression and characterization for clinical evaluation for nearly a quarter of a century. Here, we report on the identification, biochemical, biophysical, and immunological characterization of recombinant Pfs230 domains. Rabbit antibodies generated against recombinant Pfs230 domains blocked mosquito transmission of a laboratory strain and two field isolates using an ex vivo assay. A planned clinical trial of the Pfs230 vaccine is a significant step toward the potential development of a transmission blocking vaccine to eliminate malaria.Development of a malaria vaccine that effectively protects against parasite infection of both the natural host, Anopheles mosquitoes, and its secondary host, man, would effectively disrupt transmission and clinical disease. The most well known investigational vaccine against Plasmodium falciparum malaria that recently received a positive scientific opinion from the Committee for Medicinal Products for Human Use of the European Medicines Agency in July 2015, is identified as RTS,S (Mosquirix TM ). This vaccine targets the circumsporozoite protein that is present on the surface of the sporozoite, the parasite stage that infects man (1). RTS,S, a virus like-particle-based vaccine, is protective against clinical disease in about 30% of the young children who participated in a phase 3 trial (1, 2).Efforts toward development of a vaccine to disrupt parasite infection of the mosquito host, also identified as a transmission blocking vaccine have to date only been able to evaluate a sexual stage-specific protein, Pfs25, 2 which is a 25-kDa protein expressed on the zygote and ookinete surfaces. A phase 1 study demonstrated that human antibodies raised against a recombinant Pfs25 (Pfs25H) protein formulated in Montanide ISA 51, a water-in-oil adjuvant formulation, were biologically active in an ex vivo feeding assay (3), however, this formulation was not deemed suitable for a public health vaccine. More recently, in preclinical studies, Pfs25H has been shown to have enhanced immunogenic properties when chemically conjugated to a carrier molecule such as Neisseria meningitis outer membrane protein complex (4), or Pseudomonas aeruginosa ExoProtein A (EPA) (5, 6). In particular, the chemically conjugated Pfs25-EPA has the biophysical features of a nanoparticle with a diameter of about 25 m in solution, simil...

The prevalence of submicroscopic Plasmodium falciparum gametocyte carriage and multiplicity of infection in children, pregnant women and adults in a low malaria transmission area in Southern Ghana

“…Recombinant Pf s230D1M (GenBankTM accession number XP_001349600.1) was characterized from Pf s230 domain 1 ( Pf s230D1H) and non-structured domain 1–2 ( Pf s230D1-2). The recombinant protein was refolded by removing the His 6 fusion tag and expressed in Pichia pastoris using good manufacturing practice conditions and purified by RP-HPLC as previously described [ 29 ].…”

Safety and Immunogenicity of Pfs25-EPA/Alhydrogel®, a Transmission Blocking Vaccine against Plasmodium falciparum: An Open Label Study in Malaria Naïve Adults

“…For surface labeling of live parasites, a previously reported procedure [ 25 , 26 ] was followed with a few modifications. In brief, gametocyte-enriched NF54 parasites were allowed to exflagellate and fertilize by incubating in RPMI 1640 (KD Medical, Columbia, MD) containing 10% human serum for 90 min at room temperature.…”

Functional characterization of Plasmodium berghei PSOP25 during ookinete development and as a malaria transmission-blocking vaccine candidate

“…Research on TBVs has led to the identification and experimental validation of several potential TBV candidates, but only a few including Pfs48/45 [8, 9], Pfs230 [10, 11] and Pfs25 [12] in P. falciparum , and Pvs25 and Pvs28 in P. vivax [13], have been found effective in blocking parasite transmission. Investigations on the two 6-cysteine domain protein family members, Pfs48/45 and Pfs230, have shown that anti-Pfs48/45 monoclonal and polyclonal antibodies in experimental animals can effectively inhibit the transmission of P. falciparum to mosquitoes [9, 14, 15], while Pfs230-raised antibodies are sufficient to block development of the oocysts and competent to induce complement-dependent transmission-blocking (TB) activity [11].…”