MLL/AF9 Expression Causes Leukemia in Zebrafish

Abstract Background Acute Leukaemia (AL) is a genetically heterogeneous disorder caused by somatic mutations and acquired chromosomal translocations. Translocations can lead to the formation of fusion genes such as the MLL/AF9 fusion, which results from the t(9;11)(p22;q23). A better understanding the molecular pathophysiology of AML, of the mechanisms of treatment resistance, disease relapse can be achieved by developing animal models. The MLL/AF9 fusion is frequently used to model AML in mice. However, to date, no MLL/AF9 leukemia models in zebrafish have been reported. Aim Our aim was to establish a transgenic zebrafish leukemia model using the human MLL/AF9 fusion gene. Methods To generate transgenic fish, two constructs (pTol2-Runx1+23: MLL-AF9-IRES-EGFP-cmlc-GFP and pTol2-Runx1+23: MLL-AF9-IRES-mCherry-cmlc-GFP) were injected together with Tol2 transposase mRNA into one-cell stage zebrafish embryos. We used the murine Runx1+23 enhancer to direct MLL/AF9 expression to hematopoietic stem cells and EGFP or mCherry as fluorescent markers. We selected transgenic embryos 24 hours post-fertilization based on the heart marker expression (cmlc). Results 29% (100 of 340 embryos) of the transgenics reached adulthood (6 weeks). After 6 to 24 months, 77% (77) of them developed signs of sickness. They became less active with protruding eyes and hump formation on the nose. Some started bleeding from the gills and/or showed tumor formation around the abdomen and head. Sick fish were euthanized and dissected. The autopsies showed pale and dysmorphic kidneys, pale and enlarged spleens, and in some cases white granular spots on the spleen. Histological sections revealed increased kidney, spleen, and liver cellularity with massive cellular infiltration of cells in these organs. In flow cytometry, kidney marrow cells from the transgenic fish showed a different forward scatter (FSC) and side scatter (SSC) profile compared to that of the normal zebrafish kidney marrow cells. The transgenic F 0 zebrafish showed increased numbers of lymphoid cells (12 fish), precursor cells (9 fish), or myeloid cells (6 fish). Peripheral blood smears showed many intermediate-sized mononuclear cells with an increased nuclear-cytoplasmic ratio, with nuclei containing dipsersed chromatin and inconspicuous nucleoli resembling blasts. There were occasional myelocytes and no mature granulocytes. These data are consistent with the development of acute leukemia in our MLL/AF9 transgenic fish. Whole-mount in situ hybridization (WISH) was performed on F 1 embryos. RNA probes for early hematopoietic markers (gata1, scl, runx1, ikaros, cmyb, mpx, and lyz) were hybridized to 24 and 48 hpf F1 transgenic embryos. There were expression changes of these markers compared to age-matched wild-type larvae, including low expression of gata1, scl, cmyb and high expression of lyz, mpx and ikaros in the caudal hematopoietic organ. We also performed transplantation experiments with the kidney marrow cells from diseased fish to test whether the disease was transplantable. The disease was serially transplantable into secondary and tertiary recipient fish. Transplanted fish had a significantly shorter latency to disease development of only 2 to 6 weeks. The morphological evidence and the serial transplantability of the disease proves that we have in fact succeeded in establishing an MLL/AF9-driven acute leukemia model in zebrafish. The long latency and incomplete penetrance observed in our F 0 MA9 zebrafish, along with a shorter latency in the transplanted fish, suggests that additional somatic mutations are required for leukemogenesis in this model. We performed whole exome sequencing to find cooperating somatic mutations and RNASeq to identify differentially expressed genes in MA9 leukemia fish. Whole exome sequencing on six samples identified putative somatic mutations in genes such as Stat5, Cyp2j20, Ms4a17a.3, Tapbp.1 and Herc5.3, which have been reported to be mutated in human cancer. RNA-seq analysis on seven samples showed 67 differentially expressed genes with a q value < 0.05 (e.g., cxcl32b.1, myof1, ctdsp2, egr3, il2rb) and nine enriched pathways with a P-value of < 0.054 (e.g.: KRAS, TP53, MEK) in our transgenic leukemic MLL/AF9 fish. Conclusion Our MLL/AF9 zebrafish acute leukemia model will be a helpful tool to understand leukemia biology and enable testing of new therapeutic strategies. Disclosures Browett: AbbVie: Honoraria; Janssen: Membership on an entity's Board of Directors or advisory committees; MSD: Membership on an entity's Board of Directors or advisory committees.

