Exome Sequencing Identifies Recurring DNMT3A Mutations in Adult ETP-ALL

Abstract Abstract 1377 Introduction: Early T-cell precursor (ETP) ALL accounting for 10% of all T-ALL cases is of special interest because of its proposed origin from early thymic progenitors with multilineage differentiation potential. ETP-ALL is associated with a poorer outcome in pediatric and adult patients. On the molecular level, ETP-ALL is characterized by a specific immunophenotype (CD1-, CD5weak, CD8-, co-expression of stem cell and/or myeloid antigens) and distinct molecular features (expression of stem cell genes, high frequency of FLT3 mutations with absence of NOTCH1 mutations). Whereas a highly heterogeneous genetic pattern was revealed by whole genome sequencing in pediatric patients, the genetic background of adult ETP-ALL remains largely unknown. Here we investigated genetic alterations in adult ETP-ALL by whole exome sequencing and subsequently analyzed specific target genes. Patients and methods: We performed whole exome sequencing of five paired (diagnosis/remission) adult ETP-ALL patients enrolled in German Acute Lymphoblastic Leukemia Multicenter Study Group (GMALL) trials. Using exon capturing from genomic DNA, followed by 76-bp paired-end sequencing on an Illumina Genome Analyzer IIx platform, we generated at least 5 Gb of exome sequence from each ETP-ALL and remission samples. Somatic mutations were identified by comparing the ETP-ALL with the remission exome sequence, excluding all annotated polymorphisms (dbSNP130), non-coding positions and positions with evidence of a variant in the corresponding remission samples. Candidate variants were confirmed by capillary sequencing of genomic DNA. The DNMT3A mutations status was analyzed by Sanger sequencing of exons 11–23 in additional 68 adult ETP-ALL (55 male, 13 female, median age: 38 years) as well as the mutation status of the polycomb repressor complex (PRC) genes EZH2 and SUZ12. For 52 of 68 patients clinical follow-up data were available. Results: Using whole exome sequencing we found a total of 56 non-synonymous somatic mutations or indels in the five ETP-ALL patients (range: 6 to 16 per patient). Eleven mutations/indels affected cancer genes. DNMT3A (2/5) and FAT3 (2/5) were recurrently mutated in the five patients. The DNA-methyl-transferase DNMT3A is a frequent mutational target in acute myeloid leukemia (AML; 20%), whereas FAT3 (FAT, tumor suppressor homolog 3) mutations were recently reported in ovarian carcinoma (TCGA, Nature 2011). Novel mutations identified in adult ETP-ALL involved genes in epigenetic regulation (e.g. MLL2, MLL3, BMI1), and in genes previously reported to be mutated in ETP-ALL (e.g. in JAK1, ETV6, NOTCH1, DNM2). By Sanger sequencing, we screened for DNMT3A mutations in a larger cohort of adult ETP-ALL. DNMT3A mutations were present in 11 of the 68 (16%) patients, a mutation rate similar to AML. Amino acid R882 (exon 23), the most frequently mutated amino acid in AML, was mutated in five ETP-ALL. The remaining six mutations occurred in single spots, with one exception in the ZNF or the MTF domain. Patients with a DNMT3A mutation were significantly older (median: 63 vs 37 years, P=0.016). No correlation was found between DNMT3A and FLT3 mutations (27% in DNMT3A mut pts. vs. 37% in DNMT3A wt pts., P=0.41) or NOTCH1 mutations (10% in DNMT3A mut pts. vs. 16% in DNMT3A wt pts., P=0.47). In addition, we investigated genetic alterations in epigenetic regulators including members of the polycomb repressor complex (PRC). Mutations were seen in EZH2 in 4/68 (6%), SUZ12 in 1/68 (1%) and SH2B3 in 4/69 (6%) of ETP-ALL. Interestingly, patients with at least one mutation in an epigenetic regulator gene (DNMT3A, SUZ12, SH2B3, MLL2, or EZH2) showed a trend towards an inferior survival (one-year-survival: 50% vs. 85%, P=0.08). Conclusion: Adult ETP-ALL patients display a heterogenous spectrum of mutations, particularly affecting genes involved in epigenetic regulation. The spectrum is different to pediatric patients with a lower rate of polycomb repressor complex and a higher rate of DNMT3A mutations. The higher rate of DNMT3A mutations in older patients might point to a different pathogenesis compared to pediatric ETP-ALL. Like in AML, DNMT3A mutations in adult ETP-ALL show a similar frequency, within the same hot spots and are correlated with an adverse prognostic value, underscoring the myeloid character of ETP-ALL. Thus, these data may provide a rationale to use epigenetic therapy in ETP-ALL. Disclosures: Krebs: Illumina: Honoraria. Greif:Illumina: Honoraria.

