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Cancer Cytogenetics

"A subdiscipline of genetics which deals with the cytological and molecular analysis of the CHROMOSOMES, and location of the GENES on chromosomes, and the movements of chromosomes during the CELL CYCLE." (MeSH 2013)

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Cytogenetics Organisations and Resources
Recent Research Publications
Fluorescent in Situ Hybridization (FISH)
Comparative Genomic Hybridization (CGH)

Cytogenetics Organisations and Resources (8 links)

Recent Research Publications

Consoli ML, Romano A, Parrinello NL, et al.
Unusual Karyotype in Acute Myelomonocitic Leukemia: A Case Report.
Anticancer Res. 2019; 39(8):4329-4332 [PubMed] Related Publications
BACKGROUND/AIM: Acute myeloid leukemia is well characterized by chromosomal aberrations that correspond to various subtypes of acute leukemias. The t(8;21)(q22;q22) is a frequent chromosomal abnormality strongly associated with acute myeloblastic leukemia with maturation (AML-M2), but is rarely associated with other subtypes. Translocation involving a third chromosome could produce new genetic rearrangements that lead to leukemogenesis.
PATIENTS AND METHODS: Conventional cytogenetic analysis and fluorescence in situ hybridization (FISH) were performed to identify the karyotype. Reverse transcriptase polymerase chain reaction (RT-PCR) was used to detect the AML1/ETO transcript.
RESULTS/CONCLUSION: We herein report a novel rearrangement with a three-way translocation involving chromosomes 8, 21 and another unknown chromosome, in an 83-year-old female patient diagnosed as AML-M4, with an ALM1/ETO negative transcript. This is an uncommon case of AML-M4 with three-way translocation in a new variant of t(8;21) acute myeloid leukaemia. The detailed mechanism of different phenotype expression is unclear. Further study is needed to identify the leukemogenetic transformation resulting from t(8;21) translocation.

Palumbo C, Furlan M, Balzarini P, et al.
Long-term Prognostic Impact of Chromosome Abnormalities in Clear Cell Renal Cell Carcinoma.
Anticancer Res. 2019; 39(6):2757-2765 [PubMed] Related Publications
BACKGROUND/AIM: Clear cell renal cell carcinoma (ccRCC) shows variable chromosomal abnormalities. The aim of this study was to assess the prognostic role of ccRCC chromosomal abnormalities in a single-center cohort with an extended follow-up.
MATERIALS AND METHODS: A systematic cytogenetic analysis was performed in 283 consecutive surgically-treated patients for renal masses between 1997 and 2002. Kaplan-Meier and multivariable Cox regression (MCR) models were used to calculate cancer specific survival (CSS).
RESULTS: Among 174 ccRCC patients, the most common abnormality was deletion in chromosome 3 (54.6%). At a median follow-up of 119 months, 38 patients (21.8%) died from RCC. At MCR models, worse CSS was independently predicted by deletions in chromosomes 2, 19, 20 or 22 and insertions in chromosome 18.
CONCLUSION: Specific ccRCC chromosomal abnormalities are independently associated with worse CSS. Cytogenetic evaluation may direct further genetic analysis for personalized prognostic stratification.

Cortes Barrantes P, Jakobiec FA, Dryja TP
A Review of the Role of Cytogenetics in the Diagnosis of Orbital Rhabdomyosarcoma.
Semin Ophthalmol. 2019; 34(4):243-251 [PubMed] Related Publications
Rhabdomyosarcoma (RMS) is the most common sarcoma of childhood and adolescence. Approximately 10% arise in the orbit, where the embryonal type is most common variant. The alveolar variant is less frequent and has a worse prognosis. Cytogenetic studies have revealed that most alveolar rhabdomyosarcomas have translocations involving the PAX and the FOX01 genes, giving rise to fusion genes that contribute to lack of differentiation and proliferation of the tumor cells. However, approximately 20% of alveolar rhabdomyosarcomas lack translocations and have been found to behave more similarly to embryonal cases. Histopathology remains the basis of diagnosis, but cytogenetic features and molecular signatures are becoming part of the routine analysis of RMS, since they determine not only prognosis, but also management and treatment regimens. A comprehensive review of the recent published literature in relation to orbital rhabdomyosarcomas and their cytogenetic features as well as clinical and therapeutic implications will be discussed.

Du N, Bao W, Zhang K, et al.
Cytogenetic characterization of the malignant primitive neuroectodermal SK-PN-DW tumor cell line.
BMC Cancer. 2019; 19(1):412 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: The SK-PN-DW cell line was established in 1979 and is commercially available. Despite the use of this cell line as an in vitro model for functional and therapeutic studies of malignant primitive neuroectodermal tumor (PNET), there is a lack of complete information about the genetic alterations that are present at the cytogenetic level. Thus, the current study aimed to characterize the cytogenetic profile of this cell line.
METHODS: Routine G-banded chromosome analysis, fluorescence in situ hybridization, and oligonucleotide array comparative genomic hybridization assays were performed to characterize the chromosomal changes in this cell line.
RESULTS: The G-banded karyotype analysis showed that the number of chromosomes in this cell line ranged between 36 and 41. Importantly, all cells displayed a loss of chromosomes Y, 11, 13, and 18. However, some cells showed an additional loss of chromosome 10. Additionally, the observed structural changes indicated: a) unbalanced translocation between chromosomes 1 and 7; b) translocation between chromosomes 11 and 22 at breakpoints 11q24 and 22q12, which is a classical translocation that is associated with Ewing sarcoma; c) a derivative chromosome due to a whole arm translocation between chromosomes 16 and 17 at likely breakpoints 16p10 and 17q10; and d) possible rearrangement in the short arm of chromosome 18. Moreover, a variable number of double minutes were also observed in each metaphase cell. Furthermore, the microarray assay results not only demonstrated genomic-wide chromosomal imbalance in this cell line and precisely placed chromosomal breakpoints on unbalanced, rearranged chromosomes, but also revealed information about subtle chromosomal changes and the chromosomal origin of double minutes. Finally, the fluorescence in situ hybridization assay confirmed the findings of the routine cytogenetic analysis and microarrays.
CONCLUSION: The accurate determination of the cytogenetic profile of the SK-PN-DW cell line is helpful in enabling the research community to utilize this cell line for future identity and comparability studies, in addition to demonstrating the utility of the complete cytogenetic profile, as a public resource.