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Whole-Exome Sequencing In Myelodysplastic Syndromes With 5q- and Normal Karyotype

Blood ◽

10.1182/blood.v122.21.1551.1551 ◽

2013 ◽

Vol 122 (21) ◽

pp. 1551-1551

Author(s):

Vera Adema ◽

Mar Mallo ◽

Leonor Arenillas ◽

María Díez-Campelo ◽

Elisa Luño ◽

...

Keyword(s):

Bone Marrow ◽

Exome Sequencing ◽

Board Of Directors ◽

Whole Exome Sequencing ◽

Research Funding ◽

Somatic Mutations ◽

Normal Karyotype ◽

Advisory Committees ◽

Whole Exome ◽

5Q Deletion

Abstract Introduction Myelodysplastic Syndromes (MDS) are a heterogeneous group of clonal myeloid stem cells disorders with high prevalence in the elderly characterized by inefficient hematopoiesis, peripheral blood (PB) cytopenias, and an increased risk of transformation to acute myeloid leukemia (AML). The karyotype is the clinical parameter with the strongest prognostic impact according the IPSS-R (Greenberg et al., 2012). The most frequent cytogenetic alteration is the chromosome 5q deletion (del[5q]) which as a single anomaly, confers a good prognosis and predicts an excellent response to lenalidomide. Whether other genetic abnormalities routinely cooperate with del(5q) is not known. Whole-exome sequencing (WES) is a powerful tool to identify somatic mutations in protein coding genes that might cooperate with del(5q). In order to better understand the genetic basis of MDS with del(5q), we performed whole-exome sequencing (assessing 334,378 exons) of tumor-normal paired samples from 21 MDS patients. Herein we describe the preliminary findings. The analysis is ongoing and the complete results will be presented in the meeting. Methods A total of 21 patients with MDS (16 with del(5q) as a sole abnormality, 3 with del(5q) and additional alterations and 2 with normal karyotype) were included in our study. We examined a total of 25 tumor samples (21 diagnostic bone marrow (BM) samples with matched CD3+ cells as a controls, additional BM samples from 3 patients during lenalidomide treatment and 1 bone marrow sample from a del(5q) patient after AML progression). DNA was extracted from BM samples and from isolated peripheral blood CD3+ cells (magnetic-activated cell sorting (MACS), MiltenyiBiotec GmbH, Germany). The purity of CD3+ cells was assessed by FC 500 flow cytometer (Beckman Coulter, Hialeah, Fl, USA). Only DNA that fulfilled quality controls required by WES were submitted. For each diagnostic sample, we performed Conventional G-banding cytogenetics and fluorescence in situ hybridization (FISH, to confirm or dismiss 5q deletions). Whole-exome targeted capture was carried out on 3 μg of genomic DNA, using the SureSelect Human Exome Kit 51Mb version 4 (Agilent Technologies, Inc., Santa Clara, CA, USA). The captured and amplified exome library was sequenced with 100 bp paired-end reads on an Illumina HiSeq2000. Whole-exome sequencing data were analyzed using an in-house bioinformatics pipeline as previously reported. Somatic mutations identified as alterations present in tumor but not in the matched CD3+ sample were validated by Sanger sequencing. Results In our preliminary analysis of WES from 12 patients (10 patients with 5q- and 2 patients with normal karyotype), a total of 249 non-silent somatic variant candidates were identified, of which 146 were confirmed as somatic mutations. Recurrent mutations were observed in three genes (ASXL1, NBPF10 and SF3B1) in 3 different patients. Seven genes (HRNR, JAK2, POTEG, MUC5B, PHLDA, TTN, ZNF717) were mutated in two patients. Mutations in several genes known to be mutated in MDS (ASXL1, JAK2, RUNX1, SF3B1, SRSF2 and TET2) were also identified. Patients with the 5q deletion had an average of 11 mutations whereas patients with normal karyotype had a higher mean (14.5). Mutated genes identified in both groups were HRNR, JAK2, MUC5B, NBPF10 and SF3B1. No mutations in TP53 were detected in this subset. Pathway analysis of the complete list of somatically mutated genes will be carried out once all 21 patients are analyzed. The four in-treatment samples will be examined from their matched diagnostic samples. Conclusions Whole-exome sequencing of largely del(5q) MDS patient samples identified both known and previously unreported somatic mutations. Analysis of additional samples will allow a more complete description of the genes and pathways that may cooperate with del(5q) in the development and progression of MDS. Acknowledgments Financial support: This work has been supported (in part) by a grant from Instituto de Salud Carlos III, Ministerio de Sanidad y Consumo, Spain (PI 11/02010); by Red Temática de Investigación Cooperativa en Cáncer (RTICC, FEDER) (RD07/0020/2004; RD12/0036/0044); Acción COST BM0801: European Genomics and Epigenomics Study on MDS and AML; Sociedad Española de Hematología y Hemoterapia (SEHH) and MDS Celgene. Footnotes Rafael Bejar and Francesc Sole contributed equally. Disclosures: Díez-Campelo: Novartis and Celgene: Honoraria, Research Funding. Cañizo:Celgene Jansen-Cilag Arry Novartis: Membership on an entity’s Board of Directors or advisory committees, Research Funding. Sanchez:Celgene: Honoraria, Research Funding. Bejar:Genoptix: Consultancy, Honoraria, Membership on an entity’s Board of Directors or advisory committees; Celgene: Consultancy, Membership on an entity’s Board of Directors or advisory committees. Solé:Celgene: Consultancy, Membership on an entity’s Board of Directors or advisory committees, Research Funding.

