Chromosomal and microsatellite instability in sporadic gastric cancer

BACKGROUND: Gastric cancer can progress through two pathways of genomic instability: chromosomal (CIN) and microsatellite instability (MSI). It is hypothesized that these two pathways are not always independent and that some tumors show overlap between these two mechanisms. METHODS: A total of 98 sporadic gastric cancers were classified based on their MSI status, using microsatellite assay with BAT26. Evidence for CIN was investigated by identifying loss of heterozygosity (LOH) events on chromosome arms, 5q, 10p, 17p, 17q, and 18q, which are regions harboring tumor suppressor genes that are significant in gastric cancer development. RESULTS: Twelve tumors (12%) showed high-frequency MSI (MSI-H). Overall, 43 of the tumors (44%) had at least one LOH event, with most frequent chromosomal losses observed on 10p and 18q (30%, respectively), followed by 5q (21%), 17p (14%), and 17q (12%). Interestingly, overlap was observed between CIN and MSI pathways. Of 43 cancers with LOH events, four (9%) were also MSI-H. It was also found that 48% of cancers without MSI-H had no LOH events identified, comprising a subgroup of tumors that were not representative of either of these two pathways of genomic instability. CONCLUSION: These results suggest that molecular mechanisms of genomic instability are not necessarily independent and may not be fully defined by either the MSI or CIN pathways in sporadic gastric cancers.     

Microsatellite instability in gallbladder carcinoma: two independent genetic pathways of gallbladder carcinogenesis

Although the genetic basis for gallbladder carcinogenesis has not been clarified, considerable evidence has shown that genetic alterations play an important role in the development and progression of human cancers. In this study, we analyzed 30 gallbladder carcinomas to investigate the role of genetic alterations in their tumorigenesis, and to study correlations with their clinicopathological features. Tissue samples were obtained from 30 patients with gallbladder carcinoma (11 men and 19 women; mean age, 62 years; age range, 38–80 years). Genomic DNAs were extracted from fresh tumor tissue. We examined loss of heterozygosity (LOH) in the p53, APC, DCC, RB, and NM23-H1 gene regions by polymerase chain reaction (PCR)-LOH assay using an automated fluorescent DNA sequencer employing four microsatellite markers (p53, APC, DCC, NM23-H1). Five additional microsatellite markers were used for the determination of microsatellite instability (MSI). LOH was found at p53 in 9 of 15 informative cases (60%), at DCC in 10 of 22 (45%), at APC in 5 of 15 (33%), at RB in 1 of 8 (13%), and at NM23-H1 in 1 of 15 (7%). MSI was observed in 5 of 30 cases (17%) in at least one chromosomal loci of these nine microsatellite markers. None of the patients with MSI-positive tumors showed lymph node metastasis, and there was an inverse correlation between MSI and the presence of LOH in gallbladder carcinoma. These results suggest that there are two independent genetic pathways in gallbladder carcinogenesis; that is, an MSI pathway and an LOH pathway. Received: December 24, 1999 / Accepted: May 26, 2000     

Genetic characterization of colorectal cancers in young patients based on chromosomal loss and microsatellite instability

BACKGROUND: In recent studies a high frequency of microsatellite instability among colorectal cancers in young patients has been reported, but the frequency of microsatellite instability (MSI) and chromosomal instability among colorectal cancers in young patients has not yet been fully elucidated. Only one report showed an increased loss of heterozygosity (LOH) ratio at 9p locus, which harbors tumor suppressor genes p16. The LOH and MSI status among colorectal cancers in young patients was examined. METHODS: Twenty-five patients under 40 years of age diagnosed with colorectal cancer were examined for MSI and LOH using 17 microsatellite markers, and also p16 expression patterns were evaluated by immunohistochemistry and methylation status of the p16 gene was assessed by methylation-specific PCR. RESULTS: MSI was observed in only one case (4%). LOH at 2p, 5q, 9p, 11q, 17p, and 18q was observed in 41%, 59%, 42%, 35%, 46%, and 56% of cases, respectively. Eighty-three percent of patients showed p16-positive expression patterns. Fifty percent of colorectal cancers in young patients exhibited p16 methylation (3/6). CONCLUSIONS: Our study demonstrated that colorectal cancers in young patients without MSI showed a high frequency of LOH at the 9p locus. However, LOH status at 9p and p16 expression pattern did not show a significant correlation. Other tumor suppressor genes on the 9p, with the exception of p16, may play an important role in the carcinogenesis of colorectal cancers in young patients.     

