散发性结肠直肠癌肿瘤分化及转移相关基因杂合缺失分析

目的：探讨散发性结肠直肠癌患者2号染色体上可能的肿瘤转移相关基因位点。方法：以2号染色体上30个不同荧光标记的高度多态性微卫星引物对83例散发性结肠直肠癌患者基因组DNA扩增相应的微卫星位点,用ABI PRISM 377测序仪进行基因扫描,检测各位点杂合缺失率,比较与肿瘤分期、分化的关系。结果：24个位点获得有效数据,平均遗传距离为11厘摩（cM),杂合缺失率平均为15．16％,较高的有2个们点：D2S206（2q33-37)的32．08％和D2S364（2q24.2)、31．03％,其余位点的杂合缺失率均小于20．00％;D2S142（2q24.1)、D2S126（2q35)、D2S2211（2q24.2)、D2S305（2q23.3)的杂合缺失率随着肿瘤恶性程度的增加而增高,后2个位点间的缺失有相关性。结论：已知几个错配修复基因位点附近的微卫星位点并无高频杂合缺失发生,D2S2305（2q23.3)到D2S2211（2q24.2)之间区域为整体性缺失,此区域和D2S142（2q24.1)、D2S126（2q35)2个位点与肿瘤的恶性程度相关,提示存在未知的肿瘤分化和转换相关基因的可能。     

Analysis of loss of heterozygosity on chromosome 10 in patients with malignant astrocytic tumors: correlation with patient age and survival

OBJECT: The most frequent genetic abnormality in human malignant gliomas is loss of heterozygosity (LOH) on chromosome 10. Candidate genes on chromosome 10 that are associated with the prognosis of patients with anaplastic astrocytoma (AA) and glioblastoma (GBM) were evaluated. METHODS: The authors used 12 fluorescent microsatellite markers on both arms of chromosome 10 to study LOH in 108 primary astrocytic tumors. The LOH on chromosome 10 was observed in 11 (32%) of 34 AAs and 34 (56%) of 61 GBMs. No LOH was detected in 13 low-grade gliomas. Loss of heterozygosity was not detected in any AA in the seven patients younger than 35 years, but it was discovered in 41% of the patients older than 35 years. The prognostic significance of LOH at each locus was evaluated in 89 patients older than 15 years; 33 (37%) had supratentorial AAs and 56 (63%) had supratentorial GBMs. The Cox proportional hazards model, adjusted for patient age at surgery, the preoperative Karnofsky Performance Scale score, and the extent of surgical resection revealed that LOH on marker D10S209 near the FGFR2 and DMBT1 genes was significantly associated with shorter survival in patients with AA. The LOH on markers D10S215 and D10S541, which contain the PTEN/MMAC1 gene between them, was significantly associated with shorter survival in patients with GBM. CONCLUSIONS: In the present study it is found that LOH on chromosome 10 is an age-dependent event for patients with AAs and that LOH on marker D10S209 near the FGFR2 and DMBT1 loci is a significantly unfavorable prognostic factor. It is also reported that LOH on the PTEN/MMAC1 gene is a significantly unfavorable prognostic factor in patients with GBM.     

Investigation of tumor suppressor genes apart from VHL on 3p by deletion mapping in sporadic clear cell renal cell carcinoma (cRCC)

