IGF::OT::IGF ANTI FUSOBACTERIUM NUCLEATUM VACCINE FOR COLORECTAL CANCER IMMUNOPREVENTION

结直肠癌（CRC）是全球癌症相关死亡率的主要原因之一。虽然局部CRC的五年生存率高达90%，但超过一半的新发CRC病例在诊断时已经扩散到区域淋巴结和/或远处器官。结直肠癌伴远处转移的5年生存率很低，仅为13.5%。这些数据清楚地表明，如果疾病被早期发现，CRC相关的死亡率可以显着改善。CRC的风险因素包括年龄（>50岁）、种族（非裔美国人）、肥胖、吸烟、II型糖尿病、炎症性肠病史、结直肠息肉或CRC家族史以及已知会增加CRC风险的遗传性遗传综合征，如家族性腺瘤性息肉病（FAP）和Lynch综合征。FAP和Lynch综合征分别由APC基因和DNA错配修复（MMR）基因的种系突变引起。遗传性胃肠癌综合征患者的管理需要采取超出平均风险人群建议的额外措施，以最大限度地降低癌症相关发病率和死亡率的总体风险。 梭杆菌是常见的人类口腔革兰氏阴性厌氧微生物菌群，分离自牙菌斑和牙龈疾病，但很少在健康个体的结直肠中检测到。近年来，在结直肠腺瘤和结直肠癌患者的结肠组织和粪便中发现了一种特殊的致病型，具核梭杆菌（Fn）。临床证据表明，Fn的患病率从异型增生、腺瘤到CRC逐渐增加，并且Fn的较高量与具有高微卫星不稳定性（MSI-高）的CRC显著相关，这是由DNA MMR缺陷引起的，如在Lynch综合征相关的CRC中所见。Fn与CRC肿瘤发生的潜在关联已经在FAP小鼠模型中进行了检查。ApcMin/+小鼠携带Apc基因密码子850处的无义突变（Apc ε 850），口服Fn后，小肠和结肠直肠肿瘤的发病显著加速，并且多样性增加。这些数据有力地表明Fn在ApcMin/+小鼠中的肿瘤促进作用。 虽然Fn如何促进CRC肿瘤发生尚未完全阐明，但新出现的证据表明其两个毒力因子FadA和Fap 2是CRC肿瘤发生和进展的潜在增强剂。FadA粘附素表达于细菌表面，介导Fn附着和内皮细胞和上皮细胞的侵袭。最近的一项研究表明，Fn FadA与E-钙粘蛋白结合并激活Wnt/β-连环蛋白信号传导，导致人结肠癌细胞中细胞增殖增加。通过合成抑制肽破坏FadA和E-钙粘蛋白结合显著降低了Fn驱动的CRC细胞体外增殖和体内肿瘤生长。在临床环境中，已经在从人类患者获得的结肠腺瘤和CRC组织中证明了FadA表达水平的增加。Fn Fap 2蛋白是另一种毒力因子，被认为在人类Fn相关的CRC肿瘤发生中发挥作用。Fap 2结合人T细胞免疫球蛋白ITIM结构域（TIGIT），其是已知作为免疫共抑制分子起作用的免疫球蛋白超家族受体。其配体CD 155和CD 112在抗原呈递细胞、T细胞以及多种肿瘤细胞上表达。Fap 2结合NK和其它T细胞上的TIGIT保护肿瘤细胞免受NK细胞介导的细胞毒性并阻断肿瘤微环境中的效应T细胞功能。TIGIT还显示促进Treg功能。综上所述，Fn不仅可以通过激活Wnt/β-连环蛋白信号通路促进CRC肿瘤生长，而且可以通过利用TIGIT的免疫抑制功能（通过Fap 2蛋白）并有助于肿瘤免疫逃避机制来发挥肿瘤促进作用。 人乳头瘤病毒（HPV）和B肝炎病毒（HBV）的预防性疫苗与感染率的显著降低有关，预计这将分别导致宫颈癌和肝细胞癌发病率的大幅下降。与预防感染相比，从宿主中根除致癌感染因子通常具有高度挑战性，如果不是不可能的话。然而，如果病原体靶向疫苗能够增强宿主的抗病原体免疫，从而抑制病原体介导的致瘤功能或抑制感染，则可以增强宿主的免疫防御系统。 与其他革兰氏阴性细菌一样，Fn产生外膜囊泡（OMV），其中包含Fn的大部分生物内容物，但没有复制能力。OMV是天然分泌的、尺寸为20-250 nm的双层脂质膜纳米结构，并且是高度稳定和免疫原性的。OMV也可以被工程化以表达特异性抗原，同时进行遗传改变以降低内毒性。如已成功开发高效的基于OMV的抗脑膜炎球菌血清群B疫苗（即Trumenba®和Bexsero®）所证明的，Fn OMV可用作抗Fn疫苗以引发将减少Fn定殖的免疫。目前的项目将尝试开发基于Fn OMV的疫苗（Fn疫苗），这将导致肠道和结直肠中Fn定植的减少，从而降低FAP和Lynch综合征个体以及其他高风险队列中CRC的风险。

Colorectal cancer (CRC) is one of the leading causes of cancer-related mortality worldwide. While the five-year survival rate for localized CRC is excellent at 90%, more than half of new CRC cases have the disease spread to regional lymph nodes and/or distant organs at the time of diagnosis. CRC with distant metastasis has a dismal five-year survival rate of 13.5%. These data clearly indicate that CRC-related mortality can be significantly improved if the disease is detected early. Risk factors for CRC include age (>50), race (African American), obesity, cigarette smoking, type II diabetes, history of inflammatory bowel diseases, family history of colorectal polyps or CRC, and inherited genetic syndromes known to increase the CRC risks, such as familial adenomatous polyposis (FAP) and Lynch syndrome. FAP and Lynch syndrome are caused by germline mutations in the APC gene and DNA mismatch repair (MMR) genes, respectively. The management of individuals with hereditary gastrointestinal cancer syndromes requires additional measures beyond what is recommended for the average-risk population to minimize the overall risk of cancer-associated morbidity and mortality. Fusobacteria are common human oral gram-negative anaerobic microflora isolated from dental plaque and gum diseases, but rarely detectable in the colorectum of healthy individuals. In recent years, the enrichment of a specific pathotype, Fusobacterium nucleatum (Fn), has been demonstrated in the colonic tissues and stools from patients with colorectal adenomas and CRC. Clinical evidence suggests that the prevalence of Fn progressively increases from dysplasia, adenomas to CRC, and the higher amount of Fn is significantly associated with CRC with high microsatellite instability (MSI-high), which is caused by DNA MMR deficiencies as seen in Lynch syndrome-associated CRC. The potential association of Fn with CRC tumorigenesis