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We aimed at molecularly dissecting the anatomical heterogeneity of small lymphocytic lymphoma (SLL), by analysing a cohort of 12 patients for whom paired DNA from a lymph node biopsy and circulating cells, as well as plasma-circulating tumour DNA (ctDNA) was available. Notably, the analyses of the lymph node biopsy and of circulating cells complement each other since a fraction of mutations (20·4% and 36·4%, respectively) are unique to each compartment. Plasma ctDNA identified two additional unique mutations. Consistently, the different synchronous sources of tumour DNA complement each other in informing on driver gene mutations in SLL harbouring potential prognostic and/or predictive value., (© 2021 The Authors. British Journal of Haematology published by British Society for Haematology and John Wiley & Sons Ltd.)

Smith-Magenis syndrome (SMS), characterized by dysmorphic features, neurodevelopmental disorder, and sleep disturbance, is due to an interstitial deletion of chromosome 17p11.2 (90%) or to point mutations in the RAI1 gene. In this retrospective cohort, we studied the clinical, cognitive, and behavioral profile of 47 European patients with SMS caused by a 17p11.2 deletion. We update the clinical and neurobehavioral profile of SMS. Intrauterine growth was normal in most patients. Prenatal anomalies were reported in 15%. 60% of our patients older than 10 years were overweight. Prevalence of heart defects (6.5% tetralogy of Fallot, 6.5% pulmonary stenosis), ophthalmological problems (89%), scoliosis (43%), or deafness (32%) were consistent with previous reports. Epilepsy was uncommon (2%). We identified a high prevalence of obstipation (45%). All patients had learning difficulties and developmental delay, but ID range was wide and 10% of patients had IQ in the normal range. Behavioral problems included temper tantrums and other difficult behaviors (84%) and night-time awakenings (86%). Optimal care of SMS children is multidisciplinary and requires important parental involvement. In our series, half of patients were able to follow adapted schooling, but 70% of parents had to adapt their working time, illustrating the medical, social, educative, and familial impact of having a child with SMS., (© 2020 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Acute promyelocytic leukemia (APL) is generally characterized by t(15;17)(q24;q21). In some cases, the classic translocation cannot be identified by conventional methods, since the PML-RARA fusion protein results from complex, variant, or cryptic translocation. The diagnostic algorithm of APL starts with screening methods, such as flow cytometry (FC), followed by fluorescence in situ hybridization or polymerase chain reaction to confirm the diagnosis. Our aim was to develop a novel protocol for analyzing APL samples based on multidimensional dot-plots that can provide comprehensive information about several markers at the same time. The protocol included four optimized multidimensional dot-plots, which were tested by retrospective reanalysis of FC results in APL (n = 8) and non-APL (n = 12) acute myeloid leukemia (AML) cases. After predicting the potential position of hypergranular- and microgranular-type aberrant promyelocytes, the percentages of blast populations were examined within the gates in all AML cases. The percentage of blasts in each predefined gate was well above the cut-off value (95%) in APL cases in all tubes. In non-APL AML cases, the percentage of blasts in the same gates never reached the cut-off value in all investigated tubes, and even when it did in a single tube, the pattern was markedly different from that observed in APL cases. In conclusion, multidimensional dot-plots can be used for screening APL even in cryptic APL cases, although reproducibility across several laboratories would require standardization of antibodies and fluorochromes. This easy-to-use and quick method can support the diagnosis of APL and the prompt initiation of the appropriate treatment.

The Myeloma X trial (ISCRTN60123120) registered patients with relapsed multiple myeloma. Participants were randomised between salvage autologous stem cell transplantation (ASCT) or weekly cyclophosphamide following re-induction therapy. Cytogenetic analysis performed at trial registration defined t(4;14), t(14;16) and del(17p) as high-risk. The effect of cytogenetics on time to progression (TTP) and overall survival was investigated. At 76 months median follow-up, ASCT improved TTP compared to cyclophosphamide (19 months (95% confidence interval [95% CI] 16-26) vs. 11 months (9-12), hazard ratio [HR]: 0·40, 95% CI: 0·29-0·56, P < 0·001), on which the presence of any single high-risk lesion had a detrimental impact [likelihood ratio test (LRT): P = 0·011]. ASCT also improved OS [67 months (95% CI 59-not reached) vs. 55 months (44-67), HR: 0·64, 95% CI: 0·42-0·99, P = 0·0435], with evidence of a detrimental impact with MYC rearrangement (LRT: P = 0·021). Twenty-one (24·7%) cyclophosphamide patients received an ASCT post-trial, median OS was not reached (95% CI: 39-not reached) for these participants compared to 31 months (22-39), in those who did not receive a post-trial ASCT. The analysis further supports the benefit of salvage ASCT, which may still be beneficial after second relapse in surviving patients. There is evidence that this benefit reduces in cytogenetic high-risk patients, highlighting the need for targeted study in this patient group., (© 2019 The Authors. British Journal of Haematology published by British Society for Haematology and John Wiley & Sons Ltd.)

