Potential Role of STAG1 Mutations in Genetic Predisposition to Childhood Hemato-Oncological Diseases

Abstract Background. Genetic predisposition has been recently assessed in about 5-10% of pediatric tumors, including leukemias. Among genetic syndromes associated with an increased risk of developing leukemia, a possible link has been proposed for Cohesinopathies, like Cornelia de Lange Syndrome (CdLS), caused by mutations in Cohesin family genes. Somatic mutation in these genes, known for their fundamental role in cell cycle and DNA repair mechanisms, are described in myeloid malignancies (10-20% of AML, 50% of DS-AMKL, 5-15% of MDS and 10% of MPN) and solid tumours. We previously described the first germline variant of NIPBL in a CdL case with acute lymphoblastic leukemia (ALL). The identification of germline variants can provide new insights in the pathogenesis of hemato-oncological diseases. Objectives. This study aims to define the association of germline variants of the Cohesin family genes with predisposition to pediatric hemato-oncological diseases, including ALL. Functional assays have been performed on a STAG1 unique variant. Methods. We set up an NGS capture-base DNA panel, including 40 genes associated with 8 pathways associated to ALL predisposition, including Cohesin genes. Patients' bone marrow (BM) diagnostic samples have been analyzed together with a germline tissue (BM at remission or buccal brush). Bioinformatic analysis has been carried out by Sophia DDM software (Variant Fraction >5%; Coverage at least 500X; Variant Allele Frequency 1%) and variants were annotated as certainly pathogenic, potentially pathogenic or VUS. A Lymphoblastoid Cell Line (LCL) was obtained from PB of a MDS patient, then used for functional studies. Genomic stability and X-ray induced DNA-damage repair were evaluated in the STAG1-mutated LCL by Sister Chromatids Exchange (SCE) assay, cell cycle analysis (Propidium Iodide) and pH2AX staining. Results. Overall, 117 consecutive pediatric ALL patients and sporadic cases with familial recurrency of cancer or other hematological disorders have been analyzed through the NGS panel. A total of 225 mutations were identified in ALL, distributed as it follows: 77 variants in Transcription factors, 68 in Signaling-associated genes, 56 variants in RAS pathway, 44 in chromatin modifier genes, 38 variants in B-cell differentiation genes, 30 in DNA repair genes and 16 variants in genes associated to predisposition. Overall, 12/117 unique variants (5%) were found in Cohesins family genes; 11/12 were germline, while a single somatic variant was found in STAG2 in the same position as a germline variant. In details, 3 mutations were found in BRD2 gene while 2 variants were identified in STAG1, STAG2 and NIPBL genes, respectively. Only 1 variant was detected in SMC1A, SMC3 and BRD2. No mutations were detected in HDAC8 and RAD21 genes. Interestingly, the Arg1167Gln variant in STAG1 is located in a highly conserved region involved in solid tumors. A STAG1 variant (Arg1187Gln) was identified in a pediatric MDS patient, which was then further characterized in the LCL. First, we evaluated the effects on chromosomal stability, and we observed a higher number of abnormal chromatids exchanges in mutated LCL (STAG1 patient mean = 4.8 exchanges/metaphase compared to four control LCLs mean = 3.05; p <0.0001), as shown in the attached figure. Differences in cell cycle phases between STAG1-mutated LCL and controls, even after X-ray irradiation (3Gy - 6Gy), are not significant. Mutated cells showed a significantly lower capability to repair DNA after an ionizing radiation. γH2AX phosphorylation status of STAG1 mutated LCL appears higher also in basal condition (T0: 1.7X, p <0.01; T24: 2.2X, p <0.0001; T48: 2.4X, p <0.0001; MFI STAG1 over MFI control LCLs) and it remains at higher levels at 48 hours after the irradiation (T48: 3.7X, p<0.001 [3Gy]; 4.1X, p<0.0001 [6Gy]; MFI STAG1 over MFI control LCLs). These data are confirmed by the ratio of the STAG1-mutated LCL median MFI over controls (T48/T0: 3.42 vs 0.92, p <0.0001 [3Gy]; T48/T0: 2.33 vs 0.64, p <0.01 [6Gy]). Conclusion. In pediatric hematological diseases, NGS screening showed several variants among Cohesin genes, with either a potentially pathogenic or unknown significance. Our study confirms a considerable effect of a STAG1 germline variant on genetic instability that leads to oncogenesis, opening new scenarios for Cohesins' contribution to genetic predisposition to leukemias. Figure 1 Figure 1. Disclosures Biondi: Colmmune: Honoraria; Incyte: Consultancy, Other: Advisory Board; Amgen: Honoraria; Bluebird: Other: Advisory Board; Novartis: Honoraria.

