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Chromosoma, 115, 459- 467

The breakage-fusion-bridge (BFB) cycle as a mechanism for generating genetic heterogeneity in osteosarcoma.

S. Selvarajah, M. Yoshimoto, P.C. Park, G. Maire, J. Paderova, J. Bayani, G. Lim, K. Al-Romaih, J.A. Squire, M. Zielenska

Osteosarcoma (OS) is characterized by chromosomal instability and high copy number gene amplification. The breakage–fusion–bridge (BFB) cycle is a well-established mechanism of genome instability in tumors and in vitro models used to study the origins of complex chromosomal rearrangements and cancer genome amplification. To determine whether the BFB cycle could be increasing the de novo rate of formation of cytogenetic aberrations in OS, the frequency of anaphase bridge configurations and dicentric chromosomes in four OS cell lines was quantified. An increased level of anaphase bridges and dicentrics was observed in all the OS cell lines. There was also a strong association between the frequencies of anaphase bridges, dicentrics, centrosomal anomalies, and multipolar mitotic figures in all the OS cell lines, indicating a possible link in the mechanisms that led to the structural and numerical instabilities observed in OS. In summary, this study has provided strong support for the role of the BFB cycle in generating the extensive structural chromosome aberrations, as well as cell-to-cell cytogenetic variation observed in OS, thus conferring the genetic diversity for OS tumor progression.

Int J Med Sci, 3, 124- 129

Low temperature tolerance of human embryonic stem cells.

B.C. Heng, K.J. Vinoth, H. Liu, M.P. Hande, T. Cao

This study investigated the effects of exposing human embryonic stem cells (hESC) to 4oC and 25oC for extended durations of 24h and 48h respectively. Cell survivability after low temperature exposure was assessed through the MTT assay. The results showed that hESC survivability after exposure to 25oC and 4oC for 24h was 77.3 ± 4.8 % and 64.4 ± 4.4 % respectively (significantly different, P < 0.05). The corresponding survival rates after 48h exposure to 25oC and 4oC was 71.0 ± 0.5 % and 69.0 ± 2.3 % respectively (not significantly different, P > 0.05). Spontaneous differentiation of hESC after low temperature exposure was assessed by morphological observations under bright-field and phase-contrast microscopy, and by immunocytochemical staining for the pluripotency markers SSEA-3 and TRA-1-81. hESC colonies were assigned into 3 grades according to their degree of spontaneous differentiation: (1) Grade A which was completely or mostly undifferentiated, (2) Grade B which was partially differentiated, and (3) Grade C which was mostly differentiated. In all low temperature exposed groups, about 95% of colonies remain undifferentiated (Grade A), which was not significantly different (P > 0.05) from the unexposed control group maintained at 37oC. Additionally, normal karyotype was maintained in all low temperature-exposed groups, as assessed by fluorescence in situ hybridization (FISH) of metaphase spreads with telomere and centromere-specific PNA probes. Further analysis with m-FISH showed that chromosomal translocations were absent in all experimental groups. Hence, hESC possess relatively high-tolerance to extended durations of low temperature exposure, which could have useful implications for the salvage of hESC culture during infrequent occurrences of incubator break-down and power failure.

Pharmacogen Genom, 16, 87- 99

Cytogenmetic biomarkers, urinary metabolites and metabolic gene polymorphisms in workers exposed to styrene.

L. Migliore, A. Naccarati, F. Coppedè, E. Bergamaschi, G. De Palma, A. Voho, P. Manini, H. Järventaus, A. Mutti, H. Norppa, A. Hirvonen