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A homozygous mutation of GNRHR in a familial case diagnosed with polycystic ovary syndrome

Acta Endocrinologica ◽

10.1530/eje-16-0968 ◽

2017 ◽

Vol 176 (5) ◽

pp. K9-K14 ◽

Cited By ~ 5

Author(s):

Sandrine Caburet ◽

Ronit Beck Fruchter ◽

Bérangère Legois ◽

Marc Fellous ◽

Stavit Shalev ◽

...

Keyword(s):

Exome Sequencing ◽

Whole Exome Sequencing ◽

Sanger Sequencing ◽

Polycystic Ovary ◽

Missense Variant ◽

Genetic Alterations ◽

Homozygous Mutation ◽

Gonadotropin Secretion ◽

Whole Exome ◽

A Genome

Context PCOS is a heterogeneous condition characterized by hyperandrogenism and chronic anovulation and affects about 10% of women. Its etiology is poorly known, but a dysregulation of gonadotropin secretion is one of its hallmarks. Objective As the etiology of PCOS is unclear, we have performed a genome-wide analysis of a consanguineous family with three sisters diagnosed with PCOS. Methods Whole-exome sequencing and Sanger sequencing confirmation. Results Whole-exome sequencing allowed the detection of the missense variant rs104893836 located in the first coding exon of the GNRHR gene and leading to the p.Gln106Arg (p.Q106R) substitution. Sanger sequencing of all available individuals of the family confirmed that the variant was homozygous in the three affected sisters and heterozygous in both parents. Conclusions This is the first description of a GNRHR gene mutation in patients diagnosed with PCOS. Although we do not exclude a possible interaction of the identified variant with the genetic background and/or the environment, our result suggests that genetic alterations in the hypothalamo–pituitary axis may play role in the pathogenesis of PCOS.

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Whole-exome sequencing identifies somatic mutations of BCOR in acute myeloid leukemia with normal karyotype

Blood ◽

10.1182/blood-2011-07-365320 ◽

2011 ◽

Vol 118 (23) ◽

pp. 6153-6163 ◽

Cited By ~ 152

Author(s):

Vera Grossmann ◽

Enrico Tiacci ◽

Antony B. Holmes ◽

Alexander Kohlmann ◽

Maria Paola Martelli ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Exome Sequencing ◽

