Cancer Genetics

Cytogenetic profile of a representative cohort of young adults with de novo acute myéloblastic leukaemia in Morocco Publication date: October 2019 Source: Cancer Genetics, Volume 238 Author(s): Nisrine Khoubila, Mounia Bendari, Nezha Hda, Mouna Lamchahab, Meryem Qachouh, Mohamed Rachid, Asmaa Quessar Abstract Background We analyzed the cytogenetic characteristics of a representative population of young adults with de novo acute myéloblastic leukemia (AML) treated in a single center institution. Methods Patients with de novo AML included were aged between 20 and 60 years. Cytogenetic analysis was done at diagnosis. Twenty cells were analyzed, although examination of lower numbers of metaphases was also acceptable if an abnormal clone was detected. Findings From 1/1/04 to 31/12/2014, among 1315 adult patients, 1055 (80%) patients were aged between 20 and 60 years. Karyotype was done in 927 (88%) patients and cytogenetic analysis failed in 32 cases (3·4%). Clonal abnormalities were observed in 520 (58%) patients. 175 (19·5%) were classified in the favorable group, 609 were in the intermediate group and 111 (12·5%) were in the adverse group. The most frequent chromosomal abnormality observed was t(8;21) in 112 (12·5%). Thirty three (3·7%) patients had t(15;17) and 30 (3·3%) had inv16, trisomy 8 was found in 47 (5·2%), 11q23 rearrangements in 32 (3·6%), 67 (7·4%) had a complex karyotype, −5/del(5q) and −7/del(7q) were seen in, respectively, 11(1%) and 27 (3%). Monosomy occurred in 115 (13%) patients, 70 (8%) responded to the definition of monosomal karyotypes. Interpretation This is the largest study done in Morocco, Africa and Middle East. Epidemiological studies involving different ethnic populations and geographic regions of the world should help unfold the true nature of environmental and genetic interplay in the development of AML. Our cytogenetic profile reveals some particularities when compared to other populations; it does not seem to be similar neither to eastern or western countries.

Down-regulation of STIP1 regulate apoptosis and invasion of glioma cells via TRAP1/AKT signaling pathway Publication date: September 2019 Source: Cancer Genetics, Volume 237 Author(s): Hongwei Yin, ZhiTong Deng, Xuetao Li, YanYan Li, Wenhao Yin, Guozhen Zhao, Dongyang Jiang, Chunming Sun, Youxin Zhou Abstract Background In recent years, many studies have confirmed that STIP1 (phosphorylation-induced protein 1) is involved in the development and progression of various tumors. However, its potential role in glioma progression and the underlying mechanisms of glioma development remain unclear. Methods We analyzed the expression of STIP1 in 35 human glioma tissue specimens of different grades, using 6 normal brain tissues for comparison. We transfected U87 and U251 cell lines with small interfering RNA (siRNA) to downregulate STIP1, and set up a negative control group and a blank group for comparison. The MTT assay was used to detect cell proliferation, and cell cycle progression and apoptosis were analyzed through flow cytometry. Transwell experiments were employed to detect the invasion and migration of STIP1-depleted and control U87 and U251 cells and western blotting was used to detect the expression of TRAP1/Akt pathway proteins. In addition, immunohistochemical analysis was used to reveal differences in expression and localization between transplanted tumor specimens of each group. Results We observed a high expression of STIP1 in glioblastoma, MTT assay revealed a decreased cell proliferation rate in the STIP1-downregulated cells. Cell cycle analysis revealed an increased proportion of cells in G1 phase, as well as an increase in apoptosis, upon STIP1 downregulation. Western blotting showed that TRAP1, pAkt, and MMP2 expression was decreased upon STIP1 downregulation. In addition, TRAP1, ki-67, and MMP2 displayed a decreased expression in vivo. Conclusions STIP1 is highly expressed in glioblastoma compared to normal brain tissues. Downregulation of STIP1 in glioma cells reduces cell proliferation rate and invasion and increases cell apoptosis.