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Whole-Exome Sequencing in Myelodysplastic Syndromes with 5q Deletion

Blood ◽

10.1182/blood.v124.21.4635.4635 ◽

2014 ◽

Vol 124 (21) ◽

pp. 4635-4635

Author(s):

Vera Adema ◽

Laura Palomo ◽

María Díez-Campelo ◽

Mar Mallo ◽

Leonor Arenillas ◽

...

Keyword(s):

Exome Sequencing ◽

Board Of Directors ◽

Whole Exome Sequencing ◽

Myelodysplastic Syndromes ◽

Genomic Dna ◽

Research Funding ◽

Sequencing Data ◽

Advisory Committees ◽

Whole Exome ◽

Increased Risk

Abstract INTRODUCTION Myelodysplastic syndromes (MDS) are a heterogeneous group of clonal myeloid stem cell disorders that are highly prevalent in elderly populations. MDS are characterized by inefficient hematopoiesis, peripheral blood (PB) cytopenias, and increased risk of transformation to acute myeloid leukemia (AML; 20–30% of patients with MDS). Around 50% of MDS patients carry at least one karyotypic aberration. The interstitial deletion of the long arm of chromosome 5 ([del(5q)] is the most common aberration, accounting for almost 30% of abnormal MDS karyotype. Various studies supports a favorable prognosis of MDS with isolated del(5q) with an excellent response to lenalidomide treatment. In order to describe the molecular events associated with MDS and del(5q) we performed whole-exome sequencing (WES)(assessing 334,378 exons) of tumor-normal paired samples from 20 MDS patients to unravel the genetic basis of MDS with del(5q). The analysis is ongoing and the complete results will be presented in the meeting. METHODS A total of 50 samples from 20 patients with MDS, with del(5q) were collected. For each diagnostic sample, we performed Conventional G-banding cytogenetics and fluorescence in situ hybridization (FISH, to confirm or dismiss del(5q)) and SNP arrays with Cytoscan HD (Affymetrix). These samples included: 20 tumor samples at diagnosis, 20 control samples and 10 samples after diagnosis, during lenalidomide treatment (5) or at the moment of relapse (5) in order to compare the genetic status before and during the treatment. Genomic DNA from tumor cells was obtained from bone marrow (BM) samples or from PB granulocytes. As a source of constitutional DNA we used CD3+T cells from each patient by isolating by magnetic-activated cell sorting. WES targeted capture was carried out on 7μg of genomic DNA, using the SureSelect Human Exome Kit 51Mb version 4.Libraries were sequenced on an Illumina HiSeq2000. Sequencing data will be analyzed using an in-house bioinformatics pipeline as previously reported. RESULTS Our preliminary analysis of these 20 new patients by WES confirmed our previous analyses with mutations in well described genes as ASXL1, JAK2 and TET2, but not in genes RUNX1, SF3B1 and SRSF2. In those patients we found two patients with missense mutation in TP53, one of the patients had an isolated del(5q) and is receiving lenalidomide treatment, and the other one had a complex karyotype. According to our prior analyses, in which 249 non-silent somatic variants were detected, we look forward to validate these mutations in this new series of patients. CONCLUSIONS We envision to validate these previous results with the new sequencing data of more patients with MDS and del(5q). We expect to measure somatic mutations that vary in abundance after lenalidomide treatment, potentially identifying mutations associated with resistance or relapse. ACKNOWLEDGEMENTS: This work has been supported (in part) by a grants from Instituto de Salud Carlos III, Ministerio de Sanidad y Consumo, Spain (PI 11/02010); by Red Temática de Investigación Cooperativa en Cáncer (RTICC, FEDER) (RD07/0020/2004; RD12/0036/0044); 2014 SGR225 (GRE) Generalitat de Catalunya; Fundació Internacional Josep Carreras; Obra Social “la Caixa”; Sociedad Española de Hematología y Hemoterapia (SEHH)and Celgene Spain. FOOTNOTES Rafael Bejar and Francesc Sole contributed equally. Disclosures Díez-Campelo: Novartis, Celgene: Honoraria, Research Funding. Xicoy:Celgene: Honoraria. Cañizo:Celgene, Jansen-Cilag, Arry, Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. sanchez-Garcia:Celgene: Honoraria, Research Funding. Bejar:Celgene: Membership on an entity's Board of Directors or advisory committees; Genoptix Medical Laboratory: Consultancy, Honoraria, Licensed IP, no royalties Patents & Royalties, Membership on an entity's Board of Directors or advisory committees. Sole:Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding.

