Intratumoral activation and enhanced chemotherapeutic effect of oxazaphosphorines following cytochrome P-450 gene transfer: development of a combined chemotherapy/cancer gene therapy strategy.
Intratumoral activation and enhanced chemotherapeutic effect of oxazaphosphorines following cytochrome P-450 gene transfer: development of a combined chemotherapy/cancer gene therapy strategy.
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发表时间：
1995-02
期刊：
影响因子：
11.2
通讯作者：
L. Chen;D. Waxman
中科院分区：
文献类型：
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作者：
L. Chen;D. Waxman
环磷酰胺及其异环磷酰胺是由肝细胞色素P-450酶催化的细胞周期非特异性烷基化试剂。这些恶唑磷抗癌药物的治疗效果受到宿主毒性的限制，宿主毒性是由肝脏中形成的活性药物代谢物的系统分布引起的。由于肿瘤细胞通常不具有激活恶唑磷的能力，我们研究了将编码口恶唑磷激活的主要催化剂CYP2B1的基因导入肿瘤细胞是否使细胞对环磷酰胺和异环磷酰胺的细胞毒性作用敏感。与亲本9L细胞或转导大肠杆菌β-半乳糖苷酶基因的9L细胞相比，稳定转基因的9L胶质肉瘤细胞对环磷酰胺和异环磷酰胺的细胞毒性高度敏感。细胞色素P450 2B1酶抑制剂美曲拉酮可保护表达该酶的9L细胞免受恶唑磷的细胞毒作用，表明这些细胞对化疗药物的敏感性是细胞内前体药物激活的直接结果。此外，表达CYP2B1的9L细胞可增强环磷酰胺和异环磷酰胺对共培养的阴性9L肿瘤细胞的细胞毒作用。这种“旁观者效应”不需要细胞间的接触，因此可能具有将细胞毒性药物代谢物分布到实体瘤内的大范围内的治疗优势。体内实验采用Fischer 344大鼠植入S.C.研究表明，与肝脏细胞色素P-450依赖的药物单独激活相比，在9L肿瘤细胞内表达细胞色素P450基因具有显著的治疗优势；环磷酰胺治疗可完全抑制细胞色素P450阳性肿瘤的生长，而对细胞色素P450阴性肿瘤仅有轻微的生长延迟效应。这些研究表明，药物激活的CYP基因可能有助于利用已有的癌症化疗药物开发新的化疗/基因联合治疗癌症的策略。
Cyclophosphamide and its isomer ifosfamide are cell cycle-nonspecific alkylating agents that undergo bioactivation catalyzed by liver cytochrome P-450 enzymes. The therapeutic efficacy of these oxazaphosphorine anticancer drugs is limited by host toxicity resulting from the systemic distribution of activated drug metabolites formed in the liver. Since tumor cells ordinarily do not have the capacity to activate oxazaphosphorines, we examined whether introduction into tumor cells of a cDNA encoding CYP2B1, a major catalyst of oxazaphosphorine activation, sensitizes the cells to the cytotoxic effects of cyclophosphamide and ifosfamide. Here we show that 9L gliosarcoma cells stably transfected with a cDNA encoding rat CYP2B1 are highly sensitive to cyclophosphamide and ifosfamide cytotoxicity as compared to parental 9L cells or 9L cells transfected with an Escherichia coli beta-galactosidase gene. The CYP2B1 enzyme inhibitor metyrapone protects the CYP2B1-expressing 9L cells from oxazaphosphorine cytotoxicity, demonstrating that the chemosensitivity of these cells is a direct consequence of intracellular prodrug activation. Moreover, CYP2B1-expressing 9L cells potentiate the cytotoxic effects of cyclophosphamide and ifosfamide toward cocultured CYP2B1-negative 9L tumor cells. This "bystander effect" does not require cell-cell contact, and therefore may have the therapeutic advantage of distributing cytotoxic drug metabolites to a wide area within a solid tumor mass. In vivo experiments using Fischer 344 rats implanted s.c. with CYP2B1-expressing 9L tumor cells demonstrated that intratumoral expression of the CYP2B1 gene provides a substantial therapeutic advantage over that provided by liver cytochrome P-450-dependent drug activation alone; cyclophosphamide treatment resulted in complete growth inhibition of CYP2B1-positive tumors, whereas only a modest growth delay effect was obtained with CYP2B1-negative tumors. These studies establish that drug-activating CYP genes may be useful for the development of novel combined chemotherapy/gene therapy strategies for cancer treatment utilizing established cancer chemotherapeutic agents.