The present study comprised a biomonitoring study in 95 workers occupationally exposed to styrene and 98 unexposed controls, employing an integrated approach involving biomarkers of exposure, effect, and susceptibility. Airborne styrene was evaluated at workplace, and urinary styrene metabolites, mandelic acid (MA), phenylglyoxylic acid (PGA), vinylphenols (VPTs) and phenylhydroxyethylmercapturic acids (PHEMAs), were measured as biomarkers of internal dose. Cytogenetic alterations were evaluated by analysing the frequency of chromosomal aberrations (CAs) and micronucleated binucleated cells (MNBN) in peripheral blood lymphocytes. The micronucleus assay was coupled with centromeric fluorescence in situ hybridization to distinguish micronuclei (MN) arising from chromosomal breakage (C- MN) from those harboring whole chromosomes (C+ MN). The possible influence of genetic polymorphisms of xenobiotic-metabolizing enzymes involved in styrene biotransformation (EPHX1, GSTT1, GSTM1, GSTP1) and NAT2 on the cytogenetic endpoints was investigated. The exposed workers showed a significantly higher frequency of MNBN (13.8+/-0.5% versus 9.2+/-0.4%; P<0.001) compared to control subjects. The effect appeared to concern both C- and C+ MN. A positive correlation was seen between the frequency of C+ MN and urinary level of MA+PGA (P<0.05) and VPTs (P<0.001). Chromosome-type CAs positively correlated with airborne styrene level and VPTs (P<0.05), whereas chromatid-type CAs correlated with PHEMAs (P<0.05). Workers bearing GSTM1 null genotype showed lowered levels of PHEMAs (P<0.001). The GSTT1 null genotype was associated with increased MNBN frequencies in the exposed workers (P<0.05) and the fast activity EPHX genotype with a moderate decrease in both MNBN and CAs in the controls. Our results suggest that occupational exposure to styrene has genotoxic effects that are potentiated by the GSTT1 gene deletion. These observations may have relevance considering the risk of lymphatic and haematopoietic malignancies tentatively associated with styrene exposure.

Nucl Acids Res (ePub), 35, 0- 0

Human RAD18 is involved in S phase-specific single-strand break reapir without PCNA monoubiquitination.

N. Shiomi, M. Mori, H. Tsuji, T. Imai, H. Inoue, S. Tateishi, M. Yamaizumi, T. Shiomi

<p>Switching from a replicative to a translesion polymerase is an important step to further continue on replication at the site of DNA lesion. Recently, RAD18 (a ubiquitin ligase) was shown to monoubiquitinate proliferating cell nuclear antigen (PCNA) in cooperation with RAD6 (a ubiquitin-conjugating enzyme) at the replication-stalled sites, causing the polymerase switch. Analyzing RAD18-knockout (RAD18-/-) cells generated from human HCT116 cells, in addition to the polymerase switch, we found a new function of RAD18 for S phase-specific DNA single-strand break repair (SSBR). Unlike the case with polymerase switching, PCNA monoubiquitination was not necessary for the SSBR. When compared with wild-type HCT116 cells, RAD18-/- cells, defective in the repair of X-ray-induced chromosomal aberrations, were significantly hypersensitive to X-ray-irradiation and also to the topoisomerase I inhibitor camptothecin (CPT) capable of inducing single-strand breaks but were not so sensitive to the topoisomerase II inhibitor etoposide capable of inducing double-strand breaks. However, such hypersensitivity to CPT observed with RAD18-/- cells was limited to only the S phase due to the absence of the RAD18 S phase-specific function. Furthermore, the defective SSBR observed in S phase of RAD18-/- cells was also demonstrated by alkaline comet assay.</p>

J Invest Dermatol, 126, 2308- 2315

Are keratoacanthomas variants of squamous cell carcinomas? A comparison of chromosomal aberrations by comparative genomic hybridization.

O.P.F. Clausen, H.C.D. Aass, M. Beigi, K.J. Purdie, C.M. Proby, V.L. Brown, M. Mattingsdahl, F. Micci, S. K\olvraa, L. Bolund, P.M. De Angelis

Keratoacanthoma (KA) is a benign keratinocytic neoplasm that usually presents as a solitary nodule on sunexposed areas, develops within 6-8 weeks and spontaneously regresses after 3-6 months. KAs share features such as infiltration and cytological atypia with squamous cell carcinomas (SCCs). Furthermore, there are reports of KAs that have metastasized, invoking the question of whether or not KA is a variant of SCC. To date no reported criteria are sensitive enough to discriminate reliably between KA and SCC, and consequently there is a clinical need for discriminating markers. We screened fresh frozen material from 132 KAs and 37 SCCs for gross chromosomal aberrations by using comparative genomic hybridization (CGH). Forty-nine KAs (37.1%) and 31 SCCs (83.7%) showed genomic aberrations, indicating a higher degree of chromosomal instability in SCCs. Gains of chromosomal material from 1p, 14q, 16q, 20q, and losses from 4p were seen significantly more frequently in SCCs compared with KAs (P-values 0.0033, 0.0198, 0.0301, 0.0017, and 0.0070), whereas loss from 9p was seen significantly more frequently in KAs (P-value 0.0434). The patterns of recurrent aberrations were also different in the two types of neoplasms, pointing to different genetic mechanisms involved in their developments.

