Pregnancy Malaria Vaccine development

胎盘型疟疾是一个主要的公共卫生问题，与严重的孕产妇贫血、先兆子痫、妊娠丢失、低出生体重分娩和婴儿死亡有关。PM是由结合受体硫酸软骨素A（CSA）的寄生虫在胎盘中的隔离引起的。以前的体外实验表明，寄生虫与CSA的结合可以被抗体抑制，抗体来自获得特异性免疫力的经产妇女CSA结合寄生虫，或抗体免疫动物介导的寄生虫与CSA结合的抗原。 自本年度刊发以来，我们报告二零二零财政年度的以下进展： 1. Doritchamou JYA，莫里森R，Renn JP，Ribeiro JMC，Duan J，Fried M，Duffy PE.胎盘疟疾疫苗候选抗原VAR 2CSA在某些恶性疟原虫株中显示非典型结构域结构。通信生物学2019年12月6日;2：457。doi：10.1038/s42003-019-0704-z。 在这里，我们确定了非典型的扩展或截短的VAR 2CSA细胞外结构，并确认了一个扩展的结构在马里母体分离，使用一种新的蛋白质片段组装方法的RNA-seq和DNA-seq数据。扩展结构包括常规NTS-DBL 1X-6结构域结构下游的一个或两个额外的DBL结构域，与var 2csa和非var 2csa基因中的DBL最接近。总体而言，4/82株分离株显示非典型VAR 2CSA结构。表达延伸的VAR 2CSA的母体分离物与CSA结合，但其重组VAR 2CSA与CSA的结合不如VAR 2CSANF 54，并且对天然获得的产次依赖性抗体显示出较低的反应性。我们的蛋白片段序列组装方法揭示了影响抗原反应性和功能的非典型VAR 2CSA结构域架构，并应告知基于VAR 2CSA的疫苗的设计。 我们在本报告所述期间未公布的进展包括以下进展： 于2020财政年度，我们继续在哺乳动物悬浮细胞（HEK 293）中使用全长PfVAR 2CSA的7个不同等位基因的重组版本进行研究。 使用这些宝贵的试剂，我们的特点是抗原性和等位基因的全长VAR 2CSA的第一次差异。所有七种VAR 2CSA重组体以剂量依赖性方式与CSA结合，并且来自马里孕妇的血浆表现出对这些抗原的反应性，其随着产次增加而增加，表明正确折叠的蛋白质。然而，等位基因变体在其CSA结合水平和血清反应性方面存在显著差异。我们的研究结果表明，正确折叠和功能VAR 2CSA变体显示生物物理和免疫学差异，可能有助于自然发生的PM严重程度和获得保护性免疫的差异。

Placental malaria (PM) is a major public health problem associated with severe maternal anemia, preeclampsia, pregnancy loss, low birthweight delivery and infant mortality. PM is caused by sequestration in the placenta of parasites that bind the receptor chondroitin sulfate A (CSA). Previous ex vivo experiments have shown that parasite binding to CSA can be inhibited by antibodies from multigravida women who acquired specific immunity to CSA-binding parasites, or by antibodies from animals immunized with antigens that mediate parasite binding to CSA. From our publication this year, we report the following advances in FY2020: 1. Doritchamou JYA, Morrison R, Renn JP, Ribeiro JMC, Duan J, Fried M, Duffy PE. Placental malaria vaccine candidate antigen VAR2CSA displays atypical domain architecture in some Plasmodium falciparum strains. Communications Biology. 2019 Dec 6;2:457. doi: 10.1038/s42003-019-0704-z. Here, we identified atypical extended or truncated VAR2CSA extracellular structures and confirmed one extended structure in a Malian maternal isolate, using a novel protein fragment assembly method for RNA-seq and DNA-seq data. Extended structures included one or two additional DBL domains downstream of the conventional NTS-DBL1X-6 domain structure, with closest similarity to DBL in var2csa and non-var2csa genes. Overall, 4/82 isolates displayed atypical VAR2CSA structures. The maternal isolate expressing an extended VAR2CSA bound to CSA, but its recombinant VAR2CSA bound less well to CSA than VAR2CSANF54 and showed lower reactivity to naturally acquired parity-dependent antibody. Our protein fragment sequence assembly approach revealed atypical VAR2CSA domain architectures that impact antigen reactivity and function, and should inform the design of VAR2CSA-based vaccines. Our unpublished progress during this reporting period includes the following advances: In FY2020, we continued studies using recombinant version of 7 different alleles of full-length PfVAR2CSA in mammalian suspension cells (HEK293). Using these invaluable reagents, we have characterized allelic differences in antigenicity and adhesiveness of full-length VAR2CSA for the first time. All seven VAR2CSA recombinants bound to CSA in a dose-dependent manner, and plasma from Malian pregnant women demonstrated reactivity to these antigens that increased with parity, suggesting properly folded protein. However, allelic variants differed significantly in their CSA-binding levels and their serum reactivities. Our findings indicate correctly folded and functional VAR2CSA variants display biophysical and immunological differences that may contribute to naturally occurring differences in PM severity and acquisition of protective immunity.