Survival Mechanisms of Cancer-initiating (Stem) Cells in Ph+ Leukemia

描述（由申请人提供）：该项目的总体目标是确定白血病起始（干细胞）自我更新和存活的分子机制，这些细胞可能是人类费城染色体阳性（Ph+）白血病的治愈性治疗靶点。由BCR-ABL癌基因诱导的Ph+白血病，包括慢性髓性白血病（CML）和b细胞急性淋巴细胞白血病（B-ALL），是最常见的骨髓增生性恶性肿瘤之一。同种异体骨髓移植是治疗慢性粒细胞白血病的唯一有效方法，但对大多数患者来说并不可行。包括甲磺酸伊马替尼（格列卫）在内的BCR-ABL酪氨酸激酶抑制剂对慢性粒细胞白血病患者非常有效，但对更晚期的CML细胞危象和Ph+ B-ALL患者无效。此外，临床对伊马替尼产生耐药性，而且伊马替尼不能完全根除白血病干细胞，这表明使用BCR-ABL激酶抑制剂（如伊马替尼）作为单一药物不能阻止最终的疾病进展或治愈CML。需要新的治疗策略，特别是对于老年CML患者。我们的总体假设是，这些白血病干细胞的自我更新和存活是由不同的分子途径介导的，而不是那些刺激白血病细胞增殖的分子途径，靶向白血病干细胞对Ph+白血病的根治性治疗至关重要。本项目利用高效、准确的Ph+白血病小鼠模型来鉴定和表征白血病干细胞，主要有三个具体目的：1)确定BCR-ABL转导的骨髓细胞的哪些造血谱系转移Ph+白血病，影响白血病干细胞功能，以及对BCR-ABL激酶抑制剂敏感。白血病小鼠将接受BCR-ABL激酶抑制剂伊马替尼或BMS-354825治疗，随后使用多色流式细胞仪分析白血病干细胞，以识别和分类不同的细胞系，以进行疾病的过继转移，并检查干细胞的存活/自我更新。2)确定2-catenin被BCR-ABL激活的造血谱系，检测Src激酶是否参与2-catenin在这些谱系中的激活，并对这些细胞的存活产生积极影响，确立2-catenin在BCR-ABL白血病发生中的作用。Src激酶缺陷小鼠和Src激酶抑制剂以及2-catenin条件敲除小鼠将用于研究2-catenin在BCR-ABL白血病发生中的作用。3)确定新型抗干细胞药物BMS-214662对表达BCR-ABL的造血干细胞存活的抑制机制，并检测该化合物与BCR-ABL激酶抑制剂联合治疗Ph+白血病是否具有治愈性。白血病小鼠将单独使用BMS-214662或与双BCR- ABL/Src激酶抑制剂BMS-254825一起治疗，然后分析小鼠白血病干细胞的存活/自我更新。这项研究将有助于了解白血病干细胞的生物学特性，并为Ph+白血病开发新的抗干细胞策略。

DESCRIPTION (provided by applicant): The overall goal of this project is to identify molecular mechanisms for self-renewal and survival of leukemic initiating (stem) cells that may be targets for curative therapy of human Philadelphia chromosome-positive (Ph+) leukemias. Ph+ leukemias induced by the BCR-ABL oncogene, including chronic myeloid leukemia (CML) and the B-cell acute lymphoblastic leukemia (B-ALL), are among the most common myeloproliferative malignancies. Allogeneic bone marrow transplantation, the only established curative therapy for CML, is not feasible for most patients. The BCR-ABL tyrosine kinase inhibitors including imatinib mesylate (Gleevec) are highly effective in treating chronic phase CML patients, but not more advanced CML blast crisis and Ph+ B-ALL patients. Moreover, clinical resistance to imatinib develops and imatinib does not completely eradicate leukemic stem cells, suggesting that use of a BCR-ABL kinase inhibitor such as imatinib as a single agent will not prevent eventual disease progression or cure CML. New therapeutic strategies are needed, especially for older CML patients. Our overall hypothesis is that self-renewal and survival of these leukemic stem cells are mediated by different molecular pathways than those that stimulate leukemic cell proliferation, and targeting leukemic stem cells is essential to curative therapy of Ph+ leukemias. This project uses the efficient, accurate mouse models for Ph+ leukemias to identify and characterize leukemic stem cells by focusing on three specific aims: 1) To determine which hematopoietic lineages of BCR-ABL-transduced bone marrow cells transfer Ph+ leukemias, affect leukemic stem cell function, and are sensitive to BCR-ABL kinase inhibitors. Leukemic mice will be treated with a BCR-ABL kinase inhibitor imatinib or BMS-354825, followed by the analysis of leukemic stem cells using multicolor FACS to identify and sort different cell lineages for adoptive transfer of the diseases and for examination of survival/self-renewal of the stem cells. 2) To determine the hematopoietic lineages in which 2-catenin is activated by BCR-ABL, to test whether Src kinases are involved in 2-catenin activation in these lineages and have a positive effect on survival of these cells, and to establish the role of 2-catenin in BCR-ABL leukemogenesis. Src kinase-deficient mice and Src kinase inhibitors, and 2-catenin conditional knockout mouse will be used to address the role of 2-catenin in BCR-ABL leukemogenesis. 3) To determine the inhibitory mechanism of a novel anti-stem cell agent BMS-214662 on survival of BCR-ABL-expressing HSCs and to test whether this compound in combination with a BCR-ABL kinase inhibitor provides a curative therapy for Ph+ leukemias. Leukemic mice will be treated with BMS-214662 alone or together with a dual BCR- ABL/Src kinase inhibitor BMS-254825, followed by the analysis of survival/self-renewal of leukemic stem cells in mice. This study will help to understand the biology of leukemia stem cells and develop novel anti-stem cell strategies for Ph+ leukemias.