Prostate Cancer Cells (LNCaP) Generated after Long-Term Interleukin 6 (IL-6) Treatment Express IL-6 and Acquire an IL-6 Partially Resistant Phenotype

• Published in: Clinical cancer research 2001-09, Vol.7 (9), p.2941-2948
• Main Authors: HOBISCH, Alfred, RAMONER, Reinhold, FUCHS, Dietmar, GODOY-TUNDIDOR, Sonia, BARTSCH, Georg, KLOCKER, Helmut, CULIG, Zoran
• Format: Article
• Language: English
• Subjects: Androgens - metabolism; Androgens - pharmacology; Binding, Competitive; Biological and medical sciences; Cell Count; Cell Cycle - drug effects; Cell Division - drug effects; Cell physiology; Cell transformation and carcinogenesis. Action of oncogenes and antioncogenes; Dose-Response Relationship, Drug; Drug Resistance, Neoplasm; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation, Neoplastic; Humans; Interleukin-6 - genetics; Interleukin-6 - metabolism; Interleukin-6 - pharmacology; Male; Metribolone - metabolism; Metribolone - pharmacology; Molecular and cellular biology; Prostate-Specific Antigen - drug effects; Prostate-Specific Antigen - secretion; Prostatic Neoplasms - drug therapy; Prostatic Neoplasms - genetics; Prostatic Neoplasms - pathology; RNA, Messenger - genetics; RNA, Messenger - metabolism; Signal Transduction; Tritium; Tumor Cells, Cultured - drug effects; Tumor Cells, Cultured - metabolism
• ISSN: 1078-0432; 1557-3265

Purpose : The levels of interleukin-6 (IL-6) are frequently elevated in sera from patients with advanced prostate carcinoma. Our main objective was to investigate changes in responsiveness to IL-6 and/or androgen that occur in LNCaP cells after long-term treatment with IL-6. This in vitro model could be of clinical relevance because of its similarity with late-stage prostate carcinoma. Experimental Design: LNCaP human prostate cancer cells were treated with IL-6 at a concentration of 5 ng/ml. After 20 passages, the new subline LNCaP-IL-6+ has been established. Passages 20–40 are referred to as low passages (LP) and passages 41–73 as high passages (HP). LNCaP cells passaged at the same time in the absence of IL-6 were used as controls (LNCaP-IL-6−). Cells were counted after treatment with either IL-6 or the synthetic androgen methyltrienolone (R1881), and cell cycle analysis was performed. Binding of IL-6 or R1881 was assessed by radioligand binding assays. Reporter gene activity was measured by chloramphenicol acetyltransferase assay. Prostate-specific antigen in LNCaP-IL-6+ supernatants was measured by an enzyme immunoassay. Expression of IL-6 mRNA and protein was assessed by reverse transcription-PCR and ELISA, respectively. Results: The basal proliferation rate in HP LNCaP-IL-6+ cells was higher than that in LNCaP-IL-6− cells. IL-6 inhibited proliferation of LNCaP-IL-6− cells but not that of either LP or HP of LNCaP-IL-6+ cells. This inability to elicit a growth-inhibitory response was associated with lack of effect on cell cycle distribution in the LNCaP-IL-6+ subline. In parallel, IL-6 binding decreased gradually during long-term IL-6 treatment and, in HP, reached only 33% of the levels measured in controls. Binding of radiolabeled androgen increased 2-fold in HP LNCaP-IL-6+ cells. Reporter gene assays revealed that R1881, at nanomolar concentrations, was a more potent androgen receptor activator in LNCaP-IL-6+ than in LNCaP-IL-6− cells. However, androgen- and IL-6-induced prostate-specific antigen secretion decreased in long-term IL-6-treated cells. IL-6 cDNA fragments were detected by reverse transcription-PCR in HP LNCaP-IL-6+ cells but not in controls or LP. IL-6 protein was first detected in passage 36 of LNCaP-IL-6+ cells, and it increased in HP. Conclusions: Long-term treatment of LNCaP human prostate cancer cells with IL-6 leads to abolishment of inhibitory growth response. In contrast to control cells, the LNCaP-IL-6+ subline expresses IL-6 m