胆管癌染色体DNA拷贝数的变化研究

目的探讨胆管癌全基因组DNA拷贝数的变化特征,寻找与胆管癌相关的癌基因和抑癌基因的染色体候选区域。方法采用比较基因组杂交方法分析18例胆管癌组织基因组的不平衡,即DNA拷贝数的扩增和丢失。结果胆管癌常见的染色体DNA扩增区域是8q、20q、5p、17q;染色体DNA缺失区域为3p、18q、17p、8p、9p。结论胆管癌中存在多条染色体拷贝数的改变,3p、5p、8p、8q、9p、17p、17q、18q、20q等部位可能分别存在与胆管癌密切相关的癌基因和抑癌基因。     

Chromosomal imbalances revealed in primary rhabdomyo- sarcomas by comparative genomic hybridization

Background Previous cytogenetic studies revealed rhabdomyosarcoma. We profiled chromosomal imbalances aberrations varied among the three subtypes of n the different subtypes and investigated the relationships between clinical parameters and genomic aberrations. Methods Comparative genomic hybridization was used to investigate genomic imbalances in 25 cases of primary rhabdomyosarcomas and two rhabdomyosarcoma cell lines. Specimens were reviewed to determine histological type, pathological grading and clinical staging. Results Changes involving one or more regions of the genome were seen in all rhabdomyosarcomal patients. For rhabdomyosarcoma, DNA sequence gains were most frequently （〉30%） seen in chromosomes 2p, 12q, 6p, 9q, 10q, lp, 2q, 6q, 8q, 15q and 18q; losses from 3p, 11p and 6p. In aggressive alveolar rhabdomyosarcoma, frequent gains were seen on chromosomes 12q, 2p, 6p, 2q, 4q, 10q and 15q; losses from 3p, 6p, lq and 5q. For embryonic rhabdomyosarcoma, frequent gains were on 7p, 9q, 2p, 18q, lp and 8q; losses only from 11p. Frequently gained chromosome arms of translocation associated with rhabdomyosarcoma were 12q, 2, 6, 10q, 4q and 15q; losses from 3p, 6p and 5q. The frequently gained chromosome arms of nontranslocation associated with rhabdomyosarcoma were 2p, 9q and 18q, while 11p and 14q were the frequently lost chromosome arms. Gains on chromosome 12q were significantly correlated with translocation type. Gains on chromosome 9q were significantly correlated with clinical staging. Conclusions Gains on chromosomes 2p, 12q, 6p, 9q, 10q, lp, 2q, 6q, 8q, 15q and 18q and losses on chromosomes 3p, 11p and 6p may be related to rhabdomyosarcomal carcinogenesis. Furthermore, gains on chromosome 12q may be correlated with translocation and gains on chromosome 9q with the early stages of rhabdomyosarcoma.     

Novel chromosomal alterations detected in primary nasopharyngeal carcinoma by comparative genomic hybridization

Objective　To gain a better understanding of genetic changes in Cantonese nasopharyngeal carcinoma (NPC). Methods　Comparative genomic hybridization (CGH) was performed on 17 primary nasopharyngeal carcinomas. Results　A novel copy number gain an chromosome 4q and loss of chromosome 1p were found at a high frequency (>50%). Conclusions　Current analysis revealed a comprehensive profile of the chromosomal regions showing gain of chromosomes 4q, 12q, and 1q as well as loss of chromosomes 1p, 3p, 11q, 14q, 15q, 13q, Xq, 9q, 10p, 10q, and 16q. Frequently altered loci may encode oncogenes or tumor suppressor genes involved in the development of primary NPC.     

