The two recombinantly Panobinostat nmr produced vaccine antigens, RTS,S and TRAP, were manufactured by GlaxoSmithKline (GSK) Biologicals (Rixensart, Belgium). The RTS,S vaccine antigen has been described [12]. The TRAP antigen is a recombinant protein produced in, and purified from, the culture supernatant of insect cells (Spodoptera frugiperda Sf9 cell line) infected with a recombinant baculovirus (AcMNPV). The baculovirus expresses a truncated form of the TRAP gene derived from P. falciparum

strain NF54 (clone 3D7). The final purified antigen consists of a 493 amino acid long polypeptide comprising amino acids 26 (arginine/R) to 511 (lysine/K) of the authentic TRAP protein, extended at its carboxy terminal end by the addition of 7 histidine residues. The antigens (RTS,S/TRAP or TRAP) were presented as lyophilized pellets in single dose vials. Just before administration, each pellet was reconstituted with liquid AS02 Adjuvant System [12]. Subjects received 50 μg RTS,S or 25 μg TRAP or both 50 μg of RTS,S and 25 μg of TRAP together with 50 μg MPL, and 50 μg QS21 in an oil/water emulsion as a 0.5 mL dose, by intramuscular injection. Local and systemic adverse events (AEs) were

systematically assessed using standardized criteria as previously reported [2] (see Supplementary Appendix). All unsolicited reports of AEs occurring within 30 days, and of reactogenicity within 4 days, of vaccination were recorded. Serious AEs (SAEs) were collected PAK6 throughout the study. Hematological and biochemical tests for safety evaluation were performed and any clinically significant values Perifosine noted. Antibodies (IgG) against the CS central repeat tetrapeptide epitopes were measured using ELISA with recombinant R32LR as the capture antigen as described previously [35] and [36]. Antibodies against TRAP were measured by ELISA using the vaccine antigen as the capture antigen, and expressed as titers. For both studies,

the peripheral blood mononuclear cells (PBMCs) were separated from heparinized whole blood on a density gradient and stored in liquid nitrogen as described previously [37]. Lymphoproliferative (LP) results were expressed as stimulation indices (SI*) which are the ratio between the quantities of 3H-thymidine incorporated by the cells in the presence of a specific antigen and the ones incorporated by the cells cultured in medium alone (for assay methodologies, see the Supplementary Appendix). IFN-γ and IL-5 secretion by whole PBMC was measured in supernatant harvested from antigen-stimulated PBMC after 120 h by commercial ELISA kit (respectively IFN-γ EASIA®; Medgenix, Fleurus, Belgium or Biosource International, Camarillo, CA). Further detail is provided in the Supplementary Appendix. ELISPOT assays were conducted as previously described (see Supplementary Appendix) [5] and [38].