Cytogenetic Patterns Research Articles

Intratumoral patterns of clonal evolution in gliomas

Few studies have explored the patterns of clonal evolution in gliomas. Here, we investigate the cytogenetic patterns of intratumoral clonal evolution of gliomas and their impact on tumor histopathology and patient survival. Cytogenetic analysis of 90 gliomas was performed in individual tumor cells (>200 cells/tumor) using multicolor (N = 16 probes) interphase-FISH. Overall, chromosome gains were more frequent than chromosome losses. Gains of chromosome 7 and/or EGFR amplification were detected in 91% of the cases, whereas del(9p21) (77%) and del(10q23) (78%) were the most frequent chromosome losses. Virtually, all cases (99%) showed >or=2 tumor cell clones, with higher numbers among high- versus low-grade gliomas (p = 0.001). Nine different cytogenetic patterns were found in the ancestral tumor clones. In most gliomas, ancestral clones showed abnormalities of chromosome 7, 9p, and/or 10q and cytogenetic evolution consisted of acquisition of additional abnormalities followed by tetraploidization. Conversely, early tetraploidization was associated with low-grade astrocytomas-2/3 pilocytic and 3/6 grade II diffuse astrocytomas-and combined loss of 1p36/19q13 with oligodendrogliomas, respectively; both aberrations were associated with a better patient outcome (p = 0.03). Overall, our results support the existence of different pathways of intratumoral evolution in gliomas.

Cytogenetic profile of de novo acute myeloid leukemia: a study based on 1432 patients in a single institution of China

Geographic heterogeneity of cytogenetic patterns in hematological malignancies has been reported earlier, but few systematic studies of cytogenetic abnormalities in acute myeloid leukemia (AML) patients are available. We examined the karyotypic patterns in 1432 de novo AML patients from a single center in China and compared our data with reports from other regions of the world. Chromosomal abnormalities were detected in 746 (58%) cases. The most frequent cytogenetic abnormality was t(15;17), detected in 14% of successful cases, followed by t(8;21) in 8%, and t(9;22), +21 and +8 each in 2%. The mean age of patients with a translocation karyotype was significantly younger than that of patients with normal, deletion or trisomy karyotypes. A higher incidence of AML M3 and a lower frequency of M4 were observed in the Asian population and the frequencies of certain cytogenetic aberrations were different from those in the earlier reports. Population-based age-specific incidences of AML were calculated and compared with those in western reports.

Recovery of Normal Cytogenetic Records in Mussels After Cessation of Pollutant Effluents in Puerto Madryn (Patagonia, Argentina)

The capacity of coast recovering after interruption of pollution sources has been explored employing mussels as biomarkers. In an area polluted by sewage sludge in Puerto Madryn (Argentina), abnormally high cytogenetic records (micronuclei) had been detected in the mussel Mytilus edulis, even higher than those obtained in this and other mussel species (Brachydontes rodriguezi, Aulacomya atra atra, Perumytilus purpuratus) sampled from heavily polluted industrial areas, and much higher than those recorded in samples from unpolluted areas of the same region. Normal cytogenetic patterns were recovered in Puerto Madryn less than 1 year after cessation of sewage sludge discharges, without additional treatment of the affected area. This discovery opens the possibility of considering restored coastal areas for aquaculture purposes instead of endangering natural populations in virgin areas.

Biogeography and comparative cytogenetics between two populations of Hoplias malabaricus (Bloch, 1794) (Ostariophysi: Erythrinidae) from coastal basins in the State of Bahia, Brazil

The species Hoplias malabaricus is a predator fish found in nearly all cis-Andean basins. From a cytogenetic point of view, this species comprises, at least, seven differentiated karyomorphs. Several localities have been formerly analyzed in Brazil, however, some regions, such as Bahia State, remain underrepresented. Recently, the Brazilian Environment Ministry classified both Itapicuru and Contas river basins (entirely located within Bahia territory) as priority conservation areas, whose biodiversity status lacks enough information. Therefore, the goal of the present work was to characterize, cytogenetically, populations of H. malabaricus from both basins, by using conventional staining, Ag-NOR and C-banding techniques. All specimens presented a diploid number of 2n = 40 with metacentric/submetacentric chromosomes, without differences between sexes, thereby representing the so-called "karyomorph F". The first metacentric pair presented a remarkably larger size in relation to the other pairs. The NORs were multiple, comprising the terminal region on long arms of two chromosomal pairs in both populations. However, the C-banding pattern was somewhat distinguishable between samples. Although sharing heterochromatic blocks at centromeric region of all chromosomes, the population from Itapicuru River basin appeared to have some more conspicuous blocks than those observed in the population from Contas River basin. The similar karyotype observed in both populations suggests a common geological history between them. The present results represent an advance in the knowledge about the cytogenetic pattern of H. malabaricus populations from poorly studied basins.

