Myeloma stem cell and cancer testis antigens

描述(申请人提供)：多发性骨髓瘤(MM)是一种常见的血液系统癌症，标准治疗很难治愈。尽管接受免疫调节剂和/或化疗的骨髓瘤患者可能会经历完全的临床反应，但大多数人最终会复发。包括异基因干细胞移植在内的更积极的治疗可能是治愈的，但清髓性调节的高发病率和死亡率使许多患者无法使用。该项目的目标是开发一种以免疫疗法为靶向的骨髓瘤干细胞的临床前模型，从长远来看，该模型可以发展为一种新的临床治疗策略，将改善骨髓瘤患者的预后。我们和其他人之前已经描述了一组骨髓瘤干细胞，具有克隆形成能力，对化疗药物高度耐药，并可以启动肿瘤形成。干细胞用来限制其在化疗中暴露的机制之一是表达能够泵出药物的运输蛋白。干细胞的这一特性可用于其纯化，因为在流式细胞仪分析中，相同的转运体排出Hoechst染料形成不同的侧群。这些细胞可能在疾病复发中发挥关键作用。我们现在建议针对这些骨髓瘤干细胞进行针对癌症睾丸抗原(CTA)的免疫治疗。CTA在多发性骨髓瘤中异常表达，由于其免疫原性和在正常组织中的限制性表达，使其不会形成T细胞耐受，使CTA成为骨髓瘤的潜在抗原靶点。此外，最近研究发现，一些CTA包括NRAGE、NY-ESO、MAGE-1和SSX在人骨髓间充质干细胞(MSCs)中表达。这表明CTA的表达可能不仅是配子发生的标志，也可能是干细胞的标志。基于这些观察，我们假设骨髓瘤干细胞优先表达CTA，因此容易被CTA特异性T细胞杀伤。此外，CTA转录部分受甲基化调控，因此去甲基化药物可能用于增强低表达肿瘤中CTA的表达，增加这些肿瘤细胞对CTA特异性T细胞杀伤的敏感性。我们的假设得到了我们的初步数据的支持，即来自人骨髓瘤细胞系RPMI8226的骨髓瘤干细胞比成熟的骨髓瘤细胞有更高的CTA表达，包括PRAME、CT7和NY-ESO1(C.1节)。此外，我们已经产生了PRAME特异性细胞毒性T细胞(CTL)，可以杀死PRAME阳性的骨髓瘤细胞。在目标1中，我们将分析一大组骨髓瘤肿瘤样本，以确定CTA在原代骨髓瘤干细胞中的表达是否保留和/或丰富。在目标2中，我们将使用小鼠骨髓瘤模型来确定针对CTA的CTL是否可以直接杀死骨髓瘤干细胞。 公共卫生相关性：该项目的长期目标是通过使用细胞毒性T细胞靶向在肿瘤发生和疾病复发中发挥关键作用的癌症干细胞来改善多发性骨髓瘤(MM)的预后，MM是一种目前无法治愈的常见浆细胞起源的血液系统癌症。这项建议的主要目标是为使用细胞毒性T细胞靶向骨髓瘤干细胞产生临床前数据。建立的基本模型和从这项建议中获得的知识将代表着未来设计新的骨髓瘤长期治愈方法的关键。

DESCRIPTION (provided by applicant): Multiple myeloma (MM) is a common hematological cancer that with standard treatment is rarely curable. Although myeloma patients treated with immunomodulatory agents and or chemotherapy may experience a complete clinical response, the majority will eventually relapse. More aggressive treatments including allogeneic stem cell transplantation may be curative, but the high morbidity and mortality of myeloablative conditioning precludes use in many patients. The objective of this project is to develop a preclinical model of targeting myeloma stem cells with immunotherapy that in the long-term can be developed into a novel clinical therapeutic strategy that will improve outcomes for patients with myeloma. We and others have previously described a population of myeloma stem cells that possess clonogenic potential, are highly resistant to chemotheraputics agents, and can initiate tumorigenesis. One of the mechanisms that stem cells use to limit their exposure to chemotherapy is to express transport proteins that can pump out drug. This property of stem cells can be exploited for their purification, since the same transporters efflux Hoechst dye forming a distinct side population on flow cytometric analysis. These cells are likely to play a key role in disease relapse. We now propose to target these myeloma stem cells with immunotherapy directed against cancer testis antigens (CTAs). CTAs are expressed aberrantly in multiple myeloma, and by virtue of their immunogenecity and restricted expression in normal tissue such that T-cell tolerance does not develop, makes CTAs a promising antigen target in myeloma. Additionally, it was recently reported that some CTAs including NRAGE, NY-ESO, MAGE-1, and SSX are expressed in human mesenchymal stem cells (MSCs). This suggests that CTA expression may not only be a hallmark of gametogenesis but also a stem cell marker. Based on these observations, we hypothesize that myeloma stem cells preferentially express CTA and are therefore susceptible to killing by CTA specific T-cells. In addition, CTA transcription is partially regulated by methylation, and therefore demethylating agents maybe used to enhance CTA expression in low expressing tumors and increase susceptibility of these tumor cells to CTA-specific T-cell killings. Our hypothesis is supported by our preliminary data that myeloma stem cells from human myeloma cell line, RPMI 8226, have higher expression of several CTAs, including PRAME, CT7 and NY-ESO1, than the mature myeloma cells (Section C.1). Furthermore, we have already generated PRAME specific cytotoxic T- cells (CTLs) that can kill PRAME positive myeloma cells. In aim 1 we will analyze a large panel of myeloma tumor samples to determine if CTA expression is preserved and/or enriched in primary myeloma stem cells. In aim 2 we will determine if CTLs directed to CTA can directly kill myeloma stem cells using a murine myeloma model. PUBLIC HEALTH RELEVANCE: The long-term objective of this project is to improve the outcome of multiple myeloma (MM), a currently incurable common hematological cancer of plasma cell origin, by targeting the cancer stem cell which play a key role in tumorigenesis and disease relapse using cytotoxic T-cells. The primary goal of this proposal is to generate pre- clinical data for using cytotoxic T-cells to target myeloma stem cells. The fundamental models established and knowledge obtained from this proposal will represent the key for the future design of novel methods toward the long-term cure of myeloma.