Improved Tumor Targeting of Salmonella VNP20009 via Ice-llama Antibody Guidance

描述（由申请人提供）：摘要虽然沙门氏菌VNP 20009（VNP）在动物模型中显示出肿瘤靶向的前景，但由于细菌优先在坏死核心中生长而不能到达坏死核心，因此肿瘤的再生长是显而易见的结果。 重要的外围。在转移性黑色素瘤患者的人体试验中，即使靶向本身也非常差，没有观察到肿瘤消退。我们的目标是通过赋予VNP抗体介导的肿瘤靶向的成功识别潜力来提高VNP的肿瘤靶向和肿瘤消退能力。我们之前曾尝试使用lpp-OmpA表面展示平台和难溶性scFv抗体来实现这一点，但后来发现该组合导致细菌生长停滞并限制了工程化VNP的治疗潜力。我们假设，通过冰核蛋白（INP）展示小的且高度可溶的抗CEA单结构域抗体的VNP将显示CEA阳性肿瘤的增强的靶向而不抑制细菌侵入或复制，因此将能够比亲本VNP更有效地使肿瘤消退。我们的具体目标是：1、工程化VNP以展示抗CEA sdAb而不阻碍复制，但能够在体外结合固定化CEA; 2、证明工程化VNP能够在MCF-7细胞中内化，并且还能够在三维球状细胞培养模型中进行同质靶向; 3.在乳腺癌的小鼠模型中证明工程化VNP的改进的体内肿瘤靶向，阐明肿瘤内分布和完全肿瘤消退的潜力。VNP的多功能能力，包括移动性和携带大有效载荷的能力， 通过可视化单个生物发光细胞的能力和通过抗生素容易消除细菌来平衡。因此，只要我们能够提高靶向和消退能力，VNP作为一种高度可控但用途广泛的癌症治疗剂的潜力是巨大的。

DESCRIPTION (provided by applicant): ABSTRACT While Salmonella VNP 20009 (VNP) has shown promise in tumor targeting in animal models it is becoming apparent that regrowth of tumors is the outcome since the bacteria preferentially grow in the necrotic core and fail to reach the vital periphery. In human trials of metastatic melanoma patients, even targeting itself was very poor and no tumor regression was observed. We aim to improve the tumor targeting and tumor regressive ability of VNP by conferring upon it the successful recognition potential of antibody mediated tumor targeting. We had previously attempted to do this using the lpp-OmpA surface display platform and poorly soluble scFv antibodies but have since found out that the combination caused bacterial growth arrest and restricted the therapeutic potential of engineered VNP. We hypothesize that VNP displaying small and highly soluble anti- CEA single domain antibodies via the ice nucleation protein (INP) will show enhanced targeting of CEA positive tumors without inhibiting bacterial invasion or replication and will therefore be able to regress tumors more effectively than parental VNP. Our specific aims are to; 1, Engineer VNP to display anti-CEA sdAb without impeding replication, yet capable of binding immobilized CEA in vitro; 2, Demonstrate engineered VNP is capable of being internalized in MCF-7 cells and is also capable of homogenous targeting in three- dimensional spheroidal cell culture models; 3, Demonstrate improved in vivo tumor targeting of engineered VNP in a mouse model of breast cancer, elucidating intra-tumoral distribution and potential for complete tumor regression. The multifunctional capabilities of VNP including mobility and ability to carry large payloads are well balanced by the ability to visualize single bioluminescent cells and facile elimination of the bacteria by antibiotics. As such, the potential for VNP as a highly controllable yet versatile cancer therapeutic is enormous so long as we can improve targeting and regressive capacity.