Renal Cell Carcinomas With Papillary Architecture and Clear Cell Components

Although histologic features enable an accurate diagnosis in most renal carcinomas, overlapping morphologic findings between some renal neoplasms make subclassification difficult. Some renal carcinomas show papillary architecture but are composed extensively of cells with clear cytoplasm, and it is unclear whether they should be classified as clear cell renal cell carcinomas or papillary renal cell carcinomas. We analyzed the immunohistochemical profiles and the cytogenetic patterns of 14 renal carcinomas showing papillary architecture in which there were variable amounts of cells with clear cytoplasm. The patients were 8 women and 6 men (mean age: 54 y). Immunohistochemistry and fluorescence in situ hybridization analysis distinguished 2 different groups. The first consisted of 10 renal cell carcinomas with strong immunoreactivity for alpha-methyl coenzyme A racemase, of which 9 also expressed cytokeratin 7. All of these neoplasms showed gains of chromosome 7 or 17 and chromosome Y was lost in all the male patients whereas 3p deletion was detected only in one case. In the other 4 renal cell carcinomas, cytokeratin 7 was not detected and alpha-methylacyl-CoA racemase was positive in only 1. In these neoplasms, no gain of chromosome 7 or 17 and no loss of chromosome Y were observed, whereas 3p deletion was detected in 3 of them. None of the 14 neoplasms showed immunoreactivity for TFE3. The combined use of immunohistochemistry and cytogenetics enabled us to provide a definitive diagnosis for 12 of 14 renal cell carcinomas with papillary architecture and clear cell components: 9 cases were confirmed to be papillary renal cell carcinomas and 3 cases were confirmed to be clear cell renal cell carcinomas. Despite these ancillary techniques, 2 cases remained unclassified. Our study establishes the utility of these procedures in accurately classifying the great majority of renal cell carcinomas with these findings.

Clofarabine (Clo) +/− Fludarabine (Flu) with IV Busulfan (Bu) and Allogeneic Stem Cell Transplantation (Allo-SCT) for Relapsed, Refractory Myeloid Leukemia (ML) and MDS

Fludarabine (Flu) combined with IV Busulfan (Bu) is a safe, reduced toxicity pretransplant conditioning program. To enhance the cytoreductive effect of the conditioning regimen, we decided to replace Flu with Clofarabine (Clo), a (superior) second generation nucleoside analog/antileukemic agent, whose immunosuppressive capability in support of allogeneic progenitor cell engraftment is unknown. In 2006 we initiated a trial where Clo would gradually replace Flu combined with IV Bu in a myeloablative program, aimed atretaining the safety profile, whileevaluating the engraftment-promoting capacity, and hopefullyimproving the antileukemic activity.METHODS and PATIENTS: Patients were enrolled/randomized as follows: Arm I: Clo:Flu 10:30 mg/m2, Arm II 20:20 mg/m2, Arm III 30:10 mg/m2, and Arm IV single-agent Clo at 40 mg/m2, the nucleoside analog(s) were/was infused over 60 min each once daily for 4 days on each day followed by Bu infused over 3 hours to an average daily AUC of 6,000 mcMol-min +/− 10%. So far, 20 patients have been enrolled, 12 males and 8 females, with a median age of 43 years (range: 6–57). Seven patients had CML (BC: 1, first AP: 3, second AP: 2, and late first CP: 1). Thirteen patients had AML: 5 with induction failure, 3 were in chemotherapy-refractory relapse, 3 in untreated relapse, 1 had a second PR, and, finally 1 was in a high-risk CR1 [cytogenetics of −7 and t(3;12)]. Of the 13 AML patients 1 had a favorable cytogenetic (CG) pattern, 6 were intermediate, and 4 had poor prognosis CG, while 2 had unknown CG data. GVHD-prophylaxis was based on tacrolimus and mini-MTX, and those who had an unrelated or one antig-mism. related donor received rabbit-ATG (Thymoglobulin™).RESULTS: One patient died of pneumonia, there was no other treatment-related death in the first 100 days, the main toxicity was mucositis grade 2–3 in about half the patients. Specifically there was no significant hepatic or neurologic toxicity. All 19 evaluable patients engrafted uneventfully (one had recurrent AML before day +30). Marrow and blood (T-cell) chimerism studies at day +30 revealed that groups I+II (lower Clo doses; n=9) had a median of 87% (range 17–100) donor (T-cell)-derived DNA, groups III+IV (higher Clo-doses, n=11) had a median of 100% range (64–100) donor-DNA. By day +100 both cohorts had a median of 100% donor-derived DNA, maintained beyond 6 months in all evaluable patients. 16 of the 19 patients with active disease at SCT entered CR, and 11 patients remain in clinical CR (August 2008) with a median duration of 8 mos (range, 2–19). One previously TKI-resistant CML patient (SCT in second AP) progressed by CG at 4 mos, but responded to dasatinib, with a maintained CG CR exceeding 22 mos. Median disease-free and overall survival has not been reached with 11 patients in unmaintained CR, and 12/20 being alive. The projected overall and disease-free survival at one year of 70% and 55%, respectively.CONCLUSION: We conclude that a Clo-Bu-based regimen appears safe in (relapsed, chemotherapy-refractory) high-risk patients, 2) there should be little or no concern about the immunosuppressive capability of Clo in support of allo-SCT in myeloid leukemia, and 3) (an) additional study(-ies) is/(are) warranted to fully evaluate the antileukemic efficacy of Clo combined with IV Bu + ATG as pretransplant conditioning therapy for myeloid leukemia and MDS.