Radiat Environ Biophys, 44(3), 219–224
December, 2005

Space radiation does not induce a significant increase of intrachromosomalexchanges in astronauts' lymphocytes.

M. Horstmann, M. Durante, C. Johannes, R. Pieper, G. Obe

Chromosome aberration analysis in astronauts has been used to provide direct, biologically motivated estimates of equivalent doses and risk associated to cosmic radiation exposure during space flight. However, the past studies concentrated on measurements of dicentrics and translocations, while chromosome intrachanges (inversions) have never been measured in astronauts' samples. Recent data reported in the literature suggest that densely ionizing radiation can induce a large fraction of intrachanges, thus leading to the suspicion that interchanges grossly underestimate the cosmic radiation-induced cytogenetic damage in astronauts. We have analyzed peripheral blood lymphocytes from 11 astronauts involved in short- or long-term space flights in low-Earth orbit using high-resolution multicolor banding to assess the frequency of intrachromosomal exchanges in both pre- and post-flight samples. We did not detect any inversions in chromosome 5 from a total of 2800 cells in astronauts' blood. In addition, no complex type exchanges were found in a total of 3590 astronauts' lymphocytes analyzed by multifluor fluorescence in situ hybridisation. We conclude that, within the statistical power of this study, the analysis of interchanges for biological dosimetry in astronauts does not significantly underestimate the space radiation-induced cytogenetic damage, and complex-type exchanges or intrachanges have limited practical use for biodosimetry at very low doses.

BJU International, 95, 1219- 1225

Quantitative molecular urinary cytology by fluorescence in situ hybridization: a tool for tailoring surveillance of patients with superficial bladder cancer?

M. Bollmann, H. Heller, A. Bánkfalvi, H. Griefingholt, R. Bollmann

OBJECTIVE: To determine whether it is possible to stratify patients with superficial bladder cancer into low- and high-risk groups for tumour recurrence/progression based on the chromosomal pattern detected by fluorescence in situ hybridization (FISH) in one urine cytology specimen used for follow-up testing. PATIENTS AND METHODS: Voided urine samples from 47 consecutive patients with urinary tract neoplasms (13 with no history of urothelial malignancy and 34 under follow-up after complete transurethral resection of superficial urothelial carcinoma of the bladder) were evaluated by liquid-based cytology (ThinPrep(R), CYTYC Corp., Boxborough, MA, USA) and UroVysion FISH (Vysis-Abbott, Downers Grove, IL). RESULTS: Of the 34 patients under surveillance, the UroVysion test was negative in four, 17 had loss of 9p21 sequences either alone or combined with low-frequency trisomy/ies or tetrasomy/ies of chromosomes 3, 7 and 17 in single cells (low-risk FISH), and 13 also had complex aneusomies of the remaining chromosomes (high-risk FISH). One of the four FISH-negative neoplasms, four of the 17 low-risk FISH cases and five of the 11 informative high-risk FISH-positive patients developed recurrence. Progression occurred only in patients with high-risk FISH results, showing high-frequency complex chromosomal polysomies (four of 11). CONCLUSION: The results from this pilot study indicate that the UroVysion FISH test may help to individually assess the clinical behaviour of superficial bladder cancer, based on the chromosomal pattern of exfoliated tumour cells in follow-up urinary cytology. It might be of use to identify those patients likely to progress at earlier and curable stages of disease, and lengthen the surveillance period in those with persistent or recurrent low-risk disease.

J. Histochem. Cytochem., 53, 1433- 1440

Standardization of the immunocytochemical detection of neuroblastoma cells in bone marrow.

