Molecular cytogenetic analysis of a gliosarcoma with osseous metaplasia.

Gliosarcoma, a rare glioblastoma variant, is composed of a glial and a mesenchymal component. Though the mesenchymal portion most commonly resembles a fibrosarcoma, other differentiation patterns have been observed. We present the first genomic characterisation (karyotyping followed by FISH and array comparative genomic hybridisation analysis) of a gliosarcoma with osseous metaplasia. In addition to chromosomal changes often found in gliomas (+7, -10, -13, and -22), the tumour cells also harboured a hitherto unknown t(3;21)(q13∼21;q21∼22).

Telomeric associations correlate with telomere length reduction and clonal chromosome aberrations in giant cell tumor of bone.

Giant cell tumor of bone (GCTB) is characterized cytogenetically by frequent telomeric associations (tas). To explore the mechanisms behind the formation of tas in GCTB and to investigate their karyotypic consequences, the frequencies of tas and clonal aberrations other than tas in 20 GCTBs were compared to telomere length and status, as assessed by quantitative PCR, fluorescence in situ hybridization (FISH), and expression levels of four genes involved in telomere maintenance. Based on the G-banding results, the tumors were divided into two groups, one with a high frequency of tas and one with a low frequency. Clonal aberrations were found to be restricted to the group with a high level of tas, and the same group showed a significantly larger reduction in telomere length in tumor cells compared to peripheral blood cells. Furthermore, 65 out of 66 tas analyzed by FISH were negative for telomeric sequences. The expression levels of TERT, TERF1, TERF2, and POT1 did not correlate with telomere length or the frequency of tas. Thus, the present findings provide strong support for the notion that decreased telomere length is a prerequisite for tas in GCTBs and that the clonal changes occurring in GCTBs are derived from tas.

Nonrandom chromosome abnormalities in short-term cultured primary squamous cell carcinomas of the head and neck.

We report the finding of clonal chromosome abnormalities in short-term cultures from 44 squamous cell carcinomas of the head and neck region. Eleven tumors had gain or loss of the Y chromosome, sometimes one clone with +Y and another with -Y, as the sole anomaly, whereas the remaining 33 all carried structural rearrangements and usually were cytogenetically complex with multiple aberrations. The chromosomal bands most frequently involved were, in decreasing order of frequency, 8p11-q11, 1p11-q11, 3p11-q11, 11q13, 13p11-q11, 1p13, 5p11-q11, 7p11-q11, 15p11-q11, and 14p11-q11. Almost one-half of the breakpoints were located in centromeric or juxtacentromeric bands. Recurrent aberrations included i(8q), i(5p), i(1q), del(3)(p11-12), del(5)(p11), t(1;1)(p13;q25), and der(14;15)(q10;q10). To see whether the karyotypic features of head and neck squamous cell carcinoma differ depending on exact tumor site, we added to the present series our previously published 23 karyotypically abnormal head and neck squamous cell carcinomas that had been cultured in the same way as the tumors of the present series. In the ensuing correlation analysis, tumors of the oral cavity and oropharynx and hypopharynx were found to share many features: highly complex karyotypes were frequent, often containing isochromosomes such as i(8q) and i(5p), and also rearrangements of 11q13 (often as homogeneously staining regions) and loss of genetic material from the short arms of chromosomes 3, 13, 14, and 15 were repeatedly seen. Laryngeal carcinomas, on the other hand, often had simple karyotypic changes.

Clonal chromosome-aberrations in fibrocystic breast disease-associated with increased risk of cancer.

Short-term cultures of 29 samples of fibrocystic breast disease were cytogenetically analyzed. Clonal chromosome aberrations were found in six specimens, whereas the remaining 23 had a normal karyotype. Three of the abnormal samples displayed karyotypic anomalies previously associated with breast cancer, i.e., gain of Iq, trisomy 18 and cytogenetic multiclonality. Furthermore, all cytogenetically aberrant specimens had either proliferative disease without atypia or atypical hyperplasia, features of fibrocystic disease considered risk factors for subsequent breast cancer development. The cytogenetic similarities between breast carcinomas and proliferative fibrocystic breast disease add further support for classifying certain types of fibrocystic disease as a premalignant condition. Whether cytogenetically abnormal fibrocystic lesions are the ones that subsequently progress to cancer remains to be elucidated.