ObjectivesTo investigate the most recurrent deletion loci on 3p12-p26 by deletion mapping studies by PCR-LOH and BAC array-FISH in sporadic conventional renal cell carcinoma (cRCC) and further, to evaluate the their clinicopathologic significance in cRCC. Comparative allelotyping studies in cRCC and major epithelial carcinomas (MEC) such as lung, breast, and bladder tumors were also carried out to investigate the specificity of the targeted loci in cRCC.Subjects and methodsA total of 40 c-RCC patients were enrolled in this study, categorized in to 2 groups: group I comprises of patients of stages I and II and group II includes patients at stages III and IV. Loss of heterozygosity (LOH) studies were performed by PCR using 15 microsatellite markers of region 3p12-p26 on paired normal-tumor tissues. The recurrent LOH loci found in 27 cRCC tumors were further validated by BAC array-FISH using 23 serially mapped BAC clones. Simultaneously, the allelic deletion status of fragile histidine triad (FHIT) gene was studied by FISH in cRCC and major epithelial carcinoma (MEC) tumors. The numerical aberrations of chromosome 3 were also studied using the centromere enumeration probe (CEP) probe for chromosome 3 to validate the observed allelic losses by BAC array-FISH in cRCC as well as MECs.ResultsOur study revealed 3 affected regions of LOH on 3p in cRCC: 3p12.2-p14.1, 3p14.2-p21.1, and 3p24.2-p26.1 in both group I (stages I and II) and group II (stage III and IV). Comparative allelotyping studies revealed that except for LOH loci D3S2406 (20%), D3S1766 (14%), and D3S1560 (20%), remaining affected loci revealed retention of heterozygosity (ROH) in breast carcinomas. Lung and bladder tumors revealed ROH at all affected LOH loci. FISH with FHIT gene probe revealed deletions in cRCC (88%), breast (30%), and lung tumors (10%). FHIT gene deletions frequency was almost equal in both groups I and II (>70%), whereas a locus 3p13 (D3S2454) revealed the highest LOH in group II (83%) patients in comparison to group I (16%). BAC array-FISH studies in cRCC identified 15 recurrent deletion loci at crucial regions, 3p12.2, 3p14.2, 3p21.3, and 3p24.2-p26 with long continuous deletion of 3p14.1-p26.1 exclusively in patients of stages III and IV. Validation of LOH loci in breast carcinomas by BAC array-FISH with BAC clones mapped at these loci revealed comparatively lower deletion frequency for RP11-59E22 (3p12.2) (30%), RP11-759B7(3p21.1) (12%), and RP11-57D6 (3p25.2, proximal to VHL) (15%) than cRCC.ConclusionMolecular cytogenetic studies by BAC array-FISH was found to be more sensitive over LOH. Deletion patterns on 3p explored that deletion of FHIT and flanking loci may occur as an initiating event followed by deletions at 3p12.2, 3p21.31–3p21.32, and 3p24.2–3p26.1 in the initial stage of development of disease, while continuous large deletions of 3p21.3–3p26.1 and 3p14.1–3p26.1 occur as progressive deletion due to genetic instability. Lack of VHL along with flanking loci in 50% cRCC patients that included both groups I and II supported the hypothesis of both VHL dependent and VHL independent pathways in cRCC tumorigenesis. Comparative allelotyping studies in cRCC and MECs indicated association of specific targeted loci including VHL in cRCC. Further expansion of these studies with characterization of the genes at targeted loci and correlation with clinical outcome will explore the prognostic significance and also provide an insight into the mechanisms of tumor suppressive pathways in genitourinary cancers such as CRCC.     

胃肠间质瘤中9p21区域杂合子丢失及P16蛋白表达研究

目的 探讨染色体9p21区域杂合子丢失（LOH）及相关的P16INK4A（CDKN2A）抑癌基因的表达与胃肠间质瘤（GIST）侵袭行为及预后的关系.方法 采用微卫星分析方法检测51例GIST标本中9p21区域D9S1751、D9S1846、D9S942和D9S1748 4个微卫星位点的LOH现象,并采用免疫组织化学技术,对D9S942位点相邻的CDKN2A抑癌基因产物P16蛋白的表达,分析P16蛋白表达缺失与GIST临床病理特征和预后的关系.结果 51例GIST标本中有2例9p21区域4个微卫星位点均为纯合子（无效信息）,其余49例9p21区域的LOH率：D9S1751为37.0%（10/27）、D9S1846为37.5%（12/32）、D9S942为42.1%（16/38）、D9S1748为24.2%（8/33）,总LOH率为63.3%（31/49）.P16蛋白在GIST标本中的阴性表达率为41.2%（21/51）,阳性率为58.8%（30/51）.高度与低或极低度侵袭风险组P16阴性表达率分别为60%（12/20）和23.5%（4/17）,差异有统计学意义（P＜0.05）.P16阴性与阳性表达组5年生存率分别为70.8%和92.0%,差异有统计学意义（P＜0.05）.结论 9p21区域的LOH在GIST中普遍存在 CDKN2A抑癌基因可能参与GIST的发生发展 P16蛋白与GIST侵袭风险及预后关系密切.     