应用比较基因组杂交技术对卵巢癌耐药机制的研究

目的探讨卵巢癌化疗耐药患者组织中染色体的变化特征,寻找或定位与卵巢癌化疗耐药密切相关的基因,以便对其耐药机制进一步研究。方法采用比较基因组杂交技术分析卵巢癌化疗耐药组（浆液性卵巢癌6例、黏液性卵巢癌3例、子宫内膜样卵巢癌2例、透明细胞癌1例,共12例）和化疗敏感组（浆液性卵巢癌5例、黏液性卵巢癌3例、子宫内膜样卵巢癌2例、透明细胞癌2例,共12例）两组癌细胞基因组的不平衡,即DNA丢失或扩增。结果化疗耐药患者组织中最常见的染色体DNA拷贝数增加的部位是3q,8q,20q,11q,17q,2p,1q;常见的染色体DNA拷贝数缺失的部位是9p,5q,10q,3p。其中17q的增加,耐药组为66．7％,敏感组为16．7％;1q的增加,耐药组为66、7％,敏感组为16．7％;3p的丢失,耐药组为50．0％,敏感组为8．3％,在化疗耐药患者组织中表现更加明显。结论卵巢癌耐药患者组织中细胞染色体基因组最明显的改变为17q,1q的增加以及3p的丢失,这些部位可能存在与卵巢癌化疗耐药密切相关的癌基因及抑癌基因。     

河南林州市居民贲门癌组织的比较基因组杂交分析

目的：探讨贲门癌高发区河南林州市贲门癌组织中的基因组变化，寻找相关未知基因。方法：采用比较基因组杂交的方法（comparative genomic hybridization,CGH)分析10例原发性贲门癌组织基因组变化。结果；CGH分析显示1q(4/10),11q(3/10),3q(2/10),8q(3/10),8p(2/10)为出现频率较高的扩增区，9q(3/10)的缺失可能是贲门癌的特征性变化。结论：1q,11q,3q,8q,8p,9q可能存在与贲门癌相关的未知基因。     

Chromosomal and Genetic Analysis of a Human Lung Adenocarcinoma Cell Line OM

Background:

Lung cancer has become the leading cause of death in many regions. Carcinogenesis is caused by the stepwise accumulation of genetic and chromosomal changes. The aim of this study was to investigate the chromosome and gene alterations in the human lung adenocarcinoma cell line OM.

Methods:

We used Giemsa banding and multiplex fluorescence in situ hybridization focusing on the human lung adenocarcinoma cell line OM to analyze its chromosome alterations. In addition, the gains and losses in the specific chromosome regions were identified by comparative genomic hybridization (CGH) and the amplifications of cancer-related genes were also detected by polymerase chain reaction (PCR).

Results:

We identified a large number of chromosomal numerical alterations on all chromosomes except chromosome X and 19. Chromosome 10 is the most frequently involved in translocations with six different interchromosomal translocations. CGH revealed the gains on chromosome regions of 3q25.3-28, 5p13, 12q22-23.24, and the losses on 3p25-26, 6p25, 6q26-27, 7q34-36, 8p22-23, 9p21-24, 10q25-26.3, 12p13.31-13.33 and 17p13.1-13.3. And PCR showed the amplification of genes: Membrane metalloendopeptidase (MME), sucrase-isomaltase (SI), butyrylcholinesterase (BCHE), and kininogen (KNG).

Conclusions:

The lung adenocarcinoma cell line OM exhibited multiple complex karyotypes, and chromosome 10 was frequently involved in chromosomal translocation, which may play key roles in tumorigenesis. We speculated that the oncogenes may be located at 3q25.3-28, 5p13, 12q22-23.24, while tumor suppressor genes may exist in 3p25-26, 6p25, 6q26-27, 7q34-36, 8p22-23, 9p21-24, 10q25-26.3, 12p13.31-13.33, and 17p13.1-13.3. Moreover, at least four genes (MME, SI, BCHE, and KNG) may be involved in the human lung adenocarcinoma cell line OM.     

