Abstract
Background: Pancreatic Cancer is an extremely aggressive disease with a bad outcome owing to its late detection, high rate of metastasis and self-renewal post-surgical and chemotherapeutic intervention. In cancer the balance between differentiation and self-renewal is tightly regulated by SOX2. Over expression of SOX2, as in pancreatic cancer, can tilt the balance towards self-renewal and thus increase the population of these undifferentiated “stem cells”.
Methods: A CRISPR based OGT knockout kit was purchased from Origene and a knockout cell line was generated with S2VP10 cells. For the tumor initiation studies OGTi (OGT knockout cells) were implanted subcutaneously at a limiting dilution from 500,000 cells to 500 cells. Site directed mutagenesis was performed using theQuick-change Lightning from Agilent.
Results: In the current study we show for the first time that SOX2 undergoes a type of post translational modification known as O GlcNAcylation. We also show for the first time that in humans this modification happens at Serine 246 and mutating this particular site via a side directed mutagenesis leads to a loss of O GlcNAcylation of SOX2.O GlcNAc modification of SOX2 further leads to its activation which then leads to its stabilization in the nucleus. A CRISPR-OGT knockout in pancreatic cancer cell line S2VP10 resulted in a delayed tumor initiation. We further showed that mutation of this site (S246A) prevents the modification of Sox2 and its downstream activity. Our study also demonstrated that targeting OGT in vivo with a small molecule inhibitor OSMI, results in decreased tumor burden, delayed tumor progression and a decreased expression of SOX2 in pancreatic cancer cells.
Conclusion: Our study highlights for the first time that the O-GlcNAc transferase dependent SOX2 glycosylation has a profound effect on the transcriptional activity of SOX2 and is instrumental in determining self-renewal in pancreatic cancer which can be targeted therapeutically.
Citation Format: Nikita S. Sharma, Vineet K. Gupta, Patricia Dauer, Roey Hadad, Kousik K. kesh, Vikas Dudeja, Ashok Saluja, Sulagna Banerjee. O GlcNAcylation of SOX2 regulates its tumor initiation properties in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4498.
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