Zemanová Z, Michalová K, Březinová J
Implication of cytogenetic and molecular cytogenetic analysis in diagnosis of hematological malignancies in the era of the new sequencing techniques.
Cas Lek Cesk. 2019; 158(1):22-27 [PubMed] Related Publications
In patients with hematological malignancies one of the most substantial findings is the karyotype of bone marrow cells at the time of diagnosis. The detection of clonal chromosome aberrations in diagnostic samples not only confirms a neoplastic or premalignant process but also provides important diagnostic and prognostic information essential for precise disease classification and choice of suitable therapy. Karyotype analysis during the disease course also allows monitoring of the treatment success reflected as well in the revised WHO classification where patients are often classified into the different diagnostic subtypes based on the finding of specific chromosome and/or genetic changes. Recently, also increases the number of advanced treatment approaches that directly or indirectly target the genetic aberrations present in tumor cells. Despite the large development of new sequencing technologies in recent years, cytogenetic analysis supplemented by the molecular cytogenetic methods still remains a very important part of diagnostics of hematological malignancies.

Rekhi B, Mridha A, Kattoor J
Small round cell lesions of the bone: Diagnostic approach, differential diagnoses and impact on treatment.
Indian J Pathol Microbiol. 2019 Apr-Jun; 62(2):199-205 [PubMed] Related Publications
Small round cell lesions of the bone encompass a heterogeneous group of tumors and tumor-like lesions, including Ewing sarcoma, small cell osteosarcoma, mesenchymal chondrosarcoma, neuroblastoma, non-Hodgkin's lymphoma (NHL), "Ewing-like" undifferentiated round cell sarcomas, metastasizing small cell carcinoma, along with plasma cell dyscrasia and Langerhan's cell histiocytosis. At the same time, there are tumor mimics, for example, chronic osteomyelitis, which has overlapping radiologic features with Ewing sarcoma and a primary intraosseous NHL. An exact diagnosis necessitates integration of clinical, radiologic, pathologic, and ancillary test results, including immunohistochemical and molecular results. Currently, there are several immunohistochemical markers and specific molecular signatures, driving most of these tumors, available, for an exact diagnosis. This review focuses on a pragmatic approach towards uncovering specific small round cell lesions of the bone, emphasizing upon integration of traditional morphology with ancillary techniques, including immunohistochemical markers and molecular techniques, the latter, especially in cases of Ewing sarcoma, Ewing-like undifferentiated round cell sarcoma, mesenchymal chondrosarcoma, and neuroblastoma. Subsequent to the diagnostic approach, including an impact on treatment, individual intraosseous round cell lesions have been described in detail. The references include updated articles from PUBMED.

Schittenhelm MM, Walter B, Tsintari V, et al.
Alternative splicing of the tumor suppressor ASPP2 results in a stress-inducible, oncogenic isoform prevalent in acute leukemia.
EBioMedicine. 2019; 42:340-351 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: Apoptosis-stimulating Protein of TP53-2 (ASPP2) is a tumor suppressor enhancing TP53-mediated apoptosis via binding to the TP53 core domain. TP53 mutations found in cancers disrupt ASPP2 binding, arguing for an important role of ASPP2 in TP53-mediated tumor suppression. We now identify an oncogenic splicing variant, ASPP2κ, with high prevalence in acute leukemia.
METHODS: An mRNA screen to detect ASPP2 splicing variants was performed and ASPP2κ was validated using isoform-specific PCR approaches. Translation into a genuine protein isoform was evaluated after establishing epitope-specific antibodies. For functional studies cell models with forced expression of ASPP2κ or isoform-specific ASPP2κ-interference were created to evaluate proliferative, apoptotic and oncogenic characteristics of ASPP2κ.
FINDINGS: Exon skipping generates a premature stop codon, leading to a truncated C-terminus, omitting the TP53-binding sites. ASPP2κ translates into a dominant-negative protein variant impairing TP53-dependent induction of apoptosis. ASPP2κ is expressed in CD34+ leukemic progenitor cells and functional studies argue for a role in early oncogenesis, resulting in perturbed proliferation and impaired induction of apoptosis, mitotic failure and chromosomal instability (CIN) - similar to TP53 mutations. Importantly, as expression of ASPP2κ is stress-inducible it defines a novel class of dynamic oncogenes not represented by genomic mutations.
INTERPRETATION: Our data demonstrates that ASPP2κ plays a distinctive role as an antiapoptotic regulator of the TP53 checkpoint, rendering cells to a more aggressive phenotype as evidenced by proliferation and apoptosis rates - and ASPP2κ expression results in acquisition of genomic mutations, a first initiating step in leukemogenesis. We provide proof-of-concept to establish ASPP2κ as a clinically relevant biomarker and a target for molecule-defined therapy. FUND: Unrestricted grant support from the Wilhelm Sander Foundation for Cancer Research, the IZKF Program of the Medical Faculty Tübingen, the Brigitte Schlieben-Lange Program and the Margarete von Wrangell Program of the State Ministry Baden-Wuerttemberg for Science, Research and Arts and the Athene Program of the excellence initiative of the Eberhard-Karls University, Tübingen.

