Timed sequential therapy of acute leukemia with flavopiridol: in vitro model for a phase I clinical trial.
Timed sequential therapy of acute leukemia with flavopiridol: in vitro model for a phase I clinical trial.
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发表时间：
2003
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通讯作者：
J. Karp;D. Ross;Weidong Yang;M. Tidwell;Yuetong Wei;J. Greer;D. Mann;T. Nakanishi;J. Wright;A. D. Colevas
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作者：
J. Karp;D. Ross;Weidong Yang;M. Tidwell;Yuetong Wei;J. Greer;D. Mann;T. Nakanishi;J. Wright;A. D. Colevas
目的：成人急性白血病患者的生存率仍然不令人满意，需要新的治疗方法。黄吡醇调节细胞周期进程，抑制转录，诱导细胞凋亡。我们设计了一种急性白血病定时顺序治疗的体外模型，以确定黄匹吡醇是否能：(a)触发新鲜急性白血病细胞凋亡；(b)使存活的白血病细胞进入增殖状态，从而使这些细胞产生与s期相关的1- β - d -阿拉伯糖醛基胞嘧啶（ara-C）的细胞毒性。实验设计：采用Ficoll Hypaque沉淀法富集20例复发和难治性急性白血病成人骨髓细胞，培养成原细胞。细胞于第0天在黄酮吡醇250 nM中培养24 h，与黄酮吡醇分离24 h，然后在ara-C 1 microM中再培养72 h (F（250)A(1)）。碘化丙啶染色后细胞凋亡和细胞周期期分布。用双醋酸荧光素和碘化丙啶双细胞荧光法测定暴露于ara-C 72 h后的细胞存活率。结果黄吡醇在培养后24小时内诱导人白血病细胞凋亡增加4.3倍。随后去除黄匹吡醇导致第2天处于S期的细胞比例增加1.7倍。暴露于ara-C 72小时后，F(250)A(1)培养液的平均存活率为35.6%，与单独使用黄酮吡醇相比(F(250)A(0)为56.1%；P = 0.0003)和单独使用ara-C (F(0)A(1)，占65.2%；P < 0.00001)。结论黄吡醇可诱导难治性急性白血病患者骨髓母细胞凋亡。此外，黄吡醇预处理增加了ara-C的促凋亡和细胞毒性作用。序列FP(250)A(1)治疗急性髓性白血病和急性淋巴细胞白血病优于单独使用任何一种药物。这些发现支持了一项定时序贯治疗的临床试验，其中黄吡醇用于细胞减少，随后启动剩余的白血病细胞以增强周期依赖性药物的细胞毒性。
PURPOSE The survival of adults with acute leukemias remains unsatisfactory and requires new treatment approaches. Flavopiridol modulates cell cycle progression, inhibits transcription, and induces apoptosis. We designed an in vitro model of timed sequential therapy for acute leukemia to determine whether flavopiridol can: (a). trigger apoptosis in fresh acute leukemia; and (b). recruit surviving leukemic cells to a proliferative state, thereby priming such cells for the S-phase-related cytotoxicity of 1-beta-D-arabinofuranosylcytosine (ara-C). EXPERIMENTAL DESIGN Bone marrow cells from 20 adults with relapsed and refractory acute leukemias were enriched for blasts by Ficoll Hypaque sedimentation. Blasts were cultured on day 0 in flavopiridol 250 nM for 24 h, removed from flavopiridol for 24 h, and then cultured in ara-C 1 microM for an additional 72 h (F(250)A(1)). Apoptosis and cell cycle phase distribution were estimated from cells stained with propidium iodide. Cell survival was determined after the 72 h ara-C exposure by double cytofluorescence assay with fluorescein diacetate and propidium iodide. RESULTS Flavopiridol induced a 4.3-fold increase in apoptosis in human leukemia samples within the first 24 h of culture. Subsequent removal of flavopiridol led to a 1.7-fold increase in the proportion of cells in S phase by day 2. Mean survival in F(250)A(1) cultures after 72 h exposure to ara-C was 35.6% compared with flavopiridol alone (F(250)A(0), 56.1%; P = 0.0003) and ara-C alone (F(0)A(1), 65.2%; P < 0.00001). CONCLUSIONS Flavopiridol induces apoptosis in marrow blasts from patients with refractory acute leukemias. Furthermore, flavopiridol pretreatment increases the proapoptotic and cytotoxic effects of ara-C. The advantage of sequential FP(250)A(1) over either agent alone is seen for both acute myelogenous leukemia and acute lymphoblastic leukemia. These findings support a clinical trial of timed sequential therapy where flavopiridol is given for cytoreduction and subsequent priming of remaining leukemic cells for enhanced cycle-dependent drug cytotoxicity.