Microsatellite instability in Ewing tumor is not associated with loss of mismatch repair protein expression

Only few clinical factors predict the prognosis of patients with Ewing tumors. Unfavorable outcome is associated with primary metastatic disease, age > 15 years, tumor volume above 200 ml, and the histological response to chemotherapy. The aim of this study was to elucidate the prevalence and clinical impact of microsatellite instability (MSI) together with the relation between MSI and mismatch repair protein expression in Ewing tumors. DNA from 61 primary Ewing tumors and 11 Ewing tumor cell lines was extracted and microsatellite analysis for the detection of instability or loss of heterozygosity was performed for the five markers of the Bethesda panel BAT25, BAT26, D5S346, D2S123, and D17S250, which represents the established marker panel for the analysis of hereditary non-polyposis colorectal carcinoma (HNPCC) patients. In addition, single nucleotide repeat regions of the two tumor genes BAX and transforming growth factor receptor II (TGFBR2) were also included. All of the 61 samples were suitable for LOH analysis and 55 for the determination of MSI-status. LOH of these microsatellite markers was detected in 9 of the 61 patients (14.8%). Over all, genetic instability, i.e. MSI and/or LOH, was detected in 17 tumors (27.9%). One out of the 11 tumor cell lines (STA ET1) was characterized by instability of all the five Bethesda markers, while from primary tumor samples, only one showed MSI in more than one microsatellite marker (D5S346 and D17S250, MSI-high). Eight of the fifty-five patients (14.5%) showed instability of one microsatellite locus (MSI-low). No instability was detected in BAT26, D2S123, BAX and TGFBR2. There was no significant correlation between MSI and loss of expression of mismatch repair proteins MLH1, MSH2, or MSH6. The impairment of the p53 signaling pathway (expression of TP53 and/or MDM2 by immunohistochemistry) was significantly associated with reduced overall survival (15 of 49 patients (30.6%), P = 0.0410, log-rank test). We conclude that MSI is not prevalent in Ewing tumor and that the nature of instability differs from the form observed in colorectal carcinoma, the model tumor of MSI. This is documented by the different pattern of MSI (no BAT26 instability) in Ewing tumors and the lack of a strict correlation between MSI-high and loss of expression of MSH2, MSH6 and MLH1. IA and KLS contributed equally to this study.     

慢性白血病微卫星不稳定性和杂合性缺失的研究

应用PCR扩增技术检测17例慢性白血病患者不同染色体上8个微卫星位点的微卫星不稳定性（MSI）和杂合性缺失（LOH）,同时检测11例健康志愿者的MSI和LOH,以作对照。结果发现,健康对照组所选微卫星位点均未发生MSI或LOH;9例处于慢性白血病加速期的人中有7例发生至少一个位点的MSI,8例慢性期患者中仅1例发生一个位点的MSI,慢性白血病加速期的MSI发生率明显高于其慢性期（P＜0.05>,提示微卫星的遗传不稳定性可能与慢性白血病的病情进展有关。17例慢性白血病中只有1例在急变后出现LOH,提示所选位点的LOH可能不是慢性白血病的多发事件。     

The CpG island methylator phenotype and chromosomal instability are inversely correlated in sporadic colorectal cancer

BACKGROUND & AIMS: The CpG island methylator phenotype (CIMP) is one of the mechanisms involved in colorectal carcinogenesis (CRC). Although CIMP is probably the cause of high-frequency microsatellite instability (MSI-H) sporadic CRCs, its role in microsatellite stable (MSS) tumors is debated. The majority of MSS CRCs demonstrate chromosomal instability (CIN) with frequent loss of heterozygosity (LOH) at key tumor suppressor genes. We hypothesized that the majority of sporadic CRCs without CIN would be associated with CIMP. METHODS: We tested 126 sporadic CRCs for MSI and LOH and categorized tumors into MSI, LOH, or MSI-/LOH- subgroups. Methylation status was evaluated using 6 CIMP-related markers (MINT1, MINT2, MINT31, p16(INK4alpha), p14(ARF), and hMLH1) and 6 tumor suppressor genes (PTEN, TIMP3, RUNX3, HIC1, APC, and RARbeta2). BRAF V600E mutation analysis was performed using allele-specific polymerase chain reaction and DNA sequencing. RESULTS: We observed frequent methylation at all 12 loci in all CRCs. BRAF V600E mutations correlated with the MSI (P < .0001) and MSI-/LOH- (P = .03) subgroups. MSI and MSI-/LOH- tumors exhibited more promoter methylation than CRCs with LOH (P < .0001). We also found an inverse correlation between the frequencies of methylation and LOH (rho = -0.36; P < .0001). CONCLUSIONS: The associations between methylation frequencies at CIMP-related markers and MSI or MSI-/LOH- sporadic CRCs suggest that the majority of these tumors evolve through CIMP. These findings suggest that CIN and CIMP represent 2 independent and inversely related mechanisms of genetic and epigenetic instability in sporadic CRCs and confirm that MSI cancers arise as a consequence of CIMP.     

