Loss of ASS1 Expression in Myxofibrosarcoma Via Promoter Hypermethylation: Prognostic and Potential Therapeutic Implications.
Chien-Feng Li, Li-Tzong Chen, Yu-Hui Wang, Yow-Ling Shiue, Hsuan-Ying Huang. Chi-Mei Hosp., Tainan, Taiwan; Institute of Biomedical Sciences, National Sun Yat-Sen University, Kaohsiung, Taiwan; NCRI, NHRI, Tainan, Taiwan; Chang Gung Memorial Hosp., Kaohsiung, Taiwan
Background: Argininosuccinate synthase 1 (ASS1), catalyzing the penultimate step of the arginine biosynthetic pathway, is commonly downregulated in human cancers. Taking advantage of this deregulated cellular metabolism, pegylated arginine deiminase (ADI-PEG20) has emerged as a novel target therapy that renders ASS1-deficient cancer cells particularly susceptible to arginine deprivation. We herein address ASS1 expression status and its downregulating mechanism, prognostic value, and therapeutic relevance in myxofibrosarcoma (MFS).
Design: Immunoexpression of ASS1 protein was evaluated on tissue microarrays of primary MFS, yielding 90 cases with assessable results for correlation with clincopathological variables and patient survival. Methylation status of ASS1 gene promoter was examined by methylation-specific PCR (MSP). Compared with CCD966SK dermal fibroblasts, three MFS cell lines (OH931, NMFH-1, NMFH-2) were analyzed for protein and mRNA levels of ASS1 by immunoblotting and quantitative RT-PCR assays, respectively. XTT assay was performed to evaluate the susceptibility to ADI-PEG20 treatment in three MFS cell lines vs. CCD966SK fibroblasts.
Results: ASS1 deficiency was identified in 40 cases and significantly associated with increment in FNCLCC grade (p=0.005), AJCC stage (p=0.007), and mitotic activity (p=0.030), with remarkable tumor necrosis (p=0.046), and univariately with inferior metastasis-free survival (MeFS, p=0.0052) and disease-specific survival (DSS, p=0.0035). In multivariate comparisons, ASS1 deficiency independently predicted both MeFS (p=0.0213, risk ratio [RR] =4.237) and DSS (p=0.0230, RR=10.416), along with higher FNCLCC grades. In the first batch subjected to MSP, hypermethylation of ASS1 promoter was identified in 6 of 10 cases and associated with loss of ASS1 immunoexpression (p=0.004). Compared to fibroblasts, the endogenous ASS1 mRNA expression was remarkably downregulated in all 3 MFS cell lines (0-, 0.004-, and 0.055-folds), concordant with the hypermethylation of their ASS1 promoters and deficiency in protein expression. In contrast to no susceptibility seen in CCD966SK fibroblasts, ADI-PEG20 at a low concentration (0.1ug/ml) could remarkably attenuate cell viability of 3 MFS cell lines.
Conclusions: Epigenetic promoter methylation may lead to ASS1 deficiency in a significant subset of MFS, which not only confers tumor aggressiveness but also represents a novel target for arginine deprivation therapy.
Category: Bone & Soft Tissue
Monday, February 28, 2011 1:00 PM
Poster Session II # 5, Monday Afternoon