T25 Repeat in the 3′ Untranslated Region of the CASP2 Gene: A Sensitive and Specific Marker for Microsatellite Instability in Colorectal Cancer

Abstract

DNA mismatch repair deficiency is observed in about 10% to 15% of all colorectal carcinomas and in up to 90% of hereditary nonpolyposis colorectal cancer (HNPCC) patients. Tumors with mismatch repair defects acquire mutations in short repetitive DNA sequences, a phenomenon termed high-level microsatellite instability (MSI-H). The diagnosis of MSI-H in colon cancer is of increasing relevance, because MSI-H is an independent prognostic factor in colorectal cancer, seems to influence the efficacy of adjuvant chemotherapy, and is the most important molecular screening tool to identify HNPCC patients. To make MSI typing feasible for the routine pathology laboratory, highly reproducible and cost effective laboratory tests are required. Here, we describe a novel T 25 mononucleotide marker in the 3Vuntranslated region of the CASP2 gene (CAT25) that displayed a quasimonomorphic repeat pattern in normal tissue of 200 unrelated individuals of Caucasian origin. In addition, CAT25 was monomorphic also in all tested donors of African and Asian origin (n = 102 and n = 79, respectively) and thus differs from the most commonly used markers BAT25 and BAT26. Without the analysis of corresponding normal tissue, CAT25 correctly detected 56 of 57 colorectal cancer specimens classified as MSI-H by using the standard National Cancer Institute/International Collaborative Group-HNPCC marker panel. Combined with the standard markers BAT25 and BAT26 in a multiplex PCR, all MSI-H colorectal cancer samples were typed correctly. No falsepositive results were obtained in 60 non-MSI-H control colorectal cancer specimens. These data suggest that CAT25 should be included into novel marker panels for microsatellite testing thus allowing for a significant reduction of the complexity and costs of MSI typing. Moreover, CAT25 represents a highly promising marker for early detection of colorectal cancer in HNPCC germ line mutation carriers. (Cancer Res 2005; 65(18): 8072-8)

Genetics and epigenetics of small bowel adenocarcinoma: the interactions of CIN, MSI, and CIMP

“…MSI analysis was performed using the standard NCI/ICG-hereditary nonpolyposis colorectal cancer marker panel 27 and CAT25. 28 PCR products were separated on an ABI3700 automated sequencer (Applied Biosystems, Warrington, UK). Tumors were classified as MSI-H if at least two of the markers displayed a shift in product length compared with the product of non-neoplastic mucosa.…”

High density of FOXP3-positive T cells infiltrating colorectal cancers with microsatellite instability

“…Tumours were typed for MSI using the standard NCI/ ICG-HNPCC marker panel (Boland et al, 1998) and CAT25 as described earlier (Findeisen et al, 2005). For tumour staging, the UICC/AJCC TNM system was applied (American Joint Committee on Cancer, 1997).…”

Detection of Microsatellite Instability in Colorectal Cancer Using an Alternative Multiplex Assay of Quasi-Monomorphic Mononucleotide Markers

“…PCR primers for the amplification of these markers were described elsewhere. 15,16 The sense primers were chemically labeled at the 5Ј end with 5-carboxyfluorescein fluorescent dyes. PCR was performed in a total volume of 25 l using a final concentration of 200 mol/L deoxyribonucleotide triphosphates (MBI Fermentas, St. Leon-Rot, Germany), 500 nmol/L each sense and antisense primer (Eurogentec, Seraing, Belgium), 1X PCR buffer (60 mmol/L Tris-SO 4 , pH 8.9; 18 mmol/L (NH 4 )SO 4 ; 2 mmol/L MgSO 4 ) and 1 unit of Discoverase denaturing high performance liquid chromatography DNA polymerase (Invitrogen, Merelbeke, Belgium).…”

“…Four of these mononucleotide repeats (PTHL3, SEC63, HPDMPK, and U79260) showed mutation rates in 80% or more of MSI-H CRCs and were suggested as new candidate genes for diagnostic purposes. Similarly, Findeisen et al 16 described a novel T 25 mononucleotide marker in the 3Ј untranslated region of the CASP2 gene (CAT25) that displayed a quasimonomorphic repeat pattern in normal tissue and represented a highly promising candidate marker for future highthroughput MSI testing.…”