Genetic instability and aberrant DNA methylation in chronic hepatitis and cirrhosis--A comprehensive study of loss of heterozygosity and microsatellite instability at 39 loci and DNA hypermethylation on 8 CpG islands in microdissected specimens from patients with hepatocellular carcinoma

A study was conducted to examine the significance of genetic instability and aberrant DNA methylation during hepatocarcinogenesis. Genomic DNA was extracted from 196 microdissected specimens of noncancerous liver tissue that showed no marked histologic findings or findings compatible with chronic hepatitis or cirrhosis, and 80 corresponding microdissected specimens of hepatocellular carcinoma (HCC) from 40 patients. Loss of heterozygosity (LOH) and microsatellite instability (MSI) were examined by polymerase chain reaction (PCR) using 39 microsatellite markers, and DNA methylation status on 8 CpG islands was examined by bisulfite-PCR. In noncancerous liver tissues, LOH, MSI, and DNA hypermethylation were found in 15 (38%), 6 (15%), and 33 (83%) of 40 cases, respectively. The incidence of DNA hypermethylation in histologically normal liver was similar to that in chronic hepatitis and cirrhosis, although neither LOH nor MSI was found in histologically normal liver. In cancerous tissues, LOH, MSI, and DNA hypermethylation were found in 39 (98%), 8 (20%), and 40 (100%) of 40 cases, respectively. CpG islands of the p16 gene and methylated in tumor 1, 2, 12, and 31 clones were frequently methylated in cancerous tissues, although neither the thrombospondin-1 nor the human Mut L homologue (hMLH1) gene was methylated. Absence of silencing of the hMLH1 gene by DNA hypermethylation is consistent with the low incidence of MSI in HCCs. The results of this study indicate that LOH and aberrant DNA methylation contribute to hepatocarcinogenesis; DNA hypermethylation in particular, which precedes or may even cause LOH, is as an early event during hepatocarcinogenesis.     

Microsatellite instability is rare in the clinical course of myelodysplastic syndrome studied with DNA from fresh and paraffin-embedded tissues

Microsatellite instability (MSI) has been reported to occur in various types of malignant neoplasms. We performed a polymerase-chain-reaction-based assay for MSI between the initial and the most recently available (“latest”) samples from 23 patients with myelodysplastic syndrome (MDS). Of these patients, 15 were informative at more than three microsatellite loci. Seven patients showed an increase in leukemic cells while 8 patients did not during the interval between the two analyses. Only 1 of the patients, who had refractory anemia with excess blasts, which changed to acute myelogenous leukemia, showed microsatellite alteration at the analysis times. Among all 23 patients, two alterations were detected in the 42 informative paired samples that showed an increase in leukemic cells (4.8%), while none was detected in the 59 paired samples without such an increase. In total, therefore only two alterations were detected among 101 informative paired samples (2%). This indicates that MSI is rare in the clinical course of MDS irrespective of disease status, and is consequently not a critical genetic event for disease progression in most MDS patients. Received: 26 November 1997 / Accepted: 15 January 1998     

Epithelial and stromal genetic instability contributes to genesis of colorectal adenomas

