Ntege Edward H, Arisue Nobuko, Ito Daisuke, Hasegawa Tomoyuki, Palacpac Nirianne M Q, Egwang Thomas G, Horii Toshihiro, Takashima Eizo, Tsuboi Takafumi
Division of Malaria Research, Proteo-Science Center, Ehime University, Matsuyama, Ehime 790-8577, Japan.
Department of Molecular Protozoology, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan.
Vaccine. 2016 Nov 4;34(46):5612-5622. doi: 10.1016/j.vaccine.2016.09.028. Epub 2016 Sep 28.
Genetic variability in Plasmodium falciparum malaria parasites hampers current malaria vaccine development efforts. Here, we hypothesize that to address the impact of genetic variability on vaccine efficacy in clinical trials, conserved antigen targets should be selected to achieve robust host immunity across multiple falciparum strains. Therefore, suitable vaccine antigens should be assessed for levels of polymorphism and genetic diversity. Using a total of one hundred and two clinical isolates from a region of high malaria transmission in Uganda, we analyzed extent of polymorphism and genetic diversity in four recently reported novel blood-stage malaria vaccine candidate proteins: Rh5 interacting protein (PfRipr), GPI anchored micronemal antigen (PfGAMA), rhoptry-associated leucine zipper-like protein 1 (PfRALP1) and Duffy binding-like merozoite surface protein 1 (PfMSPDBL1). In addition, utilizing the wheat germ cell-free system, we expressed recombinant proteins for the four candidates based on P. falciparum laboratory strain 3D7 sequences, immunized rabbits to obtain specific antibodies (Abs) and performed functional growth inhibition assay (GIA). The GIA activity of the raised Abs was demonstrated using both homologous 3D7 and heterologous FVO strains in vitro. Both pfripr and pfralp1 are less polymorphic but the latter is comparatively more diverse, with varied number of regions having insertions and deletions, asparagine and 6-mer repeats in the coding sequences. Pfgama and pfmspdbl1 are polymorphic and genetically diverse among the isolates with antibodies against the 3D7-based recombinant PfGAMA and PfMSPDBL1 inhibiting merozoite invasion only in the 3D7 but not FVO strain. Moreover, although Abs against the 3D7-based recombinant PfRipr and PfRALP1 proteins potently inhibited merozoite invasion of both 3D7 and FVO, the GIA activity of anti-PfRipr was much higher than that of anti-PfRALP1. Thus, PfRipr is regarded as a promising blood-stage vaccine candidate for next-generation vaccines against P. falciparum.
恶性疟原虫疟疾寄生虫的基因变异性阻碍了当前疟疾疫苗的研发工作。在此，我们假设，为解决基因变异性对临床试验中疫苗效力的影响，应选择保守的抗原靶点，以在多种恶性疟原虫菌株中实现强大的宿主免疫。因此，应评估合适的疫苗抗原的多态性和基因多样性水平。我们使用来自乌干达疟疾高传播地区的总共102份临床分离株，分析了四种最近报道的新型血液期疟疾疫苗候选蛋白的多态性程度和基因多样性：Rh5相互作用蛋白（PfRipr）、糖基磷脂酰肌醇锚定的微线体抗原（PfGAMA）、棒状体相关亮氨酸拉链样蛋白1（PfRALP1）和达菲结合样裂殖子表面蛋白1（PfMSPDBL1）。此外，利用小麦胚无细胞系统，我们基于恶性疟原虫实验室菌株3D7序列表达了这四种候选蛋白的重组蛋白，免疫兔子以获得特异性抗体（Abs），并进行了功能生长抑制试验（GIA）。使用同源的3D7和异源的FVO菌株在体外证明了所产生抗体的GIA活性。PfRipr和PfRALP1的多态性均较低，但后者的基因多样性相对较高，其编码序列中有不同数量的区域存在插入和缺失、天冬酰胺和6聚体重复序列。PfGAMA和PfMSPDBL1在分离株中具有多态性且基因多样，针对基于3D7的重组PfGAMA和PfMSPDBL1的抗体仅在3D7菌株而非FVO菌株中抑制裂殖子入侵。此外，尽管针对基于3D7的重组PfRipr和PfRALP1蛋白的抗体能有效抑制3D7和FVO的裂殖子入侵，但抗PfRipr的GIA活性远高于抗PfRALP1。因此，PfRipr被认为是下一代抗恶性疟原虫疫苗的有前景的血液期疫苗候选物。
