CLINICAL PHENOTYPE TARGETED ANTIFOLATE CHEMOTHERAPY

人类白血病的药物敏感性和临床转归很高 取决于它们起源的细胞谱系。这项提议将旨在 定义与药物代谢有关的生化表型 解释了对抗叶酸表现出的谱系特异性反应 人类白血病。经典和新型抗叶酸的多谷氨酰化反应 叶基多谷氨酸合成酶(FPGS)对它们的生长至关重要 药理活性，导致细胞内滞留时间延长 并增加了细胞毒性。拟议的研究将测试 人类白血病对抗叶酸的反应取决于 在FpGS基因表达的基础上进行了表达。我们将调查生化和 已报道的临床观察的分子基础是一个谱系- 在体内白血病母细胞后，FPGS活性出现特异性增加 接触到这些药物。这些研究将确定底物的作用 对FPGS的亲和力和对关键的叶酸代谢酶的抑制，以及 非多谷氨酰化抗叶酸和天然叶酸的作用。 暴露于正常造血祖细胞后的类似研究 抗叶酸将定义FPGS在药物选择性中的潜在作用。 此外，正常人DNA甲基化和FPGS mRNA表达的变化 将对骨髓细胞和白血病母细胞进行研究。评估 这些结果、FPGS和多谷氨酰化的临床意义 相关参数将在抗叶酸的临床样本中确定 敏感和耐药白血病。胎儿宫内节育器的临床意义 抗叶酸反应也将通过生长因子诱导的测试 暴露于耐药髓系白血病细胞中的FPGS上调 这些试剂，并在可诱导表达系统转染后 将hFPGS编码为一种“酶缺陷”耐药白血病表型。 总体而言，这项建议应该为以下问题提供一个明确的答案：1.如何 FPGS在抗叶酸的临床反应中重要吗？2.什么是 新型抗叶酸对白血病细胞FPGS表达的影响 它们在敏感表型、耐药表型和正常骨表型上有区别吗？ 骨髓祖细胞？3.FPGS的家族特异性表达是一种 一种生化表型预测因子的重要临床决定因素 抗叶酸肿瘤反应？

The drug sensitivity and clinical outcome of human leukemias is highly dependent on their cell lineage of origin. This proposal will aim to define biochemical phenotypes with respect to drug metabolism that account for the lineage-specific response to antifolates exhibited by human leukemias. Polyglutamylation of classical and novel antifolates by the enzyme Folylpolyglutamate synthetase (FPGS) is essential to their pharmacological activity, resulting in prolonged intracellular retention and increased cytotoxicity. The proposed studies will test the hypothesis that the response of human leukemias to antifolates depend upon the expression of FPGS. We will investigate the biochemical and molecular basis for the reported clinical observation that a lineage- specific increase in FPGS activity occurs after in vivo leukemic blasts' exposure to these drugs. These studies will define the role of substrate affinity for FPGS and inhibition of key folate-metabolizing enzymes, and the effects of non-polyglutamylatable antifolates and natural folates. Similar studies with normal hematopoietic progenitors after exposure to antifolates will define the potential role of FPGS in drug selectivity. Further, changes in DNA methylation and FPGS mRNA expression in normal bone marrow cells and leukemic blasts will be investigated. To evaluate the clinical significance of these results, FPGS and polyglutamylation related parameters will be determined in clinical samples from antifolate sensitive and resistant leukemias. The clinical relevance of FPGS in antifolate response will also be tested by growth factor-induced upregulation of FPGS in resistant myeloid leukemic blasts' exposed to these agents, and after transfection of an inducible expression system encoding hFPGS to an "enzyme deficient" resistant leukemic phenotype. Overall, this proposal should provide a definitive answer to: 1. How important is FPGS in the clinical response to antifolates? 2. What are the effects of novel antifolates on leukemic blasts' FPGS expression, and do these differ in sensitive vs resistant phenotypes vs normal bone marrow progenitors? 3. Is the lineage-specific expression of FPGS an important clinical determinant of a biochemical phenotype predictor of antifolate tumor response?