Human Donor Stem/Progenitor Cells with Dual Gene Editing (CD33 and CLL-1) Enable "Two Hit" targeting of Acute Myeloid Leukemia"

具有双基因编辑（CD33 和 CLL-1）的人类供体干细胞/祖细胞能够实现急性髓系白血病的“两次打击”

• 批准号: 10311553
• 负责人: Abdullah M Ali
• 金额: $ 22.27万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-12-03 至 2022-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10311553
• 关键词: Ablation; Activities of Daily Living; Acute Myelocytic Leukemia; Alleles; Allogenic; Antibody-drug conjugates; Antigen Targeting; Antigens; Biological Assay; Blast Cell; Bone Marrow; CD34 gene; CRISPR/Cas technology; Cell Line; Cells; Clinical; Clinical Research; Clinical Trials; Clonal Expansion; Colony-Forming Units Assay; Disease; Disease model; Disease remission; Dose; Drug Efflux; Dysmyelopoietic Syndromes; Engineering; Engraftment; Flow Cytometry; Funding; Gemtuzumab Ozogamicin; Genes; Genetic; Genomics; Goals; Grant; Hematopoietic; Hematopoietic Stem Cell Transplantation; Hematopoietic stem cells; Human; IL3RA gene; IL6 gene; IL8 gene; Immunotherapeutic agent; Immunotherapy; In Vitro; Injections; Institutional Review Boards; Legal patent; Letters; Leukemic Cell; Lymphoid; Malignant Neoplasms; Measures; Mediating; Methods; Modeling; Monitor; Morbidity - disease rate; Mus; Mutation; Myelogenous; Myeloid Cells; Myeloproliferative disease; Myelosuppression; PTPRC gene; Patients; Phagocytosis; Phase; Postremission Therapy; Pre-Clinical Model; Preparation; Production; Publishing; Refractory; Relapse; Remission Induction; Residual Neoplasm; Resistance; Resistance development; Site; Stem cell transplant; TNF gene; Testing; Therapeutic; Toxic effect; Transplantation; Tumor Antigens; Veno-Occlusive Disease; Work; acute myeloid leukemia cell; bi-specific T cell engager; chimeric antigen receptor T cells; chronic T-cell leukemia; curative treatments; cytokine; cytokine release syndrome; deep sequencing; experimental study; high risk; improved outcome; in vitro Assay; in vivo; innovation; leukemia; macrophage; monocyte; mouse model; novel; novel strategies; novel therapeutic intervention; overexpression; patient derived xenograft model; peripheral blood; phase 1 study; pre-clinical; preservation; progenitor; relapse patients; relapse risk; resistance mechanism; stem; stem cells; therapeutic gene; translational study

