Chen Min-Wei, Vacherot Francis, De La Taille Alexandre, Gil-Diez-De-Medina Sixtina, Shen Ruoqian, Friedman Richard A, Burchardt Martin, Chopin Dominique K, Buttyan Ralph
The Department of Urology, The College of Physicians and Surgeons of Columbia University, New York, NY 10032, USA.
Oncogene. 2002 Nov 7;21(51):7861-71. doi: 10.1038/sj.onc.1205991.
In order to identify gene products associated with the development of acquired therapeutic resistance by prostate cancer cells, we created two novel apoptosis-resistant prostate cancer cell lines, LNCaP-TR (phorbol-ester [TPA]-Resistant) and LNCaP-SSR (Serum Starvation-Resistant) by repeated transient exposure of cultured human LNCaP cells to apoptotic stimuli followed by expansion of surviving cell populations. These cell lines were found to be cross-resistant to the alternative selective agent and also hormone-resistant when xenografted into castrated male immunodeficient mice. RNA from the LNCaP-TR line was comparatively screened using a subtractive hybridization-PCR procedure. This allowed us to identify a 249 bp cDNA fragment that hybridized to a 4.8 kb mRNA preferentially expressed by the apoptosis-resistant cells. Using RACE procedures, we cloned and sequenced the complete 4.8 kb cDNA. It is an unusual member of the protocadherin gene family containing two large overlapping open reading frames encoding homologous polypeptides, one having a signal sequence and the other lacking a signal sequence and we refer to it as protocadherin-PC. LNCaP cells directly transformed with protocadherin-PC cDNA were comparatively resistant to phorbol-ester induced apoptosis. Antibody recognition studies demonstrating the cytoplasmic nature of the protcadherin-PC translation product and its propensity to bind beta-catenin suggest that it might influence the apoptotic sensitivity of prostate cancer cells through a unique mechanism.
为了鉴定与前列腺癌细胞获得性治疗抗性发展相关的基因产物，我们通过将培养的人LNCaP细胞反复短暂暴露于凋亡刺激，随后扩增存活的细胞群体，创建了两种新型的抗凋亡前列腺癌细胞系，LNCaP-TR（佛波酯[TPA]抗性）和LNCaP-SSR（血清饥饿抗性）。当将这些细胞系异种移植到去势的雄性免疫缺陷小鼠中时，发现它们对替代选择剂具有交叉抗性，并且对激素也具有抗性。使用消减杂交-PCR程序对LNCaP-TR系的RNA进行了比较筛选。这使我们能够鉴定出一个249 bp的cDNA片段，该片段与抗凋亡细胞优先表达的4.8 kb mRNA杂交。使用RACE程序，我们克隆并测序了完整的4.8 kb cDNA。它是原钙黏蛋白基因家族的一个不寻常成员，包含两个大的重叠开放阅读框，编码同源多肽，一个具有信号序列，另一个缺乏信号序列，我们将其称为原钙黏蛋白-PC。用原钙黏蛋白-PC cDNA直接转化的LNCaP细胞对佛波酯诱导的凋亡具有相对抗性。抗体识别研究表明原钙黏蛋白-PC翻译产物的细胞质性质及其与β-连环蛋白结合的倾向，表明它可能通过一种独特的机制影响前列腺癌细胞的凋亡敏感性。
