Targeting SYK Tyrosine Kinase to Overcome Radiation Resistance in ALL

描述（由申请人提供）：我们提出开发一种新的策略，以克服B系急性淋巴细胞白血病（ALL）的辐射抗性，使用C-61（一种新型SYK激酶底物结合（P）位点抑制剂）靶向和破坏白血病B细胞前体中的抗凋亡SYK-STAT 3信号传导网络。我们的中心工作假设是，复发性B系ALL患者的治疗结果可以通过抑制SYK-STAT 3分子靶点来改善。这将通过在HSCT的背景下使用C-61与全身照射（TBI）组合来实现。在特定目标1下，我们将使用定量流式细胞术凋亡测定和克隆形成测定来研究SYK P位点抑制剂C-61对复发性B系ALL患者的原代ALL细胞的体外辐射抗性的影响。我们假设C- 61将显著增强辐射诱导的原代B系ALL细胞凋亡，并通过增加其辐射敏感性和损害其修复亚致死辐射损伤的能力来增加其克隆形成部分的辐射诱导死亡。在特定目标2下，我们将使用复发性B系ALL和亚致死性全身照射（TBI）的SCID小鼠异种移植模型，检查C-61对复发性B系ALL患者的原代ALL细胞的体内辐射抗性的影响。我们的假设是，C-61加TBI方案将比单独TBI更有效地改善与原代B系ALL细胞激发的SCID小鼠的无事件生存结局。在特定目标3下，我们将研究复发性B系ALL患者的原代ALL细胞的激酶表达谱与其体外和体内辐射抗性之间的关联。在旨在鉴定有助于选择最有可能从C-61获益的患者的复合生物标志物谱的努力中，我们将原代B系ALL细胞的SYK、BTK和JAK激酶的激酶蛋白表达和活性水平与它们的辐射抗性、对C-61诱导的体外放射增敏的敏感性、以及C-61诱导的体内亚致死TBI的抗白血病效力的增强。根据具体目标4，我们将在放射抗性BCL-1小鼠B系白血病的同基因小鼠HSCT模型中研究亚致死（2戈伊）和临床应用（7戈伊）总放射剂量水平下含C-61的单次剂量TBI方案的疗效和安全性。我们将在同基因BMT背景下，在接种1 × 106 BCL-1细胞的BALB/c小鼠中评价2-10戈伊剂量范围的TBI与C-61联合治疗的有效性和安全性。我们推测C-61的加入不会增加TBI的非血液学毒性，同时显著增强其抗白血病疗效。我们预计，C-61纳入到HSCT TBI方案的复发性B系ALL患者将改善他们的治疗反应和生存结局。这项研究有可能为开发采用含C-61的新型TBI方案的范式转移HSCT策略提供基础。 公共卫生相关性：目前，治疗儿童白血病的主要挑战是治愈那些尽管接受了强化化疗但癌症复发的患者。该研究的目的是开发一种有效的治疗方案，采用新的抗白血病候选药物对抗化疗和放疗耐药的儿童白血病。该研究项目的成功完成可能为儿童白血病的治疗创新提供基础。

DESCRIPTION (provided by applicant): We are proposing to develop a new strategy to overcome radiation resistance in B- lineage acute lymphoblastic leukemia (ALL) using C-61, a novel SYK kinase substrate binding (P)-site inhibitor, for targeting and disrupting the anti-apoptotic SYK-STAT3 signaling network in leukemic B-cell precursors. It is our central working hypothesis that the treatment outcome of relapsed B-lineage ALL patients can be improved by inhibition of the SYK-STAT3 molecular target. This would be accomplished by using C-61 in combination with total body irradiation (TBI) in the context of HSCT. Under Specific Aim 1, we will examine the effects of the SYK P-site inhibitor C-61 on in vitro radiation resistance of primary ALL cells from relapsed B-lineage ALL patients using quantitative flow cytometric apoptosis assays and clonogenic assays. We hypothesize that C- 61 will markedly enhance radiation-induced apoptosis of primary B-lineage ALL cells and augment radiation- induced death of their clonogenic fraction by increasing their radiation sensitivity and impairing their capacity to repair sublethal radiation damage. Under Specific Aim 2, we will examine the effects of C-61 on in vivo radiation resistance of primary ALL cells from relapsed B-lineage ALL patients using SCID mouse xenograft models of relapsed B-lineage ALL and sublethal total body irradiation (TBI). Our hypothesis is that C-61 plus TBI regimens will be more effective than TBI alone in improving the event-free survival outcome of SCID mice challenged with primary B-lineage ALL cells. Under Specific Aim 3, we will examine the association between the kinase expression profiles of primary ALL cells from relapsed B-lineage ALL patients and their in vitro as well as in vivo radiation resistance. In an effort aimed at identifying a composite biomarker profile that will help select patients most likely to benefit from C-61, we will correlate the kinase protein expression and activity levels of SYK, BTK, and JAK kinases of primary B-lineage ALL cells with their radiation resistance, sensitivity to C-61 induced radiosensitization in vitro, as well as C-61 induced potentiation of the anti-leukemic potency of sublethal TBI in vivo. Under Specific Aim 4, we will study the efficacy and safety of C-61 containing single dose TBI regimens at both sublethal (2 Gy) as well as clinically applied (7 Gy) total radiation dose levels in a syngeneic murine HSCT model of radiation-resistant BCL-1 murine B-lineage leukemia. We will evaluate the efficacy and safety of TBI at doses ranging from 2-10 Gy in combination with C-61 in BALB/c mice inoculated with 1x106 BCL-1 cells in the context of syngeneic BMT. We hypothesize that the addition of C-61 will not increase the non-hematologic toxicity of TBI, while markedly potentiating its anti-leukemic efficacy. We anticipate that the incorporation of C-61 into the pre-HSCT TBI regimens of patients with relapsed B-lineage ALL will improve their treatment response and survival outcome. The proposed research has the potential provide the foundation for the development of paradigm-shifting HSCT strategies that employ C-61 containing novel TBI regimens. PUBLIC HEALTH RELEVANCE: Currently, the major challenge in the treatment of childhood leukemia is to cure patients who experience a recurrence of their cancer despite intensive chemotherapy. The purpose of the proposed research is the development of an effective treatment program against chemotherapy- and radiation therapy-resistant childhood leukemia employing a new anti-leukemia drug candidate. The successful completion of this research project may provide the foundation for therapeutic innovation against childhood leukemia.