metallothionein and Microsatellite-Instability

Colorectal malignancies with high microsatellite instability (MSI-H), either hereditary (Lynch syndrome) or sporadic, demonstrate better prognosis and altered response to 5FU chemotherapy. It is now recommended to perform MSI testing for all new cases of colorectal cancer regardless of being categorized as hereditary or sporadic. For MSI detection, immunohistochemistry or PCR-based protocols using a cohort of various sets of STR markers are recommended. Here we aimed to evaluate a simplified protocol using just a single STR marker, MT1XT20 mononucleotide repeat, for detection of MSI in Lynch syndrome patients. A Promega five-marker MSI testing panel and immunohistochemistry (IHC) were used as the gold standard in conjunction with MT1XT20.. Colorectal patients with a positive history of familial cancers were selected by evaluating medical records. Based on Amsterdam II criteria for Lynch syndrome 20 families were short listed. DNA was extracted from formalin fixed paraffin embedded tumour and adjacent normal tissues resected from the index case in each family. Extracted DNA was subjected to MT1XT20 mononucleotide marker analysis and assessment with a commercially available five marker MSI testing kit (Promega, USA). IHC also was performed on tissue sections and the results were compared with PCR based data.. Eight (40%), seven (35%) and five (25%) cases were MSI positive using with the Promega kit, IHC and MT1XT20, respectively. Among the markers included in Promega kit, BAT26 marker showed instability in all 8 samples. NR24 and NR21 markers showed instability in 7 (87.5%), and BAT25 and MONO 27 in 6 (75%) and 5 (62.5%).. Although MT1XT20 was earlier reported as a valid standalone marker for MSI testing in CRC patients, we could not verify this in our Iranian patients. Instead BAT26 among the markers included in Promega MSI testing kit showed instability in all 8 MSI-H CRC samples. Therefore, it seems BAT26 could act well as a single marker for MSI testing in Iranian CRC patients.

Topics: Adenocarcinoma; Adult; Biomarkers, Tumor; Colorectal Neoplasms, Hereditary Nonpolyposis; Female; Follow-Up Studies; Humans; Male; Metallothionein; Microsatellite Instability; Microsatellite Repeats; Middle Aged; Neoplasm Staging; Persia; Prognosis; Young Adult

Stratifying patients defective in mismatch repair (dMMR) with high microsatellite instability (MSI-H) in colorectal cancer (CRC) is of increasing relevance and may provide a more tailored approach to CRC adjuvant therapy. Here, we describe the discovery of a new MSI marker for colorectal cancer located in the 3'-untranslated region (3'UTR, T20 mononucleotide repeat) of the metallothionein 1X gene (MT1XT20).. We studied 340 consecutive CRCs using three multiplexed polymerase chain reactions amplifying BAT25, BAT26, TGFBR2, MybT22, BAT40, MT1XT20, NR21, NR24, CAT25, D2S123, D5S346, D17S250, D18S58, CSF1PO, D7S820, and D18S51. Fragments length was evaluated by automated capillary electrophoresis.. Based on the NCI/ICG-HNPCC criteria for MSI classification, 40 CRCs were found to be MSI-high (11.8%), 46 (13.5%) CRCs were MSI-low, and 254 CRCs (74.7%) were stable (MSS). MT1XT20 showed very high sensitivity (97.3%) comparable to BAT26 (97.5%) and CAT25 (97.1%) and the best specificity (100%) as well as MybT22 and CAT25. Indeed, MT1XT20 instability was detected in 36 out of 37 cases (97.3%) of MSI-high colorectal cancers, whereas no MT1XT20 alterations were observed in 254 MSS or in 46 MSI-low cases. On the contrary, BAT40 was found to be unstable in 8/46 MSI-low cases, BAT25 in 6/46, BAT26 4/46, NR21 1/46, and NR24 in 1/45.. Our results suggest that MT1XT20 represents a sensitive and specific marker for MSI testing and could be included in a complete set of MSI markers for the confident identification of familial or sporadic dMMR patients in CRCs.

Topics: 3' Untranslated Regions; Adult; Aged; Aged, 80 and over; Biomarkers, Tumor; Colorectal Neoplasms; DNA Mismatch Repair; Female; Genetic Markers; Humans; Male; Metallothionein; Microsatellite Instability; Polymerase Chain Reaction; Sensitivity and Specificity; Tandem Repeat Sequences

Tea polyphenol has been shown to have anti-colorectal cancer and anti-gene mutation effects, although the mechanism of inhibition of microsatellite instability (MSI) colorectal cancer is not known.. Using LoVo, HCT-116, HT-29, and SW480 cells treated with an aqueous solution of tea polyphenol, cell proliferation was detected by the methyl thiazolyl tetrazolium method, changes in microsatellite sequences by the Genescan method and changes in the gene expression of LoVo cells using Illumina expression arrays.. The proliferation inhibition rate of LoVo, HCT-116, HT-29, and SW480 cells treated with tea polyphenol increased with increasing drug concentration and showed an increasing tendency with time. The proliferation inhibition rate of LoVo and HCT-116 cells with tea polyphenols was higher than that of HT-29 and SW480 cells, and there was a significant difference in the proliferation inhibition rate at 24, 72 h and 1 week. The microsatellite sequence of LoVo cells treated with tea polyphenols remained stable.. The gene expression arrays and quantitative RT-PCR suggested that tea polyphenol inhibited the gene expression of metallothionein 2A (MT2A), transcription factor (MAFA), hairy and enhancer of split 1 (HES1), and jagged1 (JAG1) nearly twofold over controls. It was also found that tea polyphenol inhibited the BAX and p38 genes with a more than twofold difference but did not significantly inhibited the NFκB pathway.. Tea polyphenol significantly inhibited the proliferation of MSI colorectal cancer signals maintained stable at the microsatellite state in MSI colorectal cancer. Tea polyphenol inhibited the gene expression of HES1, JAG1, MT2A, and MAFA but upregulated the gene expression of BAX and downregulated that of (P)38. Further research is required to investigate how these pathways are interrelated.

Topics: Basic Helix-Loop-Helix Transcription Factors; bcl-2-Associated X Protein; Calcium-Binding Proteins; Cell Line, Tumor; Cell Proliferation; Colorectal Neoplasms; Dose-Response Relationship, Drug; Flavonoids; Gene Expression Regulation, Neoplastic; Homeodomain Proteins; Humans; Intercellular Signaling Peptides and Proteins; Jagged-1 Protein; Maf Transcription Factors, Large; Membrane Proteins; Metallothionein; Microsatellite Instability; p38 Mitogen-Activated Protein Kinases; Phenols; Polyphenols; Serrate-Jagged Proteins; Tea; Transcription Factor HES-1