High-grade mucinous tubular and spindle cell carcinoma: comparative genomic hybridization study

Mucinous tubular and spindle cell carcinoma (MTSCC) has recently been integrated into the World Health Organization classification. Although MTSCC is generally a low-grade carcinoma, MTSCC with high-grade morphology has been recently reported. We present the first case of high-grade MTSCC with comparative genomic hybridization findings. A 60-year-old Japanese man presented with weight loss and general fatigue. He underwent radical nephrectomy because of the clinical diagnosis of renal cancer. Histologic examination of renal tumor showed findings of high-grade MTSCC. Comparative genomic hybridization analysis showed gain of chromosomes 1q, 7, 16, 19q, and Y and loss of chromosomes 1p, 6p, 8p, 11q (del(11)(q23)), and 13. G-band karyotype showed gain of chromosomes 2, 3, 5, 7, 12, 16, and 20 and loss of chromosome 15. Results of our molecular genetic analysis support the idea that high-grade MTSCC is a real counterpart of low-grade MTSCC. There is no evidence to designate such tumors as unclassified renal cell carcinoma.     

Genetic alterations detected by comparative genomic hybridization and recurrence rate in epithelial ovarian carcinoma

To assess the putative correlation between comparative genomic hybridization (CGH)-detectable genetic alterations in epithelial ovarian cancer and disease recurrence, conventional CGH was performed on 45 epithelial ovarian cancers: 26 tumors from sporadic, BRCA mutation noncarriers and 11 and 8 tumors from BRCA1 and BRCA2 mutation carriers, respectively. Relevant clinical data, including histology, grade, stage, size of residual tumor, recurrence, and survival, were obtained from outpatient and inpatient charts. Among the 45 cases, the most common regions involving gain of DNA copy number were 3q (n = 23; 51%), 8q (n = 21; 47%), and 1q (n = 14; 31%), and the most common regions with loss were 19 and 22 at 9 cases (20%) each, followed by 5q (n = 6; 13%). In multivariate analysis, the total number of genetic alterations was not associated with risk of recurrence, but gain in 5p was associated with a higher risk of recurrence (hazard ratio HR = 6.06, P = 0.0399), and gain in 1p as well as loss in 5q were associated with a significant decrease in recurrence (HR = 0.08, P = 0.0079, and HR = 0.10, P = 0.0143, respectively). Recurrence rate in patients with epithelial ovarian cancer is seemingly associated with specific genetic alterations detected by CGH, but the specific genes involved and the implications of these findings await further studies.     

A comprehensive karyotypic analysis on a newly established sarcomatoid hepatocellular carcinoma cell line SH-J1 by comparative genomic hybridization and chromosome painting

We first established a sarcomatoid hepatocellular carcinoma cell line, designated as SH-J1, and applied comparative genomic hybridization and fluorescence in situ hybridization (FISH) with chromosome painting probes for the characterization of the chromosomal rearrangements. In the SH-J1 cell line, the pleomorphic spindle cells were arranged in bundles of interlacing patterns and were positive in immunohistochemical staining with hepatocyte-related markers. By G-banding and FISH, the chromosomal gains were detected at 6p and 17, whereas losses were observed at 3p21-pter, 3q27-qter, 4, 6q, 13pter-q11, 16, 18, 19p13, and Y.     

Chromosomal aberrations in nasopharyngeal carcinoma analyzed by comparative genomic hybridization.

To investigate the genomic imbalances associated with nasopharyngeal carcinoma (NPC), we have performed chromosome analysis by comparative genomic hybridization (CGH) on 51 tumors, including 25 primary and 26 recurrent tumors. The most common copy number increases occurred on chromosome arms 12p (59%), 1q (47%), 17q (47%), 11q (41%), and 12q (35%). The minimal overlapping regions were at 12p12-13, 1q21-22, 17q21, 17q25, 11q13, and 12q13. The most frequent losses were from chromosome arms 3p (53%), 9p (41%), 13q (41%), 14q (35%), and 11q (29%). The minimal overlapping regions were at 3p12-14, 3p25-26, 9p21-23, 13q21-32, 14q12-21, and 11q14-23. Compared with the primary cancers, no additional chromosomal change was found in the recurrent tumors; however, the most frequent gain in the recurrent NPCs was at 11q13 (53%) instead of 12p in the primary tumors. An increase of gene alterations correlated with clinical stage. Our results provide a first comprehensive view of the genomic changes associated with NPC and reveal several new sites of genomic imbalance, indicating the possible involvement of novel oncogenes/tumor suppressor genes in the carcinogenesis of NPC.     