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Whole-Exome Sequencing to Identify Mutations in Thrombosis Modifier Genes Isolated From a Factor V Leiden-Dependent Sensitized ENU Suppressor Screen in the Mouse

Blood ◽

10.1182/blood.v118.21.1183.1183 ◽

2011 ◽

Vol 118 (21) ◽

pp. 1183-1183

Author(s):

Randal Joseph Westrick ◽

Guojing Zhu ◽

Jishu Xu ◽

Audrey C.A. Cleuren ◽

Angela Yang ◽

...

Keyword(s):

Venous Thrombosis ◽

Exome Sequencing ◽

Board Of Directors ◽

Whole Exome Sequencing ◽

Factor V Leiden ◽

Modifier Genes ◽

Factor V ◽

Advisory Committees ◽

Mutagenesis Screen ◽

Whole Exome

Abstract Abstract 1183 Only ∼10% of individuals carrying the common venous thrombosis risk factor, Factor V Leiden (FVL) will develop venous thrombosis in their lifetime. In order to identify potential FVL modifier genes, we performed a sensitized dominant ENU mutagenesis screen, based on the perinatal synthetic lethal thrombus previously observed in mice homozygous for FVL (FVQ/Q) and hemizygous for tissue factor pathway inhibitor deficiency (Tfpi+/−). The genome-wide ENU mutagenesis screen was performed by crossing ENU-treated male FVQ/Q mice with FVQ/+ Tfpi+/− females. Surviving G1 offspring were analyzed to identify survivors with the otherwise lethal FVQ/Q Tfpi+/− genotype. As proof of concept, we demonstrated that reduced tissue factor (Tf+/−) suppresses the lethal FVQ/Q Tfpi+/− phenotype, suggesting that mutations at Tf should be among the suppressor genes identified by our screen. Analysis of 7,128 G1 offspring (∼2X genome coverage) identified 98 FVQ/Q Tfpi+/− mice that survived to weaning. Fourteen FVQ/Q Tfpi+/− G1 mice exhibited successful transmission of a putative suppressor mutation to two or more FVQ/Q Tfpi+/− G2 offspring. Extensive genotyping of mice from an expanded genetic cross from one of these lines mapped a candidate suppressor locus to a chromosome 3 region encompassing the TF gene (LOD=4.93). With continued improvements in next generation sequencing technologies, we have now applied whole exome sequencing to analysis of 8 of the remaining 13 lines. The entire DNA coding region (the “exome”, totaling 49.6 Mb of DNA sequence) from a progeny-tested member of each line was captured using the Agilent SureSelect mouse exome capture system. Whole-exome sequencing using the Illumina HiSeq high-throughput sequencer yielded 12–15 gigabases of sequence data per sample, corresponding to an average of ∼200 fold sequencing coverage for each nucleotide position. Variant analysis using the Gene Analysis Toolkit revealed the presence of a small number of high confidence novel heterozygous (dominant) variants in each sample. Each of these heterozygous variants is a candidate suppressor mutation and these are presently being tested in remaining FVQ/Q Tfpi+/− mice from each respective line. Based on previous studies where ENU-induced mutations in mice have been identified, we anticipate the identification of putative exomic mutations in approximately 80% of tested suppressor lines. Identification of these mutations should provide candidate modifier genes for FVL and other human hemostatic disorders. Disclosures: Ginsburg: Portola Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Catalyst Biosciences: Membership on an entity's Board of Directors or advisory committees; Shire Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees.