散发性结直肠癌的染色体不稳定研究及其临床病理意义

目的 探讨散发性结直肠癌（sporadic colorectal cancer,SCRC）的DNA倍体异常和染色体畸变的数目、部位及其与临床病理特征之间的关系.方法 将40例SCRC肿瘤标本制备成单细胞悬液,经染色后进行流式细胞术（flow cytometry,FCM）检测,分析肿瘤细胞DNA倍体;采用比较基因组杂交技术（comparative genomic hybridization,CGH）在全基因组水平对40例SCRC进行染色体畸变检测.结果 40例SCRC患者标本中,二倍体的比例为42.5%,异倍体为57.5%,DNA倍体与TNM分期有密切关系,分期越高,异倍体的比例越高,差异有统计学意义（P＜0.05）.CGH检测的所有病例均有不同程度的染色体畸变,平均每例畸变数为7.55,常见的染色体扩增区域有：20q、12q、13q、7p等,常见缺失区域有18q、5q、4q、8p等.TNM分期中Ⅲ～Ⅳ期SCRC的染色体畸变数高于Ⅰ～Ⅱ期（6.00±2.76 vs 8.70±2.84,P＜0.05）.不同肿瘤部位、分化程度、组织学类型SCRC的DNA倍体差异无统计学意义（P ＞0.05）.二倍体肿瘤的染色体畸变数明显少于异倍体肿瘤（6.35±3.35 vs 8.43±2.59,P＜0.05）.结论 染色体不稳定在SCRC中普遍存在,是SCRC病情进展的基础,染色体畸变比DNA倍体异常更为常见.     

比较基因组杂交技术分析结直肠癌远处转移的遗传变异

目的 应用微阵列比较基因组杂交技术分析结直肠癌远处转移的遗传变异。方法 收集47例结直肠癌患者（发生/未发生远处转移=18/29）的临床数据以及新鲜组织标本,进行微阵列比较基因组杂交分析,筛选出与远处转移相关的基因组改变。进行功能富集分析和蛋白质-蛋白质相互作用（protein-protein interaction, PPI）网络分析,探索结直肠癌远处转移的分子机制。结果 47例肠癌标本都存在广泛的遗传变异。研究发现,218个DNA拷贝数改变与结直肠癌的远处转移相关,且它们主要位于17q、22q、12p。功能富集分析显示这些基因组改变与免疫反应、Wnt信号通路、同源染色体重组、趋化因子信号通路和MAPK信号通路等密切相关。通过PPI网络分析获得了39个关键基因,包括TP53、EP300、MDM2、BRCA1、MAPK1等。结论 与结直肠癌远处转移相关的遗传变异主要位于17q、22q、12p,其基因组改变与免疫反应、Wnt信号通路、MAPK信号通路等密切有关。     

Microsatellite instability and allele-specific chromosome 3p deletion in breast cancer and precancerous lesions

Objective To investigate the incidence and clinicopathologic significance of MSI and LOH on 3P in breast carcinoma and its precancerous lesions, intraductal papillary adenoma and ductal carcinoma in situ. Methods 41 paired sporadic invasive breast carcinomas, 13 archival precancerous lesion specimens of the breast and 14 couples of benign hyperplasia were collected. Twelve microsatellites on chromosomes 2p, 3p, 5q, 6q, 16q, 17q, eleven markers on chromosome 3p were amplified for MSI and LOH,[第一段]     

Mutations and altered expression of p16~(INK4a) in human pancreatic carcinoma cell lines with different potential of metastasis

Objective:To analyze the p16INK4a genomic alteration and expression status in 3 human pancreatic carcinoma cell lines with different potential of metastasis. Methods:Using PCR-SSCP, Dot-blot and immunohistochemistry, the p16INK4a genomic mutation and expression were analyzed on DNA, mRNA and protein levels in 3 human pancreatic carcinoma cell lines Patu8902, Patu8988 and SW1990, which had different potential of metastasis. Results: (1) On DNA level: there was no deletion of p16INK4a Exon Ⅰ in 3cell lines; p16INK4a Exon Ⅱ was only deleted in Patu8902 while no deletion in Patu8988 and SW1990. No insertion, microdeletion and point mutation were found in the 3 cell lines. (2) On RNA level: the expression of p16INK4a protein was negative in Patu8902, low expressed in SW1990, but highly expressed in Patu8988.(3) On protein level: P16 protein was strongly stained in Patu8988, much lower in SW1990, but not stained in Patu8902. Conclusion:The genomic type and expression of p16INK4a are quite different in 3 pancreatic carcinoma cell lines which have different potential of metastasis. It is suggested that genomic homozygous deletion and low expression of mRNA might relate to the potential of metastasis of pancreatic cell lines. In other words, dysfunction of p16INK4a might be an important mechanism in the metastasis of pancreatic carcinoma.     

