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.     

Lymphatic metastasis and nm23H1 genetic instability in Chinese colon cancer patients

AIM: To investigate the pathogenic mechanism of colon cancer at the molecular level and to elucidate the relationship between intercellular adhesion molecule-1 (ICAM-1) and nm23H1 genes and Chinese patients with colon cancer.METHODS: DNA was extracted from paraffin-embedded materials. Polymerase chain reaction-single s~and conformation polymorphism (PCR-SSCP) was used to analyze MSI and LOH. Expression of ICAM-1 was detected by Envision immuno-histochemistry. Experimental results were analyzed with Leica-Qwin computer imaging techniques and SPSS software of statistics.RESULTS: ICAM-1 expression of lymphatic endothelium was negative in normal colon and posi0ve in colon cancer respectively. The number of lymphatics positive for ICAM-1 was gradually increased with degree of cancer invasion (P&lt;0.01). In the group with metastasis of colon cancer, bhe number of lymphatics positive for ICAM-1 in lymph nodes was more than that in the group with no metastasis (P&lt;0.01). The frequency of MSI, LOH and nm23H1 protein was 26.67%, 20.00% and 53.33% in colon cancer, respectively. In TNM staging, MSI (43.75%) and nm23H1 protein (81.25%) in stages I+II weredetected more easily than the corresponding indexes (MSI:7.14%, P&lt;0.05 and nm23H,: 21.43%, P&lt;0.01) in stages III+IV. By comparison, the frequency of LOH (35.71%) in stages III+IV was more than that of LOH (6.25%, P&lt;0.05) in stages I+II. LOH exhibited a rising trend along with the Duke‘s staging, nm23H1 protein in the group of tubular adenocarcinoma (60.00%) was higher expressed than that in the group of mucoid adenocarcinoma (20.00%) (P&lt;0.01),and exhibited a rising trend with the differentiation degrees of tubular adenocarcinoma, nm23H, protein in MSI positive group was higher expressed (75%) than that in MSI negative group (45.45%, P&lt;0.05).CONCLUSION: The expression of ICAM-1 in lymphatic vessels is beneficial to the judgement of the invasion and metastasis ability of colon cancer and the anti-tumor immunity function,and shows an important clinical significance in predicting lymphatic metastasis of colon cancer. MSI and LOH may separately control the development of sporadic colon cancer with different pathways. LOH mostly arises in the late period of sporadic colon cancer and endows a high aggressive and poor prognostic phenotype. By compassion, MSI may be an early period molecule marker for sporadic colon cancer, enhanced expression of nm23H1 protein can effectively inhibit colon cancer metastasis and improve prognosis of sporadic colon cancer patients.     

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.     

The correlation of genetic instability of PINX1 gene to clinico-pathological features of gastric cancer in the Chinese population

Purpose  This project explored the influence of loss of heterozygosity (LOH) and microsatellite instability (MSI) of locus D8S277 to PINX1 expression of gastric cancer in Chinese people. Methods  LOH and MSI of locus D8S277 in 90 paraffin-embedded gastric carcinoma specimens were detected by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP). Envision immunohistochemistry was used to assess the expression of PINX1. Results  The frequency of LOH was higher in cases with lymph node metastasis than those without (18.57 vs. 0.00%, P < 0.01), and was higher in the specimens that were at TNM stage III + IV than those at stage I + II (21.43 vs. 2.94%, P < 0.01). In terms of the frequency of MSI, it was lower in cases with lymph node metastasis than those without (10.00 vs. 30.00%,P < 0.05). The positive rate of PINX1 protein was higher in samples without lymph node metastasis than those with lymph node metastasis (80.00 vs. 50.00%, P < 0.01); and was higher in the cases at TNM stage I + II than those at stage III + IV; and was lower in the cases between 40 and 60 years old than those above 60 years old (43.75 vs. 65.52%, P < 0.05). Conclusion  LOH and MSI of PINX1 may play major role in tumor development and regulate it through different pathways. Because LOH plays a major role in negative expression of PINX1, it can be regarded as a sign of gastric cancer development and MSI may affect the prognosis and tumor turnover.     