Analysis of genetic alterations in primary nasopharyngeal carcinoma by comparative genomic hybridization

To identify genetic alterations associated with the development and progression of human nasopharyngeal carcinoma (NPC), 57 tumors were analyzed by comparative genomic hybridization (CGH). In 47 cases, chromosomal imbalances were found. Several recurrent chromosomal abnormalities were identified in the present study. The most frequently detected chromosomal gains involved chromosome arms 12q (24 cases, 51%), 4q (17 cases, 36%), 3q (16 cases, 34%), 1q (15 cases, 32%), and 18q (15 cases, 32%). Common regions of gain involved 12q13--q15, 4q12--q21, and 3q21--q26. High-copy-number increases of chromosomal materials were detected in four chromosomal regions, 3q21--q26.2, 4p12--q21, 8p, and 12q14--q15. The most frequently detected loss of chromosomal materials involved chromosome arms 16q (26 cases, 55%), 14q (21 cases, 45%), 1p (20 cases, 43%), 3p (20 cases, 43%), 16p (19 cases, 40%), 11q (17 cases, 36%), and 19p (16 cases, 34%). The most common regions of loss involved 14q24--qter, 1pter--p36.1, 3p22--p21.3, 11q21--qter, and the distal region of 19p. Genomic alterations detected by CGH were compared and found to be largely consistent with those identified in banding analysis and loss of heterozygosity studies. However, several previously unrecognized recurrent alterations were also identified in the present study, including gain of 4q and 18q, and loss of 16q, 14q, and 19p. In addition, gain of 1q, 8q, 18q, and loss of 9q showed a statistically significant association with advanced clinical stages (P < 0.05). Identification of recurrent sites of chromosomal gain and loss identify regions of the genome that may contain oncogenes or tumor suppressor genes, respectively, which may be involved in the tumorigenesis of NPC. Published 2000 Wiley-Liss, Inc.     

Characterization of drug-resistant cell lines by comparative genomic hybridization.

The development of resistance to cytostatic agents is a serious obstacle to the success of cancer therapy and has been the focus of many research efforts. Traditionally, cell lines are selectively cultured in the presence of cytostatic agents and the biochemical and cytogenetic properties of the cell lines are then analyzed. In order to better understand the mechanisms by which drug resistance is mediated, we have analyzed three cell lines, each derived from the parent line K562, which are resistant to vincristine, mitoxantrone, or idarubicin, using comparative genomic hybridization (CGH). In each case, CGH successfully identified amplifications and/or deletions unique to the drug-resistant selected cell lines. Further characterization of the genetic regions identified in the CGH analysis could greatly contribute to our understanding of acquired drug resistance, and could potentially impact the clinical management of cancer.     

Detection of chromosomal imbalances in transitional cell carcinoma of the bladder by comparative genomic hybridization.

Comparative genomic hybridization (CGH) was applied for a comprehensive screening of chromosomal aberrations in 14 transitional cell carcinomas of the bladder of different grade and stage. The results were compared in a number of selected cases with those obtained by restriction fragment length polymorphism analyses and targeted fluorescence in situ hybridization. Distinct amplifications, found with CGH, were located on 3p22-24, 10p13-14, 12q13-15, 17q22-23, 18p11, and 22q11-13. These high copy number amplifications and the frequency of imbalances involving chromosome 5, occurring in 4 of 14 cases, have not yet been identified in transitional cell carcinomas. Apart from these new aberrations, imbalances were detected in 3 or more cases for chromosomes 9 and 11, as already described previously in the literature. In four tumors, the copy number of specific chromosomal regions was also analyzed by interphase cytogenetics. Although in most instances the CGH data were confirmed, in one tumor, distinct differences were observed, possibly a result of heterogeneity of the tumor cell population. Furthermore, the CGH data were compared with loss of heterozygosity as revealed by restriction fragment length polymorphism analysis in the same tumors. In 80% of informative cases, no loss was detected by restriction fragment length polymorphism or by CGH. Of the 15 cases of loss of heterozygosity, 7 showed a loss also with CGH, whereas in 8 cases no loss was observed. In summary, CGH is a fast method to obtain a comprehensive picture of chromosomal imbalances in transitional cell carcinomas, including a number of previously unknown genomic alterations such as high level amplifications.     