Altered expression of TGFB receptors and mitogenic effects of TGFB in pancreatic carcinomas.

Alteration of the transforming growth factor beta (TGFB) signalling pathway is important in pancreatic carcinogenesis, as shown by the frequent inactivation of the downstream target SMAD4. We recently analysed a series of pancreatic carcinoma cell lines with respect to alterations of five SMAD genes involved in TGFB signalling, and showed that SMAD4 was structurally rearranged in 42% of these. This pathway may, however, also be affected by alterations of genes whose products regulate the activation of TGFB as well as of TGFB receptor genes. We therefore studied the expression of UPA, UPAR, IGF2R, ALK5 (TGFBR1), TGFBR2, TGFBR3, ENG, ALK1, TGFB1, TGFB2, and TGFB3 in a series of 14 pancreatic carcinoma cell lines. We also analysed ALK5 and TGFBR2 for mutations, cell surface localisation of TGFBR2 and ENG, and TGFB1 response. No mutations of ALK5 or TGFBR2 were found. However, 4 cell lines were methylated within the ALK5 promoter region. ALK5 expression was strongly reduced in 9 cases, whereas TGFBR2 expression was increased in 12 of the cell lines. The TGFB signalling associated receptors ENG and ALK1 were co-expressed in 4 of the cell lines. There was no evidence for disruption of the UPAR-IGF2R TGFB activating pathway. The response to TGFB1 was analysed in 12 cell lines, and 6 of these (50%) showed increased proliferation. The cell lines stimulated by TGFB showed frequent mutations of SMAD4, KRAS2, and TP53, as well as frequent absence of CDKN2B expression. These results suggest that the ALK5-SMAD4 part of the TGFB signalling pathway is a major target for inactivation in pancreatic carcinomas, that the expression of TGFBR2, TGFBR3, and receptors involved in TGFB activation are maintained, and that alterations of components of the TGFB signalling pathway may be accompanied by a positive effect of TGFB on cell growth.

Cytogenetically detected clonal heterogeneity in a duodenal adenocarcinoma.

A primary duodenal adenocarcinoma, a tumor type for which no previous chromosome data existed, was cytogenetically analyzed after short-term culture. The main tumor mass was localized in the pancreatic head, but the histopathologic examination revealed its duodenal origin. A total of six abnormal, karyotypically unrelated, clones were identified. The largest exhibited clonal evolution and consisted of two subclones with massively rearranged karyotypes in the hypodiploid and hypotetraploid range. Chromosome imbalances brought about by these complex changes were gain of 1q, losses of chromosomes 6 and 9, and total or partial losses of 1p, 3p, 3q, 9p, 10p, 17p, 17q, 18q, 20p, and 20q. The remaining five smaller clones had 1-2 numerical or balanced structural chromosome aberrations. The present study thus revealed yet another epithelial tumor type characterized by karyotypically unrelated clones. For this as for other tumors, the pathogenetic significance of such cytogenetic polyclonality remains uncertain.

Cytogenetic heterogeneity in a second primary radiation-induced bladder carcinoma: ten karyotypically unrelated clones.