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Gene Mutations Detected by Whole-Exome Sequencing and Recurrent Cytogenetic Abnormalities Are Independent Events in Multiple Myeloma

Blood ◽

10.1182/blood.v118.21.1816.1816 ◽

2011 ◽

Vol 118 (21) ◽

pp. 1816-1816

Author(s):

Hervè Avet-Loiseau ◽

Graham R Bignell ◽

Cheng Li ◽

Florence Magrangeas ◽

Thierry Facon ◽

...

Keyword(s):

Multiple Myeloma ◽

Exome Sequencing ◽

Board Of Directors ◽

Whole Exome Sequencing ◽

Risk Group ◽

Gene Mutations ◽

Newly Diagnosed ◽

Advisory Committees ◽

Independent Events ◽

Whole Exome

Abstract Abstract 1816 Multiple Myeloma (MM) is characterized by several recurrent chromosomal abnormalities, some of them driving the outcome of the patients, especially t(4;14), del(17p), and hyperdiploidy. On the other hand, recent data based on whole genome sequencing showed that MM is characterized by many gene mutations, some of them being recurrent. In order to try to reconcile these two types of genetic abnormalities, we performed whole exome sequencing (WES) in 53 newly-diagnosed patients characterized by high or low risk at the chromosomal level. We selected 16 patients with del(17p) in at least 60% of the clonal plasma cells, 5 patients with del(12p), including 3 with associated t(4;14), 2 patients with t(14;16), 1 patient with both del(17p), del 12p), and t(4;14), considered as the poor risk group (24 patients), 24 patients with hyperdiploidy (including chromosome 5 gain), and 2 patients with a normal SNParray profile, considered as the good risk group (26 patients, and 3 patients with hyperdiploidy and either del(17p) and/or del(12p), considered as the uncertain risk group. A total of 3621 non-silent mutations were observed through the 53 tumor genomes. The median number of mutations per patient was 79 (31–462). We did not observe differences in the number of mutations according to cytogenetic risk. Regarding specific gene mutations, 376 genes presented 2 mutations, and 128 genes presented at least 3 mutations (3–16). Comparison with recently published sequencing data (Chapman, Nature, 2011) revealed only a few common mutated genes (NRAS 26%, KRAS 23%, TP53 13%, BRAF 11%, and FAM46C 10%). In contrast, many recurrently mutated genes were identified in this series, but not in the published one. This could be related to the relatively low number of sequenced cases in both series. Very interestingly, quite a high number of strictly identical mutations involving many genes were observed in different patients (139 cases), suggesting that those mutations are more probably driver rather than passenger events. To conclude, in this large comprehensive study, we did not find any significant correlation between recurrent chromosomal changes and gene mutations, suggesting that these two events occur independently. We cannot address the issue of the prognostic value of the gene mutations because of the low number of patients sequenced so far. This question will definitely have to be addressed in future larger series on newly diagnosed patients with MM. Disclosures: Facon: Celgene: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees. Anderson:Celgene: Membership on an entity's Board of Directors or advisory committees; Millennium: Membership on an entity's Board of Directors or advisory committees; Onyx: Membership on an entity's Board of Directors or advisory committees; Merck: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Acetylon: Membership on an entity's Board of Directors or advisory committees. Munshi:Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Millennium: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Onyx: Membership on an entity's Board of Directors or advisory committees.

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Whole Exome Sequencing from Nine Independent Sites of Extraosseous Disease in a Single Patient with Relapsed Multiple Myeloma Show That Extramedullary Disease Arise through a Combination of Branched and Parallel Evolution

Blood ◽

10.1182/blood.v128.22.2090.2090 ◽

2016 ◽

Vol 128 (22) ◽

pp. 2090-2090

Author(s):

Heather Landau ◽

Jun Fan ◽

Sergio A. Giralt ◽

Jonathan U. Peled ◽

Hani Hassoun ◽

...