Genetic alterations in epithelial ovarian tumors analyzed by comparative genomic hybridization

The genetic changes involved in the pathogenesis of ovarian carcinoma are not completely understood. To investigate this matter, we studied paraffin-embedded, microdissected tissue of 47 ovarian epithelial tumors (9 adenomas, 11 tumors of low malignant potential [LMP], 14 serous carcinomas, and 13 nonserous carcinomas) using comparative genomic hybridization (CGH). (The primary data used in this study are available at our CGH online tumor database at http://amba.charite.de/cgh.) Chromosomal imbalances were found in 1 serous adenoma and in 7 LMP tumors. In the latter the alterations appeared randomly and showed no overlap with alterations found in invasive carcinomas. Although the mean aberration number of low-grade serous carcinomas was comparable to LMP tumors, the imbalances of the former occurred with high incidence (>50%) and were found at different localizations. High-grade serous carcinomas had more than twice as much chromosomal imbalances as low-grade serous carcinomas and also had pronounced alterations. In serous carcinomas, gains were found on 3q, 6p, 7, 8q, and 20, and losses were found on 4q, 6q, 12q, 13q, and 16q. Comparing serous and nonserous carcinomas, the mean aberration number was comparable, but the number of high incidence changes was lower, and the most frequent imbalances were losses on 13q and gains on 20p. Overlapping alterations occurring in serous and nonserous carcinomas were gains on 3q and 6p, as well as losses on 4q. Chromosomal imbalances associated with poor prognosis of ovarian carcinomas were gains on 6p, 7q, and 13q and losses on 15q, 17p, 18q, and 21q. Our data indicate that serous LMP tumors and invasive carcinomas have different genetic aberrations, indicating that invasive carcinomas do not arise from preexisting serous LMP tumors. On the other hand, there are common genetic abnormalities in serous and nonserous carcinomas, suggesting that they have very early lesions in common but take different paths of further development.     

Analysis of ameloblastomas by comparative genomic hybridization and fluorescence in situ hybridization

In order to characterize the chromosomal alterations in ameloblastomas, a combination of comparative genomic hybridization (CGH) and fluorescence in situ hybridization (FISH) techniques was performed on 9 tumors. Chromosomal alterations including a gain at 1q and losses at 1pter, 10q, and 22q could be detected by CGH only in 1 tumor. Interphase FISH analysis, using centromeric probes for chromosomes 1, 10, and 22 as well as region-specific probes for 1p36 and 10q26, revealed the most frequent alterations to exist in the tumor with the abnormal CGH profile. These alterations included marked to slight increases of monosomic cells for chromosome 10 (91.5%), 10q26 (35.8%), 1p36 (24.4%), and chromosome 22 (18.8%), as well as significant elevations of trisomic cells for chromosome 1 (41.2%). Moreover, FISH analysis revealed a frequent loss of chromosome 22 in all tumors examined, except for one lesion, indicating that loss of the entire or a part of this chromosome is a common event in ameloblastomas, possibly being a predisposing factor to ameloblastoma tumorigenesis.     

Atypical endometrial hyperplasia shares genomic abnormalities with endometrioid carcinoma by comparative genomic hybridization.