Loss of heterozygosity and microsatellite instabilities of fragile histidine triad gene in gastric carcinoma

AIM: To detect the loss of heterozygosity (LOH) and microsatellite instabilities (MSI) of fragile histidine triad (FHIT) gene in gastric carcinoma and to study their association with the clinical pathological characteristics of gastric carcinoma. METHODS: LOH and MSI of FHIT gene were detected at four microsaterllite loci D3SI3H, D3S4I03, D3SI48I and D3S1234 using PCR in matched normal and cancerous tissues from 50 patients with primary gastric cancer. RESULTS: The average frequency of LOH and MSI of FHIT gene in gastric cancer was 32.4% and 26.4% respectively. LOH and MSI of FHIT gene in gastric cancer had no association with histological, Borrmann, and Lauren's classification. LOH of FHIT gene in gastric cancer was related to invasive depth. The frequency of FHIT LOH in gastric cancer with serosa-penetration was obviously higher than that in gastric cancer without serosa-penetration (73.5% vs 37.5%, P < 0.05). MSI of FHIT gene in gastric cancer was associated with the lymph node metastasis. The frequency of MSI in gastric cancer without lymph node metastasis was significantly higher than that in gastric cancer with lymph node metastasis (66.7% vs 34.3%, P < 0.05). CONCLUSION: LOH of FHIT gene is correlated with invasive depth of gastric carcinoma. MSI of FHIT gene is correlated with lymph node metastases. LOH and MSI of FHIT gene play an important role in carcinogenesis of gastric cancer.     

Genetic changes in Slovenian patients with gastric adenocarcinoma evaluated in terms of microsatellite DNA

OBJECTIVES: Adenocarcinoma of the stomach is a relatively frequent malignant disease in Slovenia. We investigated the frequency of microsatellite instability (MSI) and loss of heterozygosity (LOH) in gastric carcinomas from the Slovenian population to determine their prognostic significance. METHODS: We evaluated MSI of mismatch repair associated loci and LOH on loci associated with following tumour suppressors: APC, nm23, Rb and p53. Results of the multiplex-PCR amplifications were correlated with clinicopathological factors for 73 patients. RESULTS: LOH was found in 52% of informative samples (20.5% LOH-H; 31.5% LOH-L). We found correlation of MSI with low-frequency LOH (LOH-L) in 11% of cases and with high-frequency LOH (LOH-H) tumours in 4% of cases. LOH-H and high-frequency MSI (MSI-H) were not associated. LOH was found in APC 36%, p53 33%, Rb 24% and nm23 33% of informative samples, whereas MSI was found in 30% of samples (12% MSI-H; 18% MSI-L). LOH-H status was associated with ulceration (P=0.029). LOH-N status was associated with diagnosis at higher TNM status (0.074) and infiltrative growth (P=0.006). Interestingly, in 6% of samples we found MSI on LOH loci as well. MSI-H was associated with higher age at diagnosis (r=0.24; P=0.04), antral location (r=0.252; P=0.04), intestinal type (P=0.044), expansive growth (P=0.001), tubular type (0.014), better differentiation (P=0.01), less nodal involvement (0.006) and better survival (P=0.022). The poorest prognosis was found in patients with both low-frequency MSI (MSI-L) and low-frequency LOH (LOH-L) tumours. CONCLUSION: The experimental design presented in the study may be of potential value for clinicians: at least five relevant markers for both MSI and LOH analysis may be needed to evaluate a gastric cancer (GC) patient's clinical status.     

nm23H1 expression and its role in the evolution of non-gastrointestinal malignancies

The role of nm23H1 genetic instability is not limited to gastrointestinal malignancies. A similar close relationship exists between nm23H1 genetic instability and other non gastrointestinal systemic malignancies. For instance, in oral malignant melanomas with lymphoid metastasis, the nm23H1 expression is significantly lower in contrast to tumors with no lymphoid metas- tasis. Similarly, increased metastasis is seen in non small cell lung cancers following down regulation of nm23H1 in conjunction with KAI-1 down regulation. There is an inverse relationship between tumor stage and metastasis and nm23H1 expression in individuals with prostate carcinomas and a similar relationship ex- ists between microsatellite instability of the nm23H1 gene and ovarian carcinogenesis. For instance, nearly 70.5% of stage Ⅰ-Ⅱ ovarian tumors express nm23H1 in sharp contrast to only 25% of stage Ⅲ-Ⅳ ovarian tumors. As is clearly evident, nm23H1 has a major role in gastrointestinal and non-gastrointestinal carcinogenesis. The coming few years will hopefully see the development of new strategies by virtue of which we can alter nm23H1 expression and thus decrease the risk of metastasis in malignant tumors.     