Whole Exome Sequencing ◽

Myeloid Leukemia ◽

Somatic Mutations ◽

Normal Karyotype ◽

Genetic Alterations ◽

World Health ◽

Whole Exome ◽

Acute Myeloid

Abstract Among acute myeloid leukemia (AML) patients with a normal karyotype (CN-AML), NPM1 and CEBPA mutations define World Health Organization 2008 provisional entities accounting for approximately 60% of patients, but the remaining 40% are molecularly poorly characterized. Using whole-exome sequencing of one CN-AML patient lacking mutations in NPM1, CEBPA, FLT3-ITD, IDH1, and MLL-PTD, we newly identified a clonal somatic mutation in BCOR (BCL6 corepressor), a gene located on chromosome Xp11.4. Further analyses of 553 AML patients showed that BCOR mutations occurred in 3.8% of unselected CN-AML patients and represented a substantial fraction (17.1%) of CN-AML patients showing the same genotype as the AML index patient subjected to whole-exome sequencing. BCOR somatic mutations were: (1) disruptive events similar to the germline BCOR mutations causing the oculo-facio-cardio-dental genetic syndrome; (2) associated with decreased BCOR mRNA levels, absence of full-length BCOR, and absent or low expression of a truncated BCOR protein; (3) virtually mutually exclusive with NPM1 mutations; and (4) frequently associated with DNMT3A mutations, suggesting cooperativity among these genetic alterations. Finally, BCOR mutations tended to be associated with an inferior outcome in a cohort of 422 CN-AML patients (25.6% vs 56.7% overall survival at 2 years; P = .032). Our results for the first time implicate BCOR in CN-AML pathogenesis.

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Whole-Exome Sequencing Of Adult Philadephia-Negative Acute Lymphoblastic Leukemia From Diagnosis To Relapse After Allogeneic Hematopoietic Stem Cell Transplantation Reveals Clonal Evolution and Relapse-Associated Mutated Genes

Blood ◽

10.1182/blood.v122.21.156.156 ◽

2013 ◽

Vol 122 (21) ◽

pp. 156-156

Author(s):

Haowen Xiao ◽

Yi Luo ◽

Xiaoyu Lai ◽

Jimin Shi ◽

Yamin Tan ◽

...

Keyword(s):

Exome Sequencing ◽

Whole Exome Sequencing ◽

Somatic Mutations ◽

Lymphoblastic Leukemia ◽

Gene Mutations ◽

Genetic Alterations ◽

Sequencing Analysis ◽

Hematopoietic Stem ◽

Whole Exome ◽

Matched Samples

Abstract Introduction Although steady progress of effective chemotherapy in childhood acute lymphoblastic leukemia (ALL) carried with exceeding 80% of individuals now cured, the majority of adult patients with ALL are not cured by chemotherapy, and allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the only curative option. However, relapse remains the most leading cause of death after allo-HSCT. Adverse genetic alterations are generally accepted to be responsible for treatment failure and relapse. Several structural chromosomal alterations including rearrangement of the myeloid-lymphoid or mixed-lineage leukemia gene (MLL) and Philadelphia chromosome (Ph), have been mostly found in relapsed ALL. However, many Ph-negative (Ph-) ALL patients with normal karyotype , lacking known risk factors, also experienced relapse. The underlying pathologic determinants leading to relapse and prognostic markers in these cases remain poorly understood. More importantly, allo-HSCT is a distinct treatment option from tradtional chemotherapy and has 2 important forms to eliminate and select on malignant cells. The malignant cells that go on to causing relapse must initially survive ablation of chemotherapy before allo-HSCT and conditioning regimen in allo-HSCT. Then, after allo-HSCT, they must survive the effect of graft-versus- leukemia (GVL) reaction. Following this rationale, we hypothesized that there may be pivotal genetic causes confer leukemic cells a fitness advantage to undergo huge selective pressures and expand after allo-HSCT. To elucidate the genomic basis underlying relapse after allo-HSCT to aid to discover novel predictive biomarkers and identify therapeutic targets, we carried out the first whole-exome sequencing analysis in longitudinal matched samples from diagnosis to relapse after allo-HSCT in adult patients with the most common subtype of ALL, Ph- B-cell ALL (B-ALL). Methods Whole-exome sequencing was conducted for 9 genomic DNA samples from 3 relapsed cases with Ph- B-ALL (discovery cohort) at 3 specific time points including: diagnosis, complete remission (CR) after induction chemotherapy before allo-HSCT, relapse after allo-HSCT to discover candidate relapse-associated mutated genes. We identified putative somatic mutations by comparing each tumor ( diagnostic samples or relapsed samples) to normal (CR samples) from the same patient. To confirm candidate somatic gene mutations, screen relapse-associated gene mutations and define the frequency of somatic mutations identified by whole-exome sequencing analysis, we further carried out target genes whole coding regions sequencing in an ALL extended validation cohort including 58 adult Ph- B-ALL cases, where 27 patients experienced relapse at a median time of 6.5 (range 2-33) months after allo-HSCT and 31 patients did not relapse after allo-HSCT at a median follow-up for 34 (range 12–56) months. Results (1) We discovered novel associations of recurrently mutated genes (CREBBP, KRAS, PTPN21) with the pathogenesis of adult Ph- B-ALL relapse after allo-HSCT, which were mutated in at least two relapsed cases, but were not mutated in non- relapsed patients. (2) The generation of high-depth whole-exome sequencing data in longitudinal matched samples from diagnosis to relapse after allo-HSCT in initial 3 patients allowed us to directly assessed the evolution of somatic mutations. Our data suggested that in the progression of leukemia relapse after allo-HSCT, the relapse clone had a clear relationship to the diagnosis clone, either arising from a subclone already exsiting in the diagnostic tumor, or originating from a common preleukemic progenitor with the diagnosis clone. In the latter pattern, the relapse clone acquires new genetic alterations while retaining some but not all of the alterations found in the diagnostic tumor. In contrast, in some cases, leukemia recurrences afer allo-HSCT may be composed of second malignancies with completely distinct sets of mutations from the primary tumor. Conclusions Our study is the first to explore genetic basis of adult Ph- B-ALL from diagnosis to relapse after allo-HSCT over time, which will provide novel genetic biomarkers on risk “index” to improve individualized treatment intensification and intervention strategies, and potential therapeutic targets for Ph--ALL relapse after allo-HSCT. Disclosures: No relevant conflicts of interest to declare.

