Functional And Epigenetic Regulation Of Shp-1 In Early Stage Lung Cancer

SHP-1 tyrosine phosphatase potentiates mitogenic signalling cascades in multiple cellular pathways by dephosphorylating inhibitory phospho-tyr sites on kinases. With its central role in regulation of cell proliferation, SHP-1 is a logical candidate biomarker for malignant transformation. SHP-1 dephosphorylates a number of phospho-targets including PI3 kinase, EGFR, src family kinases (SFK), and activated cKit, hence normally has a negative impact on cell proliferation. SHP-1 protein function is highly regulated by phosphorylation with up regulation at tyr536, and down regulated by ser591 phosphorylation. The objectives of the present study are to investigate functional control of SHP- 1 and epigenetic control of expression in non small cell lung cancer (NSCLC). In our previous study, Western blot analyses revealed inhibitory ser591 phosphorylation in two of five NSCLC cell lines that expressed high levels of SHP1. We now observe that si-RNA silencing of the major SFK expressed in the EGFR constitutively activated Calu3 cells results in decreased tyr536 phosphorylation suggesting that SFK are required for SHP-1 activity. Transcription of the SHP-1 gene is regulated by two different promoters: a 3’ promoter (promoter 2) in lymphocytes and a 5’ promoter (promoter 1) in epithelial tissues. Gene silencing of SHP-1 by aberrant methylation of promoter 2 and loss of heterozygosity have been associated with pathogenesis of leukemias/lymphomas. Altered expression of SHP-1 has been reported in ovarian and breast cancer due to epigenetic changes in the 5’ promoter (promoter 1). Promoter 2 hypermethylation has been proposed as a plasma biomarker for lung cancer. Previous observations are consistent with the involvement of aberrant SHP-1 promoter 2 methylation in recurrent cases of NSCLC. Pyrosequencing analysis of 22 normal leukocytes, 19 normal lung specimens, 112 cases of early stage (Ib, IIa and IIb) NSCLC and 8 cell lines of SHP-1 promoter 1 vs promoter 2 methylation revealed 10.2% vs 5.1% in leukocytes, 32.6% vs 52.9% in normal lung 44.4% vs 58.6% in NSCLC, and 50.6% vs 96.2% in the cell lines. Hypermethylation of SHP-1 promoter 1 and promoter 2 were observed in 40.8% and 60.7% of lung tumors, respectively. Previous results showed an association of hypermethylation of promoter 2 with time to tumor recurrence in early stage patients (p=0.010). Hypermethylation of SHP-1 promoter 1 in the present study with a limited number of samples is marginally associated with shorter time to recurrence (25.5 months vs. median not reached; p=0.219). These results demonstrate tissue-specific epigenetic control of both SHP-1 promoters and suggest dysfunction of both epigenetic and post-translational regulation of signalling in early stage NSCLC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3831. doi:10.1158/1538-7445.AM2011-3831