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.     

Genetic analysis of multiple sporadic colon carcinomas from a single patient

At least two separate genetic pathways of carcinogenesis in sporadic colon cancer involving the accumulation of mutations at various genetic loci have been described. About 15% of sporadic colorectal carcinomas arise via a mechanism associated with microsatellite instability (MSI) and mutations in transforming growth factor β receptor II (TGFβRII), insulin-like growth factor II receptor (IGFIIR) and BAX, whilst the remaining 85% are associated with aneuploidy and gross chromosomal rearrangements. An 81-year-old woman had a sigmoid colon carcinoma resected and 18 months later developed two additional carcinomas of the caecum and transverse colon. To investigate whether there was a common genetic mechanism of carcinogenesis for the three lesions, MSI status was assessed, TGFβRII, IGFIIR and BAX were analysed for mutations and protein expression of transforming growth factor β1 (TGFβ1) and p53 were studied using immunohistochemistry. The caecal and transverse colonic carcinomas were both MSI positive but different mutations were identified in each lesion. No genetic abnormalities were identified in the sigmoid colonic carcinoma. This suggests that each carcinoma arose via a separate genetic mechanism of carcinogenesis. Accepted: 31 January 2000     

Ezrin和HER2在胃癌组织芯片中的表达及意义

目的:探讨Ezrin蛋白在胃癌组织中的表达,与肿瘤浸润、转移的关系及与HER2的相互作用.方法:485例原发性胃癌组织中高、中、低分化胃癌分别为19例、235例和231例;有淋巴结转移者353例;TNM分期Ⅰ、Ⅱ期166例,Ⅲ、Ⅳ期319例.另外取距肿瘤7cm的正常胃黏膜组织40例.制成8个组织芯片蜡块,用免疫组织化学方法检测石蜡包埋的胃及胃癌组织中的Ezrin和人类表皮生长因子受体2(hum an epidermal growth factor receptor 2,HER2)蛋白表达.所有患者均经外科手术治疗,病理诊断明确,术前未经放、化疗.结果:Ezrin和HER2在胃癌组织中高表达,二者均与肿瘤Lauren’s分型和肿瘤分化程度相关(χ2=17.625,χ2=20.386,均P=0.000;χ2=9.474,P=0.009,χ2=13.377,P=0.010);Ezrin同时还与组织学(日本分型)、TNM分期、浸润深度和淋巴结转移相关(χ2=37.542,P=0.000;χ2=12.237,P=0.002;χ2=21.194,P=0.002;χ2=9.868,P=0.007).Ezrin和HER2蛋白表达呈正相关(r=0.129,P=0.004).结论:Ezrin可能是预测胃癌组织浸润、转移有用的指标;联合检测Ezrin和HER2可作为判断胃癌预后、筛选高危转移患者的有效指标并有可能用于指导胃癌的个体化治疗.     

Fascin-1与消化系恶性肿瘤

侵袭转移是造成消化系恶性肿瘤患者预后不良的主要原因之一.而肿瘤细胞的运动和迁徙能力在肿瘤的侵袭转移过程中起到了重要作用.Fascin-1是一种肌动蛋白结合蛋白,其主要功能是在细胞突起内形成平行束结构,并参与细胞黏附、移动以及信号转导等.体外实验发现过表达Fascin-1能够增加细胞的侵袭和迁移能力,而抑制Fascin-1的表达则能够降低细胞的侵袭转移.在许多消化系恶性肿瘤组织中,研究发现Fascin-1的上调表达与预后不良,肿瘤的低分化,TNM分期以及淋巴结和远处器官转移密切相关.因此Fascin-1可能作为消化系恶性肿瘤新的预后预测分子及肿瘤侵袭转移可能的治疗靶点.     