Molecular Phylogeny of the Neotropical Genus Christensonella (Orchidaceae, Maxillariinae): Species Delimitation and Insights into Chromosome Evolution

Species' boundaries applied within Christensonella have varied due to the continuous pattern of variation and mosaic distribution of diagnostic characters. The main goals of this study were to revise the species' delimitation and propose a more stable classification for this genus. In order to achieve these aims phylogenetic relationships were inferred using DNA sequence data and cytological diversity within Christensonella was examined based on chromosome counts and heterochromatin patterns. The results presented describe sets of diagnostic morphological characters that can be used for species' identification. Phylogenetic studies were based on sequence data of nuclear and plastid regions, analysed using maximum parsimony and maximum likelihood criteria. Cytogenetic observations of mitotic cells were conducted using CMA and DAPI fluorochromes. Six of 21 currently accepted species were recovered. The results also support recognition of the 'C. pumila' clade as a single species. Molecular phylogenetic relationships within the 'C. acicularis-C. madida' and 'C. ferdinandiana-C. neowiedii' species' complexes were not resolved and require further study. Deeper relationships were incongruent between plastid and nuclear trees, but with no strong bootstrap support for either, except for the position of C. vernicosa. Cytogenetic data indicated chromosome numbers of 2n = 36, 38 and 76, and with substantial variation in the presence and location of CMA/DAPI heterochromatin bands. The recognition of ten species of Christensonella is proposed according to the molecular and cytogenetic patterns observed. In addition, diagnostic morphological characters are presented for each recognized species. Banding patterns and chromosome counts suggest the occurrence of centric fusion/fission events, especially for C. ferdinandiana. The results suggest that 2n = 36 karyotypes evolved from 2n = 38 through descendent dysploidy. Patterns of heterochromatin distribution and other karyotypic data proved to be a valuable source of information to understand evolutionary patterns within Maxillariinae orchids.

Treatment of high-risk MDS patients (pts) with -7/del(7q) with azacitidine (AZA) versus conventional care regimens (CCR): Effects on overall survival (OS)