Genome-wide allelotype study of primary glioblastoma multiforme

Objective To investigate the molecular genetic pathogenesis of primary glioblastoma multiforme (GBM) and identify which chromosomes or chromosomal regions of the entire genome may harbor tumor suppressor genes (TSGs) associated with GBM.Methods A high-resolution allelotype study of 21 cases of primary GBM was performed by PCR-based loss of heterozygosity （LOH）analysis. Three hundred and eighty-two fluorescent dye-labeled microsatellite markers covering all 22 autosomes were applied. The mean genetic distance between two flanking markers was about 10 cM.Results LOH was observed on all 39 nonacrocentric autosomal arms examined in this study. The LOH frequencies of 10q, 10p, 9p, 17p and 13q were the highest (>50%). Furthermore, high LOH frequencies were detected in the regions containing known TSGs including PTEN, DMBT1, p16, p15, p53 and RB; the LOH frequencies on 14q, 3q, 22q, 11p, 9q, 19q were also high (>40.5%). Our study observed the following commonly deleted regions: 9p22-23, 10p12.2-14, 10q21.3, 13q12.1-14.1, 13q14.3-31, 17p11.2-12, 17p13, 3q25.2-26.2, 11p12-13, 14q13-31, 14q32.1, 14q11.1-13, 22q13.3, 4q35, 4q31.1-31.2, 6q27 and 6q21-23.3. Conclusions The molecular pathogenesis of GBM is very complicated and associated with a variety of genetic abnormalities on many chromosomal arms. The most closely related chromosomal arms to the pathogenesis of GBM are 10q, 10p, 9p, 17p and 13q. Besides the well-known TSGs including PTEN, DMBT1, p16, p15, p53 and RB, multiple unknown TSGs associated with GBM may be present on the commonly deleted regions detected in the present study.     

Analysis of chromosomes in ascitic fluid cells of ovarian carcinoma

Chromosomes in 1620 metaphases of ascitic fluid cells in 20 cases of ovariancarcinoma were analyzed.The results showed that there were marked structuralaberrations aside from significant increase in chromosomal numerical aberrations(85.2%).In the ascitic fluid cells from 12 patients,15 types of marker chromosomes were found,among which t(6;14)(q21;q24)and t(2;6)(q35;p12)were more frequently noted witha rate of 7.84% and 7.59% respectively,which was significantly higher than that of othermarker chromosomes(P<0.01).The findings suggested that,besides t(6;14)(q21;q24),t(2;6)(q35;p12)may also be a specific marker chromosome of ovarian carcinoma.     

原发性肺癌染色体比较基因组杂交研究

目的分析原发性肺癌不同病理分类(鳞癌、腺癌和其他类型原发性肺癌)的遗传物质不平衡特征,为寻找肺癌发生发展相关基因、揭示肺癌发病机制提供线索。方法应用比较基因组杂交技术分析55例原发性肺癌患者肿瘤细胞染色体,按病理分类分为鳞状细胞癌组24例(鳞癌组),腺癌13例(腺癌组),其他组18例(腺鳞癌5例、细支气管肺泡癌8例、小细胞癌4例及非典型类癌1例),正常外周血淋巴细胞染色体DNA标本由20名健康男性志愿者提供。结果原发性肺癌鳞癌组常见染色体扩增区是2Q、5P、11Q、22Q,常见缺失区是1P、4Q、5Q、6Q、8P、9P、10Q、11P、13Q、18Q、21Q。腺癌组常见扩增区是5P、8Q、11Q,常见缺失区是10P、19。其他组中,腺鳞癌、肺泡细胞癌、小细胞癌等各有不同。结论原发性肺癌不同病理分型存在广泛的遗传物质不平衡现象,染色体基因扩增和缺失可能是不同类型肺癌发生发展的基础。     