Mizuno Y, Chinen Y, Tsukamoto T, et al.
A novel method of amplified fluorescent in situ hybridization for detection of chromosomal microdeletions in B cell lymphoma.
Int J Hematol. 2019; 109(5):593-602 [PubMed] Related Publications
Chromosomal microdeletions frequently cause loss of prognostically relevant tumor suppressor genes in hematologic malignancies; however, detection of minute deletions by conventional methods for chromosomal analysis, such as G-banding and fluorescence in situ hybridization (FISH), is difficult due to their low resolution. Here, we describe a new diagnostic modality that enables detection of chromosomal microdeletions, using CDKN2A gene deletion in B cell lymphomas (BCLs) as an example. In this method, which we refer to as amplified-FISH (AM-FISH), a 31-kb fluorescein isothiocyanate (FITC)-conjugated DNA probe encoding only CDKN2A was first hybridized with the chromosome, and then labeled with Alexa Fluor 488-conjugated anti-FITC secondary antibody to increase sensitivity. CDKN2A signals were equally identifiable by AM-FISH and conventional FISH in normal mononuclear blood cells. In contrast, when two BCL cell lines lacking CDKN2A were analyzed, CDKN2A signals were not detected by AM-FISH, whereas conventional FISH yielded false signals. Furthermore, AM-FISH detected CDKN2A deletions in two BCL patients with 9p21 microdeletions, which were not detected by conventional FISH. These results suggest that AM-FISH is a highly sensitive, specific, and simple method for diagnosis of chromosomal microdeletions.

Ridolfi M, Paudice M, Salvi S, et al.
Agar pre-embedding of small skin biopsies: real-life benefits and challenges in high throughput pathology laboratories.
J Clin Pathol. 2019; 72(6):448-451 [PubMed] Related Publications
Paraffin embedding of small, thin tissue samples requires specific expertise for optimal orientation before tissue sectioning. This study evaluates the real-life utility of the agar pre-embedding technique for small skin biopsies with regards to lengthening of work times, problems in orientation (re-embedding) and ancillary techniques (immunohistochemistry and in situ hybridisation) between two high work flow pathology laboratories, one of which routinely uses the agar pre-embedding technique and one which does not. The mean time required for pre-embedding in agar was 30.4 s, but time for paraffin embedding for agar pre-embedded samples was shorter than the traditional method (177 vs 296 s; p<0.005). The number of skin samples requiring re-embedding was significantly higher with the traditional embedding method (p<0.005). No problems in immunoreactivity were observed in all 1900 reactions performed with 17 different antibodies. Fluorescence in situ hybridisation analysis was optimised with a prolonged protease K incubation time (21 vs 18 min).

Santos MFM, Oliveira FCAC, Kishimoto RK, et al.
Pre-analytical parameters associated with unsuccessful karyotyping in myeloid neoplasm: a study of 421 samples.
Braz J Med Biol Res. 2019; 52(2):e8194 [PubMed] Free Access to Full Article Related Publications
Cytogenetics is essential in myeloid neoplasms (MN) and pre-analytical variables are important for karyotyping. We assessed the relationship between pre-analytical variables (time from collection to sample processing, material type, sample cellularity, and diagnosis) and failures of karyotyping. Bone marrow (BM, n=352) and peripheral blood (PB, n=69) samples were analyzed from acute myeloid leukemia (n=113), myelodysplastic syndromes (n=73), myelodysplastic syndromes/myeloproliferative neoplasms (n=17), myeloproliferative neoplasms (n=137), and other with conclusive diagnosis (n=6), and reactive disorders/no conclusive diagnosis (n=75). The rate of unsuccessful karyotyping was 18.5% and was associated with the use of PB and a low number of nucleated cells (≤7×103/µL) in the sample. High and low cellularity in BM and high and low cellularity in PB samples showed no metaphases in 3.9, 39.7, 41.9, and 84.6% of cases, respectively. Collecting a good BM sample is the key for the success of karyotyping in MN and avoids the use of expensive molecular techniques.

Giefing M, Siebert R
FISH and FICTION in Lymphoma Research.
Methods Mol Biol. 2019; 1956:249-267 [PubMed] Related Publications
Fluorescence in situ hybridization (FISH) is a powerful and robust technique allowing the visualization of target sequences like genes in interphase nuclei. It is widely used in routine diagnostics to identify cancer-specific aberrations including lymphoma-associated translocations or gene copy number changes in single tumor cells. By combining FISH with immunophenotyping-a technique called fluorescence immunophenotyping and interphase cytogenetic as a tool for investigation of neoplasia (FICTION)-it is moreover possible to identify a cell population of interest. Here we describe standard protocols for FISH and FICTION as used in our laboratories in diagnosis and research.

Lin T, Jin H, Gong L, et al.
Surveillance of non-muscle invasive bladder cancer using fluorescence in situ hybridization: Protocol for a systematic review and meta-analysis.
Medicine (Baltimore). 2019; 98(7):e14573 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: To evaluate the diagnostic effectiveness and predictive value of fluorescence in situ hybridization (FISH) in the surveillance of non-muscle invasive bladder cancer (NMIBC).
METHODS: PubMed/Medline, EMBASE, Web of Science, Ovid, Web of Knowledge, and Cochrane Library will be searched for studies related to the topic. The identification, inclusion, and exclusion flowcharts will be conducted according to preferred reporting items for systematic reviews and meta-analysis guidelines. The identified reports will be critically appraised according to the Newcastle-Ottawa scale, quality assessment of diagnostic accuracy studies-2 and standards for reporting of diagnostic accuracy 2015. Forest plots will be generated to display hazard ratios, sensitivities, and specificities. Pooled estimates with their 95% confidence intervals will be calculated using the bivariate model, the hierarchical summary receiver operating characteristic model and a fixed- or random-effects model.
RESULTS: This study will provide evidence and data to form a comprehensive understanding of the value of FISH in the surveillance of NMIBC.
CONCLUSION: The diagnostic efficacy of FISH will be affected by post-therapy factors. However, FISH still could facilitate the surveillance of NMIBC owing to its non-invasive feature. This study will improve the clinical decision-making and enlighten the future research of NMIBC.

