CMV/CD19 bi-Specific CAR T cells combined with CMV vaccine as post-transplantation immunotherapy for non-Hodgkin lymphoma

摘要 疾病复发是自体(自体)和异体(异体)治疗失败的主要原因 造血干细胞移植治疗非霍奇金淋巴瘤(NHL)因此， 人们对使用过继细胞免疫疗法根除有限数量的肿瘤细胞有着浓厚的兴趣。 在条件化放化疗后存活。对于不能接受HCT或复发的患者- HCT，这一策略可能会提供一种具有治疗意图的选择。嵌合抗原受体(CAR)工程T细胞 细胞已成为治疗复发/难治性CD19+恶性肿瘤的一种有前途的疗法，但其全部潜力 这种治疗的效果受到植入减弱和缺乏CAR重定向T的长期持久性的阻碍 病人体内的细胞。我们提出了一种新的方法来提高CAR T细胞的有效性和持久性 巨细胞病毒(CMV)特异性T细胞的特性及其在CMV疫苗、三联疫苗、 最近在希望之城开发并进行了临床评估。Triplex是一种多抗原重组修饰 编码3种巨细胞病毒蛋白pp65、IE1和IE2的安卡拉牛痘病毒。CMV-MVA三联体 在CMV血清阴性和血清阳性的健康志愿者中被证明是安全和强大的免疫原性 第一阶段试验，目前正在接受allHCT的患者中进行测试。我们的方法需要选择CMVpp65- 以CD19为靶向的CAR体外修饰特异性T细胞，注入双特异性CMV-CD19 CAR T 细胞进入患者体内，然后通过刺激天然的CMV特异性T细胞受体诱导体内扩张 (TCR)使用三联注射。我们的战略将缩短汽车T细胞的制造时间， 防止输注的细胞产品过早衰老的优点，并使产品更 很快就能买到。此外，在我们的汽车设计中，截短型表皮生长因子受体(EGFRt)将作为 既是一个跟踪标记，也是一个体内安全开关，用于消融CAR T细胞。建议的战略是 旨在促进增殖，延长持久性和增强过继的抗淋巴瘤活性 通过天然的CMVpp65特异性TCR重新刺激这些细胞来转移CMV-CD19 CAR T细胞。 这应该会改善NHL患者的无进展存活率。它的创新之处在于 建议用一种新的体内扩增取代输注细胞产品的体外扩增 策略。在特定目标1(SA1)中，我们将验证该双特异性CMV-CD19汽车T细胞制造平台 在cGMP条件下生产临床分级和规模化细胞产品。在具体的目标2和3中，我们将测试 CMV-CD19CAR T细胞疗法和三联疫苗治疗复发性非霍奇金淋巴瘤的初步研究 自体红细胞移植或淋巴滤过性移植(SA2)或异基因红细胞移植(SA3)后的难治性B细胞NHL。这件事的意义 方法是我们将在体内控制过继输注的CMV-CD19 CAR T细胞的扩增和 消融。该项目是提高汽车效率的方法的原则性证明， 控制T细胞的扩张，这可能会在多种治疗环境中应用于多种疾病。

SUMMARY Disease relapse is a leading cause of treatment failure after autologous (auto) and allogeneic (allo) hematopoietic stem cell transplantation (HCT) for patients with non-Hodgkin lymphoma (NHL). Accordingly, there is intense interest in using adoptive cellular immunotherapy to eradicate the limited number of tumor cells surviving after conditioning chemo-radiotherapy. For patients unable to undergo HCT or having relapsed post- HCT, this strategy may offer an option with curative intent. Chimeric Antigen Receptor (CAR)-engineered T cells have emerged as a promising treatment for relapsed/refractory CD19+ malignancies, but the full potential of this therapy is hampered by attenuated engraftment and lack of long-term persistence of CAR redirected T cells in patients. We propose a novel approach to improve efficacy and durability of CAR T cells based on properties of cytomegalovirus (CMV)-specific T cells and their expansion using a CMV vaccine, Triplex, recently developed and clinically evaluated at City of Hope. Triplex is a multi-antigen recombinant modified vaccinia Ankara (MVA) virus with genes encoding 3 CMV proteins, pp65, IE1, and IE2. CMV-MVA Triplex has proven safe and powerfully immunogenic in both CMV-seronegative and -seropositive healthy volunteers in a Phase I trial, and is now undergoing testing in alloHCT recipients. Our approach entails selecting CMVpp65- specific T cells for ex vivo modification with a CD19-targeting CAR, infusing the bi-specific CMV-CD19 CAR T cells into patients, and then inducing in vivo expansion by stimulating the native CMV-specific T cell receptor (TCR) using Triplex injections. Our strategy will shorten CAR T cell manufacturing time, with the dual advantage of preventing premature senescence of the infused cell product, and of making the product more rapidly available. Also, the truncated epidermal growth factor receptor (EGFRt) in our CAR design will act as both a tracking marker and an in vivo safety switch for ablating the CAR T cells. The proposed strategy is designed to enhance proliferation, lengthen persistence and augment the anti-lymphoma activity of adoptively transferred CMV-CD19 CAR T cells by re-stimulating these cells through the native CMVpp65-specific TCR. This should result in improved progression-free survival for NHL patients. The innovative aspect of this proposal is the substitution of in vitro expansion of the infused cell product with a novel in vivo expansion strategy. In Specific Aim 1 (SA1) we will validate this bi-specific CMV-CD19 CAR T cell manufacturing platform under cGMP conditions to produce clinical grade and scale cell products. In Specific Aims 2 and 3, we will test the CMV-CD19 CAR T cells therapy followed by Triplex vaccine in pilot studies for patients with relapsed- refractory B-cell NHL following autoHCT or lymphodepletion (SA2), or alloHCT (SA3). The significance of this approach is that we will have in vivo control of adoptively infused CMV-CD19 CAR T cell expansion and ablation. This project serves as a proof of principle for a method of enhancing CAR effectiveness and controlling T cell expansion that can potentially be applied to multiple diseases in multiple therapeutic settings.