膀胱移行细胞癌9q的杂合性丢失及抑癌基因位点的定位

目的研究膀胱移行细胞癌的９号染色体长臂（９ｑ）杂合性丢失并对抑癌基因位点进行定位。方法通过应用２５对高密度多态性微小卫星标记经ＰＣＲ扩增，６％变性聚丙烯酰胺凝胶电泳，检测膀胱移行细胞癌的９ｑ杂合性丢失，寻找膀胱移行细胞癌相关抑癌基因的位点。结果２５例病人约有９２％的肿瘤至少有一个以上位点的杂合性丢失，最常见的丢失区域为９ｑ１２～９ｑ２１、９ｑ２２和９ｑ３４，最常见的丢失位点是ＤＢＨ４４．０％、Ｄ９Ｓ１５２２７％、Ｄ９Ｓ１８１５２２７％、Ｄ９Ｓ１７６１２．０％、Ｄ９Ｓ１８３１１６．０％。绘制出膀胱移行细胞癌９ｑ杂合性丢失的染色体图谱，９ｑ杂合性丢失与肿瘤的分级、分期无相关性。结论膀胱移行细胞癌９ｑ的杂合性丢失是肿瘤发生的早期事件之一，在９ｑ３４的ＤＢＨ位点及其附近可能有与膀胱肿瘤相关的抑癌基因存在，并与肿瘤的发生有关。     

Loss of heterozygosity and microsatellite instability at chromosomal sites 1Q and 10Q in morphologically distinct regions of late stage prostate lesions

PURPOSE: We investigated the incidence of loss of heterozygosity (LOH) and microsatellite instability in sporadic prostate cancer and surrounding tissue at loci encompassing the HPC1 and PTEN genes. MATERIALS AND METHODS: Surgical specimens from 63 patients with sporadic stage T3 or T4 prostatic adenocarcinoma were analyzed for LOH and microsatellite instability. Microdissected tissue included morphologically normal foci, benign prostatic hyperplasia (BPH) and prostatic adenocarcinoma. LOH analysis was performed using 4 microsatellite markers that map in the region of the 1q24 to 25 locus of the putative prostate cancer susceptibility gene HPC1 and 4 that map in the region of the 10q23 locus of the PTEN gene. RESULTS: The incidence of LOH on 10q was consistent with that previously reported in prostatic tumors. LOH associated with the PTEN locus was recorded in morphologically normal foci, BPH and adenocarcinoma. Sequence analysis of PTEN in a limited number of lesions revealed mutations in nontumor and tumor tissue. Analysis of the DS10215 locus showed significant LOH in tumor but not in benign tissue, suggestive of a tumor suppressor gene in this region associated with prostatic neoplastic progression. In contrast, no significant LOH was observed in the same tissues at 4 loci on chromosome 1q. In this study we recorded elevated levels of microsatellite instability in benign prostatic tissue with an additional increase associated with prostatic adenocarcinoma. CONCLUSIONS: The low incidence of LOH in the region of the HPC1 locus in all prostate lesions studied suggests that this putative hereditary prostate cancer susceptibility locus does not appear to have a role in sporadic prostate cancer, at least not in the context of LOH. In contrast, analysis of the same tissues for LOH at chromosome 10q confirmed frequent alterations in this region linked to late stage prostate cancer. PTEN mutations in microdissected morphologically normal and BPH tissue showed alterations in nontumor tissue surrounding adenocarcinoma. Microsatellite instability was increased in adenocarcinomas over an elevated background recorded in surrounding tissues.     