Liu ZH, Wang K, Lin DY, et al.
Impact of the updated 2018 ASCO/CAP guidelines on HER2 FISH testing in invasive breast cancer: a retrospective study of HER2 fish results of 2233 cases.
Breast Cancer Res Treat. 2019; 175(1):51-57 [PubMed] Related Publications
OBJECTIVES: Human epidermal growth factor receptor 2 (HER2, ERBB2) is a valuable prognostic and predictive biomarker in breast cancer. Accurate assessment of HER2 status is essential in selecting the patients with invasive breast cancer who will likely response to HER2-targeted therapies. Some major modifications in the diagnostic recommendation for fluorescence in situ hybridization (FISH) have been made in the updated 2018 American Society of Clinical Oncology (ASCO)/College of American Pathologist (CAP) guideline. According to the revised guideline, concomitant IHC assays are required to arrive at the most accurate HER2 status designation after HER2 FISH equivocal results; however, little is known about its influence on the clinical practice of pathologist. The purpose of this study was to evaluate the impact of the revised 2018 ASCO/CAP guidelines on the HER2 status designation.
METHODS: We retrospectively reviewed the HER2 FISH testing results from 2233 cases of invasive breast cancer between January 2014 and December 2017. Concomitant immunohistochemistry (IHC) were performed on the same tissue blocks that were used for the FISH testing.
RESULTS: Compared to the 2013 guidelines, the HER2 status in 183 (8.2%) cases were re-defined when reassessed by the 2018 guidelines. Among these 183 cases, 175 equivocal cases according to the 2013 guideline were re-defined as HER2 negative (n = 173) or HER2 positive (n = 2). Eight previously classified as HER2 positive cases were converted to negative in the 2018 scheme, all of which were with HER2 IHC scores of 1+ or 2+. The number of cases in the negative category was 1705 according to the 2018 guidelines as opposed to 1524 by the 2013 guidelines.
CONCLUSIONS: The updated 2018 ASCO/CAP guidelines eliminated the FISH equivocal category, which can be attributed to reflex HER2 IHC, and partly ease the dilemma for clinical practice. Reflex IHC for FISH equivocal cases is of prime importance; furthermore, HER2 FISH results were converted from positivity to negativity based on the concomitant IHC results in a small percentage of cases. In all, implementation of the 2018 ASCO/CAP guidelines provides much clearer instructions and recommendations for the HER2 status designation, and thus reduces the risk of misdiagnosis.

Liu K, Yu Y, Chai X, et al.
Atypical Acute Monocytic Leukemia: a Lymphoma Presentation and Association with Complex Karyotype.
Ann Clin Lab Sci. 2018; 48(6):805-807 [PubMed] Related Publications
A 71-year-old woman presented with acute monocytic leukemia with lymphoma-like morphologic features and an unusual complex karyotype. Bone marrow (BM) aspiration revealed acute monocytic leukemia with lymphoma-like morphologic features. Thus, we called the condition a lymphoma presentation. The patient was diagnosed with acute monocytic leukemia (AML FAB M5a) according to the French-American-British (FAB) classification and the new World Health Organization (WHO) classification [1]. To our knowledge, this is the first reported case of an unusual complex karyotype in acute monocytic leukemia with a lymphoma presentation and adds to the expanding list of karyotypic abnormalities seen in acute monocytic leukemia. We present the case given its rarity, occasional misdiagnosis and poor prognosis. Whether this complex karyotype resulted in such lymphoma-like morphologic features remains to be determined. The case illustrates the importance of the morphologic features cognition and avoiding misdiagnosis. Clinical trials are available to further explore how to extend survival time and contribute to a better prognosis for patients suffering from acute monocytic leukemia with a complex karyotype.

Jin H, Lin T, Hao J, et al.
A comprehensive comparison of fluorescence in situ hybridization and cytology for the detection of upper urinary tract urothelial carcinoma: A systematic review and meta-analysis.
Medicine (Baltimore). 2018; 97(52):e13859 [PubMed] Free Access to Full Article Related Publications
OBJECTIVE: To compare the relative effectiveness of fluorescence in situ hybridization (FISH) and cytology in diagnosing upper urinary tract urothelial carcinoma (UUT-UC) and to evaluate the advantages and potential deficiencies of FISH analysis.
METHODS: We performed a complete systematic review based on studies from PubMed/Medline, Embase, Web of Science, Ovid, Web of Knowledge, and Cochrane Library. We identified 2031 patients with strict criteria in 14 individual studies between January 2005 to November 2017 in accordance to preferred reporting items for systematic reviews and meta-analysis (PRISMA) guidelines, we summarized the test performance using bivariate random effects models.
RESULTS: FISH was superior to cytology in terms of pooled sensitivities (84.0%, 95% confidence interval [CI] 74.4-90.5% vs 40.0%, 95% CI 33.6-46.7%). FISH and cytology were similar to each other in terms of pooled specificities, which were 89.5% (95% CI 85.3-92.6%) for FISH and 95.9% (95% CI 91.2-98.1%) for cytology.
CONCLUSION: We confirm the superiority of FISH over cytology in terms of sensitivity and find similar diagnostic outcomes between them based on systematic analysis. Therefore, we demonstrate that FISH is extremely sensitive while still very reliable with a relatively low error rate for diagnosing UUT-UC.

