DNA methylation of neural EGFL like 1 (NELL1) is associated with advanced disease and the metastatic state of renal cell cancer patients

Peters,Inga; Dubrowinskaja,Natalia; Hennenlotter,Jörg; Antonopoulos,Wiebke Inga; Von Klot,Christoph A.J.; Tezval,Hossein; Stenzl,Arnulf; Kuczyk,Markus A.; Serth,Jürgen

doi:10.3892/or.2018.6732

DNA methylation of neural EGFL like 1 (NELL1) is associated with advanced disease and the metastatic state of renal cell cancer patients

Authors:
- Inga Peters
- Natalia Dubrowinskaja
- Jörg Hennenlotter
- Wiebke Inga Antonopoulos
- Christoph A.J. Von Klot
- Hossein Tezval
- Arnulf Stenzl
- Markus A. Kuczyk
- Jürgen Serth
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Affiliations: Department of Urology, Hannover Medical School, D‑30625 Hannover, Germany, Department of Urology, Eberhard‑Karls University, D‑72074 Tübingen, Germany, Department of Pathology, Hannover Medical School, D‑30625 Hannover, Germany
Published online on: September 24, 2018 https://doi.org/10.3892/or.2018.6732
Pages: 3861-3868

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Abstract

Recent studies have shown that NELL1 expression is silenced epigenetically in human renal cell cancer (RCC) tissues and in RCC cell lines. However, it remains unknown whether NELL1 promoter methylation observed in clinical specimens might be associated with the clinicopathology or survival of patients with RCC. We analyzed NELL1 DNA methylation in tissues from patients with RCC and in adjacent normal renal tissues. In addition, we evaluated NELL1 methylation in cell lines derived from different urogenital tumors (prostate cancer, urothelial cancer and RCC). We performed regression analyses to determine whether NELL1 methylation is associated with clinicopathological parameters and recurrence‑free survival (RFS). This cross‑sectional study included 98 patients with RCC and 63 paired tumor and adjacent normal tissue samples. We analyzed a locus in the intron 1 region of NELL1 with pyrosequencing. We performed in silico analysis of NELL1 methylation in the TCGA Kidney Renal Clear Cell Carcinoma (KIRC) data set (n=284 patients), which served as a validation study. Statistical analyses were performed with the two‑sided paired t‑test for paired tumor and adjacent normal samples. We used logistic regression for subgroup comparisons and Cox regression for RFS comparisons. The mean methylation level was 6.8% higher in RCC tissues compared to paired adjacent normal tissues (paired t‑test, P<0.001). Methylation levels in RCC were associated with advanced disease (P=0.002), the presence of distant metastases (P=0.004), and shorter RFS (P=0.035, HR: 4.15). In silico validation with TCGA KIRC data for adjacent loci also demonstrated that high relative methylation levels were associated with adverse clinicopathology and shortened RFS. Our results suggest that NELL1 methylation contributes to RCC disease progression. This finding could provide a clinical marker to complement recent functional analyses in tumor models.

