Targeting EMP2 for the treatment of triple negative breast cancer with novel anti-EMP2 Granzyme B immunoconjugates

项目概要/摘要 免疫缀合物被设计成使用免疫缀合物将高细胞毒性剂集中递送至特定靶细胞。 单克隆抗体，潜在地提高该试剂的治疗指数。为此，我们建议 递送具有抗EMP 2 IgG 1抗体的细胞毒性蛋白质有效载荷。EMP 2或上皮膜蛋白-2， 是一种在大多数（63%; n=97）检查的浸润性乳腺癌肿瘤中高度表达的生物标志物 与健康乳腺上皮相比，在超过75%的三阴性乳腺癌中观察到EMP 2高表达。 在原发性肿瘤和转移性肿瘤中均观察到EMP 2的乳腺癌病例 病变由于工程重组抗体在癌症诊断和治疗方面具有巨大的前景，我们已经 我精心组建了一个以加州大学洛杉矶分校和德克萨斯大学医学博士安德森癌症中心为中心的研究小组 中心开发与人丝氨酸蛋白酶颗粒酶B融合的重组抗EMP 2抗体， 疗法加州大学洛杉矶分校的研究小组先前已经表明，抗EMP 2的全长IgG 1促进细胞凋亡 在许多EMP 2阳性肿瘤的体外和体内，但新的数据表明，抗EMP 2 IgG 1 可以迅速内化，我们假设它可能作为一种新的候选药物缀合。使用 颗粒酶B（GrB）作为免疫缀合物已经被提出作为“魔术子弹”，因为一旦递送至免疫缀合物， 在细胞的细胞质中，它激活细胞凋亡途径，几乎不诱导免疫原性。穆罕默达利 一个实验室已经利用人GrB作为产生重组细胞死亡诱导融合的有效载荷 蛋白质，并已清楚地表明，含有GrB的融合构建体具有令人印象深刻的和高度的 当通过抗体或生长因子细胞靶向递送至细胞质时的选择性细胞毒性作用 载波 在这项提案中，明确的目标是推进抗EMP 2抗体的科学以及GrB作为一种免疫抑制剂。 为临床试验的细胞毒性有效载荷，我们将创建和确定两个抗EMP 2 IgG 1- GrB的功效 共轭因此，具体目标提出设计和测试与抗EMP 2相关的GrB的功效 IgG 1的细胞毒性，并开始创建一个临床前包，以了解其功能亲和力， 稳定由于裸抗体在人和小鼠之间交叉反应，因此， 还将确定免疫缀合物，包括药代动力学和最大耐受剂量。给定 它在包括卵巢和子宫内膜在内的许多妇科肿瘤中的高表达，这些研究将是 重要的是将EMP 2定位为女性癌症的可行靶点。它也会，考虑到需要新的毒素 具有低免疫原性，有助于将颗粒酶B置于抗体连接毒素的最前沿。

PROJECT SUMMARY/ABSTRACT Immunoconjugates are designed to focus the delivery of highly cytotoxic agents to specific target cells using monoclonal antibodies, potentially improving the therapeutic index of the agent. To this end, we propose to deliver a cytotoxic protein payload with an anti-EMP2 IgG1 antibody. EMP2, or epithelial membrane protein-2, is a biomarker that is highly expressed in the majority (63%; n=97) of invasive breast cancer tumors examined compared to healthy mammary epithelium, and high EMP2 expression is observed in over 75% of triple negative breast cancer cases examined where EMP2 is observed in both the primary tumor as well as in metastatic lesions. As engineered recombinant antibodies hold great promise for cancer diagnostics and therapy, we have carefully assembled a research team centered at UCLA and The University of Texas MD Anderson Cancer Center to develop recombinant anti-EMP2 antibodies fused to the human serine protease granzyme B for therapy. The group at UCLA has previously shown that a full length IgG1 against EMP2 promotes apoptosis both in vitro and in vivo in a number of EMP2 positive tumors, but as new data suggests that the anti-EMP2 IgG1 can internalize rapidly, we hypothesize that it may serve as a novel candidate for drug conjugation. The use of Granzyme B (GrB) as an immunoconjugate has been proposed as a “magic bullet” as, once delivered to the cytoplasm of a cell, it activates apoptosis pathways with little to no induction of immunogenicity. The Mohamedali lab has utilized human GrB as an effective payload for the generation of recombinant cell death-inducing fusion proteins and has clearly demonstrated that GrB-containing fusion constructs have impressive and highly selective cytotoxic effects when delivered to the cytoplasm by either antibody or growth factor cell targeting carriers. In this proposal, with the explicit goal of advancing the science of anti-EMP2 antibodies as well as GrB as a cytotoxic payload for clinical testing, we will create and determine the efficacy of two anti-EMP2 IgG1- GrB conjugates. Accordingly, the specific aims propose to design and test the efficacy of GrB linked to an anti-EMP2 IgG1 for cytotoxicity as well as start creating a preclinical package to understand its functional affinity and stability. As the naked antibody cross-reacts between human and mouse, the toxicity profile of the immunoconjugate, including pharmacokinetics and maximum tolerated dosage, will also be determined. Given its high expression in a number of gynecological tumors including ovarian and endometrial, these studies will be important to position EMP2 as a viable target for cancers in women. It will also, given the need for new toxins with low immunogenicity, help position granzyme B in the forefront of antibody linked toxins.