Abstract LB-254: PRMT5 is upregulated in malignant and metastatic melanoma, and regulates expression of the MITF transcription factor

Abstract

Abstract Protein arginine methyltransferase 5 (PRMT5) is an enzyme which catalyzes the covalent attachment of methyl groups to arginine residues of various proteins. PRMT5 binds and/or methylates both nuclear (e.g. histones) and cytoplasmic (e.g. p53, CRAF) proteins to regulate cellular functions including gene expression, cell cycle, and apoptosis among others. PRMT5 has been characterized in leukemia, lymphoma, glioma, and breast cancer; however little is known regarding its role in malignant melanoma. We hypothesized that PRMT5 plays a unique role in regulating melanoma cell biology. PRMT5 expression was measured by IHC analysis of formalin-fixed samples from patients with melanoma (n=133 primary; n=66 metastases), benign nevi (n=24), and normal adjacent epidermis (n=21). PRMT5 expression was significantly elevated in melanoma samples compared to normal adjacent epidermis (p<0.001, Fisher's exact test). The nuclear and cytoplasmic localization of PRMT5 was heterogeneous within individual melanoma specimens. PRMT5 was significantly higher in the nucleus, but not the cytoplasm, of malignant tissue compared to metastatic tissue (p=0.0004 vs p=0.6, Wilcoxon rank sum). PRMT5 protein was also significantly elevated in benign melanocytic nevi (compound, junctional, and intradermal) compared to normal adjacent epidermis (p<0.0001), suggesting that elevated PRMT5 may represent an early event in tumorigenesis. Analysis of human melanoma cell lines revealed that PRMT5 expression was predominantly cytoplasmic and its expression did not differ based on B-Raf or N-Ras mutational status. Co-immunoprecipitation experiments revealed PRMT5 associated with its enzymatic co-factor MEP-50, but not cyclin D1 or STAT3, in both bulk cultured or synchronized cells, as has been reported in other cell types. Depletion of PRMT5 with siRNA led to decreased transcript and protein level of MITF-M (microphthalmia-associated transcription factor) in human melanoma cell lines with V600E-mutated B-Raf (A375, Hs294T, 1106 MEL, FO1), or wild-type B-Raf (CHL-1, Wm1366). Furthermore, depletion of PRMT5 led to a significant growth inhibition in 1106 MEL, Wm1366, and FO1 cells at 48 and 72 hours post-transfection. These data uncover important differences between the biology of PRMT5 in melanoma cells as compared to other tumor types, and are the first to demonstrate that PRMT5 is a regulator of MITF expression, a key transcription factor regulating melanoma cell biology and cell proliferation. The upregulation of PRMT5 in human melanoma specimens and cell lines further suggests that PRMT5 inhibition deserves exploration as a novel therapeutic strategy for melanoma. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-254. doi:1538-7445.AM2012-LB-254