Cytogenetic analysis of a transitional cell carcinoma (TCC) of the bladder, the tumor having developed 32 years after the patient received pelvic irradiation and interstitial radium implantation for an endometrial carcinoma, revealed the presence of 10 cytogenetically abnormal, unrelated clones. Although the tumor was poorly differentiated, all clones were pseudo- or near-diploid with rather simple balanced or unbalanced structural rearrangements or both. The chromosomes involved in structural changes more than once were chromosomes 8, 9, and 11, which were rearranged in three clones, and chromosomes 3 and 17, both rearranged in two clones. No previous TCC of the bladder with cytogenetically unrelated clones has been reported, nor has any such radiation-induced tumor with chromosomal abnormalities been described. The distinct karyotypic and clonal pattern of the case presented here is probably indicative of a carcinogenic field effect due to the previous pelvic irradiation. Postradiation bladder carcinomas thus seem to be distinct cytogenetically in addition to their known unique etiological and clinical features.

Cytogenetic heterogeneity in a clear cell hidradenoma of the skin.

Short-term cultures from a clear cell hidradenoma, a benign skin tumor for which no chromosome data exist, were cytogenetically analyzed. A total of eight unrelated aberrant clones were identified. The karyotypic profiles of two separately processed parts of the sample--a tumor nodule and seemingly normal adjacent dermal tissue--were different. Characteristic for the tumor nodule was a single abnormal clonal population consisting of three subclones: 46,XY,der(2)inv(2)(p13q23)t(2;9)(p13;q22), der(9)t(2;9)(q23;q22),t(11;19)(q21;p13),t(12;19)(q24;p13)/46,idem, inv(1)(p32q44)/92,idemx2. The adjacent tissue contained, in addition to the clone found in the tumor nodule, a spectrum of unrelated clones, the largest of which also showed clonal evolution: 45-47,XY,t(3;6)(p25;p25),t(12;17)(q15;q12),-17,+r(17)x2 [cp]/45-47,idem,inv(5)(p15q22)/90-94,idemx2. The remaining six clones found in this part were small and had simpler numerical or structural aberrations. The multiclonal pattern observed in this hidradenoma seems to reflect both cytogenetic convergence and divergence during neoplastic progression. The presence of unrelated clones may be an indication that the tumor was of multicellular origin.

Chromosomal abnormalities in two bladder carcinomas with secondary squamous cell differentiation.

Two secondary squamous cell carcinomas of the bladder (i.e., tumors that originated from primary transitional cell carcinomas) were examined cytogenetically. Both tumors showed complex karyotypes with many of the same aberrations that have formerly been described in transitional cell carcinomas. Monosomy 9, trisomy 7, and rearrangements of chromosomes 3, 8, 10, 13, and 17 were common to both tumors. Among other changes that have been implicated in bladder carcinogenesis, an isochromosome for 5p was seen in one tumor and loss of 11p material in the other. Our findings indicate that secondary squamous cell carcinomas of the bladder are karyotypically indistinguishable from advanced transitional cell carcinomas of the same organ. The putative genetic changes that steer the differentiation of the neoplastic epithelium in the direction of squamous cells thus remain unknown.

Extensive cytogenetic heterogeneity in a benign retroperitoneal schwannoma.

A benign retroperitoneal schwannoma from a patient without prior exposure to radiotherapy or chemotherapy was analyzed by chromosome banding after short-term culture. An extensive intratumor heterogeneity in the form of 29 karyotypically related as well as unrelated clones was found. The aberrant clones were diploid or near-diploid and displayed both numerical and structural changes. All chromosomes, except 11, 16, and 20, were affected. Numerical changes included trisomies X, 7, 9, 17, and 18, and monosomies 13 and 18. No clonal loss of chromosome 22, the most characteristic abnormality in schwannomas of other locations, was, however, detected. The structural aberrations resulted in a total of 58 chromosomal breakpoints, with chromosomes 18, 1, and 15 participating in rearrangements most frequently, followed by chromosomes 14, 2, and 22. A striking finding was the clonal involvement of 18p11 in eight rearrangements affecting different chromosomes, suggesting alteration of telomeric function. The molecular mechanisms underlying the observed massive polyclonality in the schwannoma, particularly the presence of cytogenetically unrelated clones, are unknown and probably heterogeneous.

Cytogenetic analysis of upper urinary tract transitional cell carcinomas.