Demyelinating Charcot-Marie-Tooth disease (CMT) is the most common subtype of CMT. It is caused mainly by 17p11.2 heterozygous duplication, but also by mutations in more than 20 genes which affect development and function of Schwann cells. To investigate the profile of genes mutated and clinical features in demyelinating CMT of Chinese descent, we collected a cohort of 44 demyelinating CMT patients and screened them using multiplex ligation-dependent probe amplification (MLPA) and targeted next-generation sequencing (NGS) technology. The MLPA technology revealed that 77.3% demyelinating CMT patients harbored 17p11.2 heterozygous duplication and 6.8% patients harbored heterozygous deletion of exon 6 of MPZ gene, that was further confirmed a novel c.674&#95;675insA mutation in MPZ gene. In the patients with 17p12 heterozygous duplication, 3 sets of independent families were discordant for the CMT phenotype within the same family. The targeted NGS technology revealed that 6 candidate mutations including 1 previously reported mutation (GDAP1: c.571C>T) and 5 novel mutations (SBF2: c.415T>C, c.619G>T, c.1258A>G; GDAP1: c.589delC; PMP22: c.318delT) were found. In conclusion, combined MLPA technique with targeted NGS, the demyelinating CMT genetic diagnostic success rate was increased., (© 2018 Peripheral Nerve Society.)

Chronic Myeloid Leukemia (CML) is myeloproliferative neoplasm characterized by Philadelphia chromosome which is a balanced translocation between chromosome 9 and 22 in 90% of cases. However, variant cytogenetic still happens in 5-10 % of cases, the importance of which is controversial as well as its response to therapy, prognosis and progression to acute leukemias. Here we report a male patient with CML and variant cytogenetic who responded to low dose of Dasatinib (50 mg daily).

Chronic lymphocytic leukemia (CLL) with 17p deletion (17p-) is associated with a lack of response to standard treatment and thus the worst possible clinical outcome. Various chromosomal abnormalities (including unbalanced translocations, deletions, ring chromosomes and isochromosomes) result in the loss of 17p and one copy of the TP53 gene. The objective of the present study was to determine whether the type of chromosomal abnormality leading to 17p- and the additional aberrations influenced the prognosis in a series of 195 patients with 17p-CLL. Loss of 17p resulted primarily from an unbalanced translocation (70%) with several chromosome partners (the most frequent being chromosome 18q), followed by deletion 17p (23%), monosomy 17 (8%), isochromosome 17q [i(17q)] (5%) and a ring chromosome 17 (2%). In a univariate analysis, monosomy 17, a highly complex karyotype (≥5 abnormalities), and 8q24 gain were associated with poor treatment-free survival, and i(17q) (P = .04), unbalanced translocations (P = .03) and 8q24 gain (P = .001) were significantly associated with poor overall survival. In a multivariate analysis, 8q24 gain remained a significant predictor of poor overall survival. We conclude that 17p deletion and 8q24 gain have a synergistic impact on outcome, and so patients with this "double-hit" CLL have a particularly poor prognosis. Systematic, targeting screening for 8q24 gain should therefore be considered in cases of 17p- CLL., (© 2017 Wiley Periodicals, Inc.)

Purpose: Chronic lymphocytic leukemia (CLL) with 17p deletion typically progresses quickly and is refractory to most conventional therapies. However, some del(17p) patients do not progress for years, suggesting that del(17p) is not the only driving event in CLL progression. We hypothesize that other concomitant genetic abnormalities underlie the clinical heterogeneity of del(17p) CLL., Experimental Design: We profiled the somatic mutations and copy number alterations (CNA) in a large group of del(17p) CLLs as well as wild-type CLL and analyzed the genetic basis of their clinical heterogeneity., Results: We found that increased somatic mutation number associates with poor overall survival independent of 17p deletion (P = 0.003). TP53 mutation was present in 81% of del(17p) CLL, mostly clonal (82%), and clonal mutations with del(17p) exhibit shorter overall survival than subclonal mutations with del(17p) (P = 0.019). Del(17p) CLL has a unique driver mutation profile, including NOTCH1 (15%), RPS15 (12%), DDX3X (8%), and GPS2 (6%). We found that about half of del(17p) CLL cases have recurrent deletions at 3p, 4p, or 9p and that any of these deletions significantly predicts shorter overall survival. In addition, the number of CNAs, but not somatic mutations, predicts shorter time to treatment among patients untreated at sampling. Indolent del(17p) CLLs were characterized by absent or subclonal TP53 mutation and few CNAs, with no difference in somatic mutation number., Conclusions: We conclude that del(17p) has a unique genomic profile and that clonal TP53 mutations, 3p, 4p, or 9p deletions, and genomic complexity are associated with shorter overall survival. Clin Cancer Res; 23(3); 735-45. ©2016 AACR., (©2016 American Association for Cancer Research.)

