Antibody Therapy of Cancer

分子生物学实验室的抗体治疗单元于2008年5月25日启动。我的实验室进行以机械为基础的转化性研究，从而开发出新的、更有效的抗癌抗体疗法。我们目前的工作重点是开发针对卵巢癌和间皮瘤的新单抗。这项工作目前部分由美国国立卫生研究院、国家癌症研究所、癌症研究中心的内部研究计划支持，部分由卵巢癌研究基金个人研究员奖(M.Ho)支持，部分由间皮瘤应用研究基金会授予Craig Kozicki荣誉(M.Ho)资助。卵巢癌和间皮瘤的生物学-癌细胞通常通过种植到间皮衬里的结构在腹膜内扩散。CA125是一种存在于大多数卵巢癌和间皮瘤细胞上的粘蛋白，而Mesothelin是一种存在于腹膜间皮细胞上的GPI锚定糖蛋白，它们之间的相互作用被认为促进了肿瘤的种植和转移。我们假设，阻断间充质蛋白-CA125的相互作用可以预防或逆转转移，并导致癌症患者总体生存率的提高。更好地了解这两种蛋白质是如何相互作用的，最终可能有助于开发这样的治疗方法。为此，我的实验室现在正在积极研究间皮蛋白和CA125分子相互作用的生物学机制。(1)我们成功地确定了CA125与间皮蛋白的结合部位。我们已经在间皮蛋白的CA125结合部位确定了参与与CA125相互作用的特定氨基酸。(2)研制了含CA125结合位点的抗体样Fc融合分子。本实验室的研究表明，Fc融合蛋白可以有效地阻断间皮蛋白与CA125的结合，提示CA125结合域可以作为拮抗剂来抑制肿瘤的进展和在腹膜腔内的扩散。(3)我们正在鉴定CA125上尚未确定的间硫蛋白结合位点，它可能包含我们先前研究中提出的一种新的N-糖链结构。抗体疗法-我们正在进行转译研究，以开发用于癌症治疗的抗体疗法。(1)我们正在分离和鉴定几种针对间硫蛋白的鼠和人单抗。我的实验室最近的研究发现，单链抗体能有效地阻断Mesothelin-CA125蛋白-蛋白质的相互作用，其表位与CA125结合位点重叠。我们正在使用抗体工程技术开发用于腹膜癌治疗的人源或人源化抗体。(2)为了评价CA125结合域Fc融合蛋白治疗腹膜癌的疗效，本实验室正在开展小鼠卵巢癌和间皮瘤腹膜异种移植模型的临床前研究。我们目前正在制造重组结构域Fc融合蛋白，并开发用于动物肿瘤成像的癌细胞系。用于临床前研究的动物方案目前正在NIH接受动物护理和使用委员会(ACUC-Bethesda)的审查。(3)建立了一种新的抗体工程方法--哺乳动物细胞展示技术。功能性单链抗体(ScFv)可以在人细胞上表达，高亲和力的scFv结合子可以从组合Fv文库中快速鉴定和分离。利用这项新技术，我们目前正在提高治疗性抗体FV的结合亲和力，以便在癌症靶向治疗中获得更好的疗效。(4)利用我们新开发的哺乳动物细胞展示技术，在人HEK-293细胞中构建了新的人FV抗体库，用于抗体的发现。(5)我们在实验室建立了一种检测卵巢癌和间皮瘤对间皮蛋白的体液(抗体)反应的酶联免疫吸附试验。在恶性胸膜间皮瘤和转移性胸腔积液的I期临床试验中，我们发现胸膜间皮间皮瘤和转移性胸腔积液有较高的抗肿瘤免疫反应率。我们将继续研究针对癌症患者的抗肿瘤免疫(特别是抗体)反应，并研究针对癌症特异性抗原的免疫反应如何与改善临床结果相关。

The Antibody Therapy Unit in the Laboratory of Molecular Biology was started on May 25, 2008. My laboratory conducts mechanistically-based, translational research leading to the development of novel and more effective anti-cancer antibody therapies. Our efforts are currently focused on developing new mAbs directed at ovarian cancer and mesothelioma. This work is currently supported in part by the Intramural Research Program of the NIH, National Cancer Institute, Center for Cancer Research, in part by the Ovarian Cancer Research Fund Individual Investigator Award (M. Ho) and in part by the Mesothelioma Applied Research Foundation Grant in Honor of Craig Kozicki (M. Ho). Biology of Ovarian Cancer and Mesothelioma - Cancer cells commonly spread within the peritoneal cavity via seeding to mesothelium-lined structures. The interaction between CA125, a mucin present on a majority of ovarian cancer and mesothelioma cells, and mesothelin, a GPI-anchored glycoprotein present on the mesothelial cells lining along the peritoneal cavity, has been suggested to facilitate implantation and metastasis of tumors. We hypothesize that blocking the mesothelin-CA125 interaction may prevent or reverse metastasis and lead to overall improved survival in cancer patients. A better understanding of how these two proteins interact may eventually aid in developing such as therapy. To this end, my laboratory is now vigorously investigating the biological mechanisms underlying the molecular interaction of mesothelin and CA125. (1) We have successfully identified the binding site on mesothelin for CA125. We have identified specific amino acids within the CA125-binding site of mesothelin that are involved in the interaction with CA125. (2) We have developed the antibody-like Fc fusion molecules containing the CA125-binding site. Studies by my laboratory have shown that the Fc fusion proteins can effectively block the binding of mesothelin to CA125, suggesting the CA125-binding domain could be used as an antagonist to inhibit tumor progression and spread in peritoneal cavity. (3) We are characterizing the yet unidentified mesothelin-binding site on CA125 which likely contains a novel N-glycan structure as suggested by our previous studies. Antibody Therapy - We are conducting translational research leading to the development of antibody therapeutics for cancer treatment. (1) We are isolating and characterizing several mouse and human mAbs specific for mesothelin. The studies by my laboratory have recently found the single chain Fv mAb that can effectively blocks the mesothelin-CA125 protein-protein interaction and whose epitope overlaps the CA125-binding site. We are using antibody engineering technologies to develop human or humanized antibodies for peritoneal cancer treatment. (2) In order to evaluate the efficacy of the CA125-binding domain Fc fusion proteins for peritoneal cancer therapy, my laboratory is in the process of initiating preclinical studies developing peritoneal xenograft models of ovarian cancer and mesothelioma in mice. We are currently making recombinant domain Fc fusion proteins and developing the cancer cell lines used for tumor imaging in animal studies. The animal protocol for preclinical studies is currently undergoing Animal Care and Use Committee (ACUC-Bethesda) review in the NIH. (3) We have established a new antibody engineering method, called mammalian cell display. Functional single chain Fv antibodies (scFv) can be expressed on human cells and high affinity scFv binders can be rapidly identified and isolated from a combinatory Fv library. Utilizing this new technology, we are currently improving binding affinity of therapeutic antibody Fvs for better efficacy in cancer targeted therapy. (4) We have initiated the construction of a new human Fv antibody library in human HEK-293 cells utilizing our new-developed mammalian cell display technology for antibody discovery. (5) We have established an ELISA assay in my laboratory to examine humoral (antibody) response to mesothelin in ovarian cancer and mesothelioma. In a phase I clinical trial of single-dose intrapleural IFN-beta gene transfer for malignant pleural mesothelioma and metastatic pleural effusions, we found high rate of antitumor immune responses directed to mesothelin. We will continue investigating the anti-tumor immune (particularly antibody) response directed to mesothelin in cancer patients and studying how the immune response against cancer-specific antigens is relevant to improved clinical outcomes.