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Haplotypes of DNA repair and cell cycle control genes, X-ray exposure, and risk of childhood acute lymphoblastic leukemia

Cancer Causes & Control ◽

10.1007/s10552-011-9848-y ◽

2011 ◽

Vol 22 (12) ◽

pp. 1721-1730 ◽

Cited By ~ 14

Author(s):

Anand P. Chokkalingam ◽

Karen Bartley ◽

Joseph L. Wiemels ◽

Catherine Metayer ◽

Lisa F. Barcellos ◽

...

Keyword(s):

Cell Cycle ◽

Dna Repair ◽

Acute Lymphoblastic Leukemia ◽

Lymphoblastic Leukemia ◽

Cell Cycle Control ◽

Childhood Acute Lymphoblastic Leukemia ◽

X Ray

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Are Pathogenic Germline Variants in Metastatic Melanoma Associated with Resistance to Combined Immunotherapy?

Cancers ◽

10.3390/cancers12051101 ◽

2020 ◽

Vol 12 (5) ◽

pp. 1101 ◽

Cited By ~ 1

Author(s):

Teresa Amaral ◽

Martin Schulze ◽

Tobias Sinnberg ◽

Maike Nieser ◽

Peter Martus ◽

...

Keyword(s):

Dna Repair ◽

Cox Regression ◽

Progression Free Survival ◽

Advanced Melanoma ◽

Germline Variants ◽

Cox Regression Analysis ◽

Dna Repair Mechanisms ◽

Germline Variant ◽

Repair Mechanisms ◽

Combined Immunotherapy

Background: Combined immunotherapy has significantly improved survival of patients with advanced melanoma, but there are still patients that do not benefit from it. Early biomarkers that indicate potential resistance would be highly relevant for these patients. Methods: We comprehensively analyzed tumor and blood samples from patients with advanced melanoma, treated with combined immunotherapy and performed descriptive and survival analysis. Results: Fifty-nine patients with a median follow-up of 13 months (inter quartile range (IQR) 11–15) were included. Interestingly, nine patients were found to have pathogenic or likely pathogenic (P/LP) germline variants in one of these genes: BRCA2, POLE, WRN, FANCI, CDKN2A, BAP1, PALB2 and RAD54B. Most of them are involved in DNA repair mechanisms. Patients with P/LP germline variants had a significantly shorter progression-free survival (PFS) and melanoma specific survival (MSS) compared to patients without P/LP germline variants (HR = 2.16; 95% CI: 1.01–4.64; p = 0.048 and HR = 3.21; 95% CI: 1.31–7.87; p = 0.011, respectively). None of the patients with a P/LP germline variant responded to combined immunotherapy. In the multivariate Cox-regression analysis, presence of a P/LP germline variant, S100B and lactate dehydrogenase (LDH) remained independently significant factors for MSS (p = 0.036; p = 0.044 and p = 0.001, respectively). Conclusions: The presence of P/LP germline variants was associated with resistance to combined immunotherapy in our cohort. As genes involved in DNA repair mechanisms are also involved in lymphocyte development and T-cell differentiation, a P/LP germline variant in these genes may preclude an antitumor immune response.

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XRRA1 Targets ATM/CHK1/2-Mediated DNA Repair in Colorectal Cancer

BioMed Research International ◽

10.1155/2017/5718968 ◽

2017 ◽

Vol 2017 ◽

pp. 1-11 ◽

Cited By ~ 1

Author(s):

Wenjun Wang ◽

Minzhang Guo ◽

Xiaojun Xia ◽

Chao Zhang ◽

Yuan Zeng ◽

...