7033 Background: -7/del(7q) is associated with poor prognosis in MDS. In a recent phase III, randomized trial, AZA significantly prolonged OS in pts with high-risk MDS (Fenaux, Blood 2007;110,817). This analysis assessed the effect of AZA on OS in this subgroup of high- risk MDS pts with -7/del(7q). Methods: Primary inclusion criterion for the Phase III study was high-risk MDS (FAB RAEB, RAEB-T, or CMMoL and IPSS Int-2 or High). Pts were randomized to AZA (75 mg/m2/d x 7d, q28d) or CCR. CCR comprised 3 treatments: BSC only (transfusions, antibiotics, G-CSF for neutropenic infection); low-dose ara-C (20 mg/m2/d x 14d, q28d); or induction chemotherapy (7+3 regimen). No erythropoietin was allowed. Results: At baseline (BL), 57 (30 AZA, 27 CCR) of 358 pts in the total population had -7/del(7q), 35% had -7/del(7q) alone and 65% had -7del(7q) as part of complex karyotype. BL characteristics were balanced in the 2 arms: 70% male; median age, 69 years. The median Kaplan Meier difference in OS for AZA vs CCR was 8.4 months, a significant improvement (3-fold) over CCR (Table). The hazard ratio (HR) was 0.33 (95% CI: 0.16–0.68) indicating a 67% reduced risk of death in the AZA arm; this compares with an HR of 0.58 for the OS improvement with AZA vs CCR with all cytogenetic subtypes in the phase III trial. At 2 years, a 4-fold OS advantage was observed in the AZA arm with 33% of pts alive vs 8% in the CCR arm (p=0.03). Secondary endpoints support the OS advantage (Table). Significantly higher IWG 2000 response rates (CR+PR) were seen in pts with -7/del(7q) alone (64% vs 11%; p=0.03) or complex (21% vs 0; p=0.02) with AZA vs CCR, and compared favorably to the overall AZA group with IPSS good and intermediate cytogenetics. AZA was generally well tolerated. Conclusion: This phase III subgroup analysis indicates the disease modifying effect of AZA extends to unfavorable cytogenetic patterns that include -7/del(7q), and suggests AZA may represent the treatment of choice for this otherwise poor prognosis subset. OS, CR, PR, HI, and transfusion independence (TI) in RBC-dependent -7/del(7q) pts at BL (n/N,%) OS (mos) CR+PR CR RBC TI HI-E HI-P AZA 13.1 13/30, 43 8/30, 27 12/21, 57 13/26, 50 10/20, 50 CCR 4.6 1/27, 4 1/27, 4 0/19, 0 0/24, 0 2/25, 8 P value 0.003 0.0005 0.03 <0.0001 <0.0001 0.002 Author Disclosure Employment or Leadership Consultant or Advisory Role Stock Ownership Honoraria Research Expert Testimony Other Remuneration Pharmion Amgen, Celgene, Pharmion Corporation Pharmion Amgen, Celgene, Cephalon, Johnson & Johnson, Novartis, Pharmion, Roche Pharmion Corporation

Risk Estimation of Neuroblastoma Patients Using Molecular Markers

The pediatric tumor neuroblastoma is a heterogeneous disease: Patients' clinical courses can range from spontaneous regression to fatal progression of the disease. Accordingly, treatment protocols vary from "wait and see" approaches to intensive multimodal therapies. Accurate risk estimation of the patients is therefore mandatory to choose the most adequate therapy. Current trials stratify by a limited number of clinical variables, such as stage of the disease and age of the patient at diagnosis, as well as molecular markers, such as amplification of the oncogene MYCN and loss of the short arm of chromosome 1. However, misclassifications of patients still occur, and thus, a precise prediction of the clinical courses remains a challenge of neuroblastoma research. In recent years, genomic alterations and gene expression profiles of this neoplasm have been characterized thoroughly. It has been shown that the diverse clinical phenotypes are reflected by both specific cytogenetic aberrations and distinct gene expression patterns. Moreover, a variety of DNA copy number changes and gene expression-based classifiers have been described that could predict the outcome of neuroblastoma patients more precisely than established prognostic variables. In this review, the recent advances in the detection and evaluation of molecular prognostic markers for neuroblastoma patients are summarized, and their current and potential contribution to risk stratification systems is discussed.

Cytogenetic analyses of two endemic fish species from the São Francisco River basin: Conorhynchus conirostris and Lophiosilurus alexandri (Siluriformes)

