Identification of a candidate gene associated with paclitaxel-resistance and application to a gene therapy in ovarian cancer.

我们通过在荷瘤小鼠体内重复给药紫杉醇，从亲代人卵巢癌细胞系中建立了紫杉醇耐药细胞系。我们在体内建立的紫杉醇耐药细胞系和体内建立的对照组之间进行了基因芯片，以确定与紫杉醇耐药相关的靶基因。(1)从体内建立的紫杉醇耐药细胞系和对照组中提取总RNA，(2)从紫杉醇耐药细胞或对照细胞皮下移植制备的肿瘤块(OM1TX6T和OM1P6T)中提取总RNA，纯化cDNA，鉴定两组之间特异性增加或减少的肿瘤相关基因表达谱。我们用来分析的基因(1)是9,063个，在抗性细胞系中表达的基因数量是对照细胞系的两倍多。此外，我们用于分析(2)的基因有8,660个，同样地，OM1TX6T表达超过OM1P6T一倍的基因数量为298个。现在我们真的在推动从这些基因中提取与紫杉醇耐药相关的基因。另一方面，第三类B-微管蛋白的过度表达作为紫杉醇耐药的机制引起了人们的关注。应用免疫组织化学方法检测卵巢癌组织中I、II、II、IV(IVa+IVb)亚型B-微管蛋白的蛋白表达水平，并探讨其与紫杉烷类药物化疗疗效的关系。结果表明，在接受紫杉烷化疗的组中，B-微管蛋白Ⅲ类亚型的高表达倾向于与短时间的无突起生存相关(p=0.081)，而在不接受紫杉烷化疗的组中没有发现这种趋势。

We established an in vivo paclitaxel-resistant cell line from the parental human ovarian cancer cell lines by repeated paclitaxel administration into tumor-bearing mice. We performed cDNA microarrays between in vivo established paclitaxel-resistant cell line and in vivo established control group to identify the target genes associated with paclitaxel-resistancy. Total RNAs were extracted (1) from in vivo established paclitaxel-resistant cell lines and control group, and (2) from tumor blocks (OM1TX6T and OM1P6T)which were made by subcutaneous transplantion of paclitaxel-resistant cells or control cells.cDNA were purified and cancer-related gene manifestation profiles which were specifically increased or decreased between two groups were identified. Genes which we used for analysis (1) were 9,063, and the number of the genes which expressed in a resistant cell lines more than double of control cell lines. In addition, genes which we used for analysis (2) were 8,660,and similarly the number of the genes which expressed OM1TX6T more than double of OM1P6T was 298. Now we really push forward extraction of genes related to paclitaxel-resistance from these genes.On the other hand, overexpression of class III B-tubulin was attracted attention as mechanism of taxane-resistance. We evaluated protein expression level of class I, II, II, IV (IVa+IVb) isotypes of B-tubulin in an ovarian cancer specimens by immunohistochemical stainig, and examined the association with responce to taxane-containing chemotherapy. As a result, in the goup which had chemotherapy with taxane, high expression of class III isotypes of B-tubulin tend to correlate with short prograssion free survival (p=0.081), while this tendency was not found in the group which had chemotherapy without taxane.