Refined mapping of allele loss at chromosome 10q23-26 in prostate cancer

BACKGROUND: Allele loss of at least two segments in 10q, one mapping to the PTEN gene and one more distal were described in prostate cancer, with loss more frequent in advanced prostate cancer. METHODS: A 63 cM region from 10q23 to q26 was studied for allele loss (LOH) in 59 prostate cancer samples using a dense map of microsatellite markers. RESULTS: LOH of at least one marker in 10q was observed in 13/59 tumors. LOH increased with grade and stage. Detailed deletion mapping identified three regions of allele loss. The first region mapped to the site of the PTEN gene, the second is defined by loss of one marker, D10S1692, in one tumor, and the third is defined between markers D10S1757 and D10S587, including DMBT, with a subregion of approximately 1.2 Mb mapping between markers D10S209 and D10S1679, lost in one tumor. CONCLUSIONS: LOH at the PTEN gene is frequent but mutations in the remaining allele were not detected by SSCP-screening. There may be more than two tumor suppressor (TS) genes mapping more distal of PTEN. The site for these putative TS genes can now be mapped with a dense set of precisely localized markers in a larger series of advanced tumors.     

乳腺癌及癌前病变3号染色体短臂杂合性缺失的研究

目的研究乳腺癌及癌前病变中3号染色体短臂（3p）杂合性缺失（loss of heterozygosity,LOH）的发生情况。方法采用聚合酶链式反应及硝酸银染色等方法检测41例原发性乳腺癌及12例癌前病变中3p的11个微卫星位点LOH发生情况;用免疫组化方法检测40例乳腺癌中雌激素受体（ER）、孕激素受体（PR）、脆性组氨酸三联体（fragile histidine triad,FHIT）及人类MutL基因的同源基因（human MutL homologue,hMLH1）的表达情况。结果97％乳腺癌患者发生3p的LOH,检出率较高的位点是D3S1295（53．1％）、D3S1029（43．6％）和D3S1038（52．5％）,分别位于3p14、3021-p22和3p25。D3S1038位点LOH及hMLH1蛋白表达与部分临床病理学参数相关（P〈0．05）。D3S1295的LOH与FHIT蛋白表达负相关（P〈0．05）。癌前病变患者3pLOH发生率为41．7％,检出率较高的位点是D3S1295（27．3％）和D3S1029（16．7％）。最小共同丢失区位于3p14-p25。结论3p14-p25区段可能有与乳腺癌发生发展相关并影响乳腺癌生物学行为的候选抑癌基因,基因的部分缺失可影响其蛋白的表达。     

Loss of P16 expression and chromosome 9p21 LOH in predicting outcome of patients affected by superficial bladder cancer

OBJECTIVES: To evaluate the prognostic role of p16 expression and loss of heterozygosity (LOH) on chromosome 9p21 in patients affected by low-grade (G1-G2) urothelial bladder cancer. METHODS: Fifty-six consecutive patients with diagnosis of urothelial bladder cancer were enrolled in this prospective study. LOH analysis was performed on a blood/tumor pair sample of each patient, by using polymerase chain reaction analysis. The D9S171 (9p21) locus on chromosome 9 was investigated. All tumors were stained immunohistochemically for p16. Data from p16 and LOH analyses were compared with follow-up data to evaluate the prognostic role of these molecular markers. RESULTS: Loss of p16 expression was found in 33 patients (58.9%) and was significantly associated with the reduced recurrence-free probability (P < 0.0001). No correlations were reported with stage (P = 0.162) or grade (P = 0.051). Forty-three patients (76.7%) showed LOH on chromosome 9p21 (D9S171). A significant association was observed between loss of p16 expression and LOH on chromosome 9p21 (P = 0.005). The Kaplan-Meier curves showed a significant correlation between recurrence-free status and p16 expression (P = 0.0001). By multivariate analysis, p16 expression (P = 0.002) and number of lesions (P = 0.002) were identified as independent tumor recurrence factors. CONCLUSIONS: Our study highlights the prognostic role of p16 in predicting the recurrence-free probability in patients affected by low-grade urothelial bladder and highlights the fact that this method could be used in everyday urologic clinical practice to better characterize the natural history of urothelial bladder carcinomas.     