喉鳞癌染色体核型的检测和分析

目的 检测和分析喉鳞癌染色体核形,探讨染色体变异与喉鳞癌基因组变异的关系,及其在肿瘤发生、发展过程中的重要作用.方法 18个来源于喉鳞状细胞癌患者的肿瘤组织进行细胞培养,收集分裂中期细胞,进行染色体"G"带染色,分析核型.并以3例来源于喉鳞癌旁正常上皮组织细胞作为阴性对照.结果 喉鳞癌染色体核型主要表现为大量复杂的、但并非任意性的染色体结构重排和数目改变:染色体丢失的机率显著高于染色体成分、数目的增加;染色体/臂成分的失衡主要表现为3p、4p、8p、18q、5q、13、21和22染色体成分的缺失,及3q、7q、8q、5p、11q13和20染色体成分的增加;大部分的染色体断裂点位于着丝粒区域.结论 染色体数目、结构的不断变异造成癌基因活化、扩增,抑癌基因的抑制和缺失,这种多基因改变的不断积累促进了喉鳞癌的发生及发展.     

肝外胆管癌瘤细胞染色体的结构畸变

目的 研究肝外胆管癌瘤细胞染色体结构畸变方式和畸变率，筛选肝外胆管癌标记染色体，定位肝外胆管癌相关基因。方法 用比较基因组杂交（CGH）和光谱核型分析（SKY）技术检测12例肝外胆管癌组织瘤细胞染色体结构畸变方式和畸变率。结果 肝外胆管癌瘤细胞多条染色体存在结构畸变，畸变方式以片段重复和丢失为主，重复集中在1q，3q，8q，15q和17q，丢失主要发生在3p，4q，6q，9p，17p和18q，其中丢失率最高的2个区段分别为3p13-p21和9p21-pter（各41．7％）。结论 肝外胆管癌瘤细胞染色体存在明显结构畸变，为进一步定位肝外胆管癌发生发展相关基因靶位点提供科学依据。     

宫颈癌多阶段多途径演进的树模型构建

目的基于宫颈癌比较基因组杂交资料构建宫颈癌的树模型,探讨宫颈癌发展演进的多途径模式。方法根据收集的58例宫颈癌完整比较基因组杂交资料,利用Desper等建立的肿瘤树模型软件平台,构建宫颈癌的树模型。结果宫颈癌树模型提示,有6个非随机的染色体扩增或者缺失事件发生在宫颈癌变过程中: 1q, 3q, 5p, 17q, 20q和 20p,其中 3q和 1q可能是宫颈癌癌变中的重要早期事件; 3q要先于 17q发生,并且两者之间可能存在先后依存关系;而且, 1q可能是一个相对独立发生的事件;这些扩增染色体上可能存在与宫颈癌发生、发展演进密切相关的瘤基因。结论基于比较基因组杂交资料构建的宫颈癌树模型,提示宫颈癌多阶段、多途径的发展演进模式,也为后续宫颈癌相关基因的探索指明了方向。     

肠型胃癌原发灶与淋巴结转移灶癌细胞染色体异常特征的比较

目的探讨胃癌发生淋巴结转移的机制,寻找和定位与胃癌转移相关基因。方法本研究应用比较基因组杂交技术分析和比较了15例肠型胃癌患者原发灶和其对应的淋巴结转移灶癌细胞染色体基因组改变特征。结果较常见染色体DNA拷贝数扩增的部位是8q,13q,20和7;较常见染色体DNA拷贝数丢失的部位是1p,17p,19,21q和22q。其中,有意义的发现是20q12-13扩增,21qcen-21丢失,4q丢失,14q22-ter丢失。结论本研究结果表明胃癌原发灶和淋巴结转移灶癌细胞染色体基因改变最显著的部位是20q扩增,及21q,4q和14q的丢失;提示在这些部位可能存在与胃癌淋巴结转移相关的基因。     