Freund JE, Liem EIML, Savci-Heijink CD, de Reijke TM
Fluorescence in situ hybridization in 1 mL of selective urine for the detection of upper tract urothelial carcinoma: a feasibility study.
Med Oncol. 2018; 36(1):10 [PubMed] Free Access to Full Article Related Publications
Kidney-sparing surgery of upper tract urothelial carcinoma (UTUC) requires a stringent follow-up with frequent ureteroscopies. Triage testing could reduce the number of follow-up ureteroscopies and hence minimize the invasiveness of follow-up. The use of urine-based markers for triage seems appealing but should be feasible with selective urine from outpatient cystoscopy to maximize the reduction of invasiveness. In this study, the feasibility of UroVysion

El-Menoufy MAM, Mourad ZI, Farahat NM
The prognostic impact of loss of chromosome 7 material detected by fluorescence in situ hybridization (FISH) in myeloid malignancies.
J Egypt Natl Canc Inst. 2018; 30(4):133-138 [PubMed] Related Publications
BACKGROUND: Monosomy 7 (-7) or deletion in its long arm [del(7q)] is among the most common chromosomal abnormalities in myeloid malignancies. There are prognostic variations between -7 and del(7q) in acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS).
AIM: To describe the clinical characteristics, response to treatment, and survival of patients with primary AML and MDS having -7 or del(7q) detected by fluorescence in situ hybridization (FISH).
PATIENTS AND METHODS: The study was conducted on 53 patients with primary AML and MDS. They were tested for chromosome 7 abnormality using FISH technique.
RESULTS: Thirty-one patients had chromosome 7 abnormality and 22 did not. Lower complete remission and higher death rates were observed in patients with -7 (47.6% and 62%, respectively) when compared to patients with del(7q) (70% and 40%, respectively) with no significant difference (p = 0.218 and 0.101, respectively). The median overall survival (OS) of patients with -7, del(7q) and normal chromosome 7 were 32.0, 43.0 and 50.0 months, respectively, with significant statistical difference (p = 0.001). This difference was evident between patients with -7 and those with normal chromosome 7 (p = 0.001), and less evident between patients with -7 and those with del(7q) (p = 0.021).
CONCLUSION: Chromosome 7 analysis has clear impact on the outcome of myeloid malignancies. The prognostic variations between -7 and del(7q) is attributed to multiple factors. Cases with del(7q) have better outcome than cases with -7. FISH provides a powerful tool for detecting and monitoring patients with chromosome 7 abnormalities.

Xu B, Shen J, Guo W, et al.
Impact of the 2018 ASCO/CAP HER2 guidelines update for HER2 testing by FISH in breast cancer.
Pathol Res Pract. 2019; 215(2):251-255 [PubMed] Related Publications
Recently, the American Society of Clinical Oncology/College of American Pathologists (ASCO/CAP) updated the guidelines on HER2 testing for invasive breast cancer. Little is known about the impact of the guidelines update. We aimed to study the impact of the 2018 ASCO/CAP HER2 testing guidelines update. We compared the HER2 FISH results interpreted by 2013 and 2018 ASCO/CAP guidelines in 331 cases of invasive breast cancers. We also analyzed the pathological features and clinical outcomes of these cases. In comparing to the 2013 ASCO/CAP guidelines, the HER2 negative rate was increased significantly from 62.5% to 75.8%(P < 0.05), and 13.3% changed from equivocal to negative by the 2018 guidelines. Our findings indicate that the guidelines update significantly increased the rate of negative results. The reclassification of the equivocal results by the 2018 guidelines is the main reason for this change. Patients with HER2 equivocal results were associated with larger tumor size and higher Ki67 index than those with negative results, while clinical outcomes were similar between them.

McHale T, Ohori NP, Cieply KM, et al.
Comparison of urinary cytology and fluorescence in situ hybridization in the detection of urothelial neoplasia: An analysis of discordant results.
Diagn Cytopathol. 2019; 47(4):282-288 [PubMed] Related Publications
BACKGROUND: We examine the performance of cytology and FISH in the detection of urothelial carcinoma (UC), and explore the reasons for discrepant results, and potential clinical implications.
METHODS: Urine samples from 89 patients were prospectively collected for simultaneous cytology and UroVysion FISH, and results correlated with concurrent biopsies and/or clinical or histologic follow-up data. Corresponding tissue biopsies, where available, were also evaluated by FISH.
RESULTS: Sensitivity and specificity of cytology and FISH for the detection of UC was 54.8% and 92% and 50% and 88%, respectively. Only one of seven false-positive urinary FISH results proved to be an "anticipatory positive" on extended follow-up. Five of eight (62.5%) high grade (HG) carcinomas with false-negative urinary FISH, were negative due to the absence/paucity of FISH-detectable changes in the tumor cells. In atypical cytology cases, the FISH result did not assist in identifying UC. There was no significant difference between an atypical cytology result and a positive FISH result, with respect to the identification of patients with UC.
CONCLUSIONS: We found urinary cytology to be more sensitivity and specific than FISH in the detection of UC, though the difference was not statistically significant. Up to 24% of HG UCs are FISH negative due to an absence of FISH-detectable abnormalities in the tumor cells. Paucity of neoplastic cells in the urine also contributes to false-negative FISH results in both HG and low grade tumors. Negative urinary FISH cannot be taken alone as indicating the absence of significant disease in patients with atypical cytology.

Bager N, Juul-Dam KL, Sandahl JD, et al.
Complex and monosomal karyotype are distinct cytogenetic entities with an adverse prognostic impact in paediatric acute myeloid leukaemia. A NOPHO-DBH-AML study.
Br J Haematol. 2018; 183(4):618-628 [PubMed] Related Publications
Data on occurrence, genetic characteristics and prognostic impact of complex and monosomal karyotype (CK/MK) in children with acute myeloid leukaemia (AML) are scarce. We studied CK and MK in a large unselected cohort of childhood AML patients diagnosed and treated according to Nordic Society for Paediatric Haematology and Oncology (NOPHO)-AML protocols 1993-2015. In total, 800 patients with de novo AML were included. CK was found in 122 (15%) and MK in 41 (5%) patients. CK and MK patients were young (median age 2·1 and 3·3 years, respectively) and frequently had FAB M7 morphology (24% and 22%, respectively). Refractory disease was more common in MK patients (15% vs. 4%) and stem cell transplantation in first complete remission was more frequent (32% vs. 19%) compared with non-CK/non-MK patients. CK showed no association with refractory disease but was an independent predictor of an inferior event-free survival (EFS; hazard ratio [HR] 1·43, P = 0·03) and overall survival (OS; HR 1·48, P = 0·01). MK was associated with a poor EFS (HR 1·57, P = 0·03) but did not show an inferior OS compared to non-MK patients (HR 1·14, P = 0·62). In a large paediatric cohort, we characterized AML with non-recurrent abnormal karyotype and unravelled the adverse impact of CK and MK on prognosis.