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NAPG mutation in family members with hereditary hemorrhagic telangiectasia in China

BMC Pulmonary Medicine ◽

10.1186/s12890-021-01524-4 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Yu Xu ◽

Yong-Biao Zhang ◽

Li-Jun Liang ◽

Jia-Li Tian ◽

Jin-Ming Lin ◽

...

Keyword(s):

Exome Sequencing ◽

Whole Exome Sequencing ◽

Hereditary Hemorrhagic Telangiectasia ◽

Sanger Sequencing ◽

Conformational Stability ◽

Sequencing Analysis ◽

Large Pedigree ◽

Whole Exome ◽

Variant Filtering ◽

Genetic Contributions

Abstract Background Hereditary hemorrhagic telangiectasia (HHT) is a disease characterized by arteriovenous malformations in the skin and mucous membranes. We enrolled a large pedigree comprising 32 living members, and screened for mutations responsible for HHT. Methods We performed whole-exome sequencing to identify novel mutations in the pedigree after excluding three previously reported HHT-related genes using Sanger sequencing. We then performed in silico functional analysis of candidate mutations that were obtained using a variant filtering strategy to identify mutations responsible for HHT. Results After screening the HHT-related genes, activin A receptor-like type 1 (ACVRL1), endoglin (ENG), and SMAD family member 4 (SMAD4), we did not detect any co-segregated mutations in this pedigree. Whole-exome sequencing analysis of 7 members and Sanger sequencing analysis of 16 additional members identified a mutation (c.784A > G) in the NSF attachment protein gamma (NAPG) gene that co-segregated with the disease. Functional prediction showed that the mutation was deleterious and might change the conformational stability of the NAPG protein. Conclusions NAPG c.784A > G may potentially lead to HHT. These results expand the current understanding of the genetic contributions to HHT pathogenesis.