Automated imaging flow cytometry integrates flow cytometry with digital microscopy to produce high-resolution digital imaging with quantitative analysis. This enables cell identification based on morphology (cell size, shape), antigen expression, quantification of fluorescence signal intensity and localisation of detected signals (i.e. surface, cytoplasm, nuclear). We describe applications of imaging flow cytometry for the diagnostic assessment of acute leukaemia. These bone marrow malignancies are traditionally diagnosed and classified by cell morphology, phenotype and cytogenetic abnormalities. Traditionally morphology is assessed by light microscopy, phenotyping by conventional flow cytometry and genetics by karyotype and fluorescence in situ hybridisation (FISH) on interphase nuclei/metaphase spreads of cells on slides. Imaging flow cytometry adds a new dimension to the diagnostic assessment of these neoplasms. We describe three specific applications: From this we conclude that imaging flow cytometry offers benefits over conventional diagnostic methods. Specifically the ability to visualise the cells of interest, the pattern and localisation of expressed antigens and assess cytogenetic abnormalities in one integrated automated high-throughput test. Imaging flow cytometry presents a new paradigm for the diagnostic assessment of leukaemia., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Acute promyelocytic leukaemia with PML-RARA fusion is usually associated with the t(15;17)(q24.1;q21.1) translocation but may also arise from complex or cryptic rearrangements. The fusion usually resides on chromosome 15 but occasionally on others. We describe a cryptic PML-RARA fusion within a novel chromosome 17 rearrangement. We performed interphase fluorescence in situ hybridisation (FISH) using a dual-fusion PML-RARA probe, followed by reverse transcriptase-polymerase chain reaction (RT-PCR) for PML-RARA, karyotyping, and metaphase FISH using RARA break-apart, locus-specific, and subtelomere probes for chromosome 17. An 850K SNP microarray was also employed. Interphase and metaphase FISH showed atypical results involving a single PML-RARA fusion, no second fusion, but instead separate diminished PML and RARA signals. RT-PCR confirmed PML-RARA fusion; however, karyotyping detected only an altered chromosome 17. Metaphase FISH showed the single fusion and diminished 5' RARA signals located unexpectedly in the subtelomeric short-arm and long-arm regions of the rearranged chromosome 17, respectively. SNP microarray revealed no copy number abnormality. This paediatric patient with PML-RARA fusion reflects a cryptic insertion that resides within a complex and novel chromosome 17 rearrangement. This rearrangement likely arose via 7 chromosome breaks with the insertion occurring first followed by sequential paracentric and then pericentric inversions., (© 2018 S. Karger AG, Basel.)

We constructed a multiple myeloma (MM)-specific gene panel for targeted sequencing and investigated 72 untreated high-risk (del17p) MM patients. Mutations were identified in 78% of the patients. While the majority of studied genes were mutated at similar frequency to published literature, the prevalence of TP53 mutation was increased (28%) and no mutations were found in FAM46C. This study provides a comprehensive insight into the mutational landscape of del17p high-risk MM. Additionally, our work demonstrates the practical use of a customized sequencing panel, as an easy, cheap and fast approach to characterize the mutational profile of MM., (© 2014 John Wiley & Sons Ltd.)

Purpose: Therapies that target overexpression of human epidermal growth factor receptor 2 (HER2) rely on accurate and timely assessment of all patients with new diagnoses. This study examines HER2 testing of primary breast cancer tissue when performed with immunohistochemistry (IHC) and additional in situ hybridization (ISH) for negative cases (IHC 0/1+). The analysis focuses on the rate of false-negative HER2 tests, defined as IHC 0/1+ with an ISH ratio ≥ 2.0, in eight pathology centers across Canada., Patients and Methods: Whole sections of surgical resections or tissue microarrays (TMAs) from invasive breast carcinoma tissue were tested by both IHC and ISH using standardized local methods. Samples were scored by the local breast pathologist, and consecutive HER2-negative IHC results (IHC 0/1+) were compared with the corresponding fluorescence or silver ISH result., Results: Overall, 711 surgical excisions of primary breast cancer were analyzed by IHC and ISH; HER2 and chromosome 17 centromere (CEP17) counts were available in all cases. The overall rate of false-negative samples was 0.84% (six of 711 samples). Interpretable IHC and ISH scores were available in 1,212 cases from TMAs, and the overall rate of false-negative cases was 1.6% (16 of 978 cases)., Conclusion: Our observation confirms that IHC is an adequate test to predict negative HER2 status in primary breast cancer in surgical excision specimens, even when different antibodies and IHC platforms are used. The study supports the American Society of Clinical Oncology/College of American Pathologists and Canadian testing algorithms of using IHC followed by ISH for equivocal cases., (© 2014 by American Society of Clinical Oncology.)

Acute promyelocytic leukemia (APL) is a distinct subset of acute myeloid leukemia (AML) associated with peculiar biologic and clinical features and requiring specific management. At the genetic level, APL is featured by a unique chromosome translocation t(15;17) which results in the PML-RARα gene fusion and chimeric protein. APL is the first example of differentiation therapy targeted to a defined genetic target i.e. PML-RARα. PML-RARα behaves as an altered retinoic acid receptor with an ability of transmitting oncogenic signaling leading to accumulation of undifferentiated promyelocytes. All-trans-retinoic acid (ATRA) induces disease remission in APL patients by triggering terminal differentiation of leukemic promyelocytes. More recently, arsenic trioxide (ATO) has been shown to contribute degradation of the PML-RARα oncoprotein through bonding the PML moiety and has shown excellent synergism with ATRA in clinical trials. Elucidating the oncogenic signaling of PML-RARα through various transcription factors and the study of APL mouse models have greatly helped to understand the molecular pathogenesis of APL. However, the precise molecular mechanism by which t(15;17) is formed and initiates leukemia remains unknown. While transforming oncogenic potential of PML-RARα has been described extensively, the mechanistic events important for the formation of t(15;17) have been taken from the model of Therapy-related APL (t-APL)., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

