Adenomatous Polyposis Coli, Mismatch Repair dan Instabilitas Mikrosatelit Kanker Kolorektal Berdasarkan Perbedaan Lokasi antara Kolon Proksimal, Distal dan Rektum

Abstract

Background: In Western countries most of colorectal cancer (CRC) cases are found in the colon (proximal and distal) rather than in the rectum. Each location can result in defferent clinical, pathological and molecular characteristics of the cancer, and most of studies have only compared between proximal and distal colon. In Indonesia the majority of this cancer is located in the rectum rather than in the colon, and a research on the adenomatous polyposis coli (APC), mismatch repair (MMR) and microsatellite instability (MSI) of CRC based on the three different locations (proximal, distal colon and rectal cancer) had never been conducted. Objectives: To investigate and study the APC protein expression (PE), MMR PE and MSI in CRC based on the three different locations, among Indonesian patients in Adam Malik Hospital, Pirngadi Hospital and other hospitals within the network of the Faculty of Medicine University of Sumatera Utara, Medan Indonesia. Methods: This prospective study was conducted from April to December 2012. The minimum number of sample size patients had been statistically calculated and decided to be 24 patients for each location, as this was a comparative study. The Fresh tissues were obtained either by colonoscopic biopsy or surgery. Inclusion criteria were adenocarcinoma confirmed histologically, and no family history of tumor or CRC. The PE of APC and MMR (MLH1, MSH2, PMS2 and MSH6) were assessed by immunohistochemistry (IHC), while MSI was determined by polymerase chain reaction (PCR) using five microsatellite markers, BAT-25, BAT-26, O2S123, O5S346 and O17S250. The primary antibodies used for IHC examination were: MLH1: Flex monoclonal mouse antibody (Clone: ES05, DAKO); MSH2: (3A2B8C); MSH6: Abeam Inc:

Cambridge MA (ab92471); PMS2: (H-300), sc-11440.lnc, and APC: (F-3): sc- 9998. Biotechnology. Dako Real Envision Rabbit/Mouse was used as Universal

Secondary Antibody. Primers and Characteristics of Microsatellite Loci Analyzed: BAT-25 (Hex), Genbank no. 9834508, U63834; BAT-26 (Tet), Genbank no. 9834505, L47575; O2S123 (Tet), Genbank no. 187953; O5S346 (Fam), Genbank no. 181171, M73547; and O17S250 (Fam), Genbank no. 177030, X54562. Results: Among the 77 CRC, 24 were located in the proximal colon, 24 in distal colon and 29 in rectum. Loss of MMR protein expression (MMR PE negative) was found in 54 out of 77 (70.1 %) CRC patients. Bivariate analysis showed that there were significant differences of MLH 1 protein expression (PE) negative between distal colon and rectal cancer (p=0.008), and MSH6 protein expression negative between proximal colon and rectal cancer (p=0.020). Multivariate analysis showed that MLH 1 protein expression negative was significantly related to the location of the cancer (p=0.006) with Odds Ratio: 0.12 (95% Cl OR: 0.026-0.547). It had 0.12 times probability to be found in distal than rectum. MLH 1 protein expression negative had 1 0 times higher probability to be found in proximal than distal colon (p=0.037). MSH6 protein expression negative was related to the locations (p=0. 026) with Odds Ratio 0.165 (95% Cl OR 0.034-0.803). It had 0.165 times probability to be found in proximal than rectal cancer, or 6.06 times higher probability to be found in rectal cancer than proximal colon cancer, and had 11 times higher probability in distal colon than proximal (p=0.043). APC protein expression negative was also significantly related to the locations (p=0.020), with Odds Ratio: 6.897 (95%CI OR: 1.359-34.995), and had 6.89 times higher probability to be found in distal than rectal cancer, with other variables controlled. MSI-High (MSI-H) was found in 29 out of 77 (37.7%), MSI-Low (MSI-L) and Microsatellite Stable (MSS) in 48 (62.3%). The proportion of MSI-H displayed a tendency to occur in proximal rather than in distal or rectal cancer, but the difference was not significant. Conclusions: This research showed that there were significant differences in the marker of MMR PE negative between proximal, distal colon and rectal cancer. MLH 1 PE negative was prominent in proximal colon cancer, MSH6 PE negative in distal colon and rectal cancer, and APC PE negative in distal colon cancer respectively. Those PE negative were significantly related to the location of the cancer. These findings suggest that the underlying carcinogenic pathway or molecular background differs according to the cancer locations, among CRC patients in this region. The cancer in each colorectal location has its specific molecular characteristics