Keyword(s):

Colorectal Cancer ◽

Cell Cycle ◽

Dna Repair ◽

Cell Cycle Arrest ◽

Human Tumor ◽

Repair Pathway ◽

X Ray ◽

Cycle Arrest ◽

Damage Response ◽

P21 Proteins

X-ray radiation resistance associated 1 (XRRA1) has been found to regulate the response of human tumor and normal cells to X-radiation (XR). Although XRRA1 overexpression is known to be involved in cancer cell response to XR, there are no reports about whether the expression of XRRA1 in tumors can adjust radioresistance. It is widely known that cell cycle arrest could cause radioresistance. We found that blocked XRRA1 expression could lead to cell cycle G2/M arrest by the regulation of cyclin A, cyclin E, and p21 proteins in colorectal cancer (CRC) and expression of XRRA1 reduced cell cycle arrest and increased cell proliferation in CRC. However, whether regulation of the cell cycle by XRRA1 can influence radioresistance is poorly characterized. Correspondingly, DNA repair can effectively lead to radioresistance. In our study, when cancer cells were exposed to drugs and ionizing radiation, low expression of XRRA1 could increase the phosphorylation of DNA repair pathway factors CHK1, CHK2, and ATM and reduce the expression of γ-H2AX, which is believed to participate in DNA repair in the nucleus. Crucially, our results identify a novel link between XRRA1 and the ATM/CHK1/2 pathway and suggest that XRRA1 is involved in a DNA damage response that drives radio- and chemoresistance by regulating the ATM/CHK1/2 pathway.

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Involvement of Death-Associated Protein Kinases In DNA Damage Responses of B-ALL Cells.

Blood ◽

10.1182/blood.v116.21.3369.3369 ◽

2010 ◽

Vol 116 (21) ◽

pp. 3369-3369

Author(s):

Magali Humbert ◽

Michaela Medova ◽

Barbara Geering ◽

Wieslawa Blank-Liss ◽

Hans-Uwe Simon ◽

...

Keyword(s):

Cell Cycle ◽

Dna Damage ◽

Dna Repair ◽

Gamma Irradiation ◽

Protein Kinases ◽

Dna Damage Response ◽

Lymphoblastic Leukemia ◽

Damage Response ◽

Dna Damage Responses

Abstract Abstract 3369 Intact DNA damage response pathways are important for genomic fidelity of cells in order to avoid tumor formation. On the other hand, inhibition of DNA repair provides an important mechanism to enhance the therapeutic efficacy of DNA damaging agents such as gamma-irradiation. Thus, it is important to identify novel players in DNA damage response that might represent novel targets for combination therapies. Death-associated protein kinases (DAPK) are serine/threonine kinases believed to be involved in cell death and autophagy mechanisms, whereby particularly the role of DAPK1 has previously been investigated. The DAPK family is composed of five members: DAPK1, DAPK2 (or DRP-1), DAPK3 (or ZIP kinase), DRAK1 and DRAK2. DAPK1 and DAPK2 share 80% homology in the catalytic domain. Generally, the role of DAPK in DNA damage responses is not well studied. To analyze the role of DAPK1 and DAPK2 in response to gamma-irradiation, we used p53 wild-type REH B-cell acute lymphoblastic leukemia (B-ALL) cells as a model. In response to irradiation, DAPK1 protein expression increased paralleled by an increased of total p53, phospho-Ser20-p53 and p21WAF1/CIP1. DAPK2 expression, however, did not increase. Since upregulation of p21WAF1/CIP1, a classical p53 target in response to DNA damage leads to cell cycle arrest, we asked whether knocking down DAPK1 or DAPK2 might affect the cell cycle. Interestingly, knocking down DAPK2 but not DAPK1 led to a significant increase of S-phase cells upon irradiation. Moreover, knocking down DAPK2 attenuated the induction of DAPK1 upon irradiation indicating a DAPK2-DAPK1 cascade in DNA damage responses. Next, given the significant role of p21WAF1/CIP1 and p53 in DNA damage responses, we tested if DAPK2 might directly participate in a novel signaling pathway by interacting with these proteins. Indeed, pull down assays revealed that p21WAF1/CIP1 and p53 are novel DAPK2 interacting proteins. Clearly, further experiments are needed to define the DAPK2-DAPK1-p53- p21WAF1/CIP1 network in DNA repair pathways. In conclusion, we identified a novel role for DAPK1 and DAPK2 in DNA damage responses of B-ALL cells and propose a novel DAPK2/DAPK1/p53/ p21WAF1/CIP1 DNA damage regulatory pathway. Disclosures: No relevant conflicts of interest to declare.

