Identification of Novel Pre-Erythrocytic Malaria Antigen Candidates for Combination Vaccines with Circumsporozoite Protein

Malaria vaccine development has been hampered by the limited availability of antigens identified through conventional discovery approaches, and improvements are needed to enhance the efficacy of the leading vaccine candidate RTS,S that targets the circumsporozoite protein (CSP) of the infective sporozoite. Here we report a transcriptome-based approach to identify novel pre-erythrocytic vaccine antigens that could potentially be used in combination with CSP. We hypothesized that stage-specific upregulated genes would enrich for protective vaccine targets, and used tiling microarray to identify P. falciparum genes transcribed at higher levels during liver stage versus sporozoite or blood stages of development. We prepared DNA vaccines for 21 genes using the predicted orthologues in P. yoelii and P. berghei and tested their efficacy using different delivery methods against pre-erythrocytic malaria in rodent models. In our primary screen using P. yoelii in BALB/c mice, we found that 16 antigens significantly reduced liver stage parasite burden. In our confirmatory screen using P. berghei in C57Bl/6 mice, we confirmed 6 antigens that were protective in both models. Two antigens, when combined with CSP, provided significantly greater protection than CSP alone in both models. Based on the observations reported here, transcriptional patterns of Plasmodium genes can be useful in identifying novel pre-erythrocytic antigens that induce protective immunity alone or in combination with CSP.

疟疾疫苗研发长期受限于传统发现手段所鉴定的抗原资源有限，且现有核心候选疫苗RTS,S——靶向感染性子孢子的环子孢子蛋白（circumsporozoite protein, CSP）——的效力仍需优化。本研究报道了一种基于转录组学的策略，用于筛选可与CSP联合应用的新型红细胞前期疫苗抗原。我们提出假说：阶段特异性上调基因可富集保护性疫苗靶点，并通过平铺式微阵列（tiling microarray）鉴定出在肝脏发育阶段较子孢子或血液发育阶段转录水平更高的恶性疟原虫（Plasmodium falciparum）基因。我们利用约氏疟原虫（Plasmodium yoelii）和伯氏疟原虫（Plasmodium berghei）的预测同源基因，针对21个基因制备了DNA疫苗，并在啮齿动物模型中采用不同递送方式，评估其对抗红细胞前期疟疾的效力。在以BALB/c小鼠感染约氏疟原虫的初步筛选实验中，我们发现16种抗原可显著降低肝脏阶段的寄生虫载量。在以C57BL/6小鼠感染伯氏疟原虫的验证性筛选实验中，我们确认了6种在两种模型中均具有保护性的抗原。其中两种抗原与CSP联合使用时，在两种模型中均展现出较单独使用CSP更显著的保护效果。基于本研究的实验结果，疟原虫基因的转录模式可用于鉴定新型红细胞前期抗原，此类抗原可单独或与CSP联合诱导保护性免疫。