The aim of this study was to establish a scoring method for ploidy analysis using silver in situ hybridisation (SISH) with a chromosome 17 centromere probe. SISH was performed using the Ventana chromosome 17 centromere probe on sections from formalin fixed, paraffin embedded archival cases of complete hydatidiform moles, partial hydatidiform moles and hydropic products of conception with previously established ploidy status (determined by flow cytometry or karyotyping). In order to determine ploidy status, a scoring method was developed based on both the average number of signals per nucleus (ASN) and the percentage of nuclei with three signals (N3S), enumerated in 50 villous cytotrophoblastic and/or stromal cells. The results of four independent observers were compared individually and collectively with previously established ploidy status. There was a highly statistically significant difference between diploid and triploid gestations for ASN (1.86 ± 0.13 and 2.70 ± 0.16 respectively, Student t-test, p < 0.0001) and for N3S (1.14 ± 1.65 and 71.59 ± 14.25 respectively, Student t-test, p < 0.0001). The sensitivity and specificity of the SISH-based assay was 99.1% and 100% respectively for ASN, and 100% and 100% respectively for N3S. A chromosome 17 centromere probe SISH-based assay can reliably distinguish between diploid and triploid gestations. This test has diagnostic utility in distinguishing partial hydatidiform moles from histological mimics.

We report the cases of 3 patients with hematological malignancies and complex karyotypes involving der(5; 17) (p10;q10), which results in the loss of 5q and 17p. Although deletions of 5q and 17p are recurrent abnormalities in hematological disease, only about 20 cases harboring der(5; 17) (p10;q10) have been reported. We address the tumorigenesis and morphological characteristics of hematological malignancies involving der(5; 17)(p10;q10), along with a review of the literature., (© 2014 S. Karger AG, Basel)

Objectives: To compare the INFORM HER2 bright-field dual in situ hybridization (DISH) DNA probe cocktail assay with the PathVysion fluorescence in situ hybridization (FISH) assay on 103 invasive breast carcinomas with a 2+ score on immunohistochemistry (IHC)., Methods: The cases were categorized as positive, equivocal, or negative for HER2 gene amplification using the 2007 American Society of Clinical Oncology/College of American Pathologists (ASCO/CAP) HER2:CEP17 ratio criteria and also based on mean HER2 gene copies/cell. The third criterion used a HER2:CEP17 ratio of 2 to categorize cases as positive or negative., Results: The agreement between FISH and DISH was 85% using the 2007 ASCO/CAP ratio criterion, 79% using the mean HER2 gene copies/cell criterion, and 92% using the 2.0 cutoff HER2:CEP17 ratio criterion. In addition, 20 known IHC 3+ breast carcinomas analyzed by DISH showed clusters of the HER2 gene consistent with unequivocal amplification., Conclusions: Despite some technical and interpretational issues associated with DISH, it compares favorably with FISH in this group of challenging breast cancer cases.

Detection of a 17p13.1 deletion (loss of TP53) or 11q22.3 deletion (loss of ATM), by fluorescence in situ hybridization (FISH), in chronic lymphocytic leukaemia (CLL) patients is associated with a poorer prognosis. Because TP53 and ATM are integral to the TP53 pathway, we hypothesized that 17p13.1- (17p-) and 11q22.3- (11q-) occurring in the same cell (clonal 17p-/11q-) would confer a worse prognosis than either 17p- or 11q-. We studied 2184 CLL patients with FISH (1995-2012) for the first occurrence of 17p-, 11q-, or clonal 17p-/11q-. Twenty (1%) patients had clonal 17p-/11q-, 158 (7%) had 17p- (including 4 with 17p- and 11q- in separate clones), 247 (11%) had 11q-, and 1759 (81%) had neither 17p- nor 11q-. Eleven of 15 (73%) tested patients with clonal 17p-/11q- had dysfunctional TP53 mutations. Overall survival for clonal 17p-/11q- was significantly shorter (1·9 years) than 17p- (3·1 years, P = 0·04), 11q- (4·8 years, P ≤ 0·0001), or neither 17p- nor 11q- (9·3 years, P ≤ 0·0001). Clonal 17p-/11q- thus conferred significantly worse prognosis, suggesting that loss of at least one copy of both TP53 and ATM causes more aggressive disease. Use of an ATM/TP53 combination FISH probe set could identify these very-high risk patients., (© 2013 John Wiley & Sons Ltd.)

Although the breast cancer susceptibility gene BRCA1 is one of the most extensively characterized genetic loci, much less is known about its upstream variable number tandem repeat element, the RNU2 locus. RNU2 encodes the U2 small nuclear RNA, an essential splicing element, but this locus is missing from the human genome assembly due to the inherent difficulty in the assembly of repetitive sequences. To fill the gap between RNU2 and BRCA1, we have reconstructed the physical map of this region by re-examining genomic clone sequences of public databases, which allowed us to precisely localize the RNU2 array 124 kb telomeric to BRCA1. We measured by performing FISH analyses on combed DNA for the first time the exact number of repeats carried by each of the two alleles in 41 individuals and found a range of 6-82 copies and a level of heterozygosity of 98%. The precise localisation of the RNU2 locus in the genome reference assembly and the implementation of a new technical tool to study it will make the detailed exploration of this locus possible. This recently neglected macrosatellite could be valuable for evaluating the potential role of structural variations in disease due to its location next to a major cancer susceptibility gene.