Analysis of loss of heterozygosity on chromosomes 10q, 11, and 16 in medulloblastomas

OBJECT: The loss of genetic material from specific chromosome loci is a common feature in the oncogenesis of tumors and is often indicative of the presence of important tumor suppressor genes at these loci. Recent molecular genetic analyses have demonstrated frequent loss of chromosomes 10q, 11, and 16 in medulloblastomas. The aim of this study was to localize the targeted deletion regions on the three aforementioned chromosomes in medulloblastomas. METHODS: Loss of heterozygosity (LOH) was examined on chromosomes 10q, 11, and 16 in a series of 22 primary and two recurrent medulloblastomas by using polymerase chain reaction-based microsatellite analysis. The DNA extracted from the tumors and corresponding normal blood samples were amplified independently in the presence of radioactively labeled microsatellite primers, resolved by denaturing gel electrophoresis and processed for autoradiography. The DNA obtained from control blood samples that displayed allelic heterozygosity at a given microsatellite locus were considered informative. Loss of heterozygosity was inferred when the allelic signal intensity of the tumor sample was reduced by at least 40%, relative to that of the constitutional control. The LOH analysis demonstrated that deletions of chromosomes 10q, 11p, and 16q are recurrent genetic events in the development of medulloblastomas. Three subchromosomal regions of loss have been identified and are localized to the deleted in malignant brain tumors 1 [DMBT1] gene site on chromosomes 10q25, 11p13-11p15.1, and 16q24.1-24.3. CONCLUSIONS: These results indicate that DMBT1 is closely associated with the oncogenesis of medulloblastomas and highlight regions of loss on chromosomes 11p and 16q for further fine mapping and cloning of candidate tumor suppressor genes that are important for the genesis of medulloblastoma.     

Loss of heterozygosity at 3p in benign lesions preceding invasive breast cancer

BACKGROUND: Loss of heterozygosity (LOH) at chromosome 3p is one of the most common genetic abnormalities identified in human cancers and has occasionally been noted in benign proliferative lesions predisposing to breast cancer. If the frequency of LOH at 3p in benign proliferative lesions correlates with the subsequent development of breast cancer, it may be possible to develop powerful tools for molecular risk assessment based on this technology. MATERIALS AND METHODS: Archival paraffin-embedded tissues from benign breast biopsies in five women who have developed breast cancer and three women who have not developed breast cancer were microdissected and allelotyped at 3p using six microsatellite markers. RESULTS: No LOH was detected in the biopsies from women who have not developed breast cancer. For women developing breast cancer, the proportion of informative loci showing LOH in the benign proliferative lesions was 0.47 as compared to 0.57 for the associated breast cancers. There was no LOH detected in epithelial DNA from a fibroadenoma. Of 15 informative loci, 4 (27%) showed LOH in both the benign proliferative lesion and the associated cancer; however, the actual parental allele lost was different in three of these four cases. CONCLUSIONS: These results suggest that there are specific patterns of genetic instability common to preneoplastic lesions and the breast cancers that subsequently develop even when the paired lesions are not clonally related. LOH analysis of benign breast epithelium may provide a tool for molecular risk assessment and a surrogate endpoint for breast cancer chemoprevention trials.     

