Immunotherapy for Malignant Mesothelioma and Lung Cancer

我们项目的总体目标是为胸部癌症患者开发更有效的治疗方法。这项工作主要分为两大类：1。利用间皮素治疗间皮瘤及相关转化研究。我们目前的研究重点是针对肿瘤分化抗原间皮素（mesothelin）的免疫治疗，该抗原在胸膜、心包和腹膜的正常间皮细胞上表达，但在几种人类肿瘤中，特别是间皮瘤、卵巢癌、肺癌和胰腺腺癌中高度表达。这种间皮素的差异表达使其成为肿瘤特异性治疗的有吸引力的候选者。我们现在的工作重点是利用不同的方法将其用于间皮瘤的治疗。其中包括抗间皮素免疫毒素（LMB-100）、抗间皮素药物偶联物（BAY 94-9343）、间皮素疫苗（CRS-207）和过继t细胞疗法（TC-210）。LMB-100是一种由抗间皮素Fv组成的免疫毒素，它与强效细菌毒素假单胞菌外毒素a的截断形式相连，假单胞菌外毒素a已被去免疫以降低其抗原性。我们最近完成了LMB-100的I期试验，并确定了其安全性和最大耐受剂量（MTD）(1)。目前，我们正在进行一项与免疫检查点抑制剂派姆单抗联合的II期研究。我们最近完成了一项I期临床试验，以确定抗间皮素抗体药物偶联物BAY 94-9343的安全性和MTD，该偶联物由人源抗间皮素单克隆抗体连接到美坦素类DM4(2)。我们还进行了一项I期临床试验，将表达活的减毒单核细胞增生李斯特菌（CRS207）的间皮素与化疗结合使用，发现恶性胸膜间皮瘤患者的总生存率有所提高(3)。我们目前正在开发靶向间皮素的过继细胞疗法，使用嵌合抗原受体（CAR） T细胞。临床开发中的大多数抗间皮素抗体靶向间皮素的膜远端区域，这可能部分解释了临床中抗间皮素CAR - T细胞治疗缺乏活性的原因。为了提高CAR - T细胞的抗肿瘤活性，我们正在开发靶向靠近肿瘤细胞表面的表位的CAR - T细胞。我的合作者Mitchell Ho博士已经鉴定出一种高亲和力的兔单克隆抗体（YP218），特异于III区，该区位于间皮素的c末端，靠近肿瘤细胞表面。我们正在动物模型中测试hYP218 CAR-T构建体对几种表达间皮素的细胞系的细胞毒性和抗肿瘤作用。同时，我们正在进行T细胞受体融合构建（TRuCs）的临床试验。与其他结构体不同，TRuCs自然地结合到天然TCR复合体中，从而充分发挥TCR驱动的T细胞激活、效应功能和调控的潜力。我们之前的研究表明，DNA修复基因的种系突变增加了胸膜间皮瘤患者对铂治疗的敏感性，并提高了患者的总生存率(4)。目前，我的实验室正在研究DNA修复基因的突变，这些突变可能易患间皮瘤并影响临床结果。我们正在招募携带这种突变的患者及其亲属进行长期随访研究，以进行早期癌症检测和预防。在实验室，我们专注于开发人类间皮瘤的体外和体内模型。我们从患者的腹水和胸水中建立了几个早期传代肿瘤细胞系。我们已经评估了这些细胞系的形态和遗传特征，并正在使用它们来研究体外药物敏感性。我们评估了这些患者来源的间皮瘤细胞系对PARP抑制剂（olaparib和talazoparib）和DNA烷基化剂替莫唑胺的遗传背景的敏感性，并表明对PARPI的敏感性与BAP1状态无关，然而，高schlafen 11和低o6 -甲基鸟嘌呤-DNA甲基转移酶表达的间皮瘤细胞系对替莫唑胺敏感(5)。此外，我们已经建立了一个人源性间皮瘤异种移植肿瘤模型，该模型使用来自患者的肿瘤细胞和来自健康供体的人pbmc进行体内研究。由于pmc人源化小鼠模型中移植物抗宿主病（GVHD）的发展限制了抗肿瘤疗效持续时间的评估，我们通过用编码hMSLN膜结合片段的hMSLN表达载体转染PD-L1阳性小鼠肺腺癌细胞系，建立了表达人间皮素的同基因免疫小鼠模型。这些细胞系被用来发展肿瘤。我们研究了LMB-100联合抗pd1抗体在两种模型中的作用，并观察到肿瘤消退。我们的研究结果发表在《科学转化医学》（Science Translational Medicine）杂志上。这些模型对于评估间皮瘤的新型治疗药物和抗肿瘤疗效的机制研究至关重要。其他正在进行的实验室研究主要集中在了解间皮瘤肿瘤免疫微环境和抗间皮瘤靶向药物治疗后的变化。2. 免疫疗法治疗肺癌。目前，我们正在对先前治疗失败的肺腺癌患者进行抗pd - l1单克隆抗体MSB0010718C的临床试验。我们的实验室最近发现约25%的转移性肺腺癌患者高表达间皮素。这些肿瘤中的间皮素表达与KRAS突变和野生型EGFR状态高度相关，并且独立地与不良预后相关。我们的假设是，K-RAS突变肺癌患者可以从间皮素定向治疗中获益。间皮素导向疗法治疗肺癌的临床试验即将开启。我们的实验室也在研究免疫检查点在恶性间皮瘤中的作用，以便针对这一途径的药物可以用于治疗间皮瘤。我们目前正在进行LMB-100联合派姆单抗治疗的NSCLC患者的II期试验。引用:1。刘建军，刘建军，李建军，等。免疫毒素LMB-100在间皮瘤和其他表达间皮素的实体瘤患者中的I期研究。巨蟹座2020（即将出版）Hassan R, Blumenschein GR, Moore KN, Santin AD, Kindler HL, Nemunaitis JJ，等。抗间皮素抗体-药物偶联Anetumab Ravtansine治疗晚期或转移性实体瘤的首次人体多中心I期剂量递增和扩展研究临床肿瘤杂志2020:JCO.19.02085Hassan R, Alley E, Kindler H, Antonia S, Jahan T, Honarmand S，等。表达间皮素的减毒单核增生李斯特菌（CRS-207）在恶性胸膜间皮瘤化疗中的临床疗效临床癌症研究2019；25:5787 - 98 4。张建军，张建军，李建军，等。恶性间皮瘤的遗传易感性和铂类化疗后的总生存率。中国科学院学报（自然科学版）；116:9008-13 5。Rathkey D, Khanal M, Murai J，张军，Sengupta M，姜强，等。间皮瘤细胞对PARP抑制剂的敏感性不依赖于BAP1，但在高schlafen 11和低o6 -甲基鸟嘌呤- dna甲基转移酶表达的细胞中，替莫唑胺增强了PARP抑制剂的敏感性。中国生物医学工程学报，2020；15:843-59 6。姜青，李建平，李建平，李建平，等。间皮瘤靶向免疫毒素LMB-100和抗pd -1抗体对间皮瘤患者和小鼠肿瘤模型的疗效增强。Science Translational Medicine 2020；12: eaaz7252

The overall goal of our program is to develop more effective therapies for patients with thoracic cancers. This work falls under two main categories: 1. Exploiting mesothelin for mesothelioma therapy and related translational research. Our current studies are focused on using immunotherapy directed against the tumor differentiation antigen mesothelin, which is expressed on normal mesothelial cells lining the pleura, pericardium and peritoneum, but is highly expressed in several human tumors especially mesothelioma, ovarian cancer, lung cancer and pancreatic adenocarcinomas. This differential expression of mesothelin makes it an attractive candidate for tumor specific therapy. Our efforts are now focused on exploiting it for mesothelioma therapy using different approaches. These include anti mesothelin