Ten primary (nine regular and one post-radiation) upper urinary tract transitional cell carcinomas (TCC), i.e., tumors of the renal pelvis and ureter, were obtained from 10 patients following nephroureterectomy and processed for cytogenetic analysis after short-term culturing. Clonal chromosomal aberrations were found in eight tumors. While 10 karyotypically related and/or unrelated clones were detected in the post-radiation tumor, cytogenetic monoclonality was seen in all other tumors. With the exception of two tumors with loss of the Y chromosome as the only change, chromosome 9 was invariably involved, either with loss of the entire chromosome or with partial loss from the short arm. Our findings indicate that the karyotypic profile of upper urinary tract TCC is identical to that of bladder TCC, an indication that the same pathogenetic mechanisms are at work in both regions.

Abnormal karyotypes in three carcinomas of the gallbladder.

Short-term cultures from three carcinomas of the gallbladder were cytogenetically analyzed. All three had an abnormal karyotype. The modal chromosome number was near- or hypertriploid in two tumors and near diploid in the third. Structural rearrangements of chromosomes 1 and 3, loss of material from the long arm of chromosome 18, and loss of chromosome 21 material were common to all three tumors and would seem to be the best candidates for nonrandom karyotypic changes in carcinomas of the gallbladder.

Psychological determinants of information searching activity.

The paper deals with the application of the activity theory in describing psychological determinants of the information searching activity. The notions of information behavior, information retrieval, information competence, information retrieval activity given in Russian and English scientific literature are compared. The research approach to the information retrieval activity based on the principles developed in the Russian theory of activity is described; and the fundamentals of G. Sukhodolsky's generalized conception of activity are presented for the first time. Analysis of empirical researches showed that specific features of information search depend on how the user evaluates information resources, information, conditions and results of search. Psychological determiners of information search may be detected as the system of evaluative alternatives, which is generated by the user during the process of his experience growth. We discovered that user's evaluation system is also related to his individual typological and personal regulative features and determines the choice of the search strategy.

Locus-specific multifluor FISH analysis allows physical characterization of complex chromosome abnormalities in neoplasia.

Novel techniques in molecular cytogenetics have radically improved the ability to characterize genetic changes in neoplastic cells. In parallel, a rapid development in high-throughput genomics has resulted in detailed physical maps of the human genome. Combining these two fields, we have developed a method for the simultaneous visualization of several physically defined segments along a chromosome. Seven YAC clones and one subtelomeric cosmid clone from chromosome 12 were labeled with unique combinations of four fluors and hybridized to metaphase chromosomes from neoplastic cells. In a uterine leiomyoma and a myxoid liposarcoma with translocations 12;14 and 12;16, the breakpoints in chromosome 12 could be localized to the HMGIC and CHOP regions, respectively. In the other tumors, more complex aberrations were visualized, including two inversions in 12q with a common breakpoint between MDM2 and D12S332 in a pleomorphic adenoma, amplification of MDM2 and CDK4 in ring chromosomes from a malignant fibrous histiocytoma, and amplification of KRAS2 together with other unbalanced rearrangements in two pancreatic adenocarcinomas. Combinatorially labeled single-copy probes may thus simultaneously provide physical localization of breakpoints and an overview of complex structural rearrangements. Genes Chromosomes Cancer 28:347-352, 2000.

Cytogenetic and fluorescence in situ hybridization analyses of chromosome 19 aberrations in pancreatic carcinomas: frequent loss of 19p13.3 and gain of 19q13.1-13.2.

Cytogenetic investigation of nine pancreatic carcinomas revealed structural rearrangements of chromosome 19 in eight cases, resulting in a high frequency of 19p losses and 19q gains. To characterize these imbalances further, we performed fluorescence in situ hybridization (FISH) analysis with 12 mapped and evenly distributed cosmids. The FISH study not only verified the cytogenetic findings but also disclosed additional chromosome 19 aberrations not detected by chromosome banding analysis. Seven carcinomas displayed 19p losses, always including 19p13.3, either through partial- or whole-arm deletions. Six cases showed gain of 19q, usually as one to two copies above the ploidy level. In one case, a high level of amplification in 19q13.1 was seen. The commonly overrepresented segment was 19q13.1-13.2. These results suggest that genes of importance in the development of pancreatic carcinomas are located in 19p13.3 and 19q13.1-13.2.