Non-invasive genotyping of metastatic colorectal cancer using circulating cell free DNA Publication date: September 2019 Source: Cancer Genetics, Volume 237 Author(s): Xuemei Shi, Dzifa Y. Duose, Meenakshi Mehrotra, Michael A. Harmon, Peter Hu, Ignacio I. Wistuba, Scott Kopetz, Rajyalakshmi Luthra Abstract Circulating cell-free DNA (ccfDNA) in plasma provides an easily accessible source of circulating tumor DNA (ctDNA) for detecting actionable genomic alterations that can be used to guide colorectal cancer (CRC) treatment and surveillance. The goal of this study was to test the feasibility of using a traditional amplicon-based next-generation sequencing (NGS) on Ion Torrent platform to detect low-frequency alleles in ctDNA and compare it with a digital NGS assay specifically designed to detect low-frequency variants (as low as 0.1%) to provide evidence for the standard care of CRC. The study cohort consisted of 48 CRC patients for whom matched samples of formalin-fixed, paraffin-embedded tumor tissue, plasma, and peripheral blood mononuclear cells were available. DNA samples from different sources were sequenced on different platforms using commercial protocols. Our results demonstrate that the ccfDNA sequencing with the traditional NGS can be reliably used in an integrated workflow to detect low-frequency somatic variants in CRC. We found a high degree of concordance between traditional NGS and digital NGS in profiling mutant alleles in ccfDNA. These findings suggest that the traditional NGS is a viable alternative to digital sequencing of ccfDNA at allele frequency above 1%. ccfDNA sequencing can not only provide real-time monitoring of CRC, but also lay the basis for its application as a clinical diagnostic test to guide personalized therapy.

A unique case of complex variant translocation of t(6;9;22)(p22;q34;q11.2), der(19) in a newly diagnosed patient with chronic myeloid leukemia Publication date: September 2019 Source: Cancer Genetics, Volume 237 Author(s): Rafiye Ciftciler, Emine Arzu Saglam, Ayten Inanc, Osman Ozcebe, Ibrahim Celalettin Haznedaroglu Abstract Chronic myeloid leukemia (CML) is a clonal myeloproliferative disorder characterized by the dysregulated production and uncontrolled proliferation of myeloid neoplastic cells. CML is associated with the fusion of BCR (on chromosome 22) and ABL1 (on chromosome 9) resulting in the BCR-ABL1 fusion gene. The translocation of chromosomes (9;22)(q34;p15) is present in almost 90–95% of patients with CML and only 5–8% CML patients have established variant complex translocation due to the participation of one or more chromosomes other than 9 and 22 chromosome. In the present study, a unique case of a pH chromosome-positive CML is reported with a new variant pH translocation involving three chromosomal aberrations 6p22, 9q34, 22q11.2 and derivation 19 which has not been described previously. The complex variant translocation with pH chromosome was 46,XY,t(6;9;22)(p22:q34;q11.2), der(19)[48]/46,XY[2]  in this newly diagnosed CML patient. Additional cytogenetic anomalies may be seen in patients which are not controlled by the tyrosine kinase inhibitor in CML patients or in accelerated/blastic phase. In this case, the patient’ treatment was switched to dasatinib because the IS-NCN could not be controlled with imatinib. In conclusion, complex translocations in unusual locations of the BCR / ABL gene appear to indicate a poor prognosis.

Single-cell cloning of human T-cell lines reveals clonal variation in cell death responses to chemotherapeutics Publication date: September 2019 Source: Cancer Genetics, Volume 237 Author(s): Kathleen Hanlon, Alex Thompson, Lorena Pantano, John N. Hutchinson, Arshed Al-Obeidi, Shu Wang, Meghan Bliss-Moreau, Jennifer Helble, Gabriela Alexe, Kimberly Stegmaier, Daniel E. Bauer, Ben A. Croker Abstract Genetic modification of human leukemic cell lines using CRISPR-Cas9 has become a staple of gene-function studies. Single-cell cloning of modified cells is frequently used to facilitate studies of gene function. Inherent in this approach is an assumption that the genetic drift, amplified in some cell lines by mutations in DNA replication and repair machinery, as well as non-genetic factors will not introduce significant levels of experimental cellular heterogeneity in clones derived from parental populations. In this study, we characterize the variation in cell death of fifty clonal cell lines generated from human Jurkat and MOLT-4 T-cells edited by CRISPR-Cas9. We demonstrate a wide distribution of sensitivity to chemotherapeutics between non-edited clonal human leukemia T-cell lines, and also following CRISPR-Cas9 editing at the NLRP1 locus, or following transfection with non-targeting sgRNA controls. The cell death sensitivity profile of clonal cell lines was consistent across experiments and failed to revert to the non-clonal parental phenotype. Whole genome sequencing of two clonal cell lines edited by CRISPR-Cas9 revealed unique and shared genetic variants, which had minimal read support in the non-clonal parental population and were not suspected CRISPR-Cas9 off-target effects. These variants included genes related to cell death and drug metabolism. The variation in cell death phenotype of clonal populations of human T-cell lines may be a consequence of T-cell line genetic instability, and to a lesser extent clonal heterogeneity in the parental population or CRISPR-Cas9 off-target effects not predicted by current models. This work highlights the importance of genetic variation between clonal T-cell lines in the design, conduct, and analysis of experiments to investigate gene function after single-cell cloning.