K. Swerts, P.F. Ambros, C. Brouzes, J.M.F. Navarro, N. Gross, D. Rampling, R. Schumacher-K., A.R. Sementa, R. Ladenstein, K. Beiske

<p>Standard cytomorphological examination of bone marrow (BM) aspirates does not appear to be sensitive enough to detect single neuroblastoma cells. The SIOPEN Neuroblastoma Bone Marrow Committee developed a sensitive and reproducible anti-GD2 immunocytochemical assay and introduced morphological and immunocytological criteria for the interpretation of results. Fixed cytospins were incubated with a commercially available anti-GD2 monoclonal antibody and an APAAP kit. Cells fulfilling all morphological and immunocytological criteria were called criteria-positive cells (CPCs). Not convincingly interpretable cells fulfilled some, but not all, criteria, and negative cells displayed only exclusion criteria. The genetic profile of doubtful cells was checked by fluorescence in situ hybridization. Ideally, 3 x 10(6) cells were analyzed to reach a 95% probability of detecting one tumor cell in 1 x 10(6) mononuclear cells. Four quality control rounds were organized to validate the method. A total of 111 quality control samples were analyzed. Two main improvements were achieved: in discordant cases, the range between the lowest and highest reported result was reduced by half, and discordant results were only found in samples with less than 10 CPCs per 1 x 10(6). This article describes the first internationally standardized protocol to detect and quantify rare neuroblastoma cells by immunocytochemistry. This method is an indispensable tool for multicenter studies evaluating the clinical significance of minimal residual disease in neuroblastoma.</p>

Biochem J, 387, 703- 710

Role of compartmentalized redox-active iron in hydrogen peroxide-induced DNA damage and apoptosis.

M. Tenopoulou, P.-T. Doulias, A. Barbouti, U. Brunk, D. Galaris

<p>Jurkat cells in culture were exposed to oxidative stress in the form of continuously generated hydrogen peroxide, obtained by the addition of glucose oxidase to the medium. This treatment induced a rapid, dose-dependent increase in the ICIP (intracellular calcein-chelatable iron pool). Early destabilization of lysosomal membranes and subsequent nuclear DNA strand breaks were also observed, as evaluated by the Acridine Orange relocation test and the comet assay respectively. Somewhat later, these effects were followed by a lowered mitochondrial membrane potential, with release of cytochrome c and apoptosis-inducing factor. These events were all prevented if cells were pretreated with the potent iron chelator DFO (desferrioxamine) for a period of time (2-3 h) long enough to allow the drug to reach the lysosomal compartment following fluid-phase endocytosis. The hydrophilic calcein, a cleavage product of calcein acetoxymethyl ester following the action of cytosolic esterases, obviously does not penetrate intact lysosomal membranes, thus explaining why ICIP increased dramatically following lysosomal rupture. The rapid decrease in ICIP after addition of DFO to the medium suggests draining of cytosolic iron to the medium, rather than penetration of DFO through the plasma membrane. Most importantly, these observations directly connect oxidative stress and resultant DNA damage with lysosomal rupture and the release of redox-active iron into the cytosol and, apparently, the nucleus.</p>

Cancer Genet Cytogenet, 163, 44- 56

Chromosomal alterations cause the high rates and wide ranges of drug resistance in cancer cells.

R. Li, R. Hehlman, R. Sachs, P. Duesberg

Conventional mutation-selection theories have failed to explain (i) how cancer cells become spontaneously resistant against cytotoxic drugs at rates of up to 10(-3) per cell generation, orders higher than gene mutation, even in cancer cells; (ii) why resistance far exceeds a challenging drug-a state termed multidrug resistance; (iii) why resistance is associated with chromosomal alterations and proportional to their numbers; and (iv) why resistance is totally dependent on aneuploidy. We propose here that cancer-specific aneuploidy generates drug resistance via chromosomal alterations. According to this mechanism, aneuploidy varies the numbers and structures of chromosomes automatically, because it corrupts the many teams of proteins that segregate, synthesize, and repair chromosomes. Aneuploidy is thus a steady source of chromosomal variation from which, in classical Darwinian terms, resistance-specific aneusomies are selected in the presence of chemotherapeutic drugs. Some of the thousands of unselected genes that hitchhike with resistance-specific aneusomies can thus generate multidrug resistance. To test this hypothesis, we determined the rates of chromosomal alterations in clonal cultures of human breast and colon cancer lines by dividing the fraction of nonclonal karyotypes by the number of generations of the clone. These rates were about 10(-2) per cell generation, orders higher than mutation. Chromosome numbers and structures were determined in metaphases hybridized with color-coded chromosome-specific DNA probes. Further, we tested puromycin-resistant subclones of these lines for resistance-specific aneusomies. Resistant subclones differed from parental lines in four to seven specific aneusomies, of which different subclones shared some. The degree of resistance was roughly proportional to the number of these aneusomies. Thus, aneuploidy is the primary cause of the high rates and wide ranges of drug resistance in cancer cells.