immunotoxin (LMB-100), an anti-mesothelin drug conjugate (BAY 94-9343), mesothelin vaccine (CRS-207) and adoptive T-cell therapy (TC-210). LMB-100 is an immunotoxin consisting of the anti-mesothelin Fv linked to a truncated form of the potent bacterial toxin, Pseudomonas exotoxin A, which has been de-immunized to decrease its antigenicity. We have recently completed the phase I trial of LMB-100 and established its safety and maximum tolerated dose (MTD) (1). Currently, we are conducting a phase II study in combination with immune checkpoint inhibitor pembrolizumab. We have recently completed a phase I clinical trial to determine the safety and MTD of the anti-mesothelin antibody drug conjugate BAY 94-9343, consisting of a humanized anti-mesothelin monoclonal antibody linked to the maytansinoid DM4 (2). We have also conducted a Phase I clinical trial with mesothelin expressing live, attenuated Listeria monocytogenes (CRS207) along with chemotherapy, and have seen improved overall survival in patients with malignant pleural mesothelioma (3). We are currently developing mesothelin-targeting adoptive cellular therapy, using chimeric antigen receptor (CAR) T cells. Majority of the anti-mesothelin antibodies in clinical development target the membrane distal region of mesothelin, that could partly account for the lack of activity of anti-mesothelin CAR T cell therapy in the clinic. To improve CAR T cell anti-tumor activity, we are developing CAR T cells that target an epitope close to the surface of tumor cells. My collaborator, Dr. Mitchell Ho, has identified a high affinity rabbit monoclonal antibody (YP218) specific for region III, which is located at the C-terminal end of mesothelin, close to the tumor cell surface. We are testing the cytotoxicity of the hYP218 CAR-T construct on several mesothelin expressing cell lines and anti-tumor effect in animal models. Parallelly, we are conducting a clinical trial of a T cell receptor fusion construct (TRuCs). Unlike other constructs, TRuCs are naturally incorporated into the native TCR complex, thus exploiting the full potential of TCR-driven T cell activation, effector function, and regulation. We have previously shown that germline mutations in DNA repair genes increases sensitivity to platinum therapy and improves overall survival in patients with pleural mesothelioma (4). Currently, my laboratory is studying mutations in DNA repair genes that could predispose to mesothelioma and influence clinical outcome. We are enrolling patients and their relatives harboring such mutations for a long term follow up study, for early cancer detection and prevention. In the laboratory, we have focused on developing in-vitro and in-vivo models of human mesothelioma. We have established several early passage tumor cell lines from ascites and pleural fluid of patients. We have evaluated the morphological and genetic characteristics of these cell lines and are using them to study in-vitro drug sensitivity. We have evaluated the sensitivity of these patient derived mesothelioma cell lines to PARP inhibitors (olaparib and talazoparib) and a DNA alkylating agent, temozolomide with respect to their genetic background and have shown that sensitivity to PARPI is independent of BAP1 status, however, mesothelioma cell lines with high-Schlafen 11 and low-O6-methylguanine-DNA methyltransferase expression are sensitive to temozolomide (5). Additionally, we have established a humanized mesothelioma xenograft tumor model with patient derived tumor cells and