Concurrent chromothripsis events in a case of TP53 depleted acute myeloid leukemia with myelodysplasia-related changes Publication date: September 2019 Source: Cancer Genetics, Volume 237 Author(s): D. Tolomeo, A. L'Abbate, A. Lonoce, P. D'Addabbo, M.F. Miccoli, C. Lo Cunsolo, P. Iuzzolino, O. Palumbo, M. Carella, V. Racanelli, T. Mazza, E. Ottaviani, G. Martinelli, G. Macchia, C.T. Storlazzi Abstract Acute myeloid leukemia with myelodysplasia-related changes (AML-MRC) is a heterogeneous hematological disorder defined by morphological, genetic, and clinical features. Patients with AML-MRC often show cytogenetic changes, which are associated with poor prognosis. Straightforward criteria for AML-MRC diagnosis and a more rigorous characterization of the genetic abnormalities accompanying this disease are needed. Here we describe an informative AML-MRC case, showing two separate, but concurrent, chromothripsis events, occurred at the onset of the tumor, and originating an unbalanced t(5;7) translocation and a derivative chromosome 12 with a highly rearranged short arm. Conversely, despite chromothripsis has been often associated with genomic amplification in cancer, in this case a large marker chromosome harboring amplified sequences from chromosomes 19 and 22 arose from a stepwise mechanism. Notably, the patient also showed a TP53 mutated status, known to be associated with an increased susceptibility towards chromothripsis and a poor prognosis. Our results indicate that multiple chromothripsis events may occur early in neoplastic transformation and act in a synergistic way with progressive chromosomal alterations to determine a dramatic impact on disease outcome, as suggested by the gene expression profile analysis.

Inherited cancer syndromes in 220 Italian ovarian cancer patients Publication date: September 2019 Source: Cancer Genetics, Volume 237 Author(s): I. Carnevali, C. Riva, A.M. Chiaravalli, N. Sahnane, E. Di Lauro, A. Viel, F. Rovera, G. Formenti, F. Ghezzi, F. Sessa, M.G. Tibiletti Abstract Background A subsets of ovarian carcinomas (OCs) are related to inherited conditions including Hereditary Breast and Ovarian Cancers (HBOC) and Lynch Syndrome (LS). The identification of inherited conditions using genetic testing might be a strategic model for cancer prevention that include benefits for the ovarian cancer patients and for their family members. Methods We describe a retrospective Italian experience for the identification of inherited conditions in 232 patients affected by OCs using both somatic and germline analyses. Results Immunohistochemical and microsatellite analyses performed on OCs identified 20 out of 101 MMR defective cancers and 15 of these were from patients carriers of the MMR germline pathogenetic variants. BRCA1 and BRCA2 testing offered to 198 OC patients revealed 67 (34%) pathogenetic variant carriers of BRCA1/2 genes. Interestingly LS patients revealed a mean age of OC onset of 45.4 years, which was significantly lower than the mean age of OCs onset of HBOC patients. Conclusions Somatic and germline analyses offered to OC patients has proved to be an efficient strategy for the identification of inherited conditions involving OC also in absence of suggestive family histories. The identification of LS and HBOC syndromes through OC patients is an effective tool for OC prevention.