高加索人和日本人胰腺癌基因组杂合性缺失比较

目的研究胰腺癌细胞基因组范围内的杂合性缺失(LOH),比较高加索人和日本人的异同。方法应用高密度单核苷酸多态性芯片和分析软件,研究12种高加索人和8种日本人胰腺癌细胞株基因组范围内的LOH。结果每一种胰腺癌细胞基因组中都有不同程度的LOH,其中T3M-4细胞频率最高(54.5%);高加索人和日本人胰腺癌细胞株不同染色体臂出现LOH频率不同,高加索人中最常见的LOH是9p(12/12,100.0%),而日本人出现频率最高的是13q(7/8,87.5%)。高频率的(50%以上)共同的LOH出现在染色体臂3p,8p,9p,13q,17p,18q和22q。结论胰腺癌全基因组范围内出现多处LOH。不同种族LOH出现可能频率不同,高频率的杂合性缺失区域可能含有新抑癌基因,为今后胰腺癌的研究提供了新的线索和方向。     

肝细胞癌基因组DNA拷贝数变异的性别差异性

目的: 比较男性与女性肝细胞癌（HCC）在基因组DNA拷贝数变异（CNA）方面的差异性。方法: 采用高分辨率微阵列比较基因组杂交技术（array-CGH）,检测 17例女性与46例男性HCC的CNA差异。结果: 染色体片段1q21.3-q22＋（76.5% vs. 37.0%,P = 0.009）、11q11＋（35.3% vs. 0.0%,P = 0.0002）、19q13.31-q13.32＋（23.5% vs. 0.0%,P = 0.004）和16p11.2－（35.3% vs. 6.5%,P = 0.009）发生率女性显著高于男性,而男性则有更高频率的11q11－（63.0% vs. 17.6%,P = 0.002）。进一步统计分析结果显示,11q11＋与19q13.31-q13.32＋（P = 0.042）及16p11.2－（P = 0.033）显著相关,而1q21.3-q22＋与19q13.31-q13.32＋（P = 0.046）显著相关。结论: CNA在性别特异性HCC演进过程中发挥重要作用。     

广泛标记系统在石蜡包埋膀胱癌组织比较基因组杂交中的应用

目的 探讨广泛标记系统（ULS）在石蜡包埋组织比较基因组杂交（CGH）中的应用价值,研究膀胱癌石蜡包埋组织中染色体基因组非平衡性情况.方法 对比实验：正常膀胱黏膜组织基因组DNA,用DNA酶进行酶切至适当大小的片段作为探针,用广泛标记系统直接标记DNA探针,并按1：1的量与经缺口平移标记的同一个组织的DNA探针进行CGH.提取膀胱移行细胞癌石蜡包埋组织中基因组DNA,经测定浓度、纯度,并经琼脂糖电泳筛选出20例适合于CGH的标本.同法进行ULS标记,并与缺口平移法标记的正常参照DNA进行CGH.结果 对比实验中,同一个正常膀胱黏膜组织应用2种标记方法进行CGH后,荧光强度相同,不需要调整探针的量.在所有膀胱癌组织中均检测到基因组DNA的畸变,其中发生频率较高的染色体增益区段或位点位于1q、3q、5p、8q、17q;缺失区段或位点位于8p、9q、11q、11p、17p.结论 膀胱癌中存在染色体基因组的不平衡.应用石蜡包埋的组织进行间接法CGH时,ULS标记系统是一种非常有价值的标记方法.这为充分利用石蜡档案进行CGH的研究提供了良好的方法.