Objective: To assess the value of the study of chromosomal alterations by fluorescent in situ hybridization in a series of patients diagnosed of urothelial carcinoma and a minimum follow up of twenty four months, as well as evaluate its putative predictive potential., Material and Methods: The overall series includes 338 samples from 98 patients with 84 episodes of urothelial carcinoma. A subgroup of 24 patients who had at least one recurrence during the follow up was used to evaluate the predictive potential of the test. Three categories were considered (positive coherent episode, negative coherent episode, and incoherent episode) depending on the relationship between the fluorescent in situ hybridization result in the concomitant study of the new episode and those of the preceding samples., Results: Fluorescent in situ hybridization showed higher sensitivity regardless of grade, negative predictive value and accuracy, while specificity and positive predictive value were superior with conventional cytology. In the recurrence, series 19/29 episodes were coherent, 11/19 were positive coherent with urothelial carcinoma all high grade and 8/19 negative coherent, most low grade., Conclusions: Fluorescent in situ hybridization test shows good sensitivity during a follow up of twenty four months and is able to predict recurrence, especially in cases of high grade. Our data demonstrate the existence of urothelial carcinomas without detectable chromosomal abnormalities by currently available methodology. The results support a multidisciplinary follow up combining fluorescent in situ hybridization, cytology and cystoscopy., (Copyright © 2012 AEU. Published by Elsevier Espana. All rights reserved.)

Standardized criteria for diagnosis and response evaluation in chronic lymphocytic leukemia (CLL) are essential to achieve comparability of results and improvement of clinical care. With the increasing range of therapeutic options, the treatment context is important when defining refractory CLL. Refractory CLL has been defined as no response or response lasting ≤ 6 months from last therapy. This subgroup has a very poor outcome, and many trials use this group as an entry point for early drug development. With the intensification of first-line regimens, the proportion of patients with refractory CLL using these criteria decreases. This has immediate consequences for recruitment of patients into trials as well as salvage strategies. Conversely, patients who are not refractory according to the traditional definition but who have suboptimal or short response to intense therapy also have a very poor outcome. In this Perspective, we discuss recent results that may lead to a reassessment of risk categories in CLL focusing on fit patients who are eligible for all treatment options. We cover aspects of the history and biologic basis for refractory CLL and will focus on how emerging data on treatment failure from large trials using chemoimmunotherapy may help to define risk groups in CLL.

The commercial sample of human DNA fragment from the choromosome 17 was used as the probe for FISH to study of the mode of its attachment to the lateral elements of synaptonemal complex (SC) in human spermatocytes. It was a 160 kb probe from the band 17p1.2, containing RAI1 gene with D17S620 marker (the probe for deletion causing Smith-Magenis syndrome). The probe made lateral chromatin protrusions, contacting with SC stained with anty-SYCP3. Different morphological configuration of lateral chromatin protrusions where observed. They depended on substages of meiotic prophase I. At zygotene, FISH probe form two sticks, c. a. 6 micro long, which was perpendicular to SC longitudinal axe, one stick at each SC side. At early pachytene, each stick transforms into a globule, one globule at each SC side again. At late pachytene each globule transformed into two crumbly globules containing short threads and clumps. At diplotene, globules finally transformed into thin DNA (chromatin) loops up to 10 micro long from the base to top with periodical thickenings (beads) along their length. As the result of this dynamics of transformation, two chromatin loops with beads were found on each side of SC of the chromosome 17. These loops most probably were the loops of sister chromatides, the full set of chromatide loops at the particular SC (bivalent) site being four in number, i. e. representing of two pair of chromatides. This study is the first one in which lateral chromatin loops in human mail meiotic prophase I are visualized as true open loop instead of that usually postulated "loops" after observation of condensed road-like or brush-like chromatin protrusion attached to the lateral elements of synaptonemal complexes. Open configuration of the loops, presumably, depends on activation of transcription during late pachytene-early diplotene. They resemble lateral loops of mini lampbrush chromosomes.

Objectives: The amplification of the gene encoding for the human epidermal growth factor receptor 2 (HER-2 oncogene), located on chromosome 17 (17q21-q22), or the overexpression of this receptor have prognostic and therapeutic implications in invasive breast cancer. An evaluation of the HER-2 status by immunohistochemistry (IHC) is performed on all invasive breast cancer cases. Fluorescent in situ hybridization (FISH) is considered as the gold standard for the detection of HER-2 gene amplification for IHC equivocal cases (score 2+). A more recent in situ hybridization technique, the dual-color chromogenic in situ hybridization (dc-CISH), has been proposed as an alternative to FISH. The aim of this study was to measure the correlation between dc-CISH and FISH for HER-2 oncogene amplification assessment in invasive breast cancer., Methods and Results: We built four tissue micro-array (TMA) blocs with 100 breast invasive cancer cases that had been previously tested by IHC for HER-2 detection: 10 score 0 cases, 10 score 3+cases, 39 score 1+and 41 score 2+cases. Both FISH and dc-CISH techniques were applied on all TMA cases as well as on two additional slides serving as controls. Interpretation of dc-CISH was carried out by a pathologist using an optical microscope. For FISH, the interpretation was done by a professional from the medical genetics department using a fluorescent microscope linked to a computer system for image capturing and analysis. The interpretation of the HER-2/CEN-17 ratio for both tests was in accordance with the values of the updated recommendations from the Canadian National Consensus Meeting on HER-2/neu testing in breast cancer and from the ASCO/CAP. Among the 100 cases initially included in the study, eight were excluded from the analysis due to sampling or technical flaws. From the 92 remaining cases, we obtained a concordance of 97.8% (90/92 cases) between the two techniques (Kappa coefficient 0.97, 95% confidence interval). The correlation coefficient (rho) between ratios was estimated at 0.57., Conclusion: This study shows a strong concordance between FISH and dc-CISH techniques and indicates that dc-CISH is a good alternative method for HER-2 gene amplification assessment in breast cancer., (Copyright © 2011 Elsevier Masson SAS. All rights reserved.)

