Prognostic significance of MST1R dysregulation in renal cell tumors

Abstract

Macrophage stimulating 1 receptor (MST1R) is a C-MET proto-oncogene family receptor tyrosine kinase. Promoter methylation patterns determine transcription of MST1R variants as hypermethylation of a region upstream of transcription start site (TSS) is associated with lack of MST1R long transcript (MST1R long) and expression of a short transcript with oncogenic potential. Thus, we aimed to investigate MST1R variant transcript regulation in renal cell tumors (RCT) and assess their prognostic potential. We found, in a series of 120 RCT comprising the four main subtypes (clear cell, papillary and chromophobe renal cell carcinoma, and oncocytoma), that higher methylation levels close to TSS were associated with total MST1R expression levels (MST1R total) in primary tumors (p=0.049) and renal cancer cell lines. After demethylating treatment, MST1R long/MST1R total ratio increased, as expected, in two renal cell carcinoma cell lines tested. However, in primary tumors with hypermethylation upstream of TSS, a decrease in MST1R long/MST1R total ratio was not detected, although higher expression ratio of nuclear factor-κB was apparent. Furthermore, survival analysis demonstrated that MST1R long/MST1R total ratio was independently associated with shorter disease-specific and disease-free survival, whereas MST1R total expression associated with shorter disease-specific survival. In conclusion, although promoter methylation patterns seem to determine MST1R global transcription regulation in renal cell carcinoma, other mechanisms might contribute to deregulate MST1R variant expression in RCT. Nevertheless, MST1R total expression and MST1R long/MST1R total ratio modulate the biological and clinical aggressiveness of renal cell carcinoma, as depicted by its prognostic significance, a finding that requires validation in a larger independent series.

Keywords: MST1R; MST1R expression; MST1R promoter methylation; RON; Renal cell tumors; epigenetics.

Figure 1

MST1R promoter methylation in renal cell tumors (RCT): bisulfite sequencing of MST1R promoter in 5 cases (A) and QMSP methylation levels in two distinct regions, one upstream TSS (MST1R _up) and another closer to TSS (MST1R _TSS), in 120 cases. White squares: CpG unmethylated; gray squares: CpG partially methylated. NK: normal kidney; RO: renal oncocytoma; chRCC: chromophobe renal cell carcinoma; pRCC: papillary renal cell carcinoma; ccRCC: clear cell renal cell carcinoma.

Figure 2

Expression levels in tumors hypermethylated or not at MST1R _up (upstream area of MST1R promoter): MST1R _long/total ratio in RCTs (n=120) (A1) and in ccRCC (n=30) (B1), and NF-κB expression ratio in RCTs (A2) and ccRCC (B2).

Figure 3

MST1R methylation and expression levels in ccRCC cell lines. A: MST1R _TSS methylation level (A1) and MST1R _total expression level (A2) in three cell lines. B: MST1R _TSS methylation level (B1) and MST1R _total expression level (B2) in 769-P and 786-O cell lines before and after demethylating treatment with 1 µM 5-aza-2’deoxycytidine for 72 h. C. MST1R _up methylation level (C1) and MST1R _long/total ratio (C2) in 769-P and 786-O cell lines before and after demethylating treatment with 1 µM 5-aza-2’deoxycytidine for 72 h.

Figure 4

Kaplan-Meier analysis for disease-specific survival in 60 RCC patients, according to MST1R _total expression level (A) and MST1R _long/total ratio (B), and for disease-free survival in 60 RCC patients, according to MST1R _long/total ratio (C). The results presented were categorized using third quartile (75^th percentile) value as cutoff.