BACKGROUND: Previously, we indicated that stromal genetic instability might contribute to tumorigenesis of both sporadic and ulcerative colitis associated colorectal adenocarcinomas. Considering the established adenoma-adenocarcinoma sequence, in this study we analysed genetic instability in colorectal adenoma cells and surrounding stroma. METHODS: In 164 colorectal tumours (34 hyperplastic polyps, 38 tubular adenomas with low grade dysplasia (TA-L), 51 tubular adenomas with high grade dysplasia (TA-H), and 41 invasive carcinomas), epithelial and stromal genetic instability with National Cancer Institute standard microsatellite markers and chromosome 17 (Chr17) markers, were analysed by a combination of laser capture microdissection and GeneScan approaches. RESULTS: While frequencies of both loss of heterozygosity (LOH) and microsatellite instability (MSI) were extremely low in hyperplastic polyps, LOH in tubular adenomas was detected in both epithelial (TA-L 13.2%, TA-H 27.5%) and stromal (5.3% and 5.9%, respectively) elements, along with MSI (5.3% and 13.7%, and 5.3 and 5.9%, respectively). Frequencies of epithelial alterations were higher in TA-H than in TA-L, and greatest in the carcinoma group. On the other hand, frequencies of stromal LOH or MSI were almost constant (5.3% approximately 17.1%, 5.3% approximately 17.1%, respectively) in adenomas and invasive carcinomas. In addition, p53 was found to be significantly overexpressed in a greater proportion of TA-L with LOH than in those without genetic instability. CONCLUSION: The results indicate the presence of genetic alterations in stroma from an early stage of carcinogenesis, accompanied by stepwise increasing genetic instability of epithelia with progression to cancer. Thus microenvironmental changes due to genetic alteration in Chr17 markers in stromal cells may play an important role in colon adenoma and adenocarcinoma development.     

High level of microsatellite instability but not hypermethylation of mismatch repair genes in therapy-related and secondary acute myeloid leukaemia and myelodysplastic syndrome

Microsatellite instability (MSI) is associated with defects in the DNA mismatch repair (MMR) system, such as mutation or epigenetic silencing of the genes by promoter hypermethylation. We investigated the presence of MSI and promoter hypermethylation of hMLH1 and hMSH2 genes in 82 patients (68 acute myeloid leukaemia, AML; 14 myelodysplastic syndromes, MDS). Twelve separate microsatellite loci, including three mononucleotide repeat markers, were used. Mutator phenotype (RER+) was detected in 20 AML (29.4%) and 3 MDS (21.4%) patients. RER+ rate was much higher in the therapy-related and secondary cases compared with the de novo cases. Three out of 7 (42.9%) secondary (s-AML) and 8 out of 17 (47.1%) therapy-related (t-AML) showed RER+ in comparison with 9 out of 44 (20.5%) de novo cases. Similar rates were detected in MDS patients (2/2 therapy-related and 1/12 de novo). The promoter hypermethylation was found in three hMLH1 (3.7%) and two hMSH2 (2.4%) genes. All these five patients had AML and were older than 60 years of age. Two of them had s-AML and one had t-AML. RER+ was detected in three of these five patients. Our data suggest that genetic instability is associated with AML and MDS, especially t-AML and s-AML. In addition, our results indicate that the hMSH2 and hMLH1 promoter hypermethylation is not a common event in these malignancies, but may play a role in the development of AML in elderly patients.     

Relationship between nm23H1 genetic instability and clinical pathological characteristics in Chinese digestive system cancer patients

AIM： To study the relationship between nm23H1 gene genetic instability and its clinical pathological characteristics in Chinese digestive system cancer patients.
METHODS： Polymerase chain reaction-single strand conformation polymorphism （PCR-SSCP） was used to analyze the microsatellite instability （MSI） and loss of heterozygosity （LOH）. Immunohistochemistry was employed to detect the expression of nm23H1.
RESULTS： The MSI was higher in TNM stage Ⅰ ＋ Ⅱ than in stage Ⅲ＋ Ⅳ of gastric, colonic and gallbladder carcinomas. The LOH was higher in TNM stage Ⅲ＋ Ⅳ than in stage Ⅰ＋ Ⅱ of gastric, colonic and hepatocellular carcinomas. Lymphatic metastasis was also observed. The expression of nm23H1 protein was lower inTNM stage Ⅲ ＋ Ⅳ than in stage Ⅰ ＋ Ⅱ of these tumors and in patients with lymphatic metastasis.The nm23H1 protein expression was higher in the LOH negative group than in the LOH positive group.
CONCLUSION： MSI and LOH may independently control the biological behaviors of digestive system cancers. MSI could serve as an early biological marker of digestive system cancers. Enhanced expression of nm23H1 protein could efficiently inhibit cancer metastasis and improve its prognosis. LOH mostly appears in late digestive system cancer.     

