Zhuang Sen H, Hung Y Elizabeth, Hung Laura, Robey Robert W, Sackett Dan L, Linehan W Marston, Bates Susan E, Fojo Tito, Poruchynsky Marianne S
Medical Oncology Branch, Center for Cancer Research, National Cancer Institute, Bethesda, MD 20892, USA.
Clin Cancer Res. 2007 Dec 15;13(24):7480-6. doi: 10.1158/1078-0432.CCR-06-2883.
Microtubule-stabilizing agents, such as taxanes, have been shown to be effective anticancer drugs. alpha-Tubulin, a basic unit of microtubules, can undergo several posttranslational modifications after assembly into stabilized microtubules, including acetylation and detyrosination. These modifications have been observed in cell cultures after exposure to microtubule stabilizers. Our objective was to develop a straightforward and dependable assay to show tubulin target engagement in tumor tissue after treatment of patients with ixabepilone(BMS-247550; Ixempra).
Levels of posttranslationally modified alpha-tubulin were assessed in lysates of cultured malignant cell lines, as well as in both tumor tissue and peripheral blood mononuclear cells derived from patients before and after treatment with ixabepilone. Modification-specific antibodies permitted quantitative Western blot analysis.
In cultured cell lines, the levels of detyrosinated (glu-terminated) and acetylated alpha-tubulin increased after microtubule stabilization induced by ixabepilone. ixabepilone treatment also induced a 2-fold to 25-fold increase in detyrosinated alpha-tubulin levels in 11 of 13 serial biopsies and a 2-fold to 100-fold increase in acetylated alpha-tubulin in 11 of 12 serial biopsies obtained from patients receiving ixabepilone. Overall, little or no difference in tubulin modifications were observed between the before and after ixabepilone treatment in lysates from their peripheral blood mononuclear cells at the time point examined.
Assessing the levels of detyrosinated and/or acetylated alpha-tubulin seems to provide a simple and reliable assay to show target engagement by the microtubule-stabilizing agent ixabepilone. Such analyses may provide further understanding of therapeutic success or failure of microtubule-stabilizing agents in cancer therapy.
微管稳定剂，如紫杉烷类，已被证明是有效的抗癌药物。α-微管蛋白是微管的基本单位，在组装成稳定的微管后可经历多种翻译后修饰，包括乙酰化和去酪氨酸化。在细胞培养物中暴露于微管稳定剂后已观察到这些修饰。我们的目标是开发一种直接且可靠的检测方法，以显示接受伊沙匹隆（BMS - 247550；Ixempra）治疗的患者肿瘤组织中微管蛋白靶点的结合情况。
在培养的恶性细胞系裂解物中，以及在接受伊沙匹隆治疗前后患者的肿瘤组织和外周血单核细胞中，评估翻译后修饰的α-微管蛋白水平。修饰特异性抗体允许进行定量蛋白质印迹分析。
在培养的细胞系中，伊沙匹隆诱导微管稳定后，去酪氨酸化（谷氨酸末端）和乙酰化的α-微管蛋白水平升高。伊沙匹隆治疗还使13例系列活检中的11例去酪氨酸化α-微管蛋白水平增加了2倍至25倍，12例系列活检中的11例接受伊沙匹隆治疗患者的乙酰化α-微管蛋白水平增加了2倍至100倍。总体而言，在所检查的时间点，接受伊沙匹隆治疗前后其外周血单核细胞裂解物中微管蛋白修饰的差异很小或没有差异。
评估去酪氨酸化和/或乙酰化α-微管蛋白的水平似乎提供了一种简单可靠的检测方法，以显示微管稳定剂伊沙匹隆的靶点结合情况。此类分析可能有助于进一步了解微管稳定剂在癌症治疗中的治疗成败。