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Exome Sequencing of Late Recurrence T-Cell Acute Lymphoblastic Leukemia in Children Confirms Second Leukemia and Exposes Predisposition Candidate Genes

Blood ◽

10.1182/blood.v118.21.755.755 ◽

2011 ◽

Vol 118 (21) ◽

pp. 755-755

Author(s):

Esmé Waanders ◽

Vincent H.J. van der Velden ◽

Tomasz Szczepanski ◽

Lisenka E.L.M. Vissers ◽

Joep de Ligt ◽

...

Keyword(s):

Exome Sequencing ◽

Genetic Predisposition ◽

Lymphoblastic Leukemia ◽

Conservation Score ◽

Splice Junction ◽

Regulatory Sequences ◽

Nonsense Mutations ◽

Germline Variants ◽

Missense Variants ◽

Frame Shift

Abstract Abstract 755 Second hematologic malignancies in non-syndromic children without a pronounced family history for cancer may be mistaken for relapses or therapy-related malignancies. Recently, we characterized diagnosis and presumed relapse samples of 22 patients with very late disease recurrences (>2.5 years), and identified 8 patients with leukemic presentations that were fully discordant at the level of TCR-rearrangements and DNA copy number aberrations (J Clin Oncol 2011; 29:1643-9). One of these patients showed a germline deletion comprising the recombination activating genes RAG1 and RAG2, and regulatory sequences of LMO2, genes frequently affected somatically in T-ALL, suggesting a genetic predisposition to leukemia. In the current study, we performed exome sequencing to assess whether consecutive leukemic presentations in such patients are indeed fully discordant, also at the sequence level, and to identify candidate pathogenic germline variants that point at a genetic predisposition. We sequenced the exomes in samples obtained from 2 consecutive leukemic presentations, and intermittent complete remissions, from 2 patients with very late disease recurrences (>2.5 years) and discordant leukemic presentations. We found on average 26,600 variants per exome. Recurrent variants recorded in the dbSNP and/or 1000 Genomes databases, or those present in our in-house database (>300 exomes) were excluded, resulting in an average of 989 private variants per exome. We divided these variants into 3 groups (i) somatic variants shared between the consecutive leukemic samples but not detected in remission (ii) somatic variants present in only one of the leukemic samples and (iii) germline variants present in the remission samples of the patients. All candidate somatic variants shared between two consecutive leukemic samples were re-sequenced by Sanger sequencing and were shown to be either present in all three samples, and thus originally missed in the remission sample, or falsely detected in one or more leukemic samples. Therefore, we conclude that in both patients no somatic variants were shared between the first and second leukemic presentations, which confirms that these patients suffered from clonally unrelated second T-ALLs. From all somatic variants present in only one of the leukemic samples, we focused on variants in exons or splice junction sites. We found 4 nonsense mutations, 9 frame-shift mutations, 12 in-frame in/dels and 7 non-synonymous missense variants with a high interspecies conservation score (PhyloP>3.0), mostly affecting genes implicated in oncogenesis like PTEN, TET3, CDKN2C, CD109, and GLRX2. Each leukemic sample harbored 2–11 of these putative deleterious variants. In the germline of the two patients, we identified 314 and 190 non-synonymous unknown variants in exons or splice junction sites, respectively. Among these were 12 nonsense mutations, 7 canonical splice-site mutations, 20 frame-shift mutations, 11 in-frame in/dels and 143 non-synonymous missense variants at highly conserved positions (PhyloP>3.0). Filtering of these variants for known T-ALL associated genes resulted in several interesting novel candidate predisposing genes such as, among others, RANBP17 and HOXC13. Sequencing of the entire RANBP17 open reading frame in a cohort of 24 sporadic T-ALL samples revealed that this gene was somatically affected in one of them. In conclusion, we confirmed by exome sequencing that consecutive leukemic presentations in patients with late T-ALL recurrences may be fully discordant and thus represent independent leukemia occurrences, most likely caused by predisposing germline abnormalities. Disclosures: No relevant conflicts of interest to declare.

