Gao A C, Lou W, Isaacs J T
The Johns Hopkins Oncology Center, James Buchanan Brady Urological Institute, Department of Urology, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21231, USA.
Cancer Res. 1998 Apr 1;58(7):1391-4.
Previously, we have demonstrated that GBX genes, a homeobox-containing human family of DNA-binding transcription factors consisting of GBX1 and GBX2, are overexpressed in a panel of human prostatic cancer cell lines (ie., TSU-pr1, PC3, DU145, and LNCaP) compared to normal prostate. In the present studies, specific primer sets were designed for reverse transcription-PCR detection of the expression of GBX1 versus GBX2 in human prostate cancer. These studies demonstrated that the GBX2 gene, but not the GBX1 gene, is consistently overexpressed in this panel of human prostate cancer cell lines compared to normal human prostate. Using a quantitative-competitive PCR analysis, GBX2 mRNA was expressed as 3 x 10(3) copies/microg RNA in normal prostate tissue and 4 x 10(4) copies/microg RNA in the immortalized normal neonatal prostate epithelial cell line 267B-1, as compared to 6 x 10(5), 5 x 10(5), 3 x 10(5), and 1 x 10(5) copies/microg RNA in TSU-pr1, DU145, LNCaP, and PC3 prostate cancer cell lines, respectively. To examine the importance of GBX2 expression for prostate cancer malignancy, GBX2-overexpressing TSU-pr1 and PC3 human prostatic cancer cells were transfected with a eukaryotic expression vector containing an antisense GBX2 homeobox domain cDNA. Stable transfectant clones with 5-10-fold decreased levels of GBX2 mRNA expression were obtained. When tested in vitro, the clonogenic ability of the GBX2 antisense transfectants was reduced by approximately 50% in both cell lines. When implanted s.c. into nude mice, the tumorigenicity of the antisense GBX2 transfectants from both human prostatic cancer cell lines was inhibited by more than 70% compared to the parental cells. These results suggest that expression of GBX2 gene is required for malignant growth of human prostate cells.
此前，我们已经证明，GBX基因是一个含同源异型盒的人类DNA结合转录因子家族，由GBX1和GBX2组成，与正常前列腺相比，在一组人类前列腺癌细胞系（即TSU-pr1、PC3、DU145和LNCaP）中过表达。在本研究中，设计了特异性引物组用于逆转录-PCR检测人前列腺癌中GBX1与GBX2的表达。这些研究表明，与正常人类前列腺相比，GBX2基因而非GBX1基因在这组人类前列腺癌细胞系中持续过表达。使用定量竞争PCR分析，GBX2 mRNA在正常前列腺组织中的表达量为3×10³拷贝/μg RNA，在永生化的正常新生儿前列腺上皮细胞系267B-1中为4×10⁴拷贝/μg RNA，而在TSU-pr1、DU145、LNCaP和PC3前列腺癌细胞系中的表达量分别为6×10⁵、5×10⁵、3×10⁵和1×10⁵拷贝/μg RNA。为了研究GBX2表达对前列腺癌恶性程度的重要性，用含有反义GBX2同源异型盒结构域cDNA的真核表达载体转染过表达GBX2的TSU-pr1和PC3人前列腺癌细胞。获得了GBX2 mRNA表达水平降低5至10倍的稳定转染克隆。在体外测试时，两个细胞系中GBX2反义转染子的克隆形成能力均降低了约50%。当皮下接种到裸鼠体内时，来自两个人前列腺癌细胞系的反义GBX2转染子的致瘤性与亲本细胞相比受到了70%以上的抑制。这些结果表明，GBX2基因的表达是人类前列腺细胞恶性生长所必需的。
