Abstract 1540: CCCTC-binding factor (CTCF) mutation in endometrioid endometrial carcinoma (EEC): Master epimutator contributing to biologic aggressiveness
Authors: Walker Christopher J. , Miranda Mario A. , Mutch David G. , Goodfellow Paul 
Published: May 2015

Endometrial cancer is the most common gynecologic malignancy in the US. At present, therapies for recurrent or advanced stage disease are largely ineffective. Understanding the genetic and epigenetic factors that contribute to biologic aggressiveness holds promise for new treatment strategies. CTCF, a master regulator of gene expression, was identified as a significantly mutated gene in endometrial cancers by TCGA. Our group identified a hot spot mutation in CTCF in microsatellite instability positive (MSI) endometrial cancers not reported by TCGA. It is noteworthy that among cancer types analyzed to date, EEC is the only malignancy in which CTCF mutation is a frequent defect. CTCF has thousands of binding sites throughout the human genome and is capable of direct activation of genes, enhancer binding and far-reaching chromatin modification, and controls expression of numerous critical regulators of cellular growth/survival (e.g. c-myc, p53, p19/ARF, p16/INK4A, BRCA1, p27, E2F1). However, the role CTCF plays in EEC is largely unknown. Our analysis of a large cohort of EEC offers further insights into CTCF's role in EEC.

Targeted ultra-deep sequencing of 540 primary ECC samples (966X mean coverage) identified 97 different variants (excluding SNPs, low quality variant calls, and synonymous variants) in 136 samples (25.2%). Mutations were more common in tumors with MSI than tumors with intact DNA mismatch repair (40% and 14% respectively p&lt;.0001). Sixty-five of the 97 variants identified (67%) are novel and sequencing of matched tumor/normal DNA confirmed variants are somatic mutations. Most mutated samples (79%) harbored stop, frameshift or splice site variants. As we have shown that CTCF frameshift and stop mutations are subject to nonsense-mediated decay, these data strongly implicate CTCF as a haploinsufficient tumor suppressor. Mutation was not associated with age, stage, grade and disease-free or overall survival. However, among patients who recurred (n=67) the recurrence free interval (RFI) was shorter for mutation carriers (n=15) than for patients whose tumors had no mutation (8.5 months and 17.9 months respectively, p=0.02).

Because we saw frequent loss of function mutations we undertook studies to determine if CTCF deletion is also a feature of EECs. Q-PCR for 149 samples revealed 8 cases with heterozygous CTCF deletion (5%), and in one tumor a 1741bp homozygous deletion (chr16: 67643579-67645330). Two patients with CTCF deletion recurred and both had short RFIs (&lt;wild-type mean). We are currently validating CTCF deletion in 33 of the 540 samples analyzed by deep sequencing (6.1%) that had low normalized CTCF read depth (&gt;1 SD below the mean).

Investigation of the effects of CTCF mutation on gene expression in primary tumors is ongoing. Taken together, our analyses serve to contextualize the effects of CTCF mutations in EEC and implicate loss of CTCF in biologic aggressiveness.

Citation Format: Christopher J. Walker, Mario A. Miranda, David G. Mutch, Paul Goodfellow. CCCTC-binding factor (CTCF) mutation in endometrioid endometrial carcinoma (EEC): Master epimutator contributing to biologic aggressiveness. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1540. doi:10.1158/1538-7445.AM2014-1540