Endometrial hyperplasia is a common disorder that is now observed with increasing frequency in women treated with hormonal replacement therapy or with tamoxifen. This study was undertaken to determine whether genomic features of various forms of endometrial hyperplasias would allow their classification as a benign, premalignant, or malignant abnormality. Comparative genomic hybridization (CGH) was performed on endometrial glands microdissected by laser capture microscope from 19 archival endometrial samples, comprising 5 normal endometria, 1 polyp, 2 simple hyperplasias, 5 hyperplasias with nuclear abnormalities (atypical hyperplasias), and 4 low-grade and 2 high-grade endometrioid carcinomas, 1 with squamous component (adenoacanthoma). Genomic DNA, extracted from the glands and the squamous component in 1 case, was amplified by degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR) and compared with sex-matched DNA by CGH. No genomic imbalances were observed in the normal samples, the polyp, or the simple hyperplasias. However, in atypical hyperplasia, regardless of the level of cytologic atypia, genomic abnormalities were observed that also occurred in endometrioid carcinomas. Chromosomes 1, 8, and 10 were most often affected. The results are compared with molecular genetic abnormalities recently reported in these lesions. This study strongly suggests that atypical endometrial hyperplasias are closely related to endometrioid carcinoma and should be considered precancerous lesions, contrary to simple hyperplasia, which is a benign disorder. The squamous component of one of the high-grade carcinomas showed genetic abnormalities similar to those of endometrioid carcinoma and therefore does not represent squamous metaplasia but is an integral part of the malignant process.     

比较基因组杂交研究鼻咽癌遗传变异

目的 了解鼻咽癌遗传学改变的特征。方法 应用比较基因组杂交检测20例鼻咽癌基因组的不平衡即DNA的丢失或扩增。结果 鼻咽癌常见的扩增的染色体是1q、2、3q、7q、8q、12;常见的缺失的染色体为3p、9p、11q、16q。结论 鼻咽癌细胞中存在多条染色体拷贝数的改变,由此引起相应瘤基因的扩增和抑癌基因的丢失可能参与了鼻咽癌的发生、发展。     

Molecular cytogenetic analysis of non-small cell lung carcinoma by spectral karyotyping and comparative genomic hybridization

The overall pattern of chromosomal changes detected by spectral karyotype (SKY) analysis of two cell lines of each major histological subtype of NSCLC, namely squamous cell carcinoma (SQCC) and adenocarcinoma (ADC), indicated a greater degree of chromosomal rearrangement, than was present or predicted by either comparative genomic hybridization (CGH) or G-banding analysis alone. To investigate these observations, CGH was used to screen DNA derived from 8 primary tumors and 15 cell lines. The results indicated that the most frequently gained chromosome arms were 5p (70%), 8q (65%), 15q (52%), 20q (48%), 1q (43%), 19q (39%), 3q (35%), and 11q (35%). Chromosomal losses were less frequently observed, and included 18q (39%), 9 (35%), 6q (30%), 13q (21%), 5q12-q32 (17%), and 19p (17%). Amplifications were found on 2p23-p24, 3q24-q27, 5p, 6cen-p21.1, 6q26, 7p21, 7q31, 8q, 11q13-qter, 20q12-q13.2. Comparison between CGH findings of the two major histological subtypes showed that gains at 1q22-q32.2, 15q, 20q, and losses at 6q, 13q, and 18q was common in ADCs, whereas SQCCs exhibited gains/amplifications at 3q. Distal 8q was gained by CGH in 65% of tumors of both subtypes. Low level MYCC amplification was confirmed by direct fluorescence in situ hybridization (FISH) analysis. The pattern of overall chromosomal changes detected using combinations of molecular cytogenetic analytical methods suggests that it will be easier to detect recurrent subtype-dependent aberrations in NSCLC.     

Recurrent chromosome alterations in hepatocellular carcinoma detected by comparative genomic hybridization

Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide and has a very poor prognosis. Fifty primary HCC cases have been analyzed in the present study to explore the association between genomic alteration in primary HCC and clinical features. Several recurrent chromosomal abnormalities were identified in this study. The most frequently detected chromosomal gains involved chromosome arms 1q (33/50 cases, 66%), 8q (24/50 cases, 48%), and 20q (10/50 cases, 20%). High-copy-number amplifications involving 1q (4 cases), 8q (3 cases), and 20q (3 cases) were detected, and a minimum overlapping amplified region at 1q12-q22 was identified. The most frequently detected loss of chromosomal material involved 16q (35/50 cases, 70%), 17p (26/50 cases, 52%), 19p (21/50 cases, 42%), 4q (20/50 cases, 40%), 1p (18/50 cases, 36%), 8p (16/50 cases, 32%), and 22q (14/50 cases, 28%). The associations between genomic alterations detected in the present study and clinical features including clinical stage, tumor size, HBV infection, chronic liver disease, and liver cirrhosis were explored. Our CGH results suggest that the gain of 20q and deletion of 8p are late genetic alterations in HCC, because the incidence of these alterations was obviously increased in the advanced clinical stages. Another finding showed that loss of 8p and gain of 8q and 20q are associated with tumor size. The recurrent gain and loss of chromosomal regions identified in this study provide candidate regions that may contain oncogenes or tumor suppressor genes respectively involved in HCC development and progression.     

Recurrent chromosomal abnormalities in hepatocellular carcinoma detected by comparative genomic hybridization

Comparative genomic hybridization (CGH) was used to evaluate and map genomic aberrations in 50 hepatocellular carcinomas (HCCs) from patients chronically infected with hepatitis B virus (HBV). CGH clearly detected nonrandom genomic imbalances. Losses were most prevalent on chromosome regions 4q (70%), 8p (65%), 16q (54%), 17p (51%), 13q and 6q (37% each), and 1p (30%). The most frequent gains occurred on 8q (60%), 1q (58%), and 6p and 17q (33% each). In a few cases, sequence amplifications were detected that were mapped to bands 11q12, 12p11, 14q12, and 19q13.1. This study represents the first analysis of primary liver cancers by CGH, and it confirms the presence of previously known chromosomal aberrations in HCC and highlights new quantitative abnormalities and sequence amplifications. These findings should lead to the characterization of new loci involved in liver cancer pathogenesis. Genes Chromosom. Cancer 18:59–65, 1997. © 1997 Wiley-Liss, Inc.     

Intratumoral heterogeneity in breast carcinoma revealed by laser-microdissection and comparative genomic hybridization

To evaluate the potential cytogenetic heterogeneity in breast carcinoma, several small cell groups (each consisting of 20 to 50 cells) were investigated within paraffin sections. By laser-microdissection, three to seven cell groups were taken per case. The DNA was amplified by degenerate oligonucleotide primed PCR (DOP-PCR), and the samples were analyzed by CGH for chromosomal gains and losses. Two ductal invasive breast carcinomas, one of them with two lymphnode metastases, were investigated. To compare the results from the small samples, CGH was also performed on DNA isolated from the tumorous regions of three to five serial sections (10⁷ to 10⁶ cells). The aberrations observed in the microdissected tumor samples were multiple and involved up to 14 different chromosomal or subchromosomal regions. The most frequent changes were gains on chromosomes 12q (14/20) and 20q (16/20), and loss on 13q (12/20). Some aberrations have rarely been detected (e.g., loss on 2p, gain on 8q). Comparing chromosomal imbalances in primary tumors and lymph node metastases, more consistent changes were found between the primary tumor and its corresponding metastases than between both primary tumors. The laser-microdissected samples in general showed more chromosomal aberrations than DNA isolated from several tumor sections. Our CGH results were confirmed by fluorescence in situ hybridization (FISH) for the chromosomal regions of centromere 1 and 20, and 20q13. In addition, microsatellite analyses on 31 samples confirmed our CGH findings for selected chromosome regions 2p and 11q. It can be concluded that there is a distinct intratumoral heterogeneity in primary breast tumors as well as in the corresponding lymph node metastases. The combination of microdissection and CGH enabled us to detect cytogenetic aberrations from important clones which are missed when analyzing DNA extracted from large cell numbers.     

