KiSS-1 methylation and protein expression patterns contribute to diagnostic and prognostic assessments in tissue specimens for colorectal cancer

Abstract

KISS1 is a metastasis suppressor lost in several solid malignancies. We evaluated the clinical relevance of KiSS-1 methylation and its protein expression in colorectal cancer. The epigenetic silencing of KiSS-1 by hypermethylation was tested in colon cancer cells (n = 5) before and after azacytidine treatment. KiSS-1 methylation was evaluated by methylation-specific PCR in colorectal cancer cells, and normal, benign, and tumor tissues (n = 352) were grouped in a training set (n = 62) and two independent validation cohorts (n = 100 and n = 190). KiSS-1 protein expression was analyzed by immunohistochemistry on tissue arrays. KiSS-1 hypermethylation correlated with transcript and protein expression loss, being increased in vitro by azacytidine. Methylation rates were 53.1, 70.0, and 80.0 % in the training and validation sets, respectively. In the training set, KiSS-1 methylation rendered a diagnostic accuracy of 72.7 % (p = 0.002). Combination of KiSS-1 methylation and serum CEA (p = 0.001) increased the prognostic utility of CEA alone (p = 0.022). In the first validation set, KiSS-1 methylation correlated with tumor grade (p = 0.011), predicted recurrence (p = 0.009), metastasis (p = 0.004), disease-free (p = 0.034), and overall survival (p = 0.015). In the second validation cohort, KiSS-1 methylation predicted disease-specific survival (p = 0.030). In the training set, cytoplasmic KiSS-1 expression was significantly higher in nonneoplastic biopsies as compared to colorectal tumors (p < 0.0005). In the validation set, loss of cytoplasmic expression correlated with tumor stage (p = 0.007), grade (p = 0.035), recurrence (p = 0.017), and disease-specific survival (p = 0.022). KiSS-1 was revealed epigenetically modified in colorectal cancer. The diagnostic and prognostic utility of KiSS-1 methylation and expression patterns suggests their assessment for the clinical management of colorectal cancer patients.

Aberrant methylated EDNRB can act as a potential diagnostic biomarker in sporadic colorectal cancer while KISS1 is controversial

“…30,31 It has been reported that EDNRB and KISS1 as the tumor suppressor and tumor metastasis suppressor genes, respectively, are silenced in various cancers such as colorectal cancer, which is caused by specific CpG islands' hypermethylation. [13][14][15] Regarding EDNRB, which encodes a G-protein coupled receptor, it is implicated that hypermethylation of CpG dinucleotides in 5 0 flanking region happens during CRC progression. 13 Indeed, this aberrant methylation is associated with the EDNRB downregulation and silencing that may be involved in the cell proliferation, 14,15 In the present study, EDNRB hypermethylation in CRC confirmed quantitatively for the first time by MS-HRM assay, concordant with what previously found in Chen et al study.…”

“…[13][14][15] Regarding EDNRB, which encodes a G-protein coupled receptor, it is implicated that hypermethylation of CpG dinucleotides in 5 0 flanking region happens during CRC progression. 13 Indeed, this aberrant methylation is associated with the EDNRB downregulation and silencing that may be involved in the cell proliferation, 14,15 In the present study, EDNRB hypermethylation in CRC confirmed quantitatively for the first time by MS-HRM assay, concordant with what previously found in Chen et al study. 13 Methylation specific PCR (MSP) results in their study, which is a non-quantitative technique, showed that EDNRB hypermethylation frequency in CRC tumor tissues was higher than adjacent normal samples (92.86 versus 59.52, P D 0.001) and this aberrant methylation was correlated with EDNRB downregulation.…”

“…Although it has been suggested that KISS1 gene expression levels is reduced in various cancers, 14 which has unfortunately not been assayed herein, Kostakis et al proposed that the KISS1 expression downregulation could be an early event outcome when colonic cells transform toward malignancy, yet the KISS1 expression is being upregulated persistently during tumor progression, especially during expansion in size and invasive properties. 25 Hence, considering the reports indicating decrease of KISS1 gene expression in various cancers, it seems possible that this reduction also could be resulted from the other key mechanisms involved in modulation of gene expression such as chromatin remodeling, post-transcriptional and post-translational modifications in addition to the gene-specific methylation which is not observed in KISS1 in present study.…”

“…For this reason, several studies have been performed and some of them non-quantitatively showed that EDNRB and KISS1 genes are subjected to methylation alterations in colorectal cancer. [13][14][15] Endothelin receptor type B (EDNRB) gene, which is mapped to chromosome region 13q22, encodes a receptor which is a member of the G-protein coupled receptor superfamily and involves normally in the development of embryonic and enteric ganglia. 13,16,17 The interaction of this receptor with endothelins leads to the releasing of InsP 3 and Ca 2C as second messengers.…”

“…25 It is demonstrated that KISS1 expression is low or reduced in transcriptional level in bladder cancer and CRC, which may be influenced by hypermethylation of CpG rich regions in 5 0 upstream and 3 0 downstream of KISS1 gene. 14,15,26 It is believed that kisspeptins serve as metastasis suppressors in some cancers, 25 however, this inhibition function is not clearly understood. Nonetheless, KISS1R activation through kisspeptins may lead to suppression of ERK activation, which culminates with a reduction in matrix metalloproteinase 9 (MMP-9) expression and ultimately inhibition of tumor metastasis.…”