human PBMCs from healthy donor for in-vivo studies. As the development of Graft Versus Host Disease (GVHD) in the PBMC-humanized mouse model limits assessment of duration of anti-tumor efficacy, we have developed a human mesothelin expressing syngeneic immunocompetent mouse model by transfecting PD-L1 positive mouse lung adenocarcinoma cell line with a hMSLN expressing vector encoding the membrane bound fragment of hMSLN. These cell lines were used to develop tumor. We have studied the effect of LMB-100 in combination with anti-PD1 antibody in both the models and have seen tumor regression. We have published our findings in Science Translational Medicine (6). These models are essential to evaluate novel therapeutic agents for mesothelioma and for the mechanistic studies of anti-tumor efficacy. Other ongoing laboratory studies are focused on understanding the mesothelioma tumor immune micro-environment and changes following treatment with anti-mesothelin targeted agents. 2. Immunotherapy to treat lung cancers. We are currently conducting clinical trial of the anti-PD-L1 monoclonal antibody MSB0010718C in patients with lung adenocarcinoma who have failed prior therapies. Our laboratory has recently shown that about 25% of patients with metastatic lung adenocarcinoma highly express mesothelin. Mesothelin expression in these tumors is highly associated with KRAS mutations and wild type EGFR status and is, independently, associated with poor prognosis. Our hypothesis is that patients with K-RAS mutant lung cancer can benefit from mesothelin directed therapies. Clinical trials of mesothelin directed therapies for treating lung cancer are about to open. Our laboratory is also studying the role of immune checkpoints in malignant mesothelioma so that drugs targeting this pathway can be exploited for treating mesothelioma. We are currently conducting a Phase II trial of NSCLC patients treated with LMB-100 in combination with pembrolizumab. References: 1. Hassan R, Alewine C, Mian I, Spreafico A, Siu LL, Gomez-Roca C, et al. Phase I Study of the Immunotoxin LMB-100 in Patients with Mesothelioma and Other Solid Tumors Expressing Mesothelin. Cancer 2020 (in press) 2. Hassan R, Blumenschein GR, Moore KN, Santin AD, Kindler HL, Nemunaitis JJ, et al. First-in-Human, Multicenter, Phase I Dose-Escalation and Expansion Study of Anti-Mesothelin Antibody-Drug Conjugate Anetumab Ravtansine in Advanced or Metastatic Solid Tumors. Journal of Clinical Oncology 2020:JCO.19.02085 3. Hassan R, Alley E, Kindler H, Antonia S, Jahan T, Honarmand S, et al. Clinical Response of Live-Attenuated, Listeria monocytogenes Expressing Mesothelin (CRS-207) with Chemotherapy in Patients with Malignant Pleural Mesothelioma. Clin Cancer Res 2019;25:5787-98 4. Hassan R, Morrow B, Thomas A, Walsh T, Lee MK, Gulsuner S, et al. Inherited predisposition to malignant mesothelioma and overall survival following platinum chemotherapy. Proc Natl Acad Sci U S A 2019;116:9008-13 5. Rathkey D, Khanal M, Murai J, Zhang J, Sengupta M, Jiang Q, et al. Sensitivity of Mesothelioma Cells to PARP Inhibitors Is Not Dependent on BAP1 but Is Enhanced by Temozolomide in Cells With High-Schlafen 11 and Low-O6-methylguanine-DNA Methyltransferase Expression. J Thorac Oncol 2020;15:843-59 6. Jiang Q, Ghafoor A, Mian I, Rathkey D, Thomas A, Alewine C, et al. Enhanced efficacy of mesothelin-targeted immunotoxin LMB-100 and anti-PD-1 antibody in patients with mesothelioma and mouse tumor models. Science Translational Medicine 2020;12:eaaz7252