J Cancer 2020; 11(16):4791-4800. doi:10.7150/jca.43665 This issue

Research Paper

NK homeobox 2.2 functions as tumor suppressor in colorectal cancer due to DNA methylation

Yuan He1,2, Xiao-Yun Liu3, Rui Gong4, Kun-Wei Peng1, Rong-Bin Liu5✉, Fang Wang3✉

1. State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou 510060, China.
2. Department of Ultrasound and Electrocardiogram, Sun Yat-sen University Cancer Center, Guangzhou 510060, China.
3. Department of Molecular Diagnostics, Sun Yat-sen University Cancer Center, Guangzhou 510060, China.
4. Department of Healthcare, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China.
5. Department of Ultrasound, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510120, China.

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Citation:
He Y, Liu XY, Gong R, Peng KW, Liu RB, Wang F. NK homeobox 2.2 functions as tumor suppressor in colorectal cancer due to DNA methylation. J Cancer 2020; 11(16):4791-4800. doi:10.7150/jca.43665. Available from https://www.jcancer.org/v11p4791.htm

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Abstract

Aim: The role of NK homeobox 2.2 (NKX2.2) in human colorectal cancer (CRC) remains to be unveiled. This study was designed to explore the epigenetic regulation and function of NKX2.2 in human CRC.

Methods: The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) datasets were used to assess the methylation data of NKX2.2 in CRC. Six CRC cell lines (HCT116, SW480, HT29, LOVO, SW1116, SW640) and 20 pairs of primary CRC tumor and normal tissues were utilized to explore the function of NKX2.2 in CRC using Sequenom EpiTYPER®, verified by cloning-based bisulfite sequencing analysis, semi-quantitative reverse transcription PCR, western blot, cell viability assessment, plate clone formation assay , and transwell assays.

Results: Bioinformatic analysis showed that NKX2.2 was significantly hypermethylated in primary tumors compared to normal tissues (p < 0.05). Our study also found that NKX2.2 methylation was upregulated (p<0.05) in tumors than normal tissues. In vitro experiments demonstrated that 5-aza-2'-deoxycytidine downregulated the methylation of NKX2.2 and retrieved its expression of mRNA and protein levels (p<0.05). No significant association was found between the NKX2.2 methylation and sex, age, tumor differentiation, TNM stage, CEA, CA199, and fecal occult blood (p>0.05). Kaplan-Meier analysis indicated that NKX2.2 hypermethylation showed a trend but not statistical significance for predicting poor overall survival in CRC patients (p=0.33). NKX2.2 overexpression suppressed cell proliferation, colony formation, and inhibited tumor invasion and migration in CRC cells (both p<0.05).

Conclusions: This study indicates that NKX2.2 is a tumor suppressor in CRC due to hypermethylation.

Keywords: Colorectal cancer, Hypermethylation, NK homeobox 2.2, Epigenetic