Two Siluriformes species endemic to the Sao Francisco River basin were characterized by conventional and differential cytogenetic analyses involving C-banding, Ag-nucleolar organizer region (NOR) and chromomycin A3 (CMA3) staining, and FISH (fluorescent in situ hybridization) with 18S and 5S rDNA probes. Conorhynchus conirostris presents a higher diploid number (2n = 60) than those detected in Pimelodidae representatives, whereas Lophiosilurus alexandri, with a karyotype of 2n = 54 chromosomes, presents a chromosomal constitution similar to that found in the family Pseudopimelodidae. Plesiomorphic characteristics such as single NORs at terminal positions are found in both species, as revealed by CMA3 and silver nitrate staining, and FISH with a 18S rDNA probe. C-banding evidenced centromeric and telomeric heterochromatic blocks distributed over most of the chromosomes with a conspicuous heterochromatin segment in a pair of submetacentric chromosomes in L. alexandri. Such karyotype data, if compared to the cytogenetic pattern of other Siluriformes species, can be partially related to their degree of endemism, favorable to the occurrence and fixation of chromosomal rearrangements. The present study in representatives from these two Siluriformes families from the Sao Francisco River contributes to a better understanding of the karyotype evolution in species of this important order of Neotropical fishes.

Chromosomal Abnormalities in Saudi Children of Jeddah City

It is attempted to estimate the incidence of chromosomal disorders, constitutional karyotype and cytogenetic patterns of chromosomal abnormalities in the Western Saudi region. Further, associations between socio-demographic, obstetrical, medical and environmental factors were also attempted. Overall incidence of chromosomal disorders was estimated at 2.8 and 2.4 for Down syndrome per thousand Saudi live births. Cytogenetic analysis revealed preponderance of numerical (92%) aberrations than structural (8%). Down syndrome was the most frequent form of the chromosomal disorders; in addition, trisomies of chromosome 13 and 18 and monosomy of X was found in lesser frequencies. A rare form of chromosomal configuration with translocation of chromosome 7 and 10, extra marker chromosome and pericentric inversion in chromosome 3 were also observed. Trisomy 21 was recorded to have high frequency (94%); other forms like Robertsonian translocation and mosaicism were also detected in lesser frequency. A significant association between advanced mother ages was confirmed, other factors have no bearing. Findings are: The estimated incidence of overall chromosomal disorders is lower, but incidence of Down syndrome is higher compared to national and international frequencies. Down syndrome is the most frequently encountered disorder and karyotypic techniques effectively help diagnosing in most of the cases.

Cytogenetic patterns in multiple myeloma after a phase of preceding MGUS

Presenting the same histological diagnosis, multiple myeloma (MM) shows a large genomic variety, resulting in variable times of overall survival. To investigate major cytogenetic categories (any 14q-translocation, t(11;14), t(4;14), 13q-deletions, 17p-deletions) and their clinical consequences in MM after a pre-existing monoclonal gammopathy (MM post-MGUS), we performed a comparative analysis of 41 patients with MM post-MGUS and 287 patients with unknown prior history MM (U-MM). In MM post-MGUS, a t(11;14) was found to be more frequent than in U-MM (24% vs. 14%) and it was associated with significantly shortened survival (24 months vs. 70 months in U-MM; P = 0.01). MM post-MGUS was further characterized by a higher frequency of 13q-deletions only (absence of all other specific abnormalities; 28% vs. 12% in U-MM; P = 0.02). A 13q-deletion only was an indicator of long survival in MM post-MGUS (median not yet reached) as opposed to U-MM (median survival, 29 months; P = 0.001). 17p-deletions were infrequent in MM post-MGUS (3% vs. 16% in U-MM; P = 0.04). Survival times for patients with t(4;14) and/or 17p-deletions and other abnormalities were similar in both MM patient cohorts. Our data suggest that t(11;14) and 13q-deletions have distinct prognostic implications in the context of MM post-MGUS.

Evaluation of the Cytogenetic Aberration Pattern in AL Amyloidosis Compared to Monoclonal Gammopathies Not Requiring Treatment: Translocation t(11;14) Is More Frequent in AL Amyloidosis.