Characterization of a rarely reported STAT5B/RARA gene fusion in a young adult with newly diagnosed acute promyelocytic leukemia with resistance to ATRA therapy Publication date: September 2019 Source: Cancer Genetics, Volume 237 Author(s): Jess F. Peterson, Rui R. He, Hassan Nayer, Raymund S. Cuevo, James B. Smadbeck, George Vasmatzis, Patricia T. Greipp, Rhett P. Ketterling, Nicole L. Hoppman, Linda B. Baughn Abstract The detection of PML/RARA or variant RARA rearrangements is critical for the diagnosis and treatment of patients with newly diagnosed acute promyelocytic leukemia (APL). While most cases of APL harboring the PML/RARA fusion respond to all-trans retinoic acid (ATRA), some variant RARA rearrangements are ATRA insensitive. Herein, we report a 27-year-old male with newly diagnosed, rapidly progressive APL and a rarely described STAT5B/RARA fusion with known resistance to ATRA therapy. While the PML/RARA dual-color dual-fusion fluorescence in situ hybridization (FISH) probe study was negative, the RARA break-apart probe study revealed an atypical RARA rearrangement in 95% of nuclei. A next generation sequencing assay, mate-pair sequencing, was subsequently performed to further characterize the RARA rearrangement and identified the RARA gene fusion partner STAT5B.

Expression deregulation of DNA repair pathway genes in gastric cancer Publication date: September 2019 Source: Cancer Genetics, Volume 237 Author(s): Shahzad Yousaf, Asad Ullah Khan, Zertashia Akram, Mahmood Akhtar Kayani, Iram Nadeem, Bilquis Begum, Ishrat Mahjabeen Abstract This study was designed to check correlation of mRNA and protein expression of BER pathway genes(XRCC1, OGG1) and a proliferation marker (Ki-67) in 100 gastric tissue samples and controls (adjacent uninvolved area). The expression was estimated usingreal time PCR and immunohistochemistry. Genomic instability was also calculated in the same study cohort using 8-OHdG assay, DNA fragmentation assay and comet assay. A significant downregulation of XRCC1 (p < 0.0001) and OGG1 (p < 0.0001) expression was observed in gastric cancer tumors vs controls. When analyzed with spearman correlation, significant positive correlation was observed between OGG1 vs XRCC1 (r = 0.319*, p < 0.02) and significant negative correlation was observed between OGG1 vs Ki-67 (r = −0.462**, p < 0.001) and XRCC1 vs Ki-67 (r = −0.589**, p < 0.001) in gastric cancer tumors. Significantly higher level of 8-OHdG, when compared to controls, was observed in gastric cancer tumors (p < 0.0001). DNA fragmentation assay and comet assay showed the formation of increased ladder patterns and comets in gastric cancer tumors when compared with controls These findings suggest that dysregulation of XRCC1, OGG1 combined with overexpression of Ki-67 may contribute to progression of gastric cancer and may help to sub-classify patients within diverse risk groups for therapeutic advantages.

Association of transcriptional levels of folate-mediated one-carbon metabolism-related genes in cancer cell lines with drug treatment response Publication date: September 2019 Source: Cancer Genetics, Volume 237 Author(s): Dong-Joon Min, Suleyman Vural, Julia Krushkal Abstract Folate-mediated one-carbon metabolism is essential for growth and survival of cancer cells. We investigated whether the response of cancer cells to antitumor treatment may be partially influenced by variation in expression of one-carbon metabolism genes. We used cancer cell line information from the Cancer Cell Line Encyclopedia and the Genomics of Drug Sensitivity in Cancer resources to examine whether variation in pretreatment expression of one-carbon metabolism-related genes was associated with response to treatment. GART, TYMS, SHMT2, MTR, ALDH2, BHMT, MAT2B, MTHFD2, NNMT, and SLC46A1 showed modest statistically significant correlations with response to a variety of antitumor agents. Higher expression levels of SLC46A1 were associated with resistance to multiple agents, whereas elevated expression of GART, TYMS, SHMT2, MTR, BHMT, and MAT2B was associated with chemosensitivity to multiple drugs. NNMT expression was bimodally distributed and showed different directions of association with various agents. Correlation of increased NNMT expression with sensitivity to dasatinib was validated in the NCI-60 cancer cell line panel. Pretreatment expression levels were correlated among many one-carbon metabolism genes. Expression of several folate genes was strongly associated with expression of multiple components of drug target pathways. Molecular mechanisms underlying associations of one-carbon metabolism gene with drug response require further investigation.