has been examined in a mouse model of FAP. ApcMin/+ mice, which carry a nonsense mutation at codon 850 of the Apc gene (Apc∆850), had a significantly accelerated onset and increased multiplicity of both small intestinal and colorectal tumors after oral inoculation of Fn. These data strongly suggested the tumor-promoting role of Fn in ApcMin/+ mice. Although how Fn contributes to CRC tumorigenesis has yet to be fully elucidated, emerging evidence points to its two virulence factors, FadA and Fap2, as potential enhancers of CRC tumorigenesis and progression. FadA adhesin, which is expressed on the bacterial surface, mediates Fn attachment and invasion of endothelial and epithelial cells. A recent study has shown that the Fn FadA binds to E-cadherin and activates Wnt/-catenin signaling, resulting in increased cell proliferation in human colon cancer cells. Disruption of FadA and E-cadherin binding by synthetic inhibitory peptides significantly reduced Fn-driven CRC cell proliferation in vitro and tumor growth in vivo. In the clinical setting, the increased level of FadA expression has been demonstrated in colonic adenoma and CRC tissues obtained from human patients. The Fn Fap2 protein is another virulence factor suggested to play a role in Fn-associated CRC tumorigenesis in humans. The Fap2 binds to human T-cell immunoglobulin ITIM domain (TIGIT), an immunoglobulin superfamily receptor known to function as an immune co-inhibitory molecule. Its ligands, CD155 and CD112, are expressed on antigen presenting cells, T cells as well as on a variety of tumor cells. Fap2-binding to TIGIT on NK and other T cells protects tumor cells from NK cell-mediated cytotoxicity and blocks effector T cell functions in the tumor microenvironment. TIGIT has also been shown to promote Treg function. Taken together, Fn may exert tumor promoting effects not only by promoting CRC tumor growth via activation of Wnt/-catenin signaling pathway, but also by exploiting the immune-suppressive function of TIGIT via the Fap2 protein and contributing to tumor immune evasion mechanisms. Prophylactic vaccines for human papillomavirus (HPV) and hepatitis B virus (HBV) have been associated with significant reductions in infection rates, which are expected to result in the dramatic decrease in the incidence of cervical cancers and hepatocellular carcinoma, respectively. Compared to the prevention of infection, the eradication of oncogenic infectious agents from the host is in general highly challenging, if not impossible. Nontheless, the host immune defense system may be fortified by pathogen-targeted vaccines, if they can boost anti-pathogen immunity and lead to the containment of infection or suppression of pathogen-mediated tumorigenic functions. As with other gram-negative bacteria, Fn produces outer membrane vesicles (OMVs), which contain much of the biological content of the Fn, but without replicative capacity. OMVs are naturally secreted, bi-layered lipid membrane nanostructures of 20-250 nm in size, and are highly stable and immunogenic. OMVs can also be engineered to express specific antigens, while genetically altered to reduce the endotoxicity. As has been demonstrated with the successful development of highly efficacious OMV based anti-meningococcal serogroup B vaccines (i.e. Trumenba® and Bexsero®), Fn OMV may be used as anti-Fn vaccines to elicit immunity that would reduce Fn colonization. The current project will attempt to develop Fn OMV based vaccines (Fn vaccines), which will result in the reduction of Fn colonization in the intestine and colorectum and in turn reduction of the CRC risk in individuals with FAP and Lynch syndrome as well as in other high-risk cohorts.