Sun Xiaorong, Ackerstaff Ellen, He Fuqiu, Xing Ligang, Hsiao Hung Tsung, Koutcher Jason A, Ling C Clifton, Li Gloria C
Department of Radiology, Shandong Key Laboratory of Radiation Oncology, Shandong Cancer Hospital and Institute, Jinan, Shandong, China.
Department of Radiation Oncology, Memorial Sloan Kettering Cancer Center, New York, NY, USA.
Oncotarget. 2015 Oct 27;6(33):34732-44. doi: 10.18632/oncotarget.5300.
Bortezomib, a novel proteasome inhibitor, has been approved for treating multiple myeloma and mantle cell lymphoma and studied pre-clinically and clinically for solid tumors. Preferential cytotoxicity of bortezomib was found toward hypoxic tumor cells and endothelial cells in vitro. The purpose of this study is to investigate the role of a pretreatment hypoxic tumor microenvironment on the effects of bortezomib in vitro and ex vivo, and explore the feasibility of dynamic contrast enhanced magnetic resonance imaging (DCE MRI) to noninvasively evaluate the biological effects of bortezomib. It was shown in vitro by Western blot, flow cytometry, and ELISA that bortezomib accumulated HIF-1α in non-functional forms and blocks its hypoxia response in human colorectal cancer cell lines. Ex vivo experiments were performed with fluorescent immunohistochemical staining techniques using multiple endogenous and exogenous markers to identify hypoxia (pimonidazole, HRE-TKeGFP), blood flow/permeability (Hoechst 33342), micro-vessels (CD31 and SMA), apoptosis (cleaved caspase 3) and hypoxia response (CA9). After bortezomib administration, overall apoptosis index was significantly increased and blood perfusion was dramatically decreased in tumor xenografts. More importantly, apoptosis signals were found preferentially located in moderate and severe pretreatment hypoxic regions in both tumor and endothelial cells. Meanwhile, DCE MRI examinations showed that the tumor blood flow and permeability decreased significantly after bortezomib administration. The present study revealed that bortezomib reduces tumor hypoxia response and blood perfusion, thus, presenting antivascular properties. It will be important to determine the hypoxic/perfusion status pre- and during treatment at further translational studies.
硼替佐米，一种新型蛋白酶体抑制剂，已被批准用于治疗多发性骨髓瘤和套细胞淋巴瘤，并已针对实体瘤进行了临床前和临床研究。体外研究发现硼替佐米对缺氧肿瘤细胞和内皮细胞具有优先细胞毒性。本研究的目的是探讨预处理缺氧肿瘤微环境对硼替佐米体外和离体效应的作用，并探索动态对比增强磁共振成像（DCE MRI）无创评估硼替佐米生物学效应的可行性。通过蛋白质免疫印迹法、流式细胞术和酶联免疫吸附测定法在体外表明，硼替佐米以无功能形式积累缺氧诱导因子-1α（HIF-1α），并阻断其在人结肠癌细胞系中的缺氧反应。采用多种内源性和外源性标记物的荧光免疫组织化学染色技术进行离体实验，以识别缺氧（匹莫硝唑、缺氧反应元件-TKeGFP）、血流/通透性（Hoechst 33342）、微血管（CD31和平滑肌肌动蛋白）、凋亡（裂解的半胱天冬酶3）和缺氧反应（碳酸酐酶9）。给予硼替佐米后，肿瘤异种移植物的总体凋亡指数显著增加，血流灌注显著降低。更重要的是，在肿瘤细胞和内皮细胞中均发现凋亡信号优先位于预处理中度和重度缺氧区域。同时，DCE MRI检查显示，给予硼替佐米后肿瘤血流和通透性显著降低。本研究表明，硼替佐米可降低肿瘤缺氧反应和血流灌注，从而呈现抗血管生成特性。在进一步的转化研究中，确定治疗前和治疗期间的缺氧/灌注状态将非常重要。
