Smythe W Roy, Mohuiddin Imran, Ozveran Mustafa, Cao Xiaobo X
Department of Thoracic and Cardiovascular Surgery, Section of Thoracic Molecular Oncology, University of Texas M.D. Anderson Cancer Center, Houston, 77030, USA.
J Thorac Cardiovasc Surg. 2002 Jun;123(6):1191-8. doi: 10.1067/mtc.2002.121684.
Malignant pleural mesothelioma is resistant to conventional therapies and to apoptosis. The bcl-2 family genes are major determinants of apoptotic homeostasis. Malignant pleural mesothelioma lines and tumors rarely express the antiapoptotic Bcl-2 protein but routinely express the antiapoptotic protein Bcl-xl and the proapoptotic proteins Bax and Bak. We have previously shown pharmacologic inhibition of bcl-xl expression in malignant pleural mesothelioma can lead to apoptosis, so we sought to determine whether antisense oligonucleotides directed at bcl-xl messenger RNA would engender apoptosis, possibly through a "forced imbalance" of bcl-2 family proteins.
Malignant pleural mesothelioma lines REN (epithelial) and I-45 (sarcomatous) were exposed to modified bcl-xl antissense oligonecleotides directed near the messenger RNA initiation sequence with and without a liposomal delivery system. Untreated cells and bcl-xl sense oligonucleotides were controls. Cell viability was measured by colorimetric assay, and apoptosis was evaluated with Hoechst staining and sub-G(1) fluorescence-activated cell sorter analysis.
Bcl-xl protein expression after antisense oligonucleotides was downwardly regulated in both cell lines relative to sense oligonucleotides (>65%). Significant cellular killing in both the I-45 and REN cell lines was achieved with antisense oligonucleotides (compared with sense oligonucleotides) without (P =.003 and.006, respectively) and with (P =.006 and.0005, respectively) liposomal delivery. Hoechst staining and sub-G(1) fluorescence-activated cell sorter analysis demonstrated apoptosis to be the mechanism of cellular death. Use of a liposomal delivery system increased therapeutic effect and allowed lower doses of antisense oligonucleotides.
Antisense oligonucleotides directed at the bcl-xl gene product engender apoptosis in mesothelioma cell lines. The therapeutic potential of inhibiting expression of this protein in mesothelioma should be evaluated.
恶性胸膜间皮瘤对传统疗法及细胞凋亡具有抗性。bcl-2家族基因是细胞凋亡稳态的主要决定因素。恶性胸膜间皮瘤细胞系和肿瘤很少表达抗凋亡蛋白Bcl-2，但通常表达抗凋亡蛋白Bcl-xl以及促凋亡蛋白Bax和Bak。我们之前已表明，对恶性胸膜间皮瘤中bcl-xl表达的药理抑制可导致细胞凋亡，因此我们试图确定针对bcl-xl信使核糖核酸的反义寡核苷酸是否会引发细胞凋亡，可能是通过bcl-2家族蛋白的“强制失衡”。
将恶性胸膜间皮瘤细胞系REN（上皮型）和I-45（肉瘤型）分别暴露于针对信使核糖核酸起始序列附近的修饰bcl-xl反义寡核苷酸，有或无脂质体递送系统。未处理的细胞和bcl-xl正义寡核苷酸作为对照。通过比色法测定细胞活力，并用Hoechst染色和亚G1期荧光激活细胞分选分析评估细胞凋亡。
相对于正义寡核苷酸，反义寡核苷酸处理后，两种细胞系中的Bcl-xl蛋白表达均下调（>65%）。在I-45和REN细胞系中，使用反义寡核苷酸（与正义寡核苷酸相比）均实现了显著的细胞杀伤，无脂质体递送时（分别为P = 0.003和0.006）以及有脂质体递送时（分别为P = 0.006和0.0005）。Hoechst染色和亚G1期荧光激活细胞分选分析表明细胞凋亡是细胞死亡的机制。脂质体递送系统的使用增强了治疗效果，并允许使用更低剂量的反义寡核苷酸。
针对bcl-xl基因产物的反义寡核苷酸可在间皮瘤细胞系中引发细胞凋亡。应评估在间皮瘤中抑制该蛋白表达的治疗潜力。
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