Development of an autologous humanized model of melanoma exploring human thymic education capacity

摘要 研究人类癌症的一个制约因素是有限的模型可用适当的人类间质和 豁免权。考虑到免疫系统在黑色素瘤中扮演的关键角色，这一警告与黑色素瘤尤其相关。 它的发展。为了解决这些局限性，我们计划研究黑色素瘤的生物学和治疗方法。 小鼠(HM)模型。我们最初开发了一种不匹配的HM(MHM；肿瘤和造血干细胞 [来自不同来源的HSCs])，现在已经生成了自体HM(肿瘤、HSCs和 间充质干细胞[MSCs]来自同一患者)。在HM荷瘤中，人类免疫和 来自患者骨髓的间充质细胞归巢于生长中的肿瘤，迁移到前 存在小鼠基质，并与人类癌细胞相互作用。在HM中生长的肿瘤更紧密 与在非HM小鼠中生长的肿瘤相似，并且基因表达漂移由 之前的通行部分被颠倒了。关键的免疫和基质通路中的信号更为突出。 在HM和非HM模型中与原始患者非常相似。免疫细胞有效地聚集起来 肿瘤特异性免疫反应，由包括T细胞在内的人类免疫细胞介导。值得注意的是，免疫 HM黑色素瘤模型对免疫治疗的反应与患者的 治疗结果。两个突出的和研究不足的问题限制了HM的更广泛应用： 肿瘤和免疫匹配的程度，这可以影响免疫反应的忠实性，并且可以 只有通过比较MHM和AHM才能欣赏。对HM的第二个批评是对人类T的异种教育 小鼠胸腺细胞上的细胞，这是由于在这样的模型中缺乏功能正常的人胸腺所致。在这 应用程序，我们将进一步解决自体HM与不匹配HM的相关性，以测试 方法产生了更具代表性的黑色素瘤模型；第二，使用相同的前体细胞 从病人身上，我们将重建一个有功能的胸腺。这项提案的首要目标是进行一项 联合临床试验，我们将完成以下工作：1)前瞻性地生成和表征AHM和 来自20名黑色素瘤患者的MHM，2)接近产生更具代表性的黑色素瘤HM模型的测试， 3)在HM中测试免疫药物(细胞毒性T淋巴细胞相关蛋白4[CTLA4]和程序化细胞 死亡蛋白1[PD-1]抑制剂)，并与临床数据相关，4)识别 CTLA4/PD-1抑制剂的耐药机制；最后作为一项探索性目标 在某些情况下，我们将从同一患者的HSCs中产生胸腺上皮，这将导致HM完全 自体黑色素瘤，胸腺和免疫系统，从而使免疫细胞教育在严格的人类 背景。这个项目将促进我们对黑色素瘤和人类肿瘤-宿主相互作用的理解 癌症，通过更好地描述黑色素瘤细胞与免疫和间质系统之间的相互作用， 从而发现了改善个性化治疗和改善结果的新方法。

SUMMARY A constraint to studying human cancer is the limited availability of models with appropriate human stroma and immunity. This caveat is particularly relevant for melanoma, given the pivotal role the immune system plays in its development. To address these limitations, we plan to study melanoma biology and therapy in a humanized mouse (HM) model. We initially developed a mis-matched HM (mHM; tumor and hematopoietic stem cells [HSCs] from different sources), and now have generated an autologous HM (aHM; tumor, HSCs and mesenchymal stem cells [MSCs] from the same patient). In HM bearing tumors the human immune and mesenchymal cells from the patient's bone marrow homed into the growing tumor, migrated into the pre- existing mouse stroma, and interacted with the human cancer cells. Tumors grown in HM more closely resembled the originator tumor than those grown in non-HM mice, and the drift in gene expression caused by prior passaging was partially reversed. Signaling in key immune and stroma pathways was more prominent and closely resembled the originator patient in HM vs. non-HM models. The immune cells mounted effective tumor-specific immune responses, mediated by human immune cells including T cells. Significantly, immune responses upon immune therapies in the HM melanoma models showed a correlation with the patient's therapy results. Two salient and under-studied issues limit the wider application of HM: The relevance of the degree of tumor and immune matching, which can affect the faithfulness of immune response, and that can only be appreciated by comparing mHM and aHM. A second criticism to HM is xenogenic education of human T cells on mouse thymic cells, resulting from the lack of a functional human thymus in such models. In this application we will further address the relevance of an autologous versus mis-matched HM to test which approach gives rise to a more representative model of melanoma; secondly, using the same precursor cells from the patient we will reconstitute a functional thymus. The overarching goal of this proposal is to conduct a co-clinical trial where we will accomplish the following: 1) prospectively generate and characterize aHM and mHM from 20 melanoma patients, 2) test which approach yields a more representative melanoma HM model, 3) test in HM the immune drugs (cytotoxic T-lymphocyte-associated protein 4 [CTLA4] and programmed cell death protein 1 [PD-1] inhibitors) that each patient received and correlate with the clinical data, 4) identify the mechanisms involved in resistance to CTLA4/PD-1 inhibitors; and finally as an exploratory Aim in selected cases, we will 5) generate thymic epithelium from the same patient's HSCs that will result in HM with a fully autologous melanoma, thymus and immune system, thus enabling immune cell education in a strictly human context. This project will advance our understanding of the tumor-host interaction in melanoma and human cancer, by better characterizing the interplay between melanoma cells and the immune and stroma systems, leading to the discovery of new approaches to improve personalization of therapy and improve outcomes.