Bischof Marc, Abdollahi Amir, Gong Ping, Stoffregen Clemens, Lipson Kenneth E, Debus J Urgen, Weber Klaus J, Huber Peter E
Department of Radiation Oncology, University of Heidelberg, Medical School, Heidelberg, Germany.
Int J Radiat Oncol Biol Phys. 2004 Nov 15;60(4):1220-32. doi: 10.1016/j.ijrobp.2004.07.689.
This is the first preclinical report evaluating a trimodal therapy consisting of irradiation, chemotherapy, and antiangiogenesis in the context of a multimodal anticancer strategy. The combination of the folate antimetabolite pemetrexed, SU5416, a receptor tyrosine kinase inhibitor of VEGFR2, and irradiation was investigated in human endothelial cells and tumor cell lines.
Primary isolated human umbilical vein endothelial cells (HUVEC), human dermal microvascular endothelial cells (HDMEC), and human glioblastoma (U87) and prostate cancer cells (PC3) were exposed to pemetrexed (2 h) alone and in combination with SU5416 (2 h). When combined with irradiation up to 8 Gy, fixed concentrations of pemetrexed (1.06 muM) and SU5416 (1.0 muM) were used. Proliferation and clonogenic assays were conducted with endothelial and tumor cells. The migration/invasion ability of endothelial cells and the ability to produce tubular structures were tested in Matrigel and tube formation assays. Apoptosis was measured by sub-G1 DNA and caspase-3 flow cytometry. To investigate underlying cell signaling, immunocytochemistry was used to detect Akt survival signaling involvement.
Triple combination using only a low-toxicity drug exposure of pemetrexed and SU5416 results in greater response than each treatment alone or than each combination of two modalities in all tested endothelial and tumor cell models. Triple combination substantially inhibits proliferation, migration/invasion, tube formation, and clonogenic survival. Triple combination also induced the highest rate of apoptosis in HDMEC and HUVEC as indicated by sub-1 G1 and caspase-3 assessment. Interestingly, triple combination therapy also reduces proliferation and clonogenic survival significantly in U87 and PC3 tumor cell lines. SU5416 potently inhibited Akt phosphorylation which could be induced by radiation and radiochemotherapy in human endothelial cells.
Our findings demonstrate the high antiendothelial/antitumoral efficacy of the concurrent administration of irradiation, chemotherapy, and angiogenesis inhibition in vitro. A potential explanation for the favorable combination would be that VEGF signaling inhibition downregulates Akt survival signaling upon activation by radiation and/or chemotherapy. The data also suggest that endothelial cell apoptosis may have an important role in the benefits of the presented therapy.
这是首篇在多模式抗癌策略背景下评估由放疗、化疗和抗血管生成组成的三联疗法的临床前报告。研究了叶酸抗代谢物培美曲塞、VEGFR2的受体酪氨酸激酶抑制剂SU5416与放疗联合应用于人类内皮细胞和肿瘤细胞系的情况。
将原代分离的人脐静脉内皮细胞（HUVEC）、人真皮微血管内皮细胞（HDMEC）、人胶质母细胞瘤（U87）和前列腺癌细胞（PC3）单独及联合SU5416（2小时）暴露于培美曲塞（2小时）。当与高达8 Gy的放疗联合时，使用固定浓度的培美曲塞（1.06 μM）和SU5416（1.0 μM）。对内皮细胞和肿瘤细胞进行增殖和克隆形成试验。在基质胶和管形成试验中检测内皮细胞的迁移/侵袭能力以及形成管状结构的能力。通过亚G1期DNA和半胱天冬酶-3流式细胞术检测细胞凋亡。为研究潜在的细胞信号传导，采用免疫细胞化学检测Akt存活信号传导的参与情况。
在所有测试的内皮细胞和肿瘤细胞模型中，仅使用低毒性药物培美曲塞和SU5416进行三联组合比单独的每种治疗或两种模式的每种组合产生更大的反应。三联组合显著抑制增殖、迁移/侵袭、管形成和克隆存活。如亚G1期和半胱天冬酶-3评估所示，三联组合在HDMEC和HUVEC中也诱导了最高的凋亡率。有趣的是，三联组合疗法在U87和PC3肿瘤细胞系中也显著降低了增殖和克隆存活。SU5416有效抑制了Akt磷酸化，而Akt磷酸化可由放疗和放化疗在人类内皮细胞中诱导产生。
我们的研究结果表明，放疗、化疗和血管生成抑制同时给药在体外具有很高的抗内皮/抗肿瘤疗效。这种有利组合的一个潜在解释可能是，VEGF信号传导抑制在放疗和/或化疗激活后下调了Akt存活信号传导。数据还表明，内皮细胞凋亡可能在所述疗法的益处中起重要作用。