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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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Identification of Two Novel Mutations in PKHD1 Gene from Two Families with Polycystic Kidney Disease by Whole Exome Sequencing

Current Genomics ◽

10.2174/1389202922666210219111810 ◽

2021 ◽

Vol 22 ◽

Author(s):

Masoud Heidari ◽

Hamid Gharshasbi ◽

Alireza Isazadeh ◽

Morteza Soleyman-Nejad ◽

Mohammad Hossein Taskhiri ◽

...

Keyword(s):

Kidney Disease ◽

Exome Sequencing ◽

Polycystic Kidney Disease ◽

Whole Exome Sequencing ◽

Novel Mutation ◽

Genetic Diagnosis ◽

Genetic Alterations ◽

Polycystic Kidney ◽

Novel Mutations ◽

Whole Exome

Background:: Polycystic kidney disease (PKD) is an autosomal recessive disorder resulting from mutations in the PKHD1 gene on chromosome 6 (6p12), a large gene spanning 470 kb of genomic DNA. Objective: The aim of the present study was to report newly identified mutations in the PKHD1 gene in two Iranian families with PKD. Materials and Methods: Genetic alterations of a 3-month-old boy and a 27-year-old girl with PKD were evaluated using whole-exome sequencing. The PCR direct sequencing was performed to analyse the co-segregation of the variants with the disease in the family. Finally, the molecular function of the identified novel mutations was evaluated by in silico study. Results: In the 3 month-old boy, a novel homozygous frameshift mutation was detected in the PKHD1 gene, which can cause PKD. Moreover, we identified three novel heterozygous missense mutations in ATIC, VPS13B, and TP53RK genes. In the 27-year-old woman, with two recurrent abortions history and two infant mortalities at early weeks due to metabolic and/or renal disease, we detected a novel missense mutation on PKHD1 gene and a novel mutation in ETFDH gene. Conclusion: In general, we have identified two novel mutations in the PKHD1 gene. These molecular findings can help accurately correlate genotype and phenotype in families with such disease in order to reduce patient births through preoperative genetic diagnosis or better management of disorders.

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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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Whole Exome Sequencing of Six Chinese Families With Hereditary Non-Syndromic Hearing Loss: A Genetic Etiology Study

10.21203/rs.3.rs-132767/v1 ◽

2020 ◽

Author(s):

Pengfei Liang ◽

Fengping Chen ◽

Shujuan Wang ◽

Qiong Li ◽

Wei Li ◽

...

Keyword(s):

Hearing Loss ◽

Exome Sequencing ◽

Whole Exome Sequencing ◽

Sanger Sequencing ◽

Large Scale ◽

Autosomal Recessive ◽

Autosomal Recessive Inheritance ◽

Chinese Families ◽

Whole Exome ◽

Syndromic Hearing Loss

Abstract Background: Hereditary non-syndromic hearing loss (NSHL) has a high genetic heterogeneity with >152 genes identified as associated molecular causes. The present study aimed to detect the possible damaging variants of the deaf probands from six unrelated Chinese families.Methods: After excluding the mutations in the most common genes, GJB2 and SLC26A4, 12 probands with prelingual deafness and autosomal recessive inheritance were evaluated by whole-exome sequencing (WES). All the candidate variants were verified by Sanger sequencing in all patients and their parents.Results: Biallelic mutations were identified in all deaf patients. Among these six families, 10 potentially causative mutations, including 3 reported and 7 novel mutations, in 3 different deafness-associated autosomal recessive (DFNB) genes (MYO15A, COL11A2, and CDH23) were identified. The mutations in MYO15A were frequent with 7/10 candidate variants. Sanger sequencing confirmed that these mutations segregated with the hearing loss of each family.Conclusions: Next-generation sequencing (NGS) approach becomes more cost-effective and efficient when analyzing large-scale genes compared to the conventional polymerase chain reaction-based Sanger sequencing, which is often used to screen common deafness-related genes. The current findings further extend the mutation spectrum of hearing loss in the Chinese population, which has a positive significance for genetic counseling.

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