Chromosome 22q a frequent site of allele loss in head and neck carcinoma

BACKGROUND: Loss of heterozygosity (LOH) correlates with inactivated tumor suppressor genes. LOH at chromosome arm 22q has been found in a variety of human neoplasms, suggesting that this region contains a tumor suppressor gene(s) other than NF2 important to tumorigenesis. The aim of this study was to evaluate the presence of LOH on chromosome 22q11.2-13 and determine whether there was a relationship between loss in this genomic region and tumor histologic parameters, anatomic site, and survival in patients with squamous cell carcinoma of the head and neck (HNSCC). METHODS: Fifty matched blood and HNSCC tumor samples taken at the time of surgical treatment were evaluated for LOH by use of four microsatellite markers mapping to 22q11.2-q13. Clinical information was available for all patients. The frequency and distribution of LOH was correlated with clinical (age, sex, use of tobacco and alcohol, site of primary tumor, clinical stage, adjuvant therapy and overall survival) and histologic parameters (histopathologic stage, tumor differentiation). RESULTS: LOH at 22q was found in 19 of 50 (38%) informative tumors. The respective incidence of allelic loss for the patients was as follows: 28% at D22S421, 10% at D22S277, 8% at D22S446, and 4% at D22S280. No statistical differences were apparent with a mean follow-up of 30 months. Laryngeal tumors showed a higher incidence of LOH compared with oral tumors. CONCLUSIONS: These results suggest that the D22S277 locus may be closely linked to a tumor suppressor gene (TSG) and involved in upper aerodigestive tract carcinogenesis. In particular, laryngeal tumors may harbor another putative TSG on 22q11.2-q12.3 that may play a role in aggressive stage III/IV disease.     

Comprehensive loss of heterozygosity analysis and identification of a novel hotspot at 3p21 in salivary gland neoplasms.

OBJECTIVES: We sought to assess loss of heterozygosity (LOH) profiles of 3p, 6q, 8q, 10q, 12q, 13q, and 17p and to identify the tumor suppressor genes involved in salivary gland neoplasms. STUDY DESIGN: LOH analysis was performed using 26 microsatellite markers by polymerase chain reaction-polyacrylamide gel electrophoresis method in 20 benign and 6 malignant salivary gland tumors. RESULTS: Overall, LOH was detected in at least one informative locus in 18 of 20 (90%) of benign tumors and in all of 6 cases of malignant tumors. High LOH frequencies were revealed at the loci D3S1307 (22%, 3p26), D3S966 (41%, 3p21), D6S255 (27%, 6q25), D8S166 (25%, 8q12), D8S199 (21%, 8q24), and D10S1765 (28%, 10q23) in benign tumors, defining the hotspot regions for putative tumor suppressor genes. CONCLUSIONS AND SIGNIFICANCE: The hotspot regions defined by the present study suggest that new tumor suppressor genes related to the development of salivary gland tumors may reside at several chromosomal loci, including loci at 3p, 6q, 8q and 10q.     

Loss of heterozygosity on chromosomes 10 and 17 in clinically localized prostate carcinoma

Loss of heterozygosity (LOH) at chromosomal loci has been associated with the presence of tumor suppressor genes at the deleted loci. Twenty-six clinically localized, Stage B prostate carcinomas were analyzed for LOH on chromosomes 10 and 17 using microsatellite markers. Two of 26 carcinomas showed LOH on 17p while one showed LOH on 17q. Chromosome 10 showed a complex pattern of LOH with monosomy (1 case), LOH on 10p (1 case), proximal 10q (1 case) and distal 10q (2 cases). Overall 29% of informative cases showed LOH on chromosome 10. These results are consistent with the presence of a tumor suppressor for prostate cancer on 17p and multiple tumor suppressor genes on chromosome 10. © 1996 Wiley-Liss, Inc.     

Molecular and clinicopathologic comparisons of head and neck squamous carcinoma variants: common and distinctive features of biological significance

To investigate, for the first time, the events associated with the phenotypic and clinical diversities of head and neck squamous carcinomas (HNSC), we performed molecular analyses on 92 primary tumors representing the entire spectrum of the morphologic subtypes using microsatellite markers at chromosome 3p, 4p, 8p, 9p, 11q, 17p, and 18q regions and correlated the results with the clinicopathologic features and patients' survival. Loss of heterozygosity (LOH) at D9S168 and D9S171 markers on chromosome 9p regions was commonly identified in all subtypes. Distinctive alterations in certain subtypes were noted at chromosomes 3p, 4p, 8p, and 11p regions. In general, less aggressive types (verrucous, papillary, and well-differentiated conventional) had a significantly lower LOH incidence than the more aggressive (basaloid, sarcomatoid, and high-grade conventional squamous carcinoma) categories. Significant association between LOH and age, stage, nodal status, and patient outcome was found. Survival analysis revealed that pathologic categorization (less versus more aggressive) and LOH at marker D11S4167 and D3S2432 are independent predictors of patients' survival. Our analysis also defined a set of limited markers that account for most of alterations within and across these tumor subtypes. Our study indicates that 1) certain genetic markers are common to all subtypes of HNSC supporting their early involvement in tumorigenesis, 2) inter- and intratumoral genetic differences evolve subsequently and may underlie their morphologic heterogeneity, 3) high incidence of LOH in certain regions characterizes aggressive tumors, 4) categorical classification and LOH at 11p and 3p regions independently correlated with patient survival, and 5) a limited set of markers identify the majority of genetic alterations in these tumors.     

