Off-The-Shelf Dual Targeted NK Cells For NHL

限制复发是非霍奇金淋巴瘤自体移植后的最大挑战。 抗原导向的CAR T细胞产品和其他靶向疗法可能正在改变这一领域，但对于高风险患者来说， 淋巴瘤，需要新的和更安全的方法。我们将研究抗原导向的， 现成的、诱导多能干细胞（iPSC）衍生的NK细胞，其被工程化用于双抗原靶向。我们 已经优化了iPSC衍生的CD 34+细胞向高效NK细胞的分化， 明尼苏达大学，现在可以在生产运行中用克隆iPSC生产数百剂 用高亲和力、不可切割的CD 16（称为hnCD 16）转导。2019年底，我们启动了临床试验， 测试这种工程化的iPSC产品（命名为FT 516）。这个项目的首要假设是， 用CAR和通过hnCD 16双重靶向NK细胞将防止自体移植后复发， 淋巴瘤和未来基因编辑改变代谢将增强过继转移。这一假设符合 该计划的主题，项目2将告知该计划关于现成NK细胞的开发， NK CAR和膜IL-15的作用，以及临床前对操纵代谢是否会 增强NK细胞治疗。这些调查得到以下具体目标的支持。目标1将测试双 使用两种作用机制ADCC和NK CAR将NK细胞靶向CD 19和CD 20，以预防 淋巴瘤自体移植后复发。我们将测试三重基因修饰的iPSC衍生的NK细胞， 用hnCD 16（NK细胞优化的CD 19 CAR）和膜结合15/IL-15 Ra转导的产物（FT 596） 融合FT 596将与利妥昔单抗联合作为维持治疗，以限制自体HCT后的复发。 该试验是FDA批准的，将首先在植入后第+30天确定MTD，然后，如果单次给药， 安全，移到植入前+7天，目标是在前100天内接种3剂，以防止复发。 目的2将测试通过ARID 5 B过表达和CD 38敲除的NK细胞代谢适应性的增强。我们 初步数据显示，适应性NK细胞和CD 38敲除iPSC衍生的NK细胞均抵抗氧化应激。 诱导细胞死亡。在适应性NK细胞中自然增加的ARID 5 B过表达介导了类似的作用。 我们将产生新的iPSC系，以构建具有ARID 5 B转基因表达或CRISPR敲除的FT 596 CD38将对这些克隆进行评估，以探索它们如何驱动NK细胞代谢的机制， 增广函数目标3将测试操纵ARID 5 B和CD 38是否会驱动代谢适应性， 增强FT 596在体内的持久性和功能。这些临床前试验将为未来的临床试验提供信息 修饰-所有这些修饰都通过延长转移的NK细胞持久性来促进增强的抗淋巴瘤活性， 力量这些代谢和效力的发现将直接告知项目1，以改善细胞靶向， 代谢，项目3细胞持久性，以及所有3个项目，以定义自主的优势， IL-15的表达。

Limiting relapse is the biggest challenge following autologous transplantation for non-Hodgkin’s lymphoma. Antigen-directed CAR T cell products and other targeted therapies may be changing the field, yet for high-risk lymphomas, new and safer approaches are needed. We will study the safety and efficacy of antigen-directed, off-the-shelf, induced pluripotent stem cell (iPSC)-derived NK cells engineered for dual-antigen targeting. We have optimized the differentiation of iPSC-derived CD34+ cells to highly potent NK cells, scaled production at the University of Minnesota, and can now produce hundreds of doses in a manufacturing run with a clonal iPSC transduced with a high-affinity, non-cleavable CD16 (called hnCD16). In late 2019, we initiated clinical trials testing this engineered iPSC product (designated FT516). The overarching hypothesis for this project is that dual targeting of NK cells with a CAR and through hnCD16 will protect against relapse after auto-transplant for lymphoma and that future gene edits to alter metabolism will enhance adoptive transfer. This hypothesis fits with the themes of this Program, and Project 2 will inform the Program on the development of off-the-shelf NK cells, NK CAR, and the role of membrane IL-15, as well as pre-clinically on whether manipulating metabolism will enhance NK cell therapy. These investigations are supported by the following Specific Aims. Aim 1 will test dual targeting of NK cells to CD19 and CD20 using two mechanisms of action, ADCC and an NK CAR, to prevent relapse after autologous transplantation for lymphoma. We will test a triple gene-modified iPSC-derived NK cell product (FT596) transduced with hnCD16, an NK cell-optimized CD19 CAR, and membrane bound 15/IL-15Ra fusions. FT596 will be combined with rituximab as maintenance therapy to limit relapse after autologous HCT. This trial is FDA-approved and will first establish an MTD at day +30 after engraftment and then, if single dosing is safe, move to day +7 before engraftment with the goal of 3 doses in the first hundred days to prevent relapse. Aim 2 will test enhancement of NK cell metabolic fitness by ARID5B overexpression and CD38 knockout. Our preliminary data shows both adaptive NK cells and CD38 knockout iPSC-derived NK cells resist oxidative stress- induced cell death. ARID5B overexpression, naturally increased in adaptive NK cells, mediates similar effects. We will generate new iPSC lines to build on FT596 with transgenic expression of ARID5B or a CRISPR knockout of CD38. These clones will be evaluated to probe the mechanisms on how they drive NK cell metabolism and augment function. Aim 3 will test whether manipulating ARID5B and CD38 will drive metabolic fitness and enhance FT596 persistence and function in vivo. These pre-clinical trials will inform future clinical trial modifications—all to promote enhanced anti-lymphoma activity by extending transferred NK cell persistence and potency. These metabolic and potency findings will directly inform Project 1 to improve cell targeting and metabolism, Project 3 for cell persistence, and all 3 Projects to define the advantages of autonomous presentation of IL-15.