Mucinous tubular and spindle cell carcinoma with aggressive histomorphology--a sarcomatoid variant

Mucinous tubular and spindle cell carcinoma (MTSCC) is a recently described renal epithelial tumor. The bland cytomorphology of the spindled component and low-grade behavior help in its differentiation from sarcomatoid renal carcinoma. Sarcomatoid change has been reported in most histologic variants of renal cell carcinoma apart from MTSCC. Herein we report a case of an MTSCC in a 72-year-old female patient with high-grade spindled areas resembling fibrosarcomatous and undifferentiated pleomorphic sarcoma patterns with metaplastic bone. This index case also demonstrates a high proliferation index and extensive necrosis representing the first documented case of sarcomatoid change in MTSCC.     

Analysis of cytogenetic aberrations in esthesioneuroblastomas by comparative genomic hybridization

Esthesioneuroblastoma (ENB) are rare tumors originating from the olfactory epithelium of the superior nasal cavity. This lesion is morphologically closely related to Ewing sarcoma and other peripheral primitive neuroectodermal tumors (pPNET). The affiliation of ENB to the pPNET family is still under discussion. Only very limited and contradictory cytogenetic data are available on ENB and only one patient has been analyzed by comparative genomic hybridization (CGH), so far. In the present study, genomic imbalances of three ENB were analyzed by CGH to evaluate (1) a recurrent pattern of imbalances, and (2) its relation to the pPNET family. The CGH analysis of three ENB revealed multiple recurrent aberrations including DNA overrepresentations of chromosomal material of the entire chromosome 19, partial gains of the long arms of chromosomes 8, 15, and 22, and deletions of the entire long arm of chromosome 4. Beside these common aberrations, several single gains and losses occurred, that is, gains on 6p, 10q, 1p, 9q, and 13q. We confirmed the former observation of amplified genetic material on chromosome 8 and found several new, currently not described recurrent genetic aberrations distinct from those described for pPNET. Our findings give evidence that ENB is not part of the pPNET family. We suggest that the combined gain of genetic material on 15q, 22q, and chromosome 8 might be indicative for ENB. To verify our findings and to define prognosis-related aberrations, a larger number of cases needs to be studied.     

Lung carcinoma with spindle cell components: sixteen cases examined by immunohistochemistry.

Sixteen cases of lung carcinoma with spindle cell components were studied by conventional histochemistry and immunohistochemistry. The epithelial components were squamous cell carcinoma in six cases, adenocarcinoma in four, adenosquamous carcinoma in five, and large cell carcinoma in one. In every case sarcomatous areas were distinctly observed and, in general, neoplastic spindle cells proliferated in close proximity to the epithelial elements. Some of the histochemical procedures suggested mesenchymal features in the stroma of the exophytic portions of three cases, but heterogeneous elements, such as bone or striated muscle, were not observed. By immunohistochemical examination the epithelial elements showed positive reactions for keratin, epithelial membrane antigen, and/or carcinoembryonic antigen to a varying degree according to the histologic types. The spindle cell elements revealed a positive immunoreaction for keratin in all but one case. Epithelial membrane antigen was demonstrated in sarcomatous areas of only five cases and carcinoembryonic antigen was not expressed at all. In contrast, vimentin was distinctly demonstrated in sarcomatous areas of five cases, although other mesenchymal markers, such as desmin, actin, and myosin, were negative. These findings indicate that the spindle cell components in these 16 cases may represent mesenchymal features with partial or complete loss of epithelial features.     

Genomic alterations in distal bile duct carcinoma by comparative genomic hybridization and karyotype analysis.

We report genomic abnormalities identified in 14 human primary common bile duct carcinomas analyzed by cytogenetics or comparative genomic hybridization, or both. Combining the results of the two methods of analysis, 11 chromosomal arms were observed to be gained in whole or in part, and 9 chromosomal arms were lost in whole or in part in at least four tumors each. The most frequently lost chromosomal regions were, in decreasing order: 18q (eight tumors); 6q and 10p (seven tumors each); 8p, 12q, and 17p (six tumors each); and 7q, 12p, and 22q (four tumors each). The most frequently gained regions were 8q and 20q (six tumors each); 12p, 17q, and Xp (five tumors each); and 2q, 6p, 7p, 11q, 13q, and 19q (four tumors each). These results are similar to those we have previously reported in pancreatic cancer and suggest that carcinomas of the common bile duct and pancreas share a number of genetic changes.