RECOMBINANT LISTERIA VACCINES FOR MELANOMA

单核细胞增生李斯特菌是一种革兰氏阳性细菌，能够进入宿主细胞，从内吞囊泡中逃逸，在细胞质内繁殖，并直接从细胞传播到细胞而不遇到细胞外环境。获得进入宿主细胞胞质溶胶的能力允许细菌分泌的蛋白质有效地通过MHC I类抗原加工和呈递途径，导致诱导CD 8+细胞毒性T细胞（CTL）。我们建立了一个表达和分泌外源抗原的遗传系统。单核细胞增多症表达淋巴细胞性脉络丛脑膜炎病毒（LCMV）核蛋白（NP）或特异性MHC I类限制性NP表位的重组疫苗株能够在接种后诱导小鼠中的LCMV特异性CD 8+细胞毒性T淋巴细胞（CTL）应答。这些菌株赋予抗病毒保护，如免疫小鼠有效清除LCMV感染的能力所示。还构建了表达E11蛋白棉尾兔乳头瘤病毒（CRP）的李斯特菌菌株。用这些重组体免疫兔可使CRPV诱导的乳头状瘤消退并保护其免于癌。这些和其他结果证明了李斯特菌疫苗菌株用于诱导抗病毒和抗肿瘤免疫的效用。我们的目标是探索重组李斯特菌菌株作为抗肿瘤疫苗的效用，并确定减毒和抗原递送的最佳策略。具体目标是：1.结构L.表达肿瘤排斥抗原的单核细胞增生疫苗株。L.将构建表达黑色素瘤相关肿瘤排斥抗原gp 100、MART 1、TRP 2或H-2K/B限制性TRP 2 181-188表位的单核细胞增生菌株。2.确定预防性或治疗性施用重组李斯特菌疫苗菌株对肿瘤建立、生长和消退的影响。两种鼠肿瘤模型将用于测试疫苗效力。3.无害李斯特菌疫苗株的构建及其与野生型和减毒李斯特菌的免疫原性比较。单核细胞增多症L.无害是李斯特菌属的非致病性成员。将L.因此，我们将尝试构建免疫原性但无毒的嵌合菌株。

Listeria monocytogenes is a gram positive bacterium that is able to enter host cells, escape from the endocytic vesicle, multiply within the cytoplasm and spread directly from cell-to-cell without encountering the extracellular milieu. The ability to gain access to the host cell cytosol allows proteins secreted by the bacterium to efficiently the MHC class I antigen processing and presentation pathway leading to the induction of CD8+ cytotoxic T cells (CTL). We developed a genetic system for expression and secretion of foreign antigens by L. monocytogenes. Recombinant vaccine strains expressing the lymphocytic choriomeningitis virus (LCMV) nucleoprotein (NP), or a specific MHC class I restricted NP epitope, are able to induce LCMV specific CD8+ cytotoxic T lymphocyte (CTL) responses in mice following vaccination. These strains confer antiviral protection as indicated by the ability of immunized mice to efficiently clear LCMV infection. Listeria strains that express the E11 protein cottontail rabbit papillomavirus (CRP) have also been constructed. Immunization of rabbits with these recombinants causes regression of CRPV induced papillomas and protection from carcinoma. These and other results demonstrate the utility of Listeria vaccine strains for inducing antiviral and anti-tumor immunity. Our objectives are to explore the utility of recombinant Listeria strains as anti-tumor vaccines and determine optimal strategies for attenuation and antigen delivery. The specific aims are to: 1. Construct L. monocytogenes vaccine strains expressing tumor rejection antigens. L. monocytogenes strains expressing the melanoma associated tumor rejection antigens gp100, MART1, TRP2, or an H-2K/b restricted TRP2 181-188 epitope will be constructed. 2. Determine the effects of prophylactic or therapeutic administration of recombinant Listeria vaccine strains on tumor establishment, growth and regression. Two murine tumor models will be used to test vaccine efficacy. 3. Construct Listeria innocua vaccine strains and compare their immunogenicity with wild type and attenuated L. monocytogenes. L. innocua is a non-pathogenic member of the Listeria genus. By transferring genes from L. monocytogenes, we will attempt to construct chimeric strains that are immunogenic yet avirulent.