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Promoter SNPs in G1/S checkpoint regulators and their impact on the susceptibility to childhood leukemia

Blood ◽

10.1182/blood-2006-02-003236 ◽

2006 ◽

Vol 109 (2) ◽

pp. 683-692 ◽

Cited By ~ 43

Author(s):

Jasmine Healy ◽

Hélène Bélanger ◽

Patrick Beaulieu ◽

Mathieu Larivière ◽

Damian Labuda ◽

...

Keyword(s):

Cell Cycle ◽

Childhood Leukemia ◽

Kinase Inhibitor ◽

Gene Dosage ◽

Lymphoblastic Leukemia ◽

Cell Cycle Checkpoint ◽

Nucleotide Polymorphisms ◽

Variable Expression ◽

Increased Risk ◽

Inhibitor Genes

AbstractMutations leading to the alteration of cell-cycle checkpoint functions are a common feature of most cancers. Because of the highly regulated nature of the cell cycle, it seems likely that variation in gene dosage of key components due to functional regulatory polymorphisms could play an important role in cancer development. Here we provide evidence of the involvement of promoter single-nucleotide polymorphisms (pSNPs) in the cyclin-dependent–kinase inhibitor genes CDKN2A, CDKN2B, CDKN1A, and CDKN1B in the etiology of childhood pre-B acute lymphoblastic leukemia (ALL). A case-control study, conducted in 240 patients with pre-B ALL and 277 healthy controls, combined with a family-based analysis using 135 parental trios, all of French-Canadian origin, were used to evaluate single-site genotypic as well as multilocus haplotypic associations for a total of 10 pSNPs. Using both study designs, we showed evidence of association between variants CDKN2A −222A, CDKN2B −593A, and CDKN1B −1608A, and an increased risk of ALL. These findings suggest that variable expression levels of cell-cycle inhibitor genes CDKN2A, CDKN2B, and CDKN1B due to regulatory polymorphisms could indeed influence the risk of childhood pre-B ALL and contribute to carcinogenesis.

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Selected genetic determinants of head and neck squamous cell carcinoma including of some genes for cell cycle control and DNA repair

Postępy Higieny i Medycyny Doświadczalnej ◽

10.5604/01.3001.0012.1150 ◽

2018 ◽

Vol 72 ◽

pp. 526-539

Author(s):

Karolina Gołąbek ◽

Jadwiga Gaździcka ◽

Zofia Ostrowska

Keyword(s):

Cell Cycle ◽

Squamous Cell Carcinoma ◽

Dna Repair ◽

Cell Carcinoma ◽

Head And Neck ◽

Squamous Cell ◽

Genetic Predisposition ◽

Cell Cycle Control ◽

Neck Squamous Cell Carcinoma ◽

Specific Gene

The head and neck squamous cell carcinoma, which are located in different anatomical structures like: oral cavity, throat and larynx, are becoming a major clinical problem due to the ever increasing number of cases and also due to difficulty with treating patients. Among the risks factors associated with that group of cancers the exogenous factors such as: exposure to carcinogens in tobacco smoke, alcohol abuse, impropriety dietary habits, poor oral hygiene as well as infection with the human papillomavirus and infection with Epstein-Barr virus, are well-known and had been described. The second potential group of the risk factors are endogenous factors such as genetic predisposition and disorder immune system. Recently, in view of technical progress of molecular biology, the aim of researches became the polymorphisms the most of genes encoding proteins with a wide variety of functions. In the present study we comprehensively discussed the problems associated with the molecular basis of development the head and neck squamous-cell carcinoma. More specifically, this study investigated the single nucleotide polymorphisms and mutations of genes involved in cell cycle control (TP53, p73, CDKN1A, CDKN2A, MDM2, E2F1, E2F2 and EGFR) and in different pathways of DNA repair (XPA, XPB, XPC, XPD, XPF, XPG, ERCC1, OGG1, XRCC1, NBS1, RAD51, BRCA2, XRCC2, XRCC3, XRCC5 and XRCC6). The results of studies indicate a mixed picture of genetic predisposition of the head and neck cancers. Nevertheless, it can be observed that a single polymorphism of specific gene moderately influence on the pathogenesis of that cancers. Commonly, coexistence of few polymorphism at the same time may increase the cancer risk significantly. It is undeniable that the issues discussed in that article should be the subject of further analysis.