Keyword(s):

Multiple Myeloma ◽

Exome Sequencing ◽

Board Of Directors ◽

Whole Exome Sequencing ◽

Research Funding ◽

Parallel Evolution ◽

High Quality ◽

Advisory Committees ◽

Extramedullary Disease ◽

Whole Exome

Abstract Introduction: Multiple myeloma (MM) is the second most common hematologic malignancy. Recent sequencing studies show evidence of massive genetic heterogeneity reflected in multiple parallel subclones already at diagnosis. Different subclones respond differently to given therapy. The role of treatment-driven subclonal skewing and intrinsic acquired mutations is poorly understood in the relapse setting. At relapse, the distribution of subclones throughout the bone marrow and at extramedullary sites is currently unknown. Methods: We performed a research autopsy on a single individual with IgG kappa MM who survived with multiple metastatic sites of disease for 10 years after diagnosis. Genomic DNA was extracted from histologically confirmed snap frozen normal tissue and nine independent sites of extraosseous disease. Whole exome sequencing (150X) was performed by the MSK Genomics Core. Data were filtered to remove germline polymorphisms and enrich for high quality somatic variants. Phylogenetics and subclonal analyses were performed using Treeomics and SCHISM. Results: The average on target coverage was 177X with 73% of bases covered a minimum of 100x. A total of 6348 somatic variants were initially identified. After filtering for high quality somatic variants, 1330 remained with 220 of these variants common to all MM samples and an average of 628 variants per sample. Annotation of high quality variants revealed potentially deleterious somatic mutations in NRAS, FAM46C, DYNC1, CREBBP, ATM, BIRC3, MGA, PPP6C and SMARCA4. Phylogenetics analyses (shown below) indicated the metastases arose through a combination of branched and parallel evolution, with the ATM mutation arising in one branch (one metastasis) that was distinct from a second clonal population containing the NRAS, FAM46C, BIRC3, CREBBP, DYNC1 and PPP6C mutations that were present in all eight other metastases. The MGA and SMARCA4 mutations further identified two additional subclones arising in from the NRAS/FAM46C mutant common ancestral clone. Clonality analyses independently supported this hierarchy by identifying NRAS and FAM46C as early clonal events and MGA and SMARCA4 as late events in the progression of this patient's disease. It also suggested a degree of subclonal mixing within each metastatic site. Conclusions: Using whole exome sequencing from nine independent sites of extraosseous disease in a single MM patient with relapse 10 years after initial diagnosis, we show that extramedullary disease arise through a combination of branched and parallel evolution. Two additional patients have also undergone research autopsy and results will be presented at the meeting. Figure Figure. Disclosures Landau: Prothena: Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees, Research Funding; Onyx/Amgen: Research Funding; Spectrum Pharmaceuticals: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy. Hassoun:Binding Site: Research Funding; Novartis: Consultancy; Celgene: Research Funding; Takeda: Consultancy, Research Funding. Korde:Medscape: Honoraria. Landgren:Medscape Myeloma Program: Honoraria; Takeda: Honoraria; Amgen: Honoraria, Research Funding; BMS: Honoraria; Merck: Honoraria; Celgene: Honoraria, Research Funding.

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Germline and somatic mutations in the pathology of pineal cyst: A whole‐exome sequencing study of 93 individuals

Molecular Genetics & Genomic Medicine ◽

10.1002/mgg3.1691 ◽

2021 ◽

Author(s):

Yuanqing Yan ◽

Rebecca Martinez ◽

Maria N. Rasheed ◽

Joshua Cahal ◽

Zhen Xu ◽

...

