Tumor downstaging with small molecule therapeutics to enhance Uveal Melanoma metastasis eradication by B7-H3 CAR T cells

项目摘要 葡萄膜黑色素瘤（UM）是一种罕见的癌症，在美国的发病率为每百万人中有5例。近一半的UM患者 由于UM细胞扩散到肝脏的高趋势，发展成孤立的肝转移。目前还没有找到治愈 转移性UM患者（mUM）。Tebentafusp是FDA批准的唯一治疗mUM的药物。然而，它表明， 在总体缓解和无进展生存期方面略有改善，其适用性仅限于40% 的患者。缺乏有效的治疗选择mUM促使我们设计一种新的组合 基于使用特异性重定向针对的嵌合抗原受体（CAR）T细胞的免疫治疗策略 B7-H3由于后者在UM细胞上高度表达，因此被选为我们免疫策略的靶点 细胞系和mUM肿瘤组织样品中，但在正常组织中具有有限的表达。 一般的经验是，基于CAR T细胞的免疫疗法在根除实体瘤方面是无效的， 临床前和临床研究。这一结果可能是由多种机制引起的，其中主要作用是 在高肿瘤负荷的宿主中使用它们发挥作用;在肿瘤中产生不利的效应物与靶（E：T）比率 微环境对CAR T细胞的抗肿瘤活性具有负面影响，如通过以下方法产生的结果所示： 临床前和临床研究。文献中的信息和我们的初步结果提供了 设计一种策略的基本原理，该策略首先应用肿瘤减积方法来减少肿瘤负荷，然后应用CAR T细胞- 基于免疫疗法根除UM转移。我们将联合使用曲美替尼，一种丝裂原活化蛋白 激酶激酶抑制剂（MEKi）和帕比司他（一种组蛋白脱乙酰酶抑制剂（HDACi）），因为这种组合具有 已显示诱导小鼠中UM肝转移的显著消退。此外，这种组合不影响B7- UM细胞的H3表达和B7-H3 CAR T细胞的抗肿瘤活性。因此，在证明B7-H3 CAR T细胞可以识别并消除MEKi和HDACi联合治疗后存活的UM细胞，我们发现， 将研究由MEKi和HDACi组合诱导的肿瘤减积是否可以增强B7-H3的能力， CAR T细胞根除NSG小鼠中的UM肝转移。由于基于CAR T细胞的治疗的一个主要限制是 以其潜在的毒性和/或细胞因子释放综合征为代表，我们在我们的CAR构建体中掺入了 诱导型胱天蛋白酶9安全开关，其允许在意外毒性的情况下快速消除CAR T细胞。 如果我们的实验产生的结果是积极的，它们将对mUM的治疗产生重大影响。

Project summary Uveal Melanoma (UM) is a rare cancer with an incidence of 5 cases per million in the United States. Nearly half UM patients develop isolated liver metastases due to the high tendency of UM cells to spread to the liver. No cure has been found for patients with metastatic UM (mUM). Tebentafusp, is the only FDA-approved therapy for mUM. However, it has shown modest improvements in terms of overall response and progression-free survival and its applicability is limited to only 40% of mUM patients. The lack of effective treatment options for mUM has prompted us to design a novel combinatorial immunotherapeutic strategy based on the use of Chimeric Antigen Receptor (CAR) T cells specifically redirected against B7-H3. The latter has been selected as the target of our immunotherapeutic strategy, since it is highly expressed on UM cell lines and mUM tumor tissue samples, but has a limited expression on normal tissues. It is a general experience that CAR T cell-based immunotherapy is not effective in eradicating solid tumors both in preclinical and clinical investigations. This result is likely caused by multiple mechanisms, among which the major role is played by their use in hosts with high tumor load; the resulting unfavorable effector to target (E:T) ratio in the tumor microenvironment has a negative impact on the antitumor activity of CAR T cells, as indicated by the results generated by preclinical and clinical investigations. The information in the literature and our preliminary results have provided the rationale to design a strategy which applies first a tumor debulking approach to reduce tumor load and then CAR T cell- based immunotherapy to eradicate UM metastases. We will use the combination of trametinib, a mitogen-activated protein kinase kinase inhibitor (MEKi) and of panobinostat, a histone deacetylase inhibitor (HDACi), since this combination has been shown to induce marked regression of UM liver metastases in mice. Furthermore, this combination does not affect B7- H3 expression by UM cells and the antitumor activity of B7-H3 CAR T cells. Therefore, after having shown that B7-H3 CAR T cells can recognize and eliminate UM cells surviving the treatment with the MEKi and HDACi combination, we will investigate whether tumor debulking induced by the MEKi and HDACi combination can enhance the ability of B7-H3 CAR T cells to eradicate UM liver metastases in NSG mice. Since one major limitation of CAR T cell-based therapy is represented by its potential toxicity and/or cytokine release syndrome, we have incorporated in our CAR construct an inducible caspase 9 safety switch which allows rapid elimination of CAR T cells in case of unexpected toxicities. If the results generated by our experiments are positive, they will have a major impact on the treatment of mUM.