Detection of frameshift mutations of RIZ in gastric cancers with microsatellite instability

AIM: To study the frameshift mutations of the retinoblastoma protein-interacting zinc finger gene RIZ in gastric cancer with microsatellite instability, and to identify two coding polyadenosine tracts of RIZ. METHODS: Frameshift mutations at (A)8 and (A)9 tracts of RIZ were detected in 70 human gastric cancer (HGC) specimens by DHPLC and DNA sequencing. Microsatellite instability (MSI) status was assessed by two mononucleotide markers, BAT26 and BAT25, by means of denaturing high-performance liquid chromatography (DHPLC). RESULTS: In 70 HGC samples, 8 (11.4%) were found positive for instabilities at BAT26 and BAT25. In 7 of the 8 cases with instabilities at both BAT26 and BAT25 (MSI-H), 1 was unstable at BAT26 but stable at BAT25. Frameshift mutations were identified in 4 (57.1%) of the 7 samples with MSI-H in the (A)9 tract of RIZ without mutations in the (A)8 tract. In contrast, frameshift mutations were found in neither of the polyadenosine tracts in 63 samples of MSI-L or MSI stable tumors. Pro704 LOH detection in 4 cases with frameshift mutations did not find LOH in these cases. CONCLUSION: Frameshift mutations of RIZ may play an important role in gastric cancers with MSI.     

Genetic instability of BRCA1 gene at locus D17S855 is related to clinicopathological behaviors of gastric cancer from Chinese population

AIM: To investigate genetic instability of gene BRCAl at locus D17S855, and their relationship with clinicopatho-logical characteristics of gastric cancer in Chinese population. METHODS: Microsatellite instability (MSI) and loss of heterozygosity (LOH) of gene BRCAl at locus D17S855 were compared between 37 samples of gastric cancer and corresponding non-cancerous gastric tissue. RESULTS: MSI at locus D17S855 was positive in 7 of 37 samples of gastric cancer (18.95%). MSI had a close relationship with TNM staging but no relation with lymph node metastasis, histological type or tumor differentiation. MSI positive frequency in TNM I II (31.58%, 6/19) was much higher than that in TNM III IV (5.56%, 1/18), (P < 0.05). LOH positive rate was 18.92% (7/37). LOH had no relationship to histological type, tumor differentiation or lymph node metastasis, but LOH positive rate in TNM III IV was 33.33% (6/18), much higher than that in TNM I II ( 5.26%, 1/19), (P < 0.05). BRCAl protein was expressed in 14 of 37 samples of gastric cancer. The positive rates of BRCAl protein in TNM I II and TNM III IV were 57.89% and 16.67%, respectively, {P < 0.05). The positive rate of BRCAl protein was 77.78% in high differentiation samples, 30.77% in middle differentiation and 12.50% in lower differentiation samples, (P < 0.05). CONCLUSION: MSI of BRCAl gene could be used as a molecular marker in early phases of sporadic gastric cancer in Chinese population. LOH occurs at later period of gastric cancer, therefore, it could be used as prognostic factor.     

Early genetic instability of both epithelial and stromal cells in esophageal squamous cell carcinomas, contrasted with Barrett's adenocarcinomas

Background We have shown previously that stromal genetic alteration may make a greater contribution to early genesis of ulcerative colitis-associated tumors than sporadic colon cancers. We assessed whether similar differences in genetic alteration might exist between squamous cell carcinomas (SCCs) and Barrett's adenocarcinomas (BACs) of the esophagus. Methods We investigated epithelial and stromal genetic instability with five National Cancer Institute standard (NCI), four chromosome 17 (Chr. 17), and six tumor suppressor gene (TSG) microsatellite markers in 26 SCC and 12 BAC cases and in 11 normal controls, using a novel combination of microdissection, polymerase chain reaction, and GeneScan. Results Frequency of epithelial loss of heterozygosity (LOH) increased in the order background mucosa, to precursor lesions, to tumors with both types of carcinoma, especially for the Chr. 17 and TSG markers, while stromal LOH was relatively high but consistent from background mucosa to carcinoma. Epithelial LOH of D17S796 demonstrated a significantly higher frequency in SCCs than in BACs, without significant variation in p53 overexpression. The frequency of microsatellite instability (MSI) showed constant high levels in both epithelium and stroma of background, dysplasia, and carcinomas in the SCC series, and rather low frequencies in the BAC series. Although epithelial hMLH1 and hMSH2 expression decreased with tumor progression, no correlation was found with the individual MSI status. Conclusions Although epithelial LOH exists similarly in both lesion types, whereas epithelial and stromal MSI may occur in a relatively early phase of SCC development, stromal MSI is rare in BACs, strongly suggesting differences in tumorigenesis between the two types.     