PROJECT SUMMARY/ABSTRACT: The goal of this study is to develop a novel therapeutic strategy for AML, a malignancy of myeloid lineage, that represents a disease with enormous unmet therapeutic need. Hematopoietic stem cell transplant (HSCT) is the only curative treatment option for patients with relapsed/refractory AML but up to 40% patients further relapse after HSCT. Targeting of AML cells using agents directed against a lineage specific antigen such as CD33 using the antibody drug conjugate Gemtuzimab Ozagamycin (GO; Mylotarg) has improved outcomes, but the use of Mylotarg is associated with severe myelosuppression. This is due to targeting of both the leukemia cells and normal myeloid cells (including stem, progenitor and myeloid cells in the donor graft) that also express CD33 (the normal myeloid cells are concomitantly killed by Mylotarg, which also has other toxicities, including veno- occlusive disease at high doses). We reasoned that by ablating CD33 expression using genomic engineering methods in donor stem/progenitor cells, we could generate stem/progenitor cells for transplant that are resistant to Mylotarg treatment, while rendering the AML cells uniquely sensitive to anti-CD33 therapy such as low- dose Mylotarg, anti-CD33 CAR-T or CD33 bi-specific T cell engagers (BiTEs). Indeed, in a recently published “proof of concept study”, we demonstrated that this strategy (either Mylotarg alone, CART-33 or both) enabled the complete killing of an engrafted human CD33+ AML cell line, while allowing fully functional hematopoietic repopulation by CD33- HSPCs. In preliminary experiments with primary human AML cells, however, we found that despite AML clearance with Mylotarg, a small fraction of AML persisted in the bone marrow after 16 weeks of treatment. These AML cells are CD33+ but also express a second lineage antigen, CLL-1+ (Clec12a). We reasoned that targeting multiple antigens will result reduce the chances of relapse. In this proposal, we plan to test the hypothesis that long-term leukemia remission is achieved when two antigens on AML cells are targeted, either serially or simultaneously. To test this hypothesis, we will measure the engraftment, differentiation, and functional potential of HSPCs that are gene-edited to ablate expression of CLL-1 antigen alone or in combination with CD33 (Aim 1). We will test this two-hit treatment approach using patient derived xenograft (PDX) models of primary human AML and also test the engraftment, differentiation, and functional potential of single (CLL-1) and double (CLL-1 and CD33) deletion HSPC in this context (Aim 2). The long-term objective of this study is to develop novel cell-based gene therapeutic strategies in combination with immunotherapeutic approaches for the treatment of AML and related cancers such myelodysplastic syndromes.

项目摘要/摘要： 这项研究的目标是开发一种新的治疗AML的策略，AML是一种髓系恶性肿瘤， 代表一种巨大的未得到满足的治疗需求的疾病。造血干细胞移植(HSCT) 复发/难治性AML患者的唯一治疗选择，但高达40%的患者进一步复发 HSCT后。使用针对谱系特异性抗原的药物如CD33靶向AML细胞 抗体药物结合物Gemtuzimab Ozagamcin(GO；Mylotarg)有改善疗效，但使用 Mylotarg与严重的骨髓抑制有关。这是由于白血病细胞和 也表达CD33的正常髓系细胞(包括供者移植物中的干细胞、祖细胞和髓样细胞) (正常的髓系细胞同时被Mylotarg杀死，Mylotarg也有其他毒性，包括静脉注射- 高剂量下的闭塞疾病)。我们通过利用基因组工程去除CD33的表达来推断 方法在捐赠者的干细胞/祖细胞中，我们可以产生耐药的干细胞/祖细胞用于移植。 Mylotarg治疗，同时使AML细胞对抗CD33治疗如Low-CD33特别敏感 剂量Mylotarg、抗CD33 CAR-T或CD33双特异性T细胞活化剂(BITES)。事实上，在最近出版的一份 “概念验证研究”，我们演示了该策略(单独使用Mylotarg、CART-33或两者都支持) 完全杀死植入的人CD33+AML细胞系，同时允许完全功能的造血 用CD33-HSPC进行再种群。 然而，在对原代人类AML细胞的初步实验中，我们发现尽管AML被清除了 Mylotarg，一小部分AML在治疗16周后在骨髓中持续存在。这些AML细胞 是CD33+，但也表达第二个谱系抗原，CLL-1+(Clec12a)。我们的理由是将多个目标 抗原会导致减少复发的机会。在这项提议中，我们计划检验这样的假设：长期 当AML细胞上的两个抗原被连续或同时靶向时，白血病的缓解就实现了。 为了验证这一假设，我们将测量HSPC的植入、分化和功能潜力 经基因编辑以单独或与CD33联合表达CLL-1抗原(目标1)。我们将测试 这种使用患者来源的异种移植(PDX)模型的两次治疗方法 检测单个(CLL-1)和双(CLL-1和CD33)细胞的植入、分化和功能潜能 删除这方面的HSPC(目标2)。这项研究的长期目标是开发新的基于细胞的基因 急性髓系白血病及相关疾病的免疫治疗策略 癌症，如骨髓增生异常综合征。