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1	Ferlay J, Soerjomataram I, Dikshit R, Eser S, Mathers C, Rebelo M, Parkin DM, Forman D and Bray F: Cancer incidence and mortality worldwide: Sources, methods and major patterns in GLOBOCAN 2012. Int J Cancer. 136:E359–E386. 2014. View Article : Google Scholar : PubMed/NCBI
2	Creighton CJ, Morgan M, Gunaratne PH, Wheeler DA, Gibbs RA, Robertson A, Chu A, Beroukhim R, Cibulskis K, Signoretti S, et al: Comprehensive molecular characterization of clear cell renal cell carcinoma. Nature. 499:43–49. 2013. View Article : Google Scholar : PubMed/NCBI
3	Cheng L, Zhang S, MacLennan GT, Lopez-Beltran A and Montironi R: Molecular and cytogenetic insights into the pathogenesis, classification, differential diagnosis, and prognosis of renal epithelial neoplasms. Hum Pathol. 40:10–29. 2009. View Article : Google Scholar : PubMed/NCBI
4	Morris MR, Ricketts C, Gentle D, Abdulrahman M, Clarke N, Brown M, Kishida T, Yao M, Latif F and Maher ER: Identification of candidate tumour suppressor genes frequently methylated in renal cell carcinoma. Oncogene. 29:2104–2117. 2010. View Article : Google Scholar : PubMed/NCBI
5	Peters I, Rehmet K, Wilke N, Kuczyk MA, Hennenlotter J, Eilers T, Machtens S, Jonas U and Serth J: RASSF1A promoter methylation and expression analysis in normal and neoplastic kidney indicates a role in early tumorigenesis. Mol Cancer. 6:492007. View Article : Google Scholar : PubMed/NCBI
6	Atschekzei F, Hennenlotter J, Jänisch S, Großhennig A, Tränkenschuh W, Waalkes S, Peters I, Dörk T, Merseburger AS, Stenzl A, et al: SFRP1 CpG island methylation locus is associated with renal cell cancer susceptibility and disease recurrence. Epigenetics. 7:447–457. 2012. View Article : Google Scholar : PubMed/NCBI
7	Onay H, Pehlivan S, Koyuncuoglu M, Kirkali Z and Ozkinay F: Multigene methylation analysis of conventional renal cell carcinoma. Urol Int. 83:107–112. 2009. View Article : Google Scholar : PubMed/NCBI
8	de Caceres Ibanez I, Dulaimi E, Hoffman AM, Al-Saleem T, Uzzo RG and Cairns P: Identification of novel target genes by an epigenetic reactivation screen of renal cancer. Cancer Res. 66:5021–5028. 2006. View Article : Google Scholar : PubMed/NCBI
9	Morris MR, Gentle D, Abdulrahman M, Clarke N, Brown M, Kishida T, Yao M, Teh BT, Latif F and Maher ER: Functional epigenomics approach to identify methylated candidate tumour suppressor genes in renal cell carcinoma. Br J Cancer. 98:496–501. 2008. View Article : Google Scholar : PubMed/NCBI
10	Dubrowinskaja N, Gebauer K, Peters I, Hennenlotter J, Abbas M, Scherer R, Tezval H, Merseburger AS, Stenzl A, Grünwald V, et al: Neurofilament Heavy polypeptide CpG island methylation associates with prognosis of renal cell carcinoma and prediction of antivascular endothelial growth factor therapy response. Cancer Med. 3:300–309. 2014. View Article : Google Scholar : PubMed/NCBI
11	Henrique R, Luís AS and Jerónimo C: The epigenetics of renal cell tumors: From biology to biomarkers. Front Genet. 3:942012. View Article : Google Scholar : PubMed/NCBI
12	Watanabe TK, Katagiri T, Suzuki M, Shimizu F, Fujiwara T, Kanemoto N, Nakamura Y, Hirai Y, Maekawa H and Takahashi EI: Cloning and characterization of two novel human cDNAs (NELL1 and NELL2) encoding proteins with six EGF-like repeats. Genomics. 38:273–276. 1996. View Article : Google Scholar : PubMed/NCBI
13	Ting K, Vastardis H, Mulliken JB, Soo C, Tieu A, Do H, Kwong E, Bertolami CN, Kawamoto H, Kuroda S, et al: Human NELL-1 expressed in unilateral coronal synostosis. J Bone Miner Res. 14:80–89. 1999. View Article : Google Scholar : PubMed/NCBI