胃癌微卫星不稳定性和抑癌基因杂合缺失

目的研究微卫星不稳和抑癌基因缺失在胃癌发生中的作用．方法采用ＰＣＲ为基础的方法,检测了５３例胃癌中６个微卫星标记突变及ＡＰＣ／ＭＣＣ和ＤＣＣ基因杂合缺失（ＬＯＨ）．结果胃癌微卫星不稳的检出率为３２１％（１７／５３）．７例（１３２％）为微卫星高频率不稳（３个以上微卫星标志）,１０例（１８９％）为微卫星低频率不稳（１或２个微卫星标记）．肠型胃癌微卫星高频率不稳的发生率（２５０％）显著高于弥漫型胃癌（３４％）（Ｐ＜００５）．高频率不稳组未发现有ＡＰＣ,ＭＣＣ和ＤＣＣ基因ＬＯＨ,微卫星高频率不稳与ＡＰＣ／ＭＣＣ和ＤＣＣ基因ＬＯＨ呈负相关．结论微卫星不稳在部分胃癌,特别是肠型胃癌早期发生中起重要作用,高频率不稳胃癌与遗传性非息肉大肠癌有共同的特点．与此相反,低频率不稳和无不稳胃癌可能通过ＬＯＨ病理途径发生     

Telomere erosion is independent of microsatellite instability but related to loss of heterozygosity in gastric cancer

AIM: To correlate the length of the telomere to microsatellite instability (MSI) and loss of heterozygosity (LOH) of APC, MCC and DCC genes in gastric carcinomas. METHODS: Telomeric restriction fragment (TRF) length of gastric cancer was measured with Southern blot. LOH of APC, MCC and DCC genes, microsatellite instability (MSI) and frameshift mutation of hMSH6, TGF-betaRII and BAX genes were analyzed by PCR-based methods. RESULTS: Sixty-eight cases of sporadic gastric carcinoma were studied for MSI using five microsatellite markers. MSI in at least one locus was detected in 17 (25%) of 68 tumors analyzed. Frameshift mutations of hMSH6, TGF-betaRII and BAX were detected in 2,6 and 3 of gastric carcinomas respectively showing high MSI (> or = 2 loci, n = 8), but none was found in those showing low MSI (only one locus, n = 9) or MSS (tumor lacking MSI or stable, n = 51). Thirty-five cases, including all high MSI and low MSI, were studied for TRF. The mean TRF length was not correlated with clinicopathological parameters. No association was observed between TRF length and MSI or frameshift mutation. On the contrary, LOH at the DCC locus was related to telomere shortening (P<0.01). This tendency was also observed in APC and MCC genes, although there was no statistical significance. CONCLUSION: The development of gastric cancer can arise through two different genetic pathways. In high MSI gastric cancers, defective mismatch repair allows mutations to accumulate and generate the high MSI phenotype. In gastric cancers showing either low MSI or MSS, multiple deletions may represent the LOH pathway. Telomere erosion is independent of high MSI phenotype but related to the LOH pathway in gastric cancer.     