Genomic disorders constitute a class of diseases that are associated with DNA rearrangements resulting from region-specific genome instability, that is, genome architecture incites genome instability. Nonallelic homologous recombination (NAHR) or crossing-over in meiosis between sequences that are not in allelic positions (i.e., paralogous sequences) can result in recurrent deletions or duplications causing genomic disorders. Previous studies of NAHR have focused on description of the phenomenon, but it remains unclear how NAHR occurs during meiosis and what factors determine its frequency. Here we assembled two patient cohorts with reciprocal genomic disorders; deletion associated Smith-Magenis syndrome and duplication associated Potocki-Lupski syndrome. By assessing the full spectrum of rearrangement types from the two cohorts, we find that complex rearrangements (those with more than one breakpoint) are more prevalent in copy-number gains (17.7%) than in copy-number losses (2.3%); an observation that supports a role for replicative mechanisms in complex rearrangement formation. Interestingly, for NAHR-mediated recurrent rearrangements, we show that crossover frequency is positively associated with the flanking low-copy repeat (LCR) length and inversely influenced by the inter-LCR distance. To explain this, we propose that the probability of ectopic chromosome synapsis increases with increased LCR length, and that ectopic synapsis is a necessary precursor to ectopic crossing-over., (Copyright © 2011 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

Background: Accurate HER2 testing is of great clinical value for the identification of breast cancer patients who are eligible for trastuzumab therapy. The aim of this study is to review breast carcinomas diagnosed from 2001 to 2007 at a Spanish National Reference Centre for HER2 testing, evaluating the agreement between HER2 immunohistochemical (IHC) tests and fluorescence in situ hybridisation (FISH) tests., Methods: Demographic and clinical information was obtained from 2751 breast carcinoma patients. HER2 IHC and FISH tests were performed both in a local laboratory and in the reference centre. The HER2 IHC0/1+, IHC2+, IHC3+ and FISH-positive patients comprised 64%, 20%, 16% and 24% of the available population, respectively (results from the reference centre). Using statistical approaches, we evaluated the agreement between: (1) HER2 IHC and FISH tests, and (2) results provided by the local and the reference laboratories., Results: The data confirmed a statistically significant relation between HER2 overexpression and amplification. We also found that instances of polysomy 17 and heterogeneous patterns of HER2 expression (heterogeneous staining distribution in different areas of the same tumour) are more frequently observed in HER2-positive tumours. Finally, since the diagnoses were made from 2001 to 2007, we could also observe a rising agreement rate between laboratories/pathologists with time., Conclusions: HER2 testing is most accurate when performed by experienced pathologists and at a high-volume reference laboratory. Polysomy 17 and HER2 heterogeneous staining patterns should also be considered for a better understanding of the variation in the anti-HER2 therapeutic response.

Multiple myeloma (MM) is a plasma cell malignancy characterized by very complex cytogenetic and molecular genetic aberrations. In newly diagnosed symptomatic patients, the modal chromosome number is usually either hyperdiploid with multiple trisomies or hypodiploid with one of several types of immunoglobulin heavy chain (Ig) translocations. The chromosome ploidy status and Ig rearrangements are two genetic criteria that are used to help stratify patients into prognostic groups based on the findings of conventional cytogenetics and fluorescence in situ hybridization (FISH). In general, the hypodiploid group with t(4;14)(p16;q32) or t(14;16)(q32;q23) is considered a high-risk group, while the hyperdiploid patients with t(11;14)(q13;q32) are considered a better prognostic group. As the disease progresses, it becomes more proliferative and develops a number of secondary chromosome aberrations. These secondary aberrations commonly involve MYC rearrangements, del(13q), del(17p), and the deletion of 1p and/or amplification of 1q. Of the secondary aberrations, del(17p) is consistently associated with poor prognosis. All of these cytogenetic aberrations and many additional ones are now identified by means of high resolution molecular profiling. Gene expression profiling (GEP), array comparative genomic hybridization (aCGH), and single-nucleotide polymorphism (SNP) arrays have been able to identify novel genetic aberration patterns that have previously gone unrecognized. With the integration of data from these profiling techniques, new subclassifications of MM have been proposed which define distinct molecular genetic subgroups. In this review, the findings from conventional cytogenetics, interphase FISH, GEP, aCGH, and SNP profiles are described to provide the conceptual framework for defining the emerging molecular genetic subgroups with prognostic significance., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Granulocytic sarcoma (GS) is composed of immature granulocytic precursors and is usually found in acute myeloid leukemia (AML) patients with t(8;21). Inv(16) is rarely associated with GS comparing with t(8;21) leukemia. Here we describe an abdominal GS patient in AML-M2 with acquired translocation between chromosomes 6 and 17 and inv (16). We have also summarized 20 reported GS cases with inv(16) and found that chloroma was most often found in abdominal lesions. Intestine maybe a tissue specific target for the expression of inv(16) leukemia. Complete physical examination and molecular diagnosis are necessary for AML patients to benefit from the diagnosis and therapeutic strategy., (Copyright 2010 Elsevier Ltd. All rights reserved.)