14	Aghaloo T, Cowan CM, Chou YF, Zhang X, Lee H, Miao S, Hong N, Kuroda S, Wu B, Ting K, et al: Nell-1-induced bone regeneration in calvarial defects. Am J Pathol. 169:903–915. 2006. View Article : Google Scholar : PubMed/NCBI
15	Maeda K, Matsuhashi S, Tabuchi K, Watanabe T, Katagiri T, Oyasu M, Saito N and Kuroda S: Brain specific human genes, NELL1 and NELL2, are predominantly expressed in neuroblastoma and other embryonal neuroepithelial tumors. Neurol Med Chir. 41:582–589. 2001. View Article : Google Scholar
16	Shah US and Getzenberg RH: Fingerprinting the diseased prostate: Associations between BPH and prostate cancer. J Cell Biochem. 91:161–169. 2004. View Article : Google Scholar : PubMed/NCBI
17	Osman I, Bajorin DF, Sun TT, Zhong H, Douglas D, Scattergood J, Zheng R, Han M, Marshall KW and Liew CC: Novel blood biomarkers of human urinary bladder cancer. Clin Cancer Res. 12:3374–3380. 2006. View Article : Google Scholar : PubMed/NCBI
18	Slovak ML, Bedell V, Hsu YH, Estrine DB, Nowak NJ, Delioukina ML, Weiss LM, Smith DD and Forman SJ: Molecular karyotypes of Hodgkin and Reed-Sternberg cells at disease onset reveal distinct copy number alterations in chemosensitive versus refractory Hodgkin lymphoma. Clin Cancer Res. 17:3443–3454. 2011. View Article : Google Scholar : PubMed/NCBI
19	Nakamura R, Oyama T, Tajiri R, Mizokami A, Namiki M, Nakamoto M and Ooi A: Expression and regulatory effects on cancer cell behavior of NELL1 and NELL2 in human renal cell carcinoma. Cancer Sci. 106:656–664. 2015. View Article : Google Scholar : PubMed/NCBI
20	Gebauer K, Peters I, Dubrowinskaja N, Hennenlotter J, Abbas M, Scherer R, Tezval H, Merseburger AS, Stenzl A, Kuczyk MA, et al: Hsa-mir-124-3 CpG island methylation is associated with advanced tumours and disease recurrence of patients with clear cell renal cell carcinoma. Br J Cancer. 108:131–138. 2013. View Article : Google Scholar : PubMed/NCBI
21	Sobin LH and Compton CC: TNM seventh edition: What's new, what's changed: Communication from the international union against cancer and the American Joint Committee on cancer. Cancer. 116:5336–5339. 2010. View Article : Google Scholar : PubMed/NCBI
22	Peters I, Eggers H, Atschekzei F, Hennenlotter J, Waalkes S, Tränkenschuh W, Grosshennig A, Merseburger AS, Stenzl A, Kuczyk MA, et al: GATA5 CpG island methylation in renal cell cancer: A potential biomarker for metastasis and disease progression. BJU Int. 110:E144–E152. 2012. View Article : Google Scholar : PubMed/NCBI
23	Colella S, Shen L, Baggerly KA, Issa JP and Krahe R: Sensitive and quantitative universal Pyrosequencing methylation analysis of CpG sites. Biotechniques. 35:146–150. 2003. View Article : Google Scholar : PubMed/NCBI
24	Team RDC: R Development Core Team. 2011. R: A language and environment for statistical computingR Foundation for Statistical Computing. Vienna, Austria: http://www.R-project.org/
25	Mori Y, Cai K, Cheng Y, Wang S, Paun B, Hamilton JP, Jin Z, Sato F, Berki AT, Kan T, et al: A genome-wide search identifies epigenetic silencing of somatostatin, tachykinin-1, and 5 other genes in colon cancer. Gastroenterology. 131:797–808. 2006. View Article : Google Scholar : PubMed/NCBI
26	Jin Z, Mori Y, Yang J, Sato F, Ito T, Cheng Y, Paun B, Hamilton JP, Kan T, Olaru A, et al: Hypermethylation of the nel-like 1 gene is a common and early event and is associated with poor prognosis in early-stage esophageal adenocarcinoma. Oncogene. 26:6332–6340. 2007. View Article : Google Scholar : PubMed/NCBI
27	Gao C and Zhang Q, Kong D, Wu D, Su C, Tong J, Chen F and Zhang Q: MALDI-TOF Mass Array analysis of Nell-1 promoter methylation patterns in human gastric cancer. Biomed Res Int. 2015:1369412015. View Article : Google Scholar : PubMed/NCBI