Cytometry, 68, 113- 120

Automatic telomere length measurements in interphase nuclei by IQ-FISH.

R. Narath, T. Lörch, K.M. Greulich-Bode, P. Boukamp, P.F. Ambros

<p>To benefit from the fluorescence-based automatic microscope (FLAME), we have adapted a PNA FISH technique to automatically determine telomere length in interphase nuclei. The method relies on the simultaneous acquisition of pan-telomeric signals and reference probe signals. We compared the quantitative figures to those for existing methods, i.e. Southern blot analysis and quantitative FISH (Q-FISH). Quantitative-FISH on interphase nuclei (IQ-FISH) allows the exact quantification of telomere length in interphase nuclei. Thus, this enables us to obtain not only exact information on the telomere length, but also morphological and topological details. The automatic measurement of large cell numbers allows the measurement of statistically relevant cell populations.</p>

Leuk Res, 29(9), 987-93

Adequate cytogenetic examination in myelodysplastic syndromes: analysis of 529 patients

C Steidl, R Steffens, W Gassmann, B Hildebrandt, R Hilgers, U Germing, L Trümper, D Haase

In myelodysplastic syndromes (MDS), the karyotype is one of the most significant prognostic markers with profound impact on differential diagnosis and therapeutic decisions. In a retrospective study, we examined karyotypes of bone marrow specimens of an oligocentric cohort comprising 529 patients with MDS to address the question how many metaphases need to be analyzed to detect even small cell clones with an appropriate expenditure. We found a statistically significant difference of the frequency of normal karyotypes in the patient group with 19 or less analyzed metaphases compared to the group with 20 or more metaphases analyzed (56% versus 47%, p=0.041). Furthermore, we demonstrate that the analysis of 25 or more metaphases can further improve the sensitivity of karyotype analysis and leads to the identification of additional clinically relevant abnormal clones or subclones in a substantial proportion of patients. In summary, our data suggest the examination of at least 20 metaphases in MDS.

Diagn Mol Pathol, 14, 77- 82

Fluorescence in situ hybridization for the detection of t(X;18)(p11.2;q11.2) in a Synovial Sarcoma tissuemicroarray using a breakapart-style probe.

J. Terry, T.S. Barry, D.E. Horsman, F.D. Hsu, A.M. Gown, D.G. Huntsman, T.O. Nielsen

Synovial sarcomas (SSs) account for 5% of soft tissue tumors and carry a balanced translocation t(X;18)(p11.2;q11.2), detectable in over 90% of cases. This translocation brings together portions of two genes: SYT and SSX. Detecting interruption of the SYT gene on chromosome 18 would be useful as a diagnostic tool. We describe a scoring method to detect disruption of SYT with breakapart probe fluorescence in situ hybridization (FISH) and the application of this method for identification of SS within a sarcoma tissue microarray. After optimization, SYT disruption was identified in 22 of 23 (96%) of known SS tumor samples but was not in 23 of 23 (100%) of non-SS sarcoma samples. Ten of 11 (91%) blinded test SS tumor samples were also correctly identified. For comparison, commercially available FISH and chromogenic in situ hybridization (CISH) probes were tested. The commercial FISH probes identified SYT disruption in 81% of the SS tumor samples but in none of the non-SS samples. The CISH probes produced signals too weak to interpret. The use of breakapart FISH probes is a relatively quick procedure for detection of synovial sarcoma translocations and can be applied to archival specimens in tissue microarrays.