To determine the clinical fate of patients with de novo deletion 17p13.1 (17p-) chronic lymphocytic leukemia (CLL), we retrospectively studied the outcome of 99 treatment-naive 17p- CLL patients from the M. D. Anderson Cancer Center (n = 64) and the Mayo Clinic (n = 35). Among 67 asymptomatic patients followed for progression, 53% developed CLL requiring treatment over 3 years. Patients who had not progressed by 18 months subsequently had stable disease, with 3 of 19 patients progressing after follow-up of up to 70 months. Risk factors for progressive disease were Rai stage of 1 or higher and unmutated immunoglobulin variable region heavy chain (IgVH). The overall survival rate was 65% at 3 years. Rai stage 1 or higher, unmutated IgVH, and 17p- in 25% or more of nuclei were adverse factors for survival. The 3-year survival rates of patients with 1 or fewer, 2, and 3 of these factors were 95%, 74%, and 22%, respectively (P < .001). Response rates to therapy with rituximab (n = 6); purine analogues and rituximab (n = 25); and purine analogues, rituximab, and alemtuzumab (n = 16) combinations were 50%, 72%, and 81%, respectively. Patients with 17p- CLL exhibit clinical heterogeneity, with some patients experiencing an indolent course. Survival can be predicted using clinical and biologic characteristics.

Therapy-related myelodysplastic syndrome (t-MDS) and therapy-related acute myelogenous leukemia (t-AML) in patients with acute promyelocytic leukemia (APL) are rare events. The cumulative exposure to chemotherapy with alkylating agents and topoisomerase II inhibitors is associated with t-AML that may develop any time after the completion of the treatment. We report the case of an acquired AML who previously received therapy for APL, after two years of being diagnosed. The diagnosis was established by morphologic findings, membrane markers, cytogenetic studies, and fluorescence in situ hybridization (FISH). To our knowledge this is the first documented case in Puerto Rico of a patient with APL that developed a t-AML without the characteristic chromosomal and morphologic findings of APL.

Semen profile and meiotic segregation products are important for assessing aneuploidy risk and risk of resulting infertility. To determine the effect of varicocelectomy on semen profile and aneuploidy frequency, we investigated semen profile and aneuploidy frequency of selected chromosomes in patients with varicocele before and after varicocelectomy. Chromosomal aneuploidy for selected chromosomes was evaluated using chromosome-specific DNA fluorescence in situ hybridisation (FISH) probes. There was a significant difference in the level of normal sperm morphology before and after varicocelectomy (P > 0.007). There were no significant differences in aneuploidy frequency of chromosomes 1, 16, 17 and 18 in sperm nuclei obtained from patients before varicocelectomy compared with 6-7 months after varicocelectomy (P > 0.05), although FISH analysis with chromosomes 17 and 18 combination showed a higher aneuploidy frequency before varicocelectomy than after varicocelectomy (7.81 +/- 9.67 versus 4.03 +/- 1.46 respectively). In conclusion, varicocele seems to affect the semen profile but minimally affects aneuploidy frequency. Varicocelectomy demonstrates a repairing effect on the semen profile and contributes to a slight decrease in aneuploidy frequency in some but not all chromosomes.

Smith-Magenis syndrome (SMS) is characterized by deletions in the short arm of chromosome 17. Systemic findings in patients with the syndrome include: dysmorphic facies and skeletal deformities. Ophthalmic findings in patients with the SMS include: strabismus, refractive errors, microcornea, iris anomalies, microphthalmos, and coloboma. A 14-year-old boy with cytogenetic studies confirming the SMS underwent a comprehensive ophthalmologic examination. The patient has a history of strabismus surgery. Clinical findings in this patient include: developmental delay, facial dysmorphism, enamel hypoplasia, short broad hands, clinodactyly, and scoliosis. Ocular findings in our patient include: myopia, iris nodules, loose zonules, and ectopia lentis. To our knowledge this is the first reported case of SMS in the Caribbean basin and the first case that report ectopia lentis in SMS. There is a possibility that lens subluxation in our patient is due to self inflicted trauma.

Mental retardation occurs in 2-3% of the general population either in isolation or in combination with facial dysmorphism and/or malformations. Chromosomal abnormalities are a most common etiology. Karyotype displays chromosome aberrations in about 10% of patients but it has a limited resolution (5 Mb). Recently, the development of new molecular cytogenetic tools, especially array CGH, allowed to detect smaller abnormalities and increased the diagnosis capability of 15-20%. Among these newly detected rearrangements, some of them are recurrent and define new recognized syndromes. We will first briefly explain the non-allelic homologous recombination (NAHR) mechanism that underlines the origin of recurrent microdeletions and microduplications. Then we will describe eight new syndromes, four microdeletions (del 17q21.31, del 3q29, del 15q24, del 2q32.3q33) and four microduplications (dup 22q11.2, dup 7q11.23, dup 17p11.2, duplication of MECP2). A better knowledge of these new recurrent chromosomal syndromes will allow to improve care for patients, to develop targeted chromosomal diagnosis and to identify genes involved in neurocognitive functions.