Meyer RG, Van Dyke DL
Analysis of Common Abnormalities Seen in Chronic Lymphocytic Leukemia Using Fluorescence In Situ Hybridization.
Methods Mol Biol. 2019; 1881:35-49 [PubMed] Related Publications
Since fluorescence in situ hybridization (FISH) was used to define a prognostic heierarchy for chronic lymphocytic leukemia (CLL) in 2000, the method has been employed widely in cytogenetics laboratories worldwide. This chapter describes techniques and trouble-shooting to maximize the efficiency of microscope slide preparation for FISH analysis in CLL.

Brajkovic S, Pelz B, Procopio MG, et al.
Microfluidics-based immunofluorescence for fast staining of ALK in lung adenocarcinoma.
Diagn Pathol. 2018; 13(1):79 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: Anaplastic lymphoma kinase (ALK) is a key oncogenic driver in lung adenocarcinoma patients and its fusion proteins are routinely assessed. The microfluidic tissue processor (MTP) device is based on a chip-confined low-volume technology allowing for rapid immunohistochemistry/immunofluorescence (IHC/IF) stainings of formalin-fixed paraffin-embedded (FFPE) or frozen tissue samples.
METHODS: A novel ALK IF protocol was developed for the MTP device using the primary mouse anti-human ALK antibody clone 5A4. FFPE tumor whole sections from 14 resected lung adenocarcinoma patients documented to be ALK positive (ALK+) by automated chromogenic IHC and/or FISH were used. MTP-derived IF immunoreactivity was measured by computerized analysis of digitalized images on individual frames of tumor epithelia and surrounding stroma, using an ImageJ plug-in.
RESULTS: The 5A4 antibody yielded saturated immunoreactivity at an incubation time of 4 min on a titration curve ranging from 2 to 32 min. Total staining time on the MTP device was 18 min including secondary IgG Alexa Fluor 647. MTP-based ALK IF confirmed all 12 cases; with epithelial signal above stromal staining based on computerized pixel-based measurement. MTP-IF (mean intensity levels 458 to 1301) and chromogenic IHC (H-score 120 to 300) showed an equal range of variation of 2.8 and 2.5 folds, respectively, and a trend for direct correlation (p-value 0.051).
CONCLUSION: The newly developed protocol for immunofluorescent detection of ALK protein with the MTP device confirms chromogenic IHC results on FFPE lung adenocarcinoma specimens. MTP-based IF is fast and reliable. We foresee this study to be a first step opening the road for further realization of microfluidic-based assays for rapid simultaneous detection of targetable oncogenic and immune-system related markers in their topographical context to investigate tumour heterogeneity and micro-environmental interactions.

Lupicki K, Elifio-Esposito S, Fonseca AS, et al.
Patterns of copy number alterations in primary breast tumors of South African patients and their impact on functional cellular pathways.
Int J Oncol. 2018; 53(6):2745-2757 [PubMed] Related Publications
Breast cancer is the most common and the leading cause of female mortality among South African (SA) women. Several non‑biological and biological risk factors may be attributed to their observed high mortality rate; however, the molecular profiles associated with their breast tumors are poorly characterized. The present study examined the patterns of genome-wide copy number alterations (CNAs) and their potential impact on functional cellular pathways targeted by cancer driver genes in patients with breast cancer from the Western Cape region of SA. Array-comparative genomic hybridization analysis, performed in 28 cases of invasive breast cancer, revealed a mean number of 8.68±6.18 CNAs per case, affecting primarily the Xp22.3 and 6p21-p25 cytobands (57.14% of the cases), followed by 19p13.3-p13.11 (35.7%), 2p25.3-p24.3, 4p16.3-p15.3, 8q11.1-q24.3 and 16 p13.3-p11.2 (32.14%). Functional enrichment analysis of genes and microRNA targets mapped in these affected cytobands revealed critical cancer-associated pathways, including fatty acid biosynthesis and metabolism, extracellular matrix-receptor interaction, hippo and tumor protein p53 signaling pathways, which are regulated by known cancer genes, including CCND1, CDKN1A, MAPK1, MDM2, TP53 and SMAD2. An inverse correlation was observed among the number of CNAs and tumor size and grade; CNAs on the 4p and 6p cytobands were also inversely correlated with tumor grade. No association was observed in the number of CNAs and/or the affected cytobands and the different ethnic groups of the SA patients, indicating that their tumor genome is affected by CNAs, irrespectively of their genetic descent. Additional genomic tumor profiling in SA and other Sub-Saharan African patients with breast cancer is required to determine the associations of the CNAs observed with prognosis and clinical outcome.

