Ascorbic acid–induced TET activation mitigates adverse hydroxymethylcytosine loss in renal cell carcinoma
Niraj Shenoy, … , Yiyu Zou, Amit Verma
Niraj Shenoy, … , Yiyu Zou, Amit Verma
Published January 31, 2019
Citation Information: J Clin Invest. 2019;129(4):1612-1625. https://doi.org/10.1172/JCI98747.
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Research Article Oncology

Ascorbic acid–induced TET activation mitigates adverse hydroxymethylcytosine loss in renal cell carcinoma

Abstract

Although clear cell renal cell carcinoma (ccRCC) has been shown to result in widespread aberrant cytosine methylation and loss of 5-hydroxymethylcytosine (5hmC), the prognostic impact and therapeutic targeting of this epigenetic aberrancy has not been fully explored. Analysis of 576 primary ccRCC samples demonstrated that loss of 5hmC was strongly associated with aggressive clinicopathologic features and was an independent adverse prognostic factor. Loss of 5hmC also predicted reduced progression-free survival after resection of nonmetastatic disease. The loss of 5hmC in ccRCC was not due to mutational or transcriptional inactivation of ten eleven translocation (TET) enzymes, but to their functional inactivation by l-2-hydroxyglutarate (L2HG), which was overexpressed due to the deletion and underexpression of L2HG dehydrogenase (L2HGDH). Ascorbic acid (AA) reduced methylation and restored genome-wide 5hmC levels via TET activation. Fluorescence quenching of the recombinant TET-2 protein was unaffected by L2HG in the presence of AA. Pharmacologic AA treatment led to reduced growth of ccRCC in vitro and reduced tumor growth in vivo, with increased intratumoral 5hmC. These data demonstrate that reduced 5hmC is associated with reduced survival in ccRCC and provide a preclinical rationale for exploring the therapeutic potential of high-dose AA in ccRCC.

Authors

Niraj Shenoy, Tushar D. Bhagat, John Cheville, Christine Lohse, Sanchari Bhattacharyya, Alexander Tischer, Venkata Machha, Shanisha Gordon-Mitchell, Gaurav Choudhary, Li-Fan Wong, LouAnn Gross, Emily Ressigue, Bradley Leibovich, Stephen A. Boorjian, Ulrich Steidl, Xiaosheng Wu, Kith Pradhan, Benjamin Gartrell, Beamon Agarwal, Lance Pagliaro, Masako Suzuki, John M. Greally, Dinesh Rakheja, R. Houston Thompson, Katalin Susztak, Thomas Witzig, Yiyu Zou, Amit Verma

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Figure 1

Loss of 5hmC is strongly associated with features of tumor aggressiveness in ccRCC.

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Loss of 5hmC is strongly associated with features of tumor aggressivenes...
(A) Correlation between median percentage positive for 5hmC and 5hmC intensity in IHC (P < 0.001). (B) Higher grade ccRCC is associated with loss of 5hmC (P < 0.001). (C) Representative photographs of low-grade and high-grade ccRCC with 5hmC IHC. (D) Loss of 5hmC correlates with higher SSIGN score, which predicts increased risk of progression of ccRCC after nephrectomy (P < 0.001). (E) Increased tumor size in ccRCC is associated with loss of 5hmC (P < 0.001). (F) Nodal metastasis in ccRCC is associated with loss of 5hmC (P < 0.001). (G) Presence of systemic metastatic disease in ccRCC is associated with loss of 5hmC (P < 0.001). (H) Presence of coagulative tumor necrosis is associated with loss of 5hmC (P < 0.001). (I) Presence of sarcomatoid differentiation is associated with loss of 5hmC (P < 0.001). Box plots have horizontal lines at the 25th percentile, the median, and the 75th percentile. The vertical lines extend to the minimum and maximum values. Associations of 5hmC expression with the clinical and pathologic features studied were evaluated using Spearman’s rank correlation coefficients, Kruskal-Wallis tests, and Wilcoxon’s rank sum tests.