Canepa Gaspar E, Molina-Cruz Alvaro, Yenkoidiok-Douti Lampouguin, Calvo Eric, Williams Adeline E, Burkhardt Martin, Peng Fangni, Narum David, Boulanger Martin J, Valenzuela Jesus G, Barillas-Mury Carolina
1Laboratory of Malaria and Vector Research, National Insti6tute of Allergy and Infectious Diseases, National Institutes of Health, Rockville, MD 20852 USA.
2Fischell Department of Bioengineering, University of Maryland, College Park, MD 20742 USA.
NPJ Vaccines. 2018 Jul 10;3:26. doi: 10.1038/s41541-018-0065-5. eCollection 2018.
Transmission-blocking vaccines are based on eliciting antibody responses in the vertebrate host that disrupt parasite development in the mosquito vector and prevent malaria transmission. The surface protein Pfs47 is present in gametocytes and female gametes. The potential of Pfs47 as a vaccine target was evaluated. Soluble full-length recombinant protein, consisting of three domains, was expressed in as a thioredoxin fusion (T-Pfs47). The protein was immunogenic, and polyclonal and monoclonal antibodies (mAb) were obtained, but they did not confer transmission blocking activity (TBA). All fourteen mAb targeted either domains 1 or 3, but not domain 2 (D2), and immune reactivity to D2 was also very low in polyclonal mouse IgG after T-Pfs47 immunization. Disruption of the predicted disulfide bond in D2, by replacing cysteines for alanines (C230A and C260A), allowed expression of recombinant D2 protein in . A combination of mAbs targeting D2, and deletion proteins from this domain, allowed us to map a central 52 amino acid (aa) region where antibody binding confers strong TBA (78-99%). This 52 aa antigen is immunogenic and well conserved, with only seven haplotypes world-wide that share 96-98% identity. Neither human complement nor the mosquito complement-like system are required for the observed TBA. A dramatic reduction in ookinete numbers and ookinete-specific transcripts was observed, suggesting that the antibodies are interacting with female gametocytes and preventing fertilization.
传播阻断疫苗的原理是在脊椎动物宿主体内引发抗体反应，从而破坏疟原虫在蚊媒中的发育，防止疟疾传播。表面蛋白Pfs47存在于配子体和雌配子中。对Pfs47作为疫苗靶点的潜力进行了评估。由三个结构域组成的可溶性全长重组蛋白在大肠杆菌中作为硫氧还蛋白融合蛋白（T-Pfs47）表达。该蛋白具有免疫原性，获得了多克隆抗体和单克隆抗体（mAb），但它们不具有传播阻断活性（TBA）。所有14种mAb都靶向结构域1或结构域3，而非结构域2（D2），并且在T-Pfs47免疫后，多克隆小鼠IgG对D2的免疫反应性也非常低。通过将半胱氨酸替换为丙氨酸（C230A和C260A）破坏D2中预测的二硫键，使得重组D2蛋白能够在大肠杆菌中表达。靶向D2的mAb组合以及该结构域的缺失蛋白，使我们能够定位一个中央52个氨基酸（aa）的区域，抗体结合该区域可赋予强传播阻断活性（78 - 99%）。这种52个氨基酸的抗原具有免疫原性且高度保守，全球仅存在7种单倍型，它们的同一性为96 - 98%。观察到的传播阻断活性既不需要人类补体也不需要蚊子补体样系统。观察到动合子数量和动合子特异性转录本显著减少，这表明抗体正在与雌配子体相互作用并阻止受精。