Fanoni D, Corti L, Alberti-Violetti S, et al.
Array-based CGH of primary cutaneous CD8+ aggressive EPIDERMO-tropic cytotoxic T-cell lymphoma.
Genes Chromosomes Cancer. 2018; 57(12):622-629 [PubMed] Related Publications
Primary cutaneous CD8+ aggressive epidermotropic cytotoxic T-cell lymphoma (pcAECyTCL) is a rare provisionally categorized cutaneous lymphoma characterized by an aggressive course. Its pathogenesis and molecular mechanisms are still unknown, and only two individual cases have so far been molecularly characterized. The aim of this study was to define the pattern of numerical chromosomal alterations in tumor samples taken from 20 patients with pcAECyTCL at the time of diagnosis by means of array-comparative genomic hybridization (a-CGH). a-CGH detected numerous genomic aberrations in all the patients and, putting these together as a whole, they affected all the chromosomes. However, no specific profile of recurrent copy number alterations (CNAs) was found. Most of the gains involved regions previously described in other aggressive cutaneous lymphomas such as 7q, 8q24.3, and 17q, whereas the most significant CNA was the loss of 9p21.3 (CDKN2A-CDKN2B), which has already been found in a variety of malignant tumors and is associated with aggressive cutaneous T-cell lymphomas. In brief, CGH analysis revealed a large number of CNAs with only few recurring regions that probably do not represent driving events. The genomic instability found in this aggressive variant of cutaneous lymphoma may therefore be a secondary event but, at the time of the diagnosis of pcAECyTCL, the genomic integrity of tumor cells is already compromised.

Ikeri NZ, Akinjo AO, Ajayi OO, Banjo AA
An audit of the diagnosis and reporting of soft tissue sarcomas at the Lagos University Teaching Hospital.
Niger J Clin Pract. 2018; 21(10):1330-1336 [PubMed] Related Publications
Background: : The effective management of patients with cancer is predicated on the right diagnoses and other relevant parameters included in the pathology report. This is particularly important in soft tissue pathology where arriving at the right diagnosis is often challenging. The aim of this study, therefore, was to perform an audit of sarcoma diagnosis and reporting in our institution.
Methods: Slides of soft tissue sarcomas diagnosed in our institution over a 5-year period were reviewed with specialist soft tissue pathologists. Ancillary immunohistochemistry and fluorescent in situ hybridization were performed where necessary. The contents of the reports were assessed using a diagnostic checklist developed by the Association of Directors of Anatomic and Surgical Pathology.
Results: Fifty-five of the 62 patients studied (88.7%) were correctly identified as sarcomas. However, the correct diagnoses were made in only 27 patients (43.6%). Kaposi sarcoma and dermatofibrosarcoma protuberans were the most recognized sarcomas, while leiomyosarcoma, myxofibrosarcoma, and malignant peripheral nerve sheath tumor were the least recognized sarcomas. The most reported parameters included the histologic type (100%) and size (89.7%), while the percentage of necrosis (0%) and the stage (0%) were the least reported parameters.
Conclusion: A pattern based approach is important for the accurate diagnosis of soft tissue sarcomas. Some essential prognostic parameters and information needed for management were not included in the histopathology reports. The adoption of a structured reporting format and multidisciplinary team meetings will help to ensure the inclusion of such important information in the pathology report.

Döhner H, Dolnik A, Tang L, et al.
Cytogenetics and gene mutations influence survival in older patients with acute myeloid leukemia treated with azacitidine or conventional care.
Leukemia. 2018; 32(12):2546-2557 [PubMed] Free Access to Full Article Related Publications
Older patients with newly diagnosed acute myeloid leukemia (AML) in the phase 3 AZA-AML-001 study were evaluated at entry for cytogenetic abnormalities, and a subgroup of patients was assessed for gene mutations. Patients received azacitidine 75 mg/m

Puripat N, Loharamtaweethong K
Phosphohistone H3 (PHH3) as a surrogate of mitotic figure count for grading in meningiomas: a comparison of PHH3 (S10) versus PHH3 (S28) antibodies.
Virchows Arch. 2019; 474(1):87-96 [PubMed] Related Publications
Mitotic figure (MF) counting is important in the evaluation of meningioma grading. Nevertheless, mitosis assessment on hematoxylin and eosin (H&E)-stained slides may be problematic because of technical factors and pathologist's experience. Phosphohistone H3 (PHH3) is a mitosis-specific antibody that has proven to facilitate mitotic count in various tumors. However, the antibody performance between PHH3 serine10 (S10) and serine28 (S28) has never been compared in these tumors before. In this study, 48 cases of meningioma (28 grade I, 14 grade II, 6 grade III) were evaluated using immunohistochemical stains for four commercially available PHH3 (S10) and S28 antibodies to identify MFs and validate PHH3 intra- and interobserver reproducibility and agreement. Two pathologists counted MFs on both H&E- and PHH3-stained slides. H&E and PHH3 MFs were highly correlated (Spearman's rho = 0.96 for PHH3 (S10)-Biocare, 0.96 for PHH3 (S10)-CST, 0.91 for PHH3 (S28)-Abcam, and 0.89 for PHH3 (S28)-Santa Cruz. The mean difference between an H&E and PHH3 mitotic count is 0.81 for PHH3 (S10)-Biocare, 0.95 for PHH3 (S10)-CST, - 0.97 for PHH3 (S28)-Abcam, and - 0.97 for PHH3 (S28)-Santa Cruz. For comparison among four PHH3 antibodies, PHH3 mitotic counts had both a good intra- and interobserver reproducibility (p > 0.05). Regarding to World Health Organization (WHO) grade, there was not a significant discrepancy in the stratification of tumor grades for all four PHH3 antibodies in terms of interobserver agreement. The Cohen's kappa coefficient (K) was 0.93 for PHH3 (S10)-Biocare, 0.82 for PHH3 (S10)-CST, 0.76 for PHH3 (S28)-Abcam, and 0.80 for PHH3 (S28)-Santa Cruz. Considering survival analyses, all five proliferation indices were univariately associated with recurrences. Increased PHH3 mitotic indices (MIs) were significantly associated with recurrence-free survival in univariate Cox proportional hazards regression analysis (p < 0.001) and remained an independent predictor in multivariate analysis (p < 0.05). The appropriate prognostic cutoff values for recurrence prediction were 5 or more per 10 high-power fields (HPFs) for PHH3 (S10) and 3 or more per 10 HPFs for PHH3 (S28).