Int J Mol Med, 16, 463- 469

Studies on the action of mitomycin C and bleomycin on telomere lengths of human lymphocyte chromosomes.

U. Wick, E. Gebhart

In order to address the problem of the action of cytostatics on chromosome ends, telomere length was measured in human lymphocyte cultures exposed to mitomycin C (MMC) and bleomycin (BLM). Telomere-specific PNA probes were used for the quantitative estimation of the relative telomere length of each individual chromosome by fluorescence in situ hybridization. A high inter-cellular and inter-individual variability of relative telomere lengths was found throughout all experiments. Different responses could be observed with respect to the action of the examined mutagens: The total average fluorescence intensity of labeled telomere repeats was decreased under the action of MMC in two of the experiments, while two revealed no significant alteration. BLM caused no significant change of total average telomeric signal intensity in four, a clear decrease in one of the six experiments, and an increase in another. Although all chromosome ends contributed to the observed trends, single telomeres were affected in a very distinct way. The highest concentration of MMC (1 microg/ml) induced significant shortening of telomeres of the chromosome arms; 2q, 3p, 5q, 7p, 10q, 11p, 13q, 17p, 18p&q, and 21q in two independent experiments. In one BLM experiment with 8 microg/ml, the most distinct decrease (p< or =0.005) of telomeric fluorescence was found at the ends of chromosome arms; 1q, 6p, 17p, 20p&q, and 22q. The increase of telomeric signal intensity affected the telomeres of some individual chromosome arms more than others, e.g. 4q, 6p, 7p, 8p, 13p, and 18q. Although the telomere length of the individual chromosome arms varied widely, clear trends could be observed with respect to the rank which was occupied by telomeric length of the various chromosome arms. The telomeres of the 1p, 3p, 4q, 5p, 12q, and 13q chromosome arms throughout all experiments were among the longest; and those of 13p, 15p, 21p, and 22p were among the shortest telomeres of the karyotype. From these data, it can be concluded that MMC affects the telomeric repeat area of chromosomes more than BLM, which mostly had no significant effect on telomere length in the performed experiments.

Cytogenet. Genome Res., 111, 41- 45

Radiosensitivity detected by the micronucleus test is not generally increased in sporadic prostate cancer patients.

D. Varga, I. Michel, B. Patino-Garcia, T. Paiss, W. Vogel, C. Maier

The micronucleus test (MNT) has shown increased micronuclei (MN) frequencies in BRCA associated and sporadic breast cancer patients, Ataxia telangiectasia and Nijmegen Breakage Syndrome patients, demonstrating a common cellular phenotype of increased radiosensitivity. Some genes, causative of these diseases, have also recently been associated with prostate cancer. In order to investigate if prostate cancer exhibits the cellular phenotype of increased radiosensitivity, we performed MNT analysis on 22 sporadic prostate cancer patients and 43 male controls. We determined the baseline MN frequency, in order to see in vivo chromosomal damage without radiation, and induced (after irradiation with 2 Gy) frequency of MN, both in binucleated cells (BNC) obtained from cultured peripheral blood lymphocytes. An automated image analysis system was used to score the MN employing two different classifiers (Classifier A and B) for detection of BNC. The mean baseline frequencies were 48/43 MN/1000 BNC (A/B) for the controls and 42/50 (A/B) for prostate cancer patients. The induced MN frequencies amounted to 107/111 MN/1000 BNC (A/B) for controls and 111/114 MN/1000 BNC (A/B) for prostate cancer patients. The obtained MN frequencies did not result in a statistically significant difference between unselected cases and controls. However, restricting the analysis to young patients (50-60 years, N = 7) and age-matched controls (N = 7) revealed marginally significant higher MN frequencies in patients. We conclude that increased radiosensitivity is not a property of prostate cancer patients in general.

Int J Radiat Biol, 26, 1707- 1713

Are telomeres a specific target for mutagenic attack by cytostatics in neoplastic cells?

