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

总结 疾病复发是自体（auto）和异体（allo）移植后治疗失败的主要原因。 造血干细胞移植（HCT）治疗非霍奇金淋巴瘤（NHL）。因此，委员会认为， 人们对使用过继性细胞免疫疗法来根除有限数量的肿瘤细胞非常感兴趣 在条件化放疗后存活。对于不能进行HCT或HCT后复发的患者， HCT，这种策略可能提供一种具有治愈目的的选择。嵌合抗原受体（CAR）-工程化T 细胞已成为复发性/难治性CD 19+恶性肿瘤的一种有希望的治疗方法，但其全部潜力 这种疗法的有效性受到减弱的植入和缺乏CAR重定向T细胞的长期持久性的阻碍。 患者体内的细胞。我们提出了一种新的方法来提高CAR T细胞的功效和耐久性， 巨细胞病毒（CMV）特异性T细胞的性质和使用CMV疫苗，Triplex， 最近在City of Hope研发并进行临床评估Triplex是一种多抗原重组修饰 安卡拉牛痘（MVA）病毒，其具有编码3种CMV蛋白pp 65、IE 1和IE 2的基因。CMV-MVA Triplex具有 在CMV血清阴性和血清阳性的健康志愿者中证明了安全性和强免疫原性， I期试验，目前正在接受alloHCT的患者中进行测试。我们的方法需要选择CMVpp 65- 用于用靶向CD 19的CAR进行离体修饰的双特异性CMV-CD 19 CAR T细胞， 细胞进入患者体内，然后通过刺激天然CMV特异性T细胞受体诱导体内扩增 (TCR)使用三重注射。我们的战略将缩短CAR T细胞的制造时间，并采用双重技术 本发明的优点在于防止输注的细胞产品的过早衰老，并且使产品更 快速可用。此外，在我们的CAR设计中，截短的表皮生长因子受体（EGFRt）将作为 用于消融CAR T细胞的跟踪标记和体内安全开关。拟议的战略是 旨在增强增殖，延长持久性和增强过继性抗淋巴瘤活性， 通过天然CMVpp 65特异性TCR再刺激这些细胞来转移CMV-CD 19 CAR T细胞。 这应该会改善NHL患者的无进展生存期。创新的一面是 建议用一种新的体内扩增方法代替输注细胞产物的体外扩增 战略在特异性目标1（SA 1）中，我们将验证该双特异性CMV-CD 19 CAR T细胞生产平台 以生产临床级和规模的细胞产品。在具体目标2和3中，我们将测试 CMV-CD 19 CAR T细胞治疗后，在复发性乳腺癌患者的试点研究中使用三联疫苗， autoHCT或淋巴细胞耗竭（SA 2）或alloHCT（SA 3）后的难治性B细胞NHL。其意义 方法是我们将在体内控制过继输注的CMV-CD 19 CAR T细胞扩增， 消融术该项目作为提高CAR有效性的方法的原理证明， 控制T细胞扩增，其可以潜在地应用于多种治疗环境中的多种疾病。

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.