Bibi F, Ali I, Naseer MI, et al.
Detection of genetic alterations in gastric cancer patients from Saudi Arabia using comparative genomic hybridization (CGH).
PLoS One. 2018; 13(9):e0202576 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: The present study was conducted to discover genetic imbalances such as DNA copy number variations (CNVs) associated with gastric cancer (GC) and to examine their association with different genes involved in the process of gastric carcinogenesis in Saudi population.
METHODS: Formalin-fixed paraffin-embedded (FFPE) tissues samples from 33 gastric cancer patients and 15 normal gastric samples were collected. Early and late stages GC samples were genotyped and CNVs were assessed by using Illumina HumanOmni1-Quad v.1.0 BeadChip.
RESULTS: Copy number gains were more frequent than losses throughout all GC samples compared to normal tissue samples. The mean number of the altered chromosome per case was 64 for gains and 40 for losses, and the median aberration length was 679115bp for gains and 375889bp for losses. We identified 7 high copy gain, 52 gains, 14 losses, 32 homozygous losses, and 10 copy neutral LOHs (loss of heterozygosities). Copy number gains were frequently detected at 1p36.32, 1q12, 1q22, 2p11.1, 4q23-q25, 5p12-p11, 6p21.33, 9q12-q21.11, 12q11-q12, 14q32.33, 16p13.3, 17p13.1, 17q25.3, 19q13.32, and losses at 1p36.23, 1p36.32, 1p32.1, 1q44, 3q25.2, 6p22.1, 6p21.33, 8p11.22, 10q22.1, 12p11.22, 14q32.12 and 16q24.2. We also identified 2 monosomy at chromosome 14 and 22, 52 partially trisomy and 22 whole chromosome 4 neutral loss of heterozygosities at 13q14.2-q21.33, 5p15.2-p15.1, 5q11.2-q13.2, 5q33.1-q34 and 3p14.2-q13.12. Furthermore, 11 gains and 2 losses at 1p36.32 were detected for 11 different GC samples and this region has not been reported before in other populations. Statistical analysis confirms significant association of H. pylori infection with T4 stage of GC as compare to control and other stages.
CONCLUSIONS: We found that high frequency of copy number gains and losses at 1p36.23, 1p32.1, 1p36.32, 3q25.2, 6p21.33 and 16q24.2 may be common events in gastric cancer. While novel CNVs at 1p36.32 harbouring PRDM16, TP73 and TP73-AS1 genes showed 11 gains and 2 losses for 11 different GC cases and this region is not reported yet in Database of Genomic Variants may be specific to Saudi population.

Kesarkar K, Tamgadge A, Peirera T, et al.
Evaluation of Mitotic Figures and Cellular and Nuclear Morphometry of Various Histopathological Grades of Oral Squamous Cell Carcinoma: Comparative study using crystal violet and Feulgen stains.
Sultan Qaboos Univ Med J. 2018; 18(2):e149-e154 [PubMed] Free Access to Full Article Related Publications
Objectives: The objectives of this study were to quantitatively estimate the number of mitotic figures (MFs) and evaluate the cellular and nuclear features of various histological grades of oral squamous cell carcinoma (OSCC) using Feulgen and 1% crystal violet stains.
Methods: This case-control study took place at the Dr D. Y. Patil Dental College & Hospital in Mumbai, Maharashtra, India, between June and December 2016. A total of 51 samples were retrieved from the hospital archives. Of these, 15 well-differentiated, 15 moderately-differentiated and six poorly-differentiated OSCC samples formed the case group while 15 samples of normal gingival
Results: The number of MFs visible per field was significantly higher in Feulgen-stained sections as compared to those stained with crystal violet (
Conclusion: The Feulgen stain is more reliable than 1% crystal violet in terms of the selective staining of MFs. Moreover, the findings of this study indicate that computer-based morphometric analysis is an effective tool for differentiating between various grades of OSCC.

Elizalde S, Laughney AM, Bakhoum SF
A Markov chain for numerical chromosomal instability in clonally expanding populations.
PLoS Comput Biol. 2018; 14(9):e1006447 [PubMed] Free Access to Full Article Related Publications
Cancer cells frequently undergo chromosome missegregation events during mitosis, whereby the copies of a given chromosome are not distributed evenly among the two daughter cells, thus creating cells with heterogeneous karyotypes. A stochastic model tracing cellular karyotypes derived from clonal populations over hundreds of generations was recently developed and experimentally validated, and it was capable of predicting favorable karyotypes frequently observed in cancer. Here, we construct and study a Markov chain that precisely describes karyotypic evolution during clonally expanding cancer cell populations. The Markov chain allows us to directly predict the distribution of karyotypes and the expected size of the tumor after many cell divisions without resorting to computationally expensive simulations. We determine the limiting karyotype distribution of an evolving tumor population, and quantify its dependency on several key parameters including the initial karyotype of the founder cell, the rate of whole chromosome missegregation, and chromosome-specific cell viability. Using this model, we confirm the existence of an optimal rate of chromosome missegregation probabilities that maximizes karyotypic heterogeneity, while minimizing the occurrence of nullisomy. Interestingly, karyotypic heterogeneity is significantly more dependent on chromosome missegregation probabilities rather than the number of cell divisions, so that maximal heterogeneity can be reached rapidly (within a few hundred generations of cell division) at chromosome missegregation rates commonly observed in cancer cell lines. Conversely, at low missegregation rates, heterogeneity is constrained even after thousands of cell division events. This leads us to conclude that chromosome copy number heterogeneity is primarily constrained by chromosome missegregation rates and the risk for nullisomy and less so by the age of the tumor. This model enables direct integration of karyotype information into existing models of tumor evolution based on somatic mutations.

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