PHORBOL ESTER INDUCED DIFFERENTIATION OF LEUKEMIC CELLS

白血病细胞的特征是成熟时停滞。 在造血发育阶段，有能力 增殖维持不变。体外培养的人白血病细胞 可以被刺激发育成非增殖细胞。因此， 白血病可以部分地被视为一种疾病。 分化与增殖一样重要。最强大的力量之一 刺激白血病细胞分化的药物是 120十四酰佛波醇13-乙酸酯(TPA)，佛波醇酯。通过 取代甘油二酯，TPA激活钙离子。 磷脂依赖激酶，蛋白激酶C。 二甘油衍生物模拟TPA刺激的细胞 反应表明，蛋白激酶C的激活可能介导了 TPA所产生的细胞效应的一部分。然而， 甘油二酯衍生物不能刺激分化 人类白血病细胞系U937。在我们的实验室里，一种非 蛋白激酶C、TPA刺激的激酶活性已被 已确认身份。蛋白激酶C与TPA刺激的蛋白激酶 活性可以通过底物的不同来区分 专一性。二酰基甘油衍生物不能 刺激分化或激活这一激酶表明 TPA刺激的激酶的激活可能在 中介TPA可促进细胞分化。 这项提议的目标是表征非蛋白激酶 C，TPA刺激的激酶，并确定这一激活 激酶在介导信号刺激中起着至关重要的作用 白血病细胞的分化。使用列 层析后，将对该激酶进行表征和比较 对其他非蛋白激酶C磷酸转移酶活性的影响 受TPA刺激。使用32P标记的完整细胞和两个 内源性底物的空间电泳法 由该激酶磷酸化将被定义。的影响 其他诱导U937分化的药物(淋巴因子、二丁三醇 营房等)将被评估对该激酶活性的影响。 更好地理解控制机制 人类白血病细胞的分化将有助于发育 专门设计的治疗方式，旨在刺激 白血病细胞的体内分化。这样的一种疗法，另外 提供另一种治疗形式，从理论上讲 避免许多毒副作用，这是目前 使用了抗肿瘤治疗。拟议中的研究还可以 提供对调解许多不同的 佛波醇酯在其他生物系统中的作用。

The leukemic cell is characterized by maturational arrest at a stage of hematopoietic development where the ability to proliferate is maintained. Human leukenic cells cultured in vitro can be stimulated to develop into a non-proliferating cell. Thus, the leukemias can be viewed, in part, as a disorder of differentiation as well as proliferation. One of the most potent agents stimulating differentiation of leukemic cells is 120tetradecanoyl phorbol 13-acetate (TPA), a phorbol ester. By substituting for diacylglycerol, TPA activates the CA2+- phospholipiddependent kinase, protein kinase C. The ability of diacyglycerol derivatives to mimic TPA stimulated cellular responses indicates that protein kinase C activation may mediate a portion of the cellular effects exerted by TPA. However, diacylglycerol derivatives are unable to stimulate differentiation of the human leukemic cell line U937. In our laboratory, a non- protein kinase C, TPA stimulated kinase activity has been identified. Protein kinase C and the TPA stimulated kinase activity can be distinguished by differences in substrate specificity. The inability of diacylglycerol derivatives to stimulate differentiation or to activate this kinase suggests that activation of the TPA stimulated kinase may be important in mediating TPA stimulated differentiation. The goal of this proposal is to characterize the nonprotein kinase C, TPA stimulated kinase and to determine if activation of this kinase is crucial in mediating the signal stimulating differentiation of the leukemic cell. Using column chromatography, this kinase will be characterized and compared to other non-protein kinase C phosphotransferase activities stimulated by TPA. Using 32P-labeled intact cells and two dimensional electrophoretic analysis, endogenous substrates phosphorylated by this kinase will be defined. The effects of other agents inducing U937 differentiation (lymphokines, dibutryl cAMP, etc.) will be evaluated for effects on this kinase activity. A better understanding of the mechanism controlling differentiation of human leukemic cells would aid in developing therapeutic modalities specifically designed to stimulate the in vivo differentiation of leukemic cells. Such a therapy, in addition to providing an alternative form of treatment, would theoretically avoid many of the toxic side effects inherent to the currently used antineoplastic therapy. The proposed research could also provide insight into the mechanisms mediating the many diverse effects exerted by phorbol esters in other biological systems.