应用激光捕获显微切割技术对肝细胞癌8号染色体短臂杂合性缺失的研究

目的探讨8p杂合性缺失（LOH）的特点及其与肝细胞癌（HCC）临床病理特征的相关性。方法选择8p上5个具有高度多态性的微卫星标记，对62例HCC组织利用激光捕获显微切割（LCM）技术进行LOH分析。结果有56．5％（35／62）的HCC患者在1个或多个基因位点发生LOH。LOH频率最高的3个位点依次为D8S298（51．1％，24／47）、D8S1771（48．8％，21／43）和D8S264（43．5％，20／46）。D8S298位点肝内转移者的LOH频率明显高于无转移者（P〈0．05）；在D8S1771位点，肿瘤直径〉3cm的LOH频率明显高于≤3cm组（P〈0．05）。结论HCC在染色体8p特定位点上LOH明显，在这些区域可能存在一个或多个与HCC发生发展相关的肿瘤抑制基因。8p上部分位点的LOH与临床病理特征有一定的相关性。     

食管鳞状细胞癌及癌前病变组织中多个染色体区域微卫星杂合性缺失的研究

目的 探讨与食管癌早期癌变的发生、发展密切相关的分子变化,寻找早期诊断的分子标记.方法 采用PCR荧光测序仪凝胶电泳分别检测34例食管癌患者手术切除的鳞状细胞癌(SCC)组织、癌前病变组织及匹配的外周血样本中16个微卫星位标的杂合性缺失状况.将轻、中度不典型增生定义为低级别不典型增生(LGD),将重度不典型增生归为高级别不典型增生(HGD).比较不同病理形态组织中16个位标的杂合缺失率.结果 16个位标的总体缺失率随组织形态的严重程度而升高,LGD中缺失率(9.8%)低于HGD(48.6%)和SCC(58.5%,P＜0.01).有8个位标,即D3S1597、D3S2452、D3S1285、D4S174、D5S2501、D9S125、D13S153和D17S786,在LGD中已呈现杂合性缺失;另发现4例患者的HGD和SCC样本分别在D3S2452、D4S174、D9S125及D17S261处出现了杂合性缺失的逆转.结论 食管鳞状上皮从不典型增生至癌变的过程中需要一系列分子变化的积累;针对某些位标的杂合性缺失分析将有助于食管癌的早期诊断;食管癌的发生可能存在遗传异质性.     

肝细胞癌微卫星不稳定和杂合性缺失分析

目的 通过检测肝细胞癌(hepatocellular carcinoma,HCC)微卫星不稳定（microsatellite instability,MSI)和杂合性缺失（loss of heterozygosity,LOH)发生频率,探讨上述遗传学改变与HCC临床病理的关系。方法 选择5个微卫星多态性标记对32例HCC进行了MSI与LOH分析。结果 32例HCC中有14例出现MSI,其中有6例出现2个位点MSI,后者癌肿转移率明显低于其他的组（P＝0．001）,在所检5个位点中D1S484和TP53 MSI发生频率最高,而D9S1604昨D8S555遗传学改变多为LOH且发生频率较低。结论 MSI是部分HCC发生、发展进程中一个重要的遗传改变,尤其在在无癌肿转移的肿瘤中。D1S484和TP53是HCC中对MSI敏感的检测位点。     

Tumor suppressor gene alterations in respiratory epithelial adenomatoid hamartoma (REAH): comparison to sinonasal adenocarcinoma and inflamed sinonasal mucosa