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Rare Variants Affecting the Fanconi Anaemia DNA Repair Genes Associate with Increased Risk for AML

Blood ◽

10.1182/blood.v128.22.41.41 ◽

2016 ◽

Vol 128 (22) ◽

pp. 41-41

Author(s):

Kyaw Ze Ya Maung ◽

Paul Leo ◽

Anna L Brown ◽

Mahmoud A Bassal ◽

Debora A. Casolari ◽

...

Keyword(s):

Dna Repair ◽

Bone Marrow Failure ◽

Subsequent Development ◽

Fanconi Anaemia ◽

Gene Variants ◽

Healthy Controls ◽

Hematopoietic Stem ◽

Germline Variants ◽

Mutational Burden ◽

Increased Risk

Abstract While there have been extensive studies to define the roles of recurrent somatic mutations in AML, the contribution of germline variants to AML initiation and progression is less well established. DNA repair disorders often predispose patients to developing myeloid malignancies. In particular, biallelic mutations affecting FANC genes cause the recessive heritable bone marrow failure syndrome Fanconi Anemia (FA), which is associated with >800-fold increased risk of progression to AML. A recent explosion of cancer predisposition studies has also revealed the importance of germlineFANC variants in elevated cancer risk (Cancer Treat Rev 2012; 38:89). To investigate the role of FANC gene variants in AML we have performed a case-control study, analyzing rare, deleterious somatic and germline variants for the 19 FANC genes in adult AML and healthy controls cohorts. Whole exome sequencing was performed on diagnosis samples from 131 adult Caucasian AML patients from two major Australian centers, and a cohort of 329 healthy females. We identified rare Tier 1 variants using a minor allele frequency (MAF) < 0.001, as reported in common dbSNP137, 1000 Genome and NHLBI-ESP project databases. Combined Annotation Dependent Depletion algorithm (CADD, Nat Genet 2014; 46: 310) >10 was used to filter for FANC gene variants with high probability of pathogenicity. Sanger sequencing of matched tumour/non-tumour DNA showed the large majority of variants tested to be germline (90%), consistent with previous studies reporting that somatic FANC genes variants are extremely rare in AML (< 1%). Overall, we identified 52 FANC gene variants in 44 cases with 34% of AML cases carrying one or more variant. For independent validation we determined the presence of somatic and germline FANC variants in the TCGA AML cohort using an identical pipeline and filtering analysis. In line with our results, we found that 36% of TCGA AML patients carry at least one germline FANC variant. We investigated known disease-causing (D-C) variants in these two AML cohorts using the FA (FAMutdb) and breast cancer (kConFab and BIC) mutation databases. We found 8 D-C FANC variants in the Australian AML cohort and 5 in the TCGA cohort, with 1 variant present in both cohorts. Moreover, the frequency of D-C variants in our cohort of females with AML (n=51) is 13.7%, while the frequency in the healthy female cohort is 4.5%, comparable to that reported in the ESP database for female European-Americans (2.1%, Hum Mol Genet 2014; 23: 6815). Accordingly, we determined that deleterious FANC germline variants confer a significant increased risk of AML (P=0.018, OR=3.3 for the Australian AML cohort). Finally, we performed mutational burden analysis to investigate enrichment of variants associated with particular FANC genes across the AML cohort. This revealed a significant enrichment of FANCL variants in AML vs healthy controls (P=0.008, Figure 1). FANCL is the enzymatic component of the FA core complex that monoubiquitinates the FANCD2/I heterodimer initiating DNA repair, and its down-regulation has been linked to AML (Oncogene 2016; doi:10.1038). Several FANCL variants, found in our AML cohort, affect the catalytic RING domain and are of particular interest. These include a D-C null variant present in 2 patients, a frame shift variant in 2 patients who presented with AML at a very early age (27 and 46 years old), and a variant affecting a critical conserved residue required for monoubiquitination of FANCD2/I. In conclusion, we show enrichment of rare potentially deleterious FANC gene mutations in AML, associated with a 3-fold increased risk of developing the disease. We hypothesise that, in hematopoietic stem/progenitor cells, these variants confer a subtle defect in interstrand cross-link repair leading to an increased accumulation of mutations and subsequent development of AML. Consistent with this there have been several reports of defective DNA damage repair and increased sensitivity to DNA damaging agents in cells from FANC carriers compared to normal controls (Nat Commun 2014; 5:5496; Mutagenesis 2009; 24:67). Importantly, it is possible to target defects in several DNA repair pathways, and our finding identifies a group of AML patients who may benefit from approaches that target defective FA and homologous recombination pathways. Figure 1. A significant increase mutational burden of FANCL was observed in our AML cohort (line represents P=0.05). Figure 1. A significant increase mutational burden of FANCL was observed in our AML cohort (line represents P=0.05). Disclosures Gill: Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees.