U. Wick, E. Gebhart

Damage to telomeres induced by cytostatic therapy theoretically could generate telomere shortening and, subsequently, induce an additional genomic instability in neoplastic cells. Model experiments were carried out to examine this hypothesis. Cells of the T-ALL derived cell line CCRF-CEM were exposed to various different concentrations of Bleomycin (BLM) or Mitomycin C (MMC) for various times. Telomere lengths of metaphase chromosomes of the exposed cells were compared with those without this exposure (controls). In addition, telomerase activity was determined with a TRAP assay under the given conditions using the BLM experiments as a model. Although slight changes of total telomere length could be found in single experiments, the differences between exposed and non-exposed cells were not significant. Also, a considerable telomerase activity was shown which, however, did not substantially differ between exposed and non-exposed cells. From these data it may be concluded that, at least in the examined cell line, telomeres are not a preferential target for this kind of mutagenic attack.

Leukemia Research, 29, 273- 281

Prognostic value of structural chromosomal rearrangements and small cell clones with high hyperdiploidy in children with acute lymphoblastic leukemia.

Z. Zemanova, K. Michalova, L. Sindelarova, P. Smisek, J. Brezinova, S. Ransdorfova, V. Vavra, A. Dohnalova, J. Stary

In this study, 107 children with acute lymphoblastic leukemia (ALL) were analysed for the presence of hyperdiploidy by cytogenetics and interphase fluorescence in situ hybridisation (I-FISH). Structural aberrations in hyperdiploid cells were investigated by multiple colour FISH (mFISH). Clones with high hyperdiploidy (>50 chromosomes) (HeH) were found in 46 patients (43%). In nine of these (20%), the abnormal clone was present in <20% of the total cell population. There was no significant difference in EFS between those patients with HeH in 2.5-20% or >20% of cells. Structural rearrangements in the HeH clone were found in 10 patients (22%). In this study, HeH karyotypes containing structural aberrations were an indication of a poor prognosis in childhood ALL.

World J Surg Oncol., 17, 35- 42

Secretory carcinoma of the breast containing the ETV6-NTRK3 fusion gene in a male: case report and review of the literature

C. Arce, D. Cortes-Padilla, D.G. Huntsman, M.A. Miller, A. Duennas-Gonzalez, A. Alvarado, V. Pérez, D. Gallardo-Rincón, F. Lara-Medina

SUMMARY: BACKGROUND: Secretory carcinoma (SC) of the breast is a rare and indolent tumor. Although originally described in children, it is now known to occur in adults of both sexes. Recently, the tumor was associated with the ETV6-NTRK3 gene translocation. CASE PRESENTATION: A 52-year-old male was diagnosed with secretory breast carcinoma and underwent a modified radical mastectomy. At 18 months the tumor recurred at the chest wall and the patient developed lung metastases. He was treated concurrently with radiation and chemotherapy without response. His tumor showed the ETV6-NTRK3 translocation as demonstrated by fluorescent in situ hybridization (FISH). CONCLUSION: SC is a rare slow-growing tumor best treated surgically. There are insufficient data to support the use of adjuvant radiation or chemotherapy. Its association with the ETV6-NTRK3 fusion gene gives some clues for the better understanding of this neoplasm and eventually, the development of specific therapies.

Nucleic Acids Research, 33, 2512- 2520

XRCC1 is required for DNA single-strand break repair in human cells.

R. Brem, J. Hall

The X-ray repair cross complementing 1 (XRCC1) protein is required for viability and efficient repair of DNA single-strand breaks (SSBs) in rodents. XRCC1-deficient mouse or hamster cells are hypersensitive to DNA damaging agents generating SSBs and display genetic instability after such DNA damage. The presence of certain polymorphisms in the human XRCC1 gene has been associated with altered cancer risk, but the role of XRCC1 in SSB repair (SSBR) in human cells is poorly defined. To elucidate this role, we used RNA interference to modulate XRCC1 protein levels in human cell lines. A reduction in XRCC1 protein levels resulted in decreased SSBR capacity as measured by the comet assay and intracellular NAD(P)H levels, hypersensitivity to the cell killing effects of the DNA damaging agents methyl methanesulfonate (MMS), hydrogen peroxide and ionizing radiation and enhanced formation of micronuclei following exposure to MMS. Lowered XRCC1 protein levels were also associated with a significant delay in S-phase progression after exposure to MMS. These data clearly demonstrate that XRCC1 is required for efficient SSBR and genomic stability in human cells.