肝细胞癌p53及nm23-H1 mRNA表达的意义

目的探讨ｐ５３,ｎｍ２３Ｈ１与原发性肝细胞癌（ＨＣＣ）发生发展的关系．方法运用原位分子杂交技术对４９例ＨＣＣ中ｐ５３和ｎｍ２３Ｈ１基因ｍＲＮＡ进行检测,并结合临床病理特征进行分析．结果ｐ５３ｍＲＮＡ杂交阳性２３例,占４６９％;ｐ５３ｍＲＮＡ过表达与肿瘤的肝内转移．包膜侵犯及Ｅｄｍｏｎｄｓｏｎ分级相关（Ｐ＜００５）;ｎｍ２３Ｈ１ｍＲＮＡ阳性表达２７例,占５５１％;ｎｍ２３Ｈ１ｍＲＮＡ表达与肿瘤肝内转移及ＴＮＭ分期呈负相关（Ｐ＜００５）;同时发现ｐ５３ｍＲＮＡ过表达和ｎｍ２３Ｈ１ｍＲＮＡ低表达在ＨＣＣ肝内转移中具有协同作用．结论ｐ５３和ｎｍ２３Ｈ１参与ＨＣＣ的发生发展,ｐ５３过表达及ｎｍ２３Ｈ１低表达提示ＨＣＣ肝内转移．     

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     

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     

Microsatellite alterations in phenotypically normal esophageal squamous epithelium and metaplasia-dysplasia-adenocarcinoma sequence

AIM： To investigate the microsatellite alterations in phenotypically normal esophageal squamous epithelium and metaplasia-dysplasia-adenocarcinoma sequence.
METHODS： Forty-one specimens were obtained from esophageal cancer （EC） patients. Histopathological assessment identified 23 squamous cell carcinomas （SCC） and 18 adenocarcinomas （ADC）, including only 8 ADC with Barrett esophageal columnar epithelium （metaplasia） and dysplasia adjacent to ADC. Paraffin-embedded normal squamous epithelium, Barrett esophageal columnar epithelium （metaplasia）, dysplasia and esophageal tumor tissues were dissected from the surrounding tissues under microscopic guidance. DNA was extracted using proteinase K digestion buffer, and DNA was diluted at 1：100, 1：1000, 1：5000, 1：10000 and 1：50000, respectively. Seven microsatellite markers （D2S123, D3S1616, D3S1300, D5S346, D17S787, D18S58 and BATRII loci） were used in this study. Un-dilution and dilution polymerase chain reactions （PCR） were performed, and microsatellite analysis was carried out.
RESULTS： No statistically significant difference was found in microsatellite instability （MSI） and loss of heterozygosity （LOH） of un-diluted DNA between SCC and ADC. The levels of MSI and LOH were high in the metaplasia-dysplasia-adenocarcinoma sequence of diluted DNA. The more the diluted DNA was, the higher the rates of MSI and LOH were at the above 7 loci, especially at D3S1616, D5S346, D2S123, D3S1300 and D18S58 loci.
CONCLUSION： The sequence of metaplasia-dysplasia-adenocarcinoma is associated with microsatellite alterations, including MSI and LOH. The MSI and LOH may be the early genetic events during esophageal carcinogenesis, and genetic alterations at the D3S1616, D5S346 and D3S123 loci may play a role in the progress of microsatellite alterations.     

nm23-H1基因在结肠癌组织中的表达及其意义

目的探讨ｎｍ２３－Ｈ１信使核糖核酸（ｍＲＮＡ）基因在结肠癌发展中的作用及其与临床有关资料的关系。方法用Ｎｏｒｔｈｅｒｎｂｌｏｔ法对１０例正常结肠组织、６例结肠息肉及２０例结肠癌中该基因的表达水平进行检测分析。结果２０例结肠癌及４例结肠息肉中ｎｍ２３－Ｈ１基因明显高于正常结肠组织，且与肿瘤的大小及分期有关，但此基因与患者的性别、年龄及肿瘤的分化程度无关。结论ｎｍ２３－Ｈ１基因在结肠癌的局部损害及转移中发挥作用     

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.     

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.