All cultured cells develop chromosome changes over time, including cultures of human embryonic stem cells (hESC), but only those cells with adaptive chromosomes changes survive. The most frequent chromosome changes in hESC cultures are trisomy 12 and trisomy 17. Cells with these trisomies are indistinguishable from normal cells by appearance and also demonstrate typical markers of pluripotency, making them difficult to identify without cytogenetic analysis. Early detection of these cells is essential since cells with trisomy 12 and 17 can replace the normal cell population in 5-10 passages. Cytogenetic analysis using G-banding is considered to be the gold standard for detecting chromosome abnormalities and, when used in combination with interphase FISH, provides a sensitive method for early detection of cytogenetic aberrations, such as full and partial trisomies of chromosomes 12 and 17. The following discussion describes the cytogenetic methods used in our laboratory to study cultured hESCs, along with recommendations for integrating these methods into a plan for routine cell line quality control.

A female patient with Down Syndrome (DS) had neonatal thrombocytosis with platelet counts exceeding 2,000 x 10(3)/microL and transient myeloproliferative disorder (TMD). Platelet counts remained elevated the first 2 months of life. A translocation located between chromosomes 17 and 11 was observed. We describe a patient with thrombocytosis and TMD showing an 11q13 translocation. The leukocytosis and thrombocytosis improved after an exchange transfusion., ((c) 2007 Wiley-Liss, Inc.)

Purpose of Review: We discuss recent findings on the genotypic alterations associated with HER2-positive breast cancer in an attempt to clarify the clinical heterogeneity observed among these tumors., Recent Findings: Molecular genetic analysis supports the distinctive nature of HER2-positive breast cancer, which is primarily driven by HER2 gene amplification. Depending on the amplicon size, a variety of genes can be coamplified and overexpressed together with HER2, some of which may contribute to tumorigenesis; the amplicon size may even predict response to trastuzumab therapy. HER2 gene amplification may further destabilize the tumor genome, facilitating the generation of additional genomic aberrations including aneuploidy. The latter might imply polysomy 17, a phenomenon that should be discriminated from true HER2 gene amplification: polysomy 17 in the absence of HER2 gene amplification is not associated with HER2 overexpression nor with the clinical characteristics of HER2-positive breast cancer., Summary: HER2 gene amplification is a complex event: it includes coamplification of other, potentially oncogenic genes and facilitates the generation of additional genomic aberrations. Further studies on these genotypic findings will be helpful to better identify the patients that might benefit from trastuzumab therapy.

Dermatofibrosarcoma protuberans (DFSP) is a soft tissue neoplasm of intermediate malignancy that is initially localized to the skin from where it can invade deep structures (fat, fascia, muscle and bone). It is the most frequent fibrohistiocytic tumor, comprising approximately 1.8 % of all soft tissue sarcomas and 0.1 % of all cancers. It has an estimated incidence of 0.8-5 cases per one million persons per year. Treatment of localized disease consists in complete surgical excision of the lesion by conventional surgery with wide margins (>3 cm) or by micrographic Mohs surgery. Although the cases of metastatic DFSP do not reach 5 % of the total, almost all of them appear after previous local relapses. The prognosis for metastatic cases is very poor with a survival of less than 2 years following detection of metastatic disease. Patients with locally advanced DFSP are not candidates for an initial radical surgical therapy therefore neoadyuvant treatment is required prior to surgery in order to reduce tumor burden. In this regard, chemotherapy and radiotherapy have not been highly efficacious so it is necessary to consider new alternatives. The demonstration of the oncogenic power of the translocation COL1A1-PDGFB in DFSP has allowed the successful introduction of drug therapy with antagonists of the PDGFB receptor for metastatic or locally advanced cases.

Rare cases of acute promyelocytic leukemia (APL) are associated with a t(11;17) translocation and a PLZF-RARalpha fusion transcript. Because of molecular specificities of the fusion protein, ATRA efficiency is often reduced in these cases. We present herein the case of an 83-year old patient which has been successfully treated by ATRA and Daunorubicin. The described quantitative RT-PCR method allowed successful monitoring and confirmation of the molecular response.

Complex variant 15;17 translocations are increasingly recognized in acute promyelocytic leukemia (APL). We report the case of a 47-year-old woman with APL harboring a novel four-way translocation. She presented with persistent bleeding after a tooth extraction. Blood cell counts at admission were hemoglobin at 9.0 g/dL, platelets at 15 x 10(9)/L, and white blood cells at 0.460 x 10(9)/L with abnormal promyelocytes. Most nucleated cells of bone marrow aspirates were abnormal promyelocytes with Auer rods. Chromosome analysis of unstimulated bone marrow cell cultures revealed a variant t(15;17) in the form of t(10;17;15;22)(q22;q21;q22;q11.2). Fluorescence in situ hybridization with a PML/RARA dual-color DNA probe showed the fusion signals on der(15) and the residual PML signals on der(22). RT-PCR showed long-form PML/RARA fusion transcripts. A complete remission was attained with a course of conventional chemotherapy including ATRA. A literature review revealed that our case is one of the very rare four-way translocations and the first report of the involvement of chromosomes 10 and 22 in a variant t(15;17).