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Hypermethylation of ZNF545 is associated with poor prognosis in patients with early-stage hepatocellular carcinoma after thermal ablation
  1. Jie Yu1,
  2. Xin Li1,
  3. Qian Tao2,
  4. Xiao-ling Yu1,
  5. Zhi-gang Cheng1,
  6. Zhi-yu Han1,
  7. Mingzhou Guo3,
  8. Ping Liang1
  1. 1Department of Interventional Ultrasound, Chinese PLA General Hospital, Beijing, China
  2. 2Department of Clinical Oncology, The Chinese University of Hong Kong, Hong Kong, Hong Kong
  3. 3Department of Gastroenterology & Hepatology, Chinese PLA General Hospital, Beijing, China
  1. Correspondence to Dr Ping Liang, Department of Interventional Ultrasound, Chinese PLA General Hospital, 28 Fuxing Road, Beijing 100853, China; liangping301{at}hotmail.com; Mingzhou Guo, M.D., Ph.D., Department of Gastroenterology & Hepatology, Chinese PLA General Hospital, #28 Fuxing Road, Beijing 100853, China; mzguo1@hotmail.com

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We read with interest the article by Wang et al1 on zinc-finger protein 545 (ZNF545) acting as a tumour suppressor gene (TSG) in gastric cancer by inhibiting rRNA transcription and its methylation as a prognostic factor for early stages of gastric cancer. Inactivation of TSGs through promoter hypermethylation also plays an important role in progression of hepatocellular carcinoma (HCC).2–4 We wish to report the results of the expression profile and epigenetic regulation of ZNF545 in HCC, and its methylation role on early-stage HCC progression after thermal ablation.

Our results showed reduced or loss of ZNF545 expression was found in seven human hepatic cancer cell lines, including SMMC7721, PRF/PCL-5, SK-hep1, HepG2, BEL7402, LAM3 and SNU449, by semiquantitative reverse transcription PCR. Expression of ZNF545 was detected in another cell of HBXF344. The methylation status of the ZNF545 promoter was examined by methylation-specific PCR (MSP). Complete methylation of the ZNF545 promoter was found in SMMC7721, PRF/PCL-5, SK-hep1, HepG2, LAM3 and SNU449 cells, partial methylation in BEL7402 and unmethylation in HBXF344. Bisulfite sequencing results were consistent with those of MSP. Restoration of ZNF545 expression could be induced by 5-aza-2-deoxycytidine in seven cell lines that showed reduced or loss of ZNF545 expression.

Based on this, 101 HCC samples were obtained with informed consent from 101 patients who underwent radiofrequency and microwave ablation for early-stage HCC within Milan criteria between October 2008 and December 2013 (ethics approval date was February 2008). Partial and dense promoter methylation of ZNF545 was detected in 56.4% (57/101) of HCC tissues, but no methylation in eight normal human hepatic tissues. Immunohistochemistry showed reduced expression of ZNF545 was significant in cancer tissue compared with adjacent tissue (p<0.001). Reduced expression was associated with promoter region hypermethylation. It suggests that ZNF545 is possibly regulated by promoter region methylation in HCC.

The size of 101 HCC lesions ranged from 1.1 to 5.0 cm (2.9±0.9 cm, mean±SD). Fifty-seven and forty-four patients were enrolled in methylation and unmethylation groups, respectively. During the follow-up ranginge from 7.6 to 59.6 months (median 28.7 months), for ZNF545 methylation group, the 1-year, 3-year and 5-year recurrence rates were 21.1%, 49.4% and 79.0%, respectively, and 27.3%, 43.4% and 43.4%, respectively, for unmethylation group. Although the result showed no statistical difference (p=0.065), methylation group showed a higher recurrence tendency. But ZNF545 methylation showed no significant difference in recurrence time (>2 years, 1–2 years and <1 year) (p=0.61). In addition, the 1-year, 3-year and 5-year overall survival rates of methylation group were 84.1%, 70.4% and 35.2%, and those of unmethylation group were 97.7%, 97.7% and 97.7% (p=0.007). Univariate analysis showed methylation group had older age (p=0.01) and poorer survival (p=0.003). Multivariate regression analyses (table 1) showed the patients with low survival rate had the increased tendency for methylation (p=0.041).

Table 1

Multivariate Cox regression analysis of correlation factors for ZNF545 methylation in patients with hepatocellular carcinoma

Thermal ablation is a promising technique for early-stage HCC therapy. However, recurrence is one of the major challenges after HCC ablation, which may lead to unfavourable outcome.4 ,5 In the present study, we creatively combined HCC ablation and ZNF545 methylation to successfully identify that ZNF545 methylation is a common epigenetic event in HCC and it may be regarded as a valuable new prognostic factor for patients with HCC. This can provide guidance in early detection of recurrent HCC and prediction of ablation effectiveness. Moreover, it will help to identify patients who would most benefit from combination of ablation and epigenetic therapy.6

In conclusion, we have identified ZNF545 as a novel potential TSG in HCC and its methylation as a valuable new prognostic factor for early-stage HCC after thermal ablation. The results will also pave the way for future HCC epigenetic therapy.

References

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Footnotes

  • JY and XL contributed equally

  • Contributors PL and MG had full access to all of the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis. Study concept and design: PL, JY, MG. Acquisition of data: PL, JY, XL, QT, X-lY, Z-gC, Z-yH and MG. Analysis and interpretation of data: PL, JY, MG. Drafting of the manuscript: JY. Critical revision of the manuscript for important intellectual content: JY, XL.

  • Funding This work was supported by one grant 7144246 from the Beijing Natural Science Foundation, and two grants 81401436 and 81430039 from the National Scientific Foundation Committee of China.

  • Competing interests None declared.

  • Patient consent Obtained.

  • Ethics approval Ethics committee of Chinese PLA general hospital.

  • Provenance and peer review Not commissioned; internally peer reviewed.

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