Keyword(s):

Exome Sequencing ◽

Whole Exome Sequencing ◽

Somatic Mutations ◽

Pineal Cyst ◽

Whole Exome

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Whole-exome sequencing reveals potential germline and somatic mutations in 60 malignant ovarian germ cell tumors

Biology of Reproduction ◽

10.1093/biolre/ioab052 ◽

2021 ◽

Author(s):

Juan Chen ◽

Yan Li ◽

Jianlei Wu ◽

Yakun Liu ◽

Shan Kang

Keyword(s):

Germ Cell ◽

Exome Sequencing ◽

Whole Exome Sequencing ◽

Peripheral Blood ◽

Copy Number ◽

Somatic Mutations ◽

Germ Cell Tumors ◽

Germline Mutations ◽

Deletion Region ◽

Whole Exome

Abstract Background Malignant ovarian germ cell tumors (MOGCTs) are rare and heterogeneous ovary tumors. We aimed to identify potential germline mutations and somatic mutations in MOGCTs by whole-exome sequencing. Methods The peripheral blood and tumor samples from these patients were used to identify germline mutations and somatic mutations, respectively. For those genes corresponding to copy number alterations (CNA) deletion and duplication region, functional annotation of was performed. Immunohistochemistry was performed to evaluate the expression of mutated genes corresponding to CNA deletion region. Results In peripheral blood, copy number loss and gain were mostly found in yolk sac tumors (YST). Moreover, POU5F1 was the most significant mutated gene with mutation frequency > 10% in both CNA deletion and duplication region. In addition, strong cytoplasm staining of POU5F1 (corresponding to CNA deletion region) was found in 2 YST and nuclear staining in 2 dysgerminomas (DG) tumor samples. Genes corresponding to CNA deletion region were significantly enriched in the signaling pathway of regulating pluripotency of stem cells. In addition, genes corresponding to CNA duplication region were significantly enriched in the signaling pathways of RIG-I-like receptor, Toll-like receptor, NF-kappa B and Jak–STAT. KRT4, RPL14, PCSK6, PABPC3 and SARM1 mutations were detected in both peripheral blood and tumor samples. Conclusions Identification of potential germline mutations and somatic mutations in MOGCTs may provide a new field in understanding the genetic feature of the rare biological tumor type in the ovary.

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Whole-Exome Sequencing Identifies Somatic Mutations Associated With Mortality in Metastatic Clear Cell Kidney Carcinoma

Frontiers in Genetics ◽

10.3389/fgene.2019.00439 ◽

2019 ◽

Vol 10 ◽

Cited By ~ 3

Author(s):

Alejandro Mendoza-Alvarez ◽

Beatriz Guillen-Guio ◽

Adrian Baez-Ortega ◽

Carolina Hernandez-Perez ◽

Sita Lakhwani-Lakhwani ◽

...

Keyword(s):

Exome Sequencing ◽

Whole Exome Sequencing ◽

Somatic Mutations ◽

Clear Cell ◽

Kidney Carcinoma ◽

Whole Exome

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TP53 and PTEN mutations were shared in concurrent germ cell tumor and acute megakaryoblastic leukemia

BMC Cancer ◽

10.1186/s12885-019-6497-0 ◽

2020 ◽

Vol 20 (1) ◽

Cited By ~ 2

Author(s):

Keiichi Akizuki ◽

Masaaki Sekine ◽

Yasunori Kogure ◽

Takuro Kameda ◽

Kotaro Shide ◽

...

Keyword(s):

Germ Cell ◽

Exome Sequencing ◽

Whole Exome Sequencing ◽

Germ Cell Tumor ◽

Somatic Mutations ◽

Cell Tumor ◽

Sequencing Analysis ◽

Male Adult ◽

Clonal Relationship ◽

Whole Exome

Abstract Background The occurrence of a mediastinal germ cell tumor (GCT) and hematological malignancy in the same patient is very rare. Due to its rarity, there have been only two reports of the concurrent cases undergoing detailed genetic analysis with whole-exome sequencing (WES), and the possible clonal relationship between the both tumors remained not fully elucidated. Methods We performed whole-exome sequencing analysis of mediastinal GCT and acute myeloid leukemia (AML) samples obtained from one young Japanese male adult patient with concurrent both tumors, and investigated the possible clonal relationship between them. Results Sixteen somatic mutations were detected in the mediastinal GCT sample and 18 somatic mutations in the AML sample. Mutations in nine genes, including TP53 and PTEN both known as tumor suppressor genes, were shared in both tumors. Conclusions All in our case and in the previous two cases with concurrent mediastinal GCT and AML undergoing with whole-exome sequencing analysis, TP53 and PTEN mutations were commonly shared in both tumors. These data not only suggest that these tumors share a common founding clone, but also indicate that associated mediastinal GCT and AML harboring TP53 and PTEN mutations represent a unique biological entity.

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