Androgen-sensitive Line LNCaP Research Articles

A functional genomics screen reveals a strong synergistic effect between docetaxel and the mitotic gene DLGAP5 that is mediated by the androgen receptor

Based on a molecular classification of prostate cancer using gene expression pathway signatures, we derived a set of 48 genes in critical pathways that significantly predicts clinical outcome in all tested patient cohorts. We tested these genes in a functional genomics screen in a panel of three prostate cancer cell lines (LNCaP, PC3, DU145), using RNA interference. The screen revealed several genes whose knockdown caused strong growth inhibition in all cell lines. Additionally, we tested the gene set in the presence of docetaxel to see whether any gene exhibited additive or synergistic effects with the drug. We observed a strong synergistic effect between DLGAP5 knockdown and docetaxel in the androgen-sensitive line LNCaP, but not in the two other androgen-independent lines. We then tested whether this effect was connected to androgen pathways and found that knockdown of the androgen receptor by si-RNA attenuated the synergy significantly. Similarly, androgen desensitized LNCaP-AI cells had a higher IC50 to docetaxel and did not exhibit the synergistic interaction. Short-term exposure to enzalutamide did not significantly alter the behaviour of parental LNCaP cells. An immunofluorescence analysis in LNCaP cells suggests that under the double insult of DLGAP5 knockdown and docetaxel, cells predominantly arrest in metaphase. In contrast, the knockdown of the androgen receptor by siRNA appears to assist cells to progress through metaphase in to anaphase, even in the presence of docetaxel. Our data suggest that DLGAP5 has a unique function in stabilizing spindle formation and surviving microtubule assault from docetaxel, in an androgen-regulated cell cycle system.

Terminal neuroendocrine differentiation of human prostate carcinoma cells in response to increased intracellular cyclic AMP.

Recent clinicopathologic studies have shown that many prostatic adenocarcinomas express focal neuroendocrine differentiation and that neuroendocrine differentiation is most apparent in advanced anaplastic tumors. While studying growth-regulatory signal transduction events in human prostate carcinoma cell lines, we found that in two of four cell lines, the androgen-sensitive line LNCaP and the highly metastatic androgen-independent line PC-3-M, elevation of cAMP through addition of cAMP analogues or phosphodiesterase inhibitors induced a markedly neuronal morphology. Also in LNCaP cells ultrastructural analysis showed that cAMP induced the appearance of neurosecretory cell-like dense-core granules. Phenotypic analysis of untreated LNCaP and PC-3-M cells showed that both cell lines express markers of the neural crest including S-100, chromogranin A, pp60c-src, and neuron-specific enolase as well as the epithelial marker KS1/4 and stage-specific embryonic antigen 4. In PC-3-M cells, cAMP markedly elevated neuron-specific enolase protein and caused an increase in the specific activity of the neuroendocrine marker pp60c-src, and in both cell lines expression of KS1/4 and stage-specific embryonic antigen 4 was down-regulated. In addition to effects on lineage markers, cAMP treatment induced G1 synchronization, growth arrest, and loss of clonogenicity, indicating terminal differentiation. Our data provide direct evidence of plasticity in the lineage commitment of adenocarcinoma of the prostate. We have shown that cell-permeant cAMP analogues can induce terminal differentiation, suggesting that hydrolysis-resistant cyclic nucleotides may present an additional approach to the treatment of advanced prostate cancer.