Respiratory epithelial adenomatoid hamartoma (REAH) is an unusual benign sinonasal glandular proliferation. REAH is not considered a neoplasm, although, no molecular evidence exists to support or refute this possibility. Microdissection of 10 cases of REAH, 9 cases of sinonasal adenocarcinoma (SNAC) and 10 cases of chronic sinusitis was performed. DNA was extracted and polymerase chain reaction performed using fluorescently labeled primers flanking known tumor suppressor genes on chromosomes 9p (CDKN2/p16), 11p (H-ras), 17p (p53), and 18q (DCC/DPC4). Polymerase chain reaction products were analyzed semiquantitatively by capillary electrophoresis. Allele ratios were calculated using the peak height from the shorter allele divided by the peak height from the longer allele. The loss of heterozygosity (LOH) ratio was calculated as the allele ratio from tumor tissue divided by the allele ratio from normal tissue. The fractional allelic loss (FAL) was calculated as the percentage of loci that harbored LOH divided by the number of loci that were informative. REAH demonstrated an intermediate FAL of 31% compared with SNAC (64%) and chronic sinusitis (2%). REAH and SNAC had the highest LOH for multiple loci located on 9p (p16) and 18q (DCC/DPC4). The molecular profile of REAH shows a mean FAL of 31%, which would be considered unusually high for a non-neoplastic entity. Appreciable allelic loss within REAH suggests the possibility that REAH may be a benign neoplasm rather than a hamartoma.     

Molecular Biology of Squamous Cell Carcinoma of the Anus: A Comparison of HIV-Positive and HIV-Negative Patients

The molecular mechanisms involved in progression of squamous cell carcinoma of the anus (SCCA) are poorly elucidated, as well as the potential role of HIV infection. Loss of heterozygosity (LOH) is one of the mechanisms responsible for inactivation of tumor suppressor genes. We hypothesized that HIV-induced immunosuppression may contribute to an alternate molecular pathway in SCCA progression, through persistence of human papillomavirus infection within the anal canal. This study was undertaken to compare the molecular biology of SCCA in HIV-positive (HIV+) and HIV-negative (HIV-) patients. We retrieved tumor specimens from 18 HIV- and 10 HIV+ patients diagnosed with SCCA in two institutions. DNA from tumor and normal tissues was extracted and then amplified by polymerase chain reaction. LOH was investigated at 14 loci: three at 18q (DCC), two at 13q (Rb), three at 17p (p53), three at 11q, one at 2p, and two at 5q (APC). LOH was defined by a tumor DNA-to-normal tissue DNA ratio of >2. HIV+ patients were younger (36 ± 7 years versus 53 ± 13 years, P = 0.001) and showed a trend toward tumors of larger size (3.7 ± 1.6 cm versus 2.6 ± 1.5 cm, P = 0.09). The median CD4⁺ count in HIV+ patients at the time of diagnosis was 74 × 10⁶/L (range, 5–900). The overall frequency of LOH was 17.3% (41 LOH of 236 informative loci). Tumors in HIV- patients were more likely to present LOH than were tumors in HIV+ patients (24.1% versus 6.6%, P = 0.0004). Differences between the two groups with regard to allelic losses were also observed at specific loci, such as 18q (41% [HIV-] versus 0% [HIV+], P = 0.05), 17p (43% versus 10%, P = 0.09), and 5q (33% versus 0%, P = 0.12). Consistent LOH on chromosomes 17p, 18q, 5q, and 11q were observed in HIV- patients with SCCA. By contrast, allelic losses at 17p, 5q, and 18q seem to be rare in tumors of HIV+ individuals. These data suggest that immunosuppression may promote SCCA progression through an alternate pathway and that persistence of HPV infection within the anal canal may play a central role in this process. Presented at the Forty-Fifth Annual Meeting of The Society for Surgery of the Alimentary Tract, New Orleans, Louisiana, May 15–19, 2004 Presented at the Forty-Fifth Annual Meeting of The Society for Surgery of the Alimentary Tract, New Orleans, Louisiana, May 15–19, 2004