Chromosome abnormalities in squamous cell carcinoma of the urethra.

Cytogenetic analysis of short-term cultured cells from a urethral squamous cell carcinoma showed the tumor to have an abnormal, karyotypically complex near-diploid clone as well as its near-tetraploid duplicate. This is the first urethral carcinoma with chromosomal abnormalities to be reported. Chromosomes Y, 2, 3, 4, 6, 7, 8, 11, and 20 were all involved in numerical and/or structural rearrangements. Of particular interest was the fact that no rearrangements of chromosomes 9 and 17, both almost ubiquitously involved in transitional cell carcinoma of the urinary tract, were seen.

Characterization of genomically amplified segments using PCR: optimizing relative-PCR for reliable and simple gene expression and gene copy analyses.

Gene amplification is one of the mechanisms for oncogene activation in solid tumors. The size of the amplified regions may vary considerably among individual tumors, and more than one gene may be affected within the same amplicon. The main objective in analyzing genomic amplifications has therefore been to map the shortest region involved and to identify genes with increased expression as a result of the increased gene copy number. To facilitate such an analysis, we have developed simple polymerase chain reaction (PCR) procedures using the internal standards beta-actin (ACTB) and L1Hs for gene expression and gene copy number analyses, respectively. We used cDNA derived from pancreatic carcinoma cell lines, and genomic DNA extracted from the same cell lines, as templates in the gene expression and in the gene copy number analyses, respectively. To determine the optimal number of PCR cycles, dilution series of the templates were made. Furthermore, competing primers were used to adjust for differences in target sequence levels. We show that by these simple means it is possible to determine optimal conditions for expression analyses. In addition, the procedure was adapted for the analysis of gene copy number changes at the genomic level using L1Hs as the internal standard. This PCR method makes it possible to produce detailed gene copy number profiles of amplified genomic regions.

Telomere shortening and mitotic dysfunction generate cytogenetic heterogeneity in a subgroup of renal cell carcinomas.

Most renal cell carcinomas (RCC) show only simple chromosomal changes. However, a more complex cytogenetic pattern has been found in a subgroup of aggressive RCC, indicating that further accumulation of chromosome changes could play a role in tumour progression. To explore the possible mechanisms behind cytogenetic evolution in RCC, a parallel assessment of chromosome mutations and mitotic segregation pattern in eight tumours was performed. In the majority of cases, no abnormalities in the cell division machinery were found and the rate of alterations in chromosome copy number, as measured by interphase FISH, was similar to that in non-neoplastic cells. This was reflected by relatively simple karyotypes, with little cytogenetic intratumour heterogeneity. In contrast, another group of tumours exhibited several cytogenetically related clones with additional structural chromosomal changes at two or more ploidy levels and a frequency of copy number alterations that was higher than in normal cells. In these cases, the telomere repeat sequences were abnormally short and chromosomal breakage-fusion-bridge events were observed at cell division, as well as multipolar configurations and supernumerary centrosomes. Abnormalities of the cell division machinery may thus contribute to the evolution of complex karyotypes and genetic intratumour heterogeneity in a subgroup of RCC.

Karyotypic characterization of urinary bladder transitional cell carcinomas.

