Safety, tolerability and immunogenicity of new formulations of the Plasmodium falciparum malaria peptide vaccine SPf66 combined with the immunological adjuvant QS-21

Abstract

SPf66 is a synthetic malaria peptide vaccine, which has been widely tested in combination with aluminium hydroxide (alum) as the adjuvant. Since this formulation is weakly immunogenic, we sought to improve its immunogenicity by using the saponin adjuvant QS-21. SPf66/QS-21 vaccines were evaluated for safety, tolerability and immunogenicity in healthy adults. The vaccines were found to be safe in 87/89 (97.8%) volunteers studied. However, two individuals developed severe vaccine allergy following the third dose of 1/3 SPf66/QS-21 formulations tested. Vaccine formulations containing QS-21 induced a 45- to over 200-fold increase in anti-SPf66 IgG titres over the alum formulation after the second and third doses, respectively. Anti-SPf66 antibody from some subjects reacted against asexual blood stage parasites, as demonstrated by immunofluorescence and immunoblotting. Antibody responses generated by the QS-21 formulations were of longer duration compared to those evoked by the alum formulation. While SPf66/alum has been found to induce only CD4+ T cell response, the QS-21 formulations exhibited the potential to also elicit SPf66-specific CD8+ responses. These observations demonstrate that the use of QS-21 can substantially enhance the immunogenicity of peptide vaccines, such as SPf66.

Introduction

The search for a malaria vaccine, especially one against Plasmodium falciparum, has been fuelled in recent years, owing to increased resistance of the parasite to anti-malarial drugs, and that of the vectors to most insecticides [1]. These factors have reinforced the view that a vaccine against P. falciparum is an urgently required tool for the prevention and control of malaria world-wide and especially in sub-Saharan Africa [2].

The malaria vaccine SPf66 was developed by Patarroyo et al. [3], [4]. SPf66 consists of a polymeric synthetic peptide incorporating amino acid sequences derived from three proteins isolated from P. falciparum infected erythrocytes. The peptide epitopes are linked by Pro-Asn-Ala-Asn-Pro (PNANP) sequences from the P. falciparum circumsporozoite protein (CSP) repeat unit, and all four peptide sequences are assembled into a 45-amino acid long monomer unit. The monomer has cysteine residues added at the C- and N-terminal ends to allow for polymerisation. Current SPf66 vaccine formulations use aluminium hydroxide (alum) as the adjuvant.

Several malaria vaccines are under clinical development, and a few have reached advanced evaluation in field trials in humans [2]. The SPf66 vaccine is the only candidate to have been evaluated extensively for safety, immunogenicity and efficacy in several countries, including Colombia [5], [6], [7], Ecuador [8], Venezuela [9], Tanzania [10], [11], [12], The Gambia [13], Thailand [14] and Brazil [15]. Results from all these trials have shown the alum-formulated vaccine to be safe in malaria-naı̈ve adults [10], [16], and semi-immune and immune children and adults [5], [6], [7], [8], [9], [10], [11], [12], [13], [14]. Efficacy evaluation of the vaccine has nevertheless led to mixed results, demonstrating some effects against P. falciparum malaria in some of the major trials [17], while producing no significant effect in Gambian and Tanzanian children under 1 year of age [12], [13] and in older children from northern Thailand [14]. Interestingly, a reduction in multiple infections was found in asymptomatic vaccine recipients compared with those in asymptomatic placebo recipients [18]. This observation was corroborated in Gambian children [19].

One commonality to all SPf66 vaccine trials has been the observation that the alum-adsorbed SPf66 vaccine is poorly immunogenic in humans. For instance, induction of measurable levels of antibody against SPf66 often requires a three-dose regimen with a large antigen dose of 2 mg for each adult immunisation. Moreover, the antibody levels evoked by vaccination are generally of short duration [16], returning to baseline levels within 15–24 months following the immunisation [11], [14], [16]. Also, SPf66-specific T cell responses tend to be very low [20]. There are a number of reasons that could explain the mixed efficacy results observed with SPf66/alum vaccines. However, given the observation that modest immune responses to SPf66 resulted in promising, although partial efficacy in some studies, we hypothesised that improving the immune responses to SPf66 could result in improved and more consistent immune protection. This has provided us with an incentive for replacing alum with a more potent adjuvant, QS-21 and analysing the immunogenicity of various formulations of QS-21-adjuvanted SPf66 vaccines.

QS-21 has been evaluated for adjuvant effects when combined with a variety of antigens, and has been shown to enhance immune responses to subunit vaccines in laboratory animals [21] and humans [22], [23], [24], [25], [26]. In human challenge studies of the P. falciparum CSP vaccine RTS,S [24], the addition of QS-21 to a vaccine formulation markedly improved the vaccine immunogenicity, and enhanced the vaccine efficacy, resulting in 50–80% protection against experimental challenge with P. falciparum sporozoites [24], [27]. QS-21 has also been shown to enhance the antibody responses against a P. falciparum CSP-derived multiple antigen peptide (MAP) vaccine in healthy adults [28]. The addition of QS-21 to alum-precipitated MAP vaccines induced higher levels of IgG anti-MAP antibody compared to the alum-bound preparation. Our recent experiments in Aotus nancymai monkeys immunised with SPf66 vaccines formulated with either alum or QS-21 showed a higher degree of protection in animals administered QS-21-containing vaccines than those immunised with alum-bound peptide (unpublished results), suggesting that an SPf66 vaccine formulated with QS-21 may be highly desirable for effective immunisation of individuals living in areas where malaria is endemic. These observations provided an incentive for the assessment of the safety, tolerability and immunogenicity of QS-21-adjuvanted SPf66 vaccines. We report here the results of the first phase I clinical trial of the SPf66/QS-21 formulations in healthy adults from Colombia.