Introduction: Cytogenetic aberrations (CA) have emerged as important pathogenetic and prognostic factors in plasma cell disorders. However, in AL amyloidosis (AL) only a few reports with small numbers of patients have been published.Methods: Using interphase FISH analysis in CD138+ cells, we evaluated the role of CA in a series of 85 AL patients as compared to 146 patients with a monoclonal gammopathy without treatment requirement in a prospective manner. Our panel included IgH translocations t(11;14), t(4;14), t(14;16) and translocation of 14q32 with an unidentified partner, gains of 1q21, 11q23 and 19q13 as well as deletions of 8p21, 13q14 and 17p13. Using these probes we could detect at least one of these aberrations in 95% of AL and in 88% of the control group. Age, gender and plasma cell content were statistically equally distributed among both groups.Results: The most frequent aberration in AL was t(11;14), which was detected in 45% of AL patients as compared to 26% of the control group (p=0,056). It was strongly associated with the lack of an intact immunoglobulin (p<0,001), thus accounting for the frequent light chain only subtype in AL. Markedly, t(11;14) was more frequently found in combination with gain 11q23 in AL than in the control group (20% versus 6%, p = 0.005). Other frequent aberrations in AL included deletion 13q14 (32%) and gain 1q21 (21%), which were observed in the control group at comparable frequencies (34% and 20%). The overlapping character of the underlying plasma cell disorder in both disease entities was also emphasized by the similarities of branching patterns of the five major CA in cluster analysis applied in 169 patients (figure 1). The relation of clinical parameters and chromosomal aberrations was also evaluated. The analyzed CA had no impact on the organ involvement pattern in AL patients.Conclusions: We observed a high frequency for t(11;14) in AL. Apart from this finding, the cytogenetic patterns known in monoclonal gammopathy of unknown significance and multiple myeloma were widely shared by AL amyloidosis. [Display omitted]

Clinical characteristics and course of chronic lymphocytic leukemia patients with the combination favorable and unfavorable cytogenetics

8086 Background: Patients with CLL have a highly variable clinical course. Genomic aberrations detected by FISH have been shown to correlate with survival and treatment free interval (TFI). Patients with the presence of 17p or 11q deletions (del) either alone or in combination with other cytogenetic abnormalities have the worst prognosis while patients with 13q del as a sole abnormality have the best prognosis. Our objective was to further investigate poor prognosis CLL patients with either del 17p or 11q to determine if the addition of the favorable 13q del influences the predicted clinical course and survival. Methods: We performed a retrospective chart review on 22 patients (pts) who had been identified by FISH as having either the combination of del 17p and 13q or del 11q and 13q. Results: 128 CLL FISH panels were performed from April of 2003 through October of 2006. Twenty-two pts (17%) had either del 17p and 13q (9%) or del 11q and 13q (9%). Historical data notes a frequency of 7% and 8% for deletions 17p and 11q, respectively, and 55% for 13q as a sole aberration. The median age was 66 yrs, the majority of whom were male (73%). Two of 22 pts (9%) presented with advanced stage disease. Splenomegaly was seen more often in the 17p/13q pts (36%) vs 11q/13q pts (9%). With a median follow up of 46 months since diagnosis, the median TFI for all patients (20 known) was 56 months. TFI was 13 months for patients with 17p/13q del; whereas TFI was not reached for patients with 11q/13q. Historical data noted a TFI for patients with deletions 17p, 11q, and 13q (as a sole aberration) as 9, 13, and 92 months respectively. The median survival from diagnosis was not reached for the group overall or for either combination of genetic abnormalities. Historical data noted, with 70 months follow up, a median survival of 108 months overall and 32, 79, and 133 months for 17p, 11q, and 13q (as a sole aberration) respectively. Conclusions: The addition of favorable cytogenetics, del 13q, in a CLL patient with an unfavorable cytogenetic pattern (either del 17p or del 11q) appears to improve the predicted clinical outcome and survival. Additional follow up and prospective studies are needed to further define which genetic subgroups help to prognosticate CLL patients. No significant financial relationships to disclose.

C-Kit/PDGFRA Gene Status Alterations Possibly Related to Primary Imatinib Resistance in Gastrointestinal Stromal Tumors

To correlate morphologic changes with molecular, biochemical, and cytogenetic profiles in gastrointestinal stromal tumor (GIST) patients before and after imatinib treatment. We investigated 132 tumor samples obtained from 35 patients with advanced disease who underwent resective surgery after imatinib treatment according to the European Organization for Research and Treatment of Cancer-Soft Tissue and Bone Sarcoma Group protocol. On the basis of imaging findings, 27 patients were responders and 8 progressors, and retaining this radiological subdivision, we analyzed posttreatment morphologic changes correlating them with molecular, biochemical, and cytogenetic analyses. On the basis of morphology (residual viable cellularity/proliferation markers), three subgroups were identified showing high, moderate, or low response. All of the progressing cases clustered in the low-response subgroup, whereas the responding cases were distributed in all three subgroups. The correlation between morphology and the molecular findings showed that secondary mutations segregated with the low-response subgroup, whereas c-Kit primary resistance mutations were randomly distributed in the three subgroups. Fluorescence in situ hybridization analysis of c-Kit/PDGFRA genes showed that all of the progressing cases were disomic. Referring to morphology, among the responding cases, a disomic pattern was mainly restricted to the high responders, whereas the moderate and low responders were aneusomic. Comparison of post-imatinib genomic profiles with the 23 available primary tumors showed that 17 cases carried the same cytogenetic pattern. Overall, 12 of the 27 primary tumors presented a gain/loss of c-Kit/PDGFRA gene copy number. Our findings show that c-Kit/PDGFRA genomic alterations were present at disease onset in 1/3 of the examined cases. They therefore represent an early event possibly related to primary imatinib resistance in GISTs.

