Laser-facilitated delivery of malarial sporozoites from the skin to liver

描述（由申请人提供）：本提案的总体目标是证明激光预照明可以增强孢子子（SPZs）的皮肤到肝脏传递，增强基于spz的疫苗。预防疟原虫感染仍然迫切需要一种有效的、获得许可的抗疟疾疫苗。迄今为止，疟疾疫苗开发人员的“黄金标准”仍然是通过辐射(g)感染spz的蚊子叮咬进行免疫接种。然而，将这种方法转化为临床面临着巨大的障碍，因为通过传染性蚊子叮咬为大量人群接种疫苗不现实，而且这种免疫途径不符合伦理。对于以SPZ为基础的疫苗，无论SPZ是通过辐照还是基因改造来减毒的，其效力都与转运到肝脏的SPZ数量相关。虽然静脉注射（IV）可能最充分地将spz输送到肝脏，但它不是临床批准的免疫途径。另一方面，皮内接种（ID）疫苗，不仅模仿自然感染，也是临床可接受的途径，但效率远低于静脉注射，因为spz从皮肤中迁移受到真皮中密集排列的网络的限制，这对于冷冻保存的gspz来说尤其如此，因为它们的活动性降低。我们发现接种部位的激光预照射在注射前将gspz的皮肤到肝脏的交通增强到与静脉注射相当的水平。光照瞬间破坏了皮肤致密的微结构，允许spz相对自由地运动，并进入真皮层的毛细血管和淋巴管。我们假设这种激光治疗可以增加基于spz的疫苗的ID递送，并且当与a-GalCer （a-半乳糖神经酰胺）联合使用时，可以进一步增强spz诱导的疟疾保护。为了验证这一假设，我们将验证激光介导的促进皮肤到肝脏的spz递送是否有效

DESCRIPTION (provided by applicant): The overall objective of this proposal is to demonstrate that laser pre-illumination can enhance skin-to-liver delivery of sporozoites (SPZs), augmenting SPZ-based vaccines. An effective, licensed anti-malarial vaccine remains an urgent need for prevention against plasmodium infection. To date, the "gold standard" for malarial vaccine developers is still immunization by bites of irradiation (g), SPZ-infected mosquitoes. However, translation of this approach into the clinic faces formidable obstacles, due to impracticality of vaccinating a large number of people by bites of infectious mosquitoes and unethic of this route of immunization. For SPZ- based vaccines, irrespective of whether the SPZs are attenuated by irradiation or genetic modification, the efficacy is correlated with the number of SPZs trafficking into the liver. While intravenous (IV) injection may be most sufficient in delivering SPZs to the liver, it is not a clinically approved route of immunization. On the othe hand, intradermal (ID) vaccination, not only mimicking natural infection but also a clinically acceptable route, is much less efficient than IV because emigration of SPZs out of the skin is restricted by the densely packed network in the dermis, which may be particularly true for cryopreserved gSPZs owing to their reduced motility. We found that laser pre-illumination of the inoculation site prior to ID enhanced skin-to-liver traffic of gSPZs to a level comparable to IV injection. The illumination transiently disrupted the dense microarchitecture of the skin, permitting relatively free movement of SPZs and their entry into the capillaries and lymphatic vessels in the dermis. We hypothesize that this laser treatment can augment ID delivery of SPZ-based vaccines and when combined with a-GalCer (a-galactosylceramide), an adjuvant for SPZ-induced immunity, can further boost SPZ-induced protection against malaria. To test this hypothesis, we will verify whether laser-mediated facilitation of skin-to-liver delivery of SPZs is SPZ dose- and/or motility-dependent by quantification of the number of SPZs in the liver 40 hr after ID injection of varying numbers of SPZs into the site of laser illumination. This dose-dependent effect will be then tested with freshly isolated or cryopreserved gSPZs or genetically attenuated SPZs. We will also validate in outbred and inbred mouse models that laser illumination followed by ID inoculation of cryopreserved gSPZs or genetically attenuated SPZs affords similar protection against virulent SPZ infections as IV vaccination and a synergistic effect between laser and a-GalCer adjuvant on the protective immunity. The preclinical study, if successful, can potentially take SPZ-based vaccines one step closer to their clinical application. PUBLIC HEALTH RELEVANCE: Brief cutaneous illumination with a safe and noninvasive laser will be employed to facilitate skin-to-liver transportation of live attenuated malarial sporozoite vaccine. Sufficient delivery of the vaccine to the liver will greatly boost the efficacy of the vaccine.