Loss of heterozygosity： An independent prognostic factor of colorectal cancer

AIM: Colorectal cancers result from the accumulation of several distinct genetic alterations. This study was to investigate the frequency and prognostic value of loss of heterozygosity (LOH) and microsatellite instability (MSI) at 14 genetic loci located near or within regions containing important genes implicated in colorectal tumorigenesis. METHODS: We studied colorectal cancers with corresponding normal mucosae in 207 patients (139 males and 68 females, mean age at the time of tumor resection 66.2±12.4 years, range 22-88 years). There were 37 right-sided colonic tumors, 85 left-sided colonic tumors and 85 rectal tumors. The distribution of tumor staging was stage Ⅰ in 25, stage Ⅱ in 73, stage Ⅲ in 68, and stage Ⅳ in 41. We analyzed the LOH and MSI of HPC1, hMSH2, hMLH1, APC, MET, P53, NH23-H1, DCC, BAT25, BAT26, D17S250, MYCL1 and D8S254 with fluorescent polymerase chain reaction and denatured gel electrophoresis. High-frequency LOH was determined to be greater than three, or more than 50% of the informative marker with LOH. High-frequency MSI (MSI-H) was determined as more than four markers with instability (>30%). Correlations of LOH and MSI with clinical outcomes and pathological features were analyzed and compared. RESULTS: The occurrence of MSI-H was 7.25%, located predominantly in the right colons (7/15) and had a higher frequency of poor differentiation (6/15) and mucin production (7/15). LOH in at least one genetic locus occurred in 78.7% of the tumors and was significantly associated with disease progression. Of the 166 potentially cured patients, 45 developed tumor recurrence within 36 mo of follow-up. Clinicopathological factors affecting 3-year disease-free survival (DFS) were TNM staging, grade of differentiation, preoperative CEA level, and high LOH status. Patients with high LOH tumors had a significantly lower DFS (50%) compared with patients with low LOH tumors (84%). Of the patients developing subsequent tumor recurrence, the number and percentage of LOH were 2.97 and 46.8% respectively, similar to the stage IV disease patients. TNM staging had the most significant impact on DFS, followed by high LOH status. CONCLUSION: Clinical manifestations of LOH and MSI are different in colorectal cancer patients. High-frequency LOH is associated with high metastatic potential of colorectal cancers.     

Prognostic significance of microsatellite alterations at tp36 in cholangiocarcinoma

AIM: To investigate loss of heterozygosity (LOH) and microsatellite instability (MSI) on the chromosomal region 1p36-pter in cholangiocarcinoma (CCA) patients and determine the association between microsatellite alterations and clinicopathological parameters.METHODS: Ten polymorphic microsatellite markers were determined for LOH and MSI using GS-3000 gel scan fragment autoanalyzer.RESULTS: Sixty-eight out of 90 cases (75.6%) showed LOH in one or more loci. LOH was found most frequently at D1S199 (40.0%), D1S507 (34.6%), D1S2845 (30.5%),and D1S2734 (30.1%). MSI was found in 34 of 90 cases (37.8%) at one or more loci. Fine mapping at 1p36 showed two distinctive regions of common loss, which were D1S2845 and the 25.5-cM region between D1S507and D1S2734, indicating the existence of putative tumor suppressor genes that is likely to play important roles in the development of CCA. Patients with LOH at D1S234 showed less lymphatic invasion (P = 0.017),whereas patients with LOH at D1S2676 exhibited more lymphatic invasion than those without (P = 0.031).LOH at D1S2845 showed a significant correlation with nerve invasion (P = 0.029). Moreover, patients who demonstrated MSI at D1S228 showed a poor prognosis (P = 0.0026).CONCLUSION: Allelic loss plays a major role in microsatellite alterations at chromosome 1p36, which may contribute to carcinogenesis and pathogenesis of liver fluke related CCA and these alterations can be used as molecular prognostic indicators for CCA patients.     

Prognostic significance of microsatellite alterations at 1p36 in cholangiocarcinoma

AIM: To investigate loss of heterozygosity (LOH) and microsatellite instability (MSI) on the chromosomal region 1p36-pter in cholangiocarcinoma (CCA) patients and determine the association between microsatellite alterations and clinicopathological parameters. METHODS: Ten polymorphic microsatellite markers were determined for LOH and MSI using GS-3000 gel scan fragment autoanalyzer. RESULTS: Sixty-eight out of 90 cases (75.6%) showed LOH in one or more loci. LOH was found most frequently at D1S199 (40.0%), D1S507 (34.6%), D1S2845 (30.5%), and D1S2734 (30.1%). MSI was found in 34 of 90 cases (37.8%) at one or more loci. Fine mapping at 1p36 showed two distinctive regions of common loss, which were D1S2845 and the 25.5-cM region between D1S507 and D1S2734, indicating the existence of putative tumor suppressor genes that is likely to play important roles in the development of CCA. Patients with LOH at D1S234 showed less lymphatic invasion (P = 0.017), whereas patients with LOH at D1S2676 exhibited more lymphatic invasion than those without (P = 0.031). LOH at D1S2845 showed a significant correlation with nerve invasion (P = 0.029). Moreover, patients who demonstrated MSI at D1S228 showed a poor prognosis (P = 0.0026). CONCLUSION: Allelic loss plays a major role in microsatellite alterations at chromosome 1p36, which may contribute to carcinogenesis and pathogenesis of liver fluke related CCA and these alterations can be used as molecular prognostic indicators for CCA patients.