World Health Organization classification in combination with cytogenetic markers improves the prognostic stratification of patients with de novo primary myelodysplastic syndromes

This study correlated chromosomal defects with French-American-British (FAB)/World Health Organization (WHO) classification subtypes, proposed a revised International Prognostic Scoring System (IPSS) cytogenetic grouping; and established which classification, when used with the IPSS cytogenetic categories, best predicted clinical outcome in the myelodysplastic syndromes (MDS). A higher prevalence of chromosomal defects and distinct defects were observed in patients with multi-lineage dysplasia and a blast cell percentage >10%. Abnormalities of the long arm of chromosome 3, del(7)(q31q35), trisomy 8, del(11)(q14q23), del(12p) and 20q- could be segregated from their respective IPSS cytogenetic categories and used to develop new cytogenetic subgroups. Clinical parameters, FAB/WHO classification, IPSS score and standard or revised cytogenetic categories were statistically relevant for overall survival (OS) and progression-free intervals (PFI) and were included within five distinct multivariate models compared by the Akaike Information Criterion. To predict OS, the best models included age, WHO classification and standard or revised IPSS cytogenetic categories; to predict PFI, the best model included the same variables and revised cytogenetic categories. In conclusion, (i) the WHO classification was associated with a more homogeneous cytogenetic pattern than the FAB classification, (ii) WHO classification and standard/revised IPSS cytogenetic categories were much more effective than IPSS for predicting MDS clinical outcome, (iii) revised cytogenetic subgroups predicted PFI more effectively than standard categories.

Cytogenetic patterns in ETV6/RUNX1‐positive pediatric B‐cell precursor acute lymphoblastic leukemia: A Nordic series of 245 cases and review of the literature

Between 1992 and 2004, 1,140 children (1 to<15 years) were diagnosed with B-cell precursor acute lymphoblastic leukemia (ALL) in the Nordic countries. Of these, 288 (25%) were positive for t(12;21)(p13;q22) [ETV6/RUNX1]. G-banding analyses were successful in 245 (85%); 43 (15%) were karyotypic failures. The modal chromosome numbers, incidence, types, and numbers of additional abnormalities, genomic imbalances, and chromosomal breakpoints in the 245 karyotypically informative cases, as well as in 152 previously reported cytogenetically characterized t(12;21)-positive ALLs in the same age group, were ascertained. The most common modal numbers among the 397 cases were 46 (67%), 47 (16%), 48 (6%), and 45 (5%). High-hyperdiploidy, triploidy, and tetraploidy were each found in approximately 1%; none had less than 40 chromosomes. Secondary chromosomal abnormalities were identified by chromosome banding in 248 (62%) of the 397 ALLs. Of these, 172 (69%) displayed only unbalanced changes, 14 (6%) only balanced aberrations, and 26 (10%) harbored both unbalanced and balanced abnormalities; 36 (15%) were uninformative because of incomplete karyotypes. The numbers of secondary changes varied between 1 and 19, with a median of 2 additional aberrations per cytogenetically abnormal case. The most frequent genomic imbalances were deletions of 6q21-27 (18%), 8p11-23 (6%), 9p13-24 (7%), 11q23-25 (6%), 12p11-13 (27%), 13q14-34 (7%), loss of the X chromosome (8%), and gains of 10 (9%), 16 (6%), and 21 (29%); no frequent partial gains were noted. The chromosome bands most often involved in structural rearrangements were 3p21 (2%), 5q13 (2%), 6q12 (2%), 6q14 (2%), 6q16 (2%), 6q21 (10%), 6q23 (6%), 6q25 (3%), 9p13 (2%), 11q13 (2%), 11q23 (2%), 12p11 (6%), 12p12 (7%), 12p13 (25%), 21q10 (6%), and 21q22 (6%). Considering that the t(12;21) is known to arise in utero and that the postnatal latency period is protracted, additional mutations are most likely necessary for overt ALL. The frequently rearranged chromosome regions may harbor genes of importance for the transformation and/or progression of an initial preleukemic t(12;21)-positive clone.

