Analysis of E-selectin Ligands of Human Acute Leukemia Cells and their Biology in Leukemogenesis

项目总结 本申请是根据NOSI NOT-CA-21-034提交的。所产生的工作来自 家长助学金(U01 CA225730)是对梅奥诊所/华盛顿大学U54资助的博士学位的补充 明尼苏达州(U54 CA224018)，我们建议执行利用资源的协作实验 从本博士论文的所有3个项目中，以增强两项赠款的影响。多发性骨髓瘤(MM)残留 不治之症与多发性骨髓瘤细胞对目前可用的联合疗法产生耐药性的机制 治疗方案和最近开发的免疫疗法仍然不完全清楚。我们的研究记录了 骨髓(BM)、间充质基质细胞(BMSCs)等非恶性细胞在BM/骨中的作用 MM、白血病和实体瘤细胞对多种药物或免疫耐药的关键作用 效应细胞。我们观察到人多发性骨髓瘤或白血病细胞在磷酸氢钙(BCP)支架中 具有人源化间充质间隔室的BM样体内系统(与体外或 传统的异种皮下移植)明显的基因组依赖性和对不同 治疗。在亲本U01下，我们认识到表面蛋白糖基化在 调控恶性造血细胞与骨髓细胞的相互作用。此外，我们最近观察到 通过基于CRISPR的表面蛋白糖基化调节因子的扰动可以促进肿瘤细胞 对药物或免疫治疗的抵抗力。这个目前的项目寻求应用我们的 BM-Like人源化体内支架模型；利用我们的资源调节表面蛋白 糖基化，我们在体内基于CRISPR的研究的经验，以及 免疫活性VK*myc基因工程小鼠模型(GEMM)从U54DrSc到(1) 检测MM细胞表面高水平的SLeX，这是造血相互作用的关键决定因素 具有BM血管生态位的细胞，对BM生态位内的体内治疗表现出更明显的抵抗力； 并应用基于CRISPR的方法系统地确定MM细胞的体内耐药机制 在BM环境中(2)抵抗药理药物的组合；以及(3)最近开发的免疫- 基础抗多发性骨髓瘤治疗。这个项目将协同应用“人性化”的翻译力量和 GEMM活体模型的协作计划和全面的CRISPR方法，提供关键 对骨髓环境中的MM细胞如何对药物组合产生耐药性的见解 或者免疫疗法。这些结果将向U54 DrSc通报新方法的临床前测试，这些方法 可预防、延缓或克服这些形式的治疗耐药性。此次协作还将提供 蓝图，特别是与表面蛋白糖基化有关的治疗抗性基因，将 加快父U01的努力；也可以应用于促进其他U54 DRSC的努力 包括起源于骨髓/骨环境或转移到骨髓/骨环境的恶性肿瘤。

PROJECT SUMMARY This application is being submitted in response to the NOSI NOT-CA-21-034. The work generated from the parent grant (U01 CA225730) is complementary to the U54 funded DRSC at Mayo Clinic/University of Minnesota (U54 CA224018) and we propose to perform collaborative experiments that utilize resources from all 3 projects of this DRSC to enhance the impact of both grants. Multiple myeloma (MM) remains incurable and the mechanisms through which MM cells develop resistance to currently available combination regimens and recently developed immunotherapies remain incompletely understood. Our studies documented that bone marrow (BM) mesenchymal stromal cells (BMSCs) and other nonmalignant cells of the BM/bone play critical roles in resistance of MM, leukemia and solid tumor cells to various pharmacological agents or immune effector cells. We observed that human MM or leukemic cells in a bicalcium phosphate (BCP) scaffold-based BM-like in vivo system with “humanized” mesenchymal stromal compartment exhibit (compared with in vitro or conventional subcutaneous xenografts) distinct genomic dependencies and decreased responses to diverse therapies. Under the parent U01, we have appreciated that surface protein glycosylation plays critical roles in regulating how malignant hematopoietic cells interact with the BM niche. Moreover, we recently observed through CRISPR-based that perturbation of surface protein glycosylation regulators can contribute to tumor cell resistance against pharmacological or immune-based treatments. This current project seeks to apply our the BM-like "humanized" scaffold in vivo model; leverage our resources on regulation of surface protein glycosylation, our experience with CRISPR-based studies in vivo and the translational relevance of the immunocompetent Vk*myc genetically engineered mouse model (GEMM) of MM from the U54 DRSC to (1) examine whether MM cells with high surface levels of sLeX, a key determinant of interaction of hematopoietic cells with the BM vascular niche, exhibit more pronounced resistance to in vivo treatments within the BM niche; and apply CRISPR-based approaches to systematically define the mechanisms of in vivo resistance of MM cells within the BM milieu (2) against combinations of pharmacological agents; and (3) recently developed immune- based anti-MM treatment. This project will synergistically apply the translational power of the “humanized” and GEMM in vivo models of the collaborating programs and comprehensive CRISPR approaches to provide key insights into how MM cells within the BM milieu develop resistance to combinations of pharmacological agents or to immune therapies. These results will inform the U54 DRSC on preclinical testing of new approaches that may prevent, delay or overcome these forms of treatment resistance. This collaboration will also provide a blueprint, especially in regard to treatment resistance genes related to surface protein glycosylation, that will accelerate the efforts of the parent U01; and may also be applied to facilitate the efforts of other U54 DRSCs that involve malignancies that originate from or metastasize to the BM/bone milieu.