Epigenetic silencing of NTSR1 is associated with lateral and noninvasive growth of colorectal tumors

// Seiko Kamimae 1, * , Eiichiro Yamamoto 1, 2, * , Masahiro Kai 1, * , Takeshi Niinuma 1, 2 , Hiro-o Yamano 3 , Masanori Nojima 4 , Kennjiro Yoshikawa 3 , Tomoaki Kimura 3 , Ryo Takagi 3 , Eiji Harada 3 , Taku Harada 1 , Reo Maruyama 1, 2 , Yasushi Sasaki 5 , Takashi Tokino 5 , Yasuhisa Shinomura 2 , Tamotsu Sugai 6 , Kohzoh Imai 7 , Hiromu Suzuki 1 1 Department of Molecular Biology, Sapporo Medical University School of Medicine, Sapporo, Japan 2 Department of Gastroenterology, Rheumatology, Clinical Immunology, Sapporo Medical University School of Medicine, Sapporo, Japan 3 Department of Gastroenterology, Akita Red Cross Hospital, Akita, Japan 4 Center for Translational Research, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan 5 Medical Genome Science, Research Institute for Frontier Medicine, Sapporo Medical University School of Medicine, Sapporo, Japan 6 Department of Molecular Diagnostic Pathology, Iwate Medical University, Morioka, Japan 7 Center for Medical Innovation, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan * These authors have contributed equally to this work Correspondence to: Hiromu Suzuki, e-mail: hsuzuki@sapmed.ac.jp Eiichiro Yamamoto, e-mail: eiichiro@xa3.so-net.ne.jp Keywords: colorectal tumor, invasion, LST, DNA methylation, biomarker Received: April 08, 2015      Accepted: August 07, 2015      Published: August 17, 2015 ABSTRACT Our aim was to identify DNA methylation changes associated with the growth pattern and invasiveness of colorectal cancers (CRCs). Comparison of the methylation statuses of large (≥20 mm in diameter along the colonic surface) noninvasive tumors (NTs) and small (<20 mm in diameter along the colonic surface) invasive tumors (ITs) using CpG island microarray analysis showed neurotensin receptor 1 ( NTSR1 ) to be hypermethylated in large NTs. Quantitative bisulfite pyrosequencing revealed that NTSR1 is frequently methylated in colorectal tumors, with large NTs exhibiting the highest methylation levels. The higher NTSR1 methylation levels were associated with better prognoses. By contrast, NTSR1 copy number gains were most frequent among small ITs. Methylation of NTSR1 was associated with the gene's silencing in CRC cell lines, whereas ectopic expression of NTSR1 promoted proliferation and invasion by CRC cells. Analysis of primary tumors composed of adenomatous and malignant portions revealed that NTSR1 is frequently methylated in the adenomatous portion, while methylation levels are generally lower in the cancerous portions. These results suggest that NTSR1 methylation is associated with lateral and noninvasive growth of colorectal tumors, while low levels of methylation may contribute to the malignant potential through activation of NTSR1 . Our data also indicate that NTSR1 methylation may be a prognostic biomarker in CRC.