Genetic Pathways in the Pathogenesis of Therapy-Related Myelodysplasia and Acute Myeloid Leukemia

In therapy-related myelodysplasia (t-MDS) and acute myeloid leukemia (t-AML), at least eight alternative genetic pathways have been defined based on characteristic recurrent chromosome abnormalities. Patients presenting as t-MDS and patients presenting as overt t-AML cluster differently in these pathways. The cytogenetic pattern depends on the type of leukemogenic therapy received: alkylating agents, topoisomerase II inhibitors, or radiotherapy. Three types of gene mutations are observed in MDS and AML: (1) Activating mutations of genes in the tyrosine kinase-RAS/BRAF signal transduction pathway, leading to increased cell proliferation (Class I mutations); (2) Inactivating mutations of genes encoding hematopoietic transcription factors, resulting in disturbed cell differentiation (Class II mutations); and (3) Inactivating mutations of the tumor suppressor gene p53. At least 14 different genes have been identified as mutated in t-MDS and t-AML, clustering differently and characteristically in the eight genetic pathways. Class I and Class II mutations are significantly associated, indicating their cooperation in leukemogenesis The chromosome aberrations and gene mutations detected in the therapy-related and in the de novo subsets of MDS and AML are identical, although the frequencies with which they are observed may differ. Hence, therapy-related and de novo MDS and AML are identical diseases and should be subclassified and treated similarly.

Chromosomal abnormalities of young multiple myeloma patients (&lt;45 yr) are not different from those of other age groups and are independent of stage according to the International Staging System

Little is known about tumor-related prognostic factors, in particular specific chromosomal abnormalities, in young patients with multiple myeloma (MM). We therefore investigated the chromosomal pattern by interphase fluorescence in situ hybridization (chromosomes 13q14, 14q32-translocations, chromosomes associated with hyperdiploidy) in 38 young patients with MM (age <45 yr) and compared the results with those observed in 69 patients with intermediate age (45-70 yr) and 64 elderly patients (age >70 yr). All chromosomal patterns were not significantly different between the three age cohorts. Similarly, standard MM parameters were equally distributed between these MM patient populations. However, survival by the International Staging System (ISS) for MM revealed marked differences between stage I/II (median survival not yet reached) and stage III (23.4 months; P < 0.0003) among young MM patients. A significant survival difference between ISS-stage I/II and ISS-stage III patients was also noted in the intermediate age group (median 65.4 months vs. 24.6 months; P = 0.0009). However, this difference disappeared among elderly MM patients (39.6 months in ISS-stage I/II vs. 32 months in ISS-stage III patients; P = 0.94), but it was unrelated to the cytogenetic pattern. Our results indicate that MM in young patients does not represent a distinct biologic entity, and that short survival of younger MM patients at ISS-stage III is independent of the molecular cytogenetic pattern.

Cytogenetic analysis of Anaecypris hispanica and its relationship with the paternal ancestor of the diploid-polyploid Squalius alburnoides complex

The karyotype of the endangered fish Anaecypris hispanica was revisited using advanced cytogenetic techniques to elucidate its putative relationship with the paternal ancestor of the hybrid complex Squalius alburnoides and to clarify some of the recently described cytogenetic patterns of the complex. The results of chromomycin A3 and Ag staining, as well as fluorescent in situ hybridization with 28S and 5S rDNA and the (TTAGGG)n telomeric probes, were compared with the patterns observed in specimens of the all-male nonhybrid lineage of S. alburnoides complex, which is considered to reconstitute the nuclear genome of the probably extinct paternal ancestor. Several cytogenetic features observed in A. hispanica specimens were indeed shared by S. alburnoides nuclear nonhybrid males, supporting the hypothesis of a close evolutionary link between A. hispanica and the paternal ancestor of the complex. The genomic rearrangements involving 28S rDNA sites previously described in the S. alburnoides complex and in its maternal ancestor (S. pyrenaicus) were not detected in A. hispanica; they are, therefore, probably due to mechanisms related to hybridization and polyploidy.