Resistance to arginine deiminase treatment in melanoma cells is associated with induced argininosuccinate synthetase expression involving c-Myc/HIF-1alpha/Sp4.

Abstract

Arginine deiminase (ADI)-based arginine depletion is a novel strategy under clinical trials for the treatment of malignant melanoma with promising results. The sensitivity of melanoma to ADI treatment is based on its auxotrophy for arginine due to a lack of argininosuccinate synthetase (AS) expression, the rate-limiting enzyme for the de novo biosynthesis of arginine. We show here that AS expression can be transcriptionally induced by ADI in melanoma cell lines A2058 and SK-MEL-2 but not in A375 cells, and this inducibility was correlated with resistance to ADI treatment. The proximal region of the AS promoter contains an E-box that is recognized by c-Myc and HIF-1alpha and a GC-box by Sp4. Through ChIP assays, we showed that under noninduced conditions, the E-box was bound by HIF-1alpha in all the three melanoma cell lines. Under arginine depletion conditions, HIF-1alpha was replaced by c-Myc in A2058 and SK-MEL-2 cells but not in A375 cells. Sp4 was constitutively bound to the GC-box regardless of arginine availability in all three cell lines. Overexpressing c-Myc by transfection upregulated AS expression in A2058 and SK-MEL-2 cells, whereas cotransfection with HIF-1alpha suppressed c-Myc-induced AS expression. These results suggest that regulation of AS expression involves interplay among positive transcriptional regulators c-Myc and Sp4, and negative regulator HIF-1alpha that confers resistance to ADI treatment in A2058 and SK-MEL-2 cells. Inability of AS induction in A375 cells under arginine depletion conditions was correlated by the failure of c-Myc to interact with the AS promoter.

• Authors: Wen-Bin Tsai, Isamu Aiba, Soo-yong Lee, Lynn Feun, Niramol Savaraj, Macus Tien Kuo
• Journal: Molecular cancer therapeutics (Mol Cancer Ther) Vol. 8 Issue 12 Pg. 3223-33 (Dec 2009) ISSN: 1538-8514 [Electronic] United States
• PMID: 19934275 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, Non-P.H.S.)

Chemical References

• Hypoxia-Inducible Factor 1, alpha Subunit
• Proto-Oncogene Proteins c-myc
• SP4 protein, human
• Sp4 Transcription Factor
• Polyethylene Glycols
• Arginine
• Hydrolases
• arginine deiminase
• Argininosuccinate Synthase

Topics

• Arginine (metabolism)
• Argininosuccinate Synthase (genetics, metabolism)
• Base Sequence
• Binding Sites (genetics)
• Cell Line, Tumor
• Chromatin Immunoprecipitation
• Drug Resistance, Neoplasm
• Gene Expression Regulation, Enzymologic (drug effects)
• Gene Expression Regulation, Neoplastic (drug effects)
• Humans
• Hydrolases (chemistry, pharmacology)
• Hypoxia-Inducible Factor 1, alpha Subunit (genetics, metabolism)
• Immunoblotting
• Melanoma (genetics, metabolism, pathology)
• Polyethylene Glycols (chemistry)
• Promoter Regions, Genetic (genetics)
• Protein Binding
• Proto-Oncogene Proteins c-myc (genetics, metabolism)
• RNA Interference
• Reverse Transcriptase Polymerase Chain Reaction
• Sp4 Transcription Factor (genetics, metabolism)