Samples from 34 primary transitional cell carcinomas (TCCs) of the bladder were short-term-cultured and processed for cytogenetic analysis after G-banding of the chromosomes. Clonal chromosome abnormalities were detected in 27 tumors and normal karyotypes in 3, and the cultures from 4 tumors failed to grow. Losses of genetic material were more common than gains, indicating that loss of tumor suppressor genes may be of major importance in TCC pathogenesis. There was no clonal heterogeneity within individual tumors, consonant with the view that TCCs are monoclonal in origin. The most striking finding was the involvement of chromosome 9 in 92% of the informative cases, as numerical loss of one chromosome copy in 15 cases, but as structural rearrangement in 8. The changes in chromosome 9 always led to loss of material; from 9p, from 9q, or of the entire chromosome. A total of 16 recurrent, unbalanced structural rearrangements were seen, of which del(1)(p11), add(3)(q21), add(5)(q11), del(6)(q13), add(7)(q11), add(11)(p11), i(13)(q10), del(14)(q24), and i(17)(q10) are described here for the first time. The karyotypic imbalances were dominated by losses of the entire or parts of chromosome arms 1p, 9p, 9q, 11p, 13p, and 17p, loss of an entire copy of chromosomes 9, 14, 16, 18, and the Y chromosome, and gains of chromosome arms 1q and 13q and of chromosomes 7 and 20. The chromosome bands and centomeric breakpoints preferentially involved in structural rearrangements were 1q12, 2q11, 5q11, 8q24, 9p13, 9q13, 9q22, 11p11, and 13p10. Rearrangements of 17p and the formation of an i(5)(p10) were associated with more aggressive tumor phenotypes. There was also a general correlation between the tumors' grade/stage and karyotypic complexity, indicating that progressive accumulation of acquired genetic alterations is the driving force behind multistep bladder TCC carcinogenesis.

Cytogenetic analysis of pancreatic carcinomas: intratumor heterogeneity and nonrandom pattern of chromosome aberrations.

Twenty-nine nonendocrine pancreatic carcinomas (20 primary tumors and nine metastases) were studied by chromosome banding after short-term culture. Acquired clonal aberrations were found in 25 tumors and a detailed analysis of these revealed extensive cytogenetic intratumor heterogeneity. Apart from six carcinomas with one clone only, 19 tumors displayed from two to 58 clones, bringing the total number of clones to 230. Karyotypically related clones, signifying evolutionary variation, were found in 16 tumors, whereas unrelated clones were present in nine, the latter finding probably reflecting a distinct pathogenetic mechanism. The cytogenetic profile of pancreatic carcinoma was characterized by multiple numerical and structural changes. In total, more than 500 abnormal chromosomes, including rings, markers, homogeneously stained regions, and double minutes, altogether displaying 608 breakpoints, were detected. This complexity and heterogeneity notwithstanding, a nonrandom karyotypic pattern can be discerned in pancreatic cancer. Chromosomes 1, 3, 6, 7, 8, 11, 12, 17, and 19 and bands 1q12, 1q21, 3q11, 6p21, 6q21, 7q11, 7q22, 7q32, 11q13, 13cen, 14cen, 17q11, 17q21, and 19q13 were most frequently involved in structural rearrangements. A total of 19 recurrent unbalanced structural changes were identified, 11 of which were not reported previously: del(1)(q11), del(3)(p11), i(3)(q10), del(4)(q25), del(11)(p13), dup(11)(q13q23), i(12)(p10), der(13;15)(q10;q10), del(18)(q12), del(18)(q21), and i(19)(q10). The main karyotypic imbalances were entire-copy losses of chromosomes 18, Y, and 21, gains of chromosomes 7, 2, and 20, partial or whole-arm losses of 1p, 3p, 6q, 8p, 9p, 15q, 17p, 18q, 19p, and 20p, and partial or whole-arm gains of 1q, 3q, 5p, 6p, 7q, 8q, 11q, 12p, 17q, 19q, and 20q. In general, the karyotypic pattern of pancreatic carcinoma fits the multistep carcinogenesis concept. The observed cytogenetic heterogeneity appears to reflect a multitude of interchangeable but oncogenetically equivalent events, and the nonrandomness of the chromosomal alterations underscores the preferential pathways involved in tumor initiation and progression.