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Associations of CDH1 germline variant location and cancer phenotype in families with hereditary diffuse gastric cancer (HDGC)

Journal of Medical Genetics ◽

10.1136/jmedgenet-2018-105361 ◽

2019 ◽

Vol 56 (6) ◽

pp. 370-379 ◽

Cited By ~ 10

Author(s):

Winifred Lo ◽

Bin Zhu ◽

Arvind Sabesan ◽

Ho-Hsiang Wu ◽

Astin Powers ◽

...

Keyword(s):

Breast Cancer ◽

Colorectal Cancer ◽

Gastric Cancer ◽

Young Age ◽

Hereditary Diffuse Gastric Cancer ◽

Diffuse Gastric Cancer ◽

Germline Variants ◽

Germline Variant ◽

Increased Risk ◽

E Cadherin

IntroductionHereditary diffuse gastric cancer (HDGC) is a cancer syndrome associated with variants in E-cadherin (CDH1), diffuse gastric cancer and lobular breast cancer. There is considerable heterogeneity in its clinical manifestations. This study aimed to determine associations between CDH1 germline variant status and clinical phenotypes of HDGC.MethodsOne hundred and fifty-two HDGC families, including six previously unreported families, were identified. CDH1 gene-specific guidelines released by the Clinical Genome Resource (ClinGen) CDH1 Variant Curation Expert Panel were applied for pathogenicity classification of truncating, missense and splice site CDH1 germline variants. We evaluated ORs between location of truncating variants of CDH1 and incidence of colorectal cancer, breast cancer and cancer at young age (gastric cancer at <40 or breast cancer <50 years of age).ResultsFrequency of truncating germline CDH1 variants varied across functional domains of the E-cadherin receptor gene and was highest in linker (0.05785 counts/base pair; p=0.0111) and PRE regions (0.10000; p=0.0059). Families with truncating CDH1 germline variants located in the PRE-PRO region were six times more likely to have family members affected by colorectal cancer (OR 6.20, 95% CI 1.79 to 21.48; p=0.004) compared with germline variants in other regions. Variants in the intracellular E-cadherin region were protective for cancer at young age (OR 0.2, 95% CI 0.06 to 0.64; p=0.0071) and in the linker regions for breast cancer (OR 0.35, 95% CI 0.12 to 0.99; p=0.0493). Different CDH1 genotypes were associated with different intracellular signalling activation levels including different p-ERK, p-mTOR and β-catenin levels in early submucosal T1a lesions of HDGC families with different CDH1 variants.ConclusionType and location of CDH1 germline variants may help to identify families at increased risk for concomitant cancers that might benefit from individualised surveillance and intervention strategies.

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Recurrent Germline Variant in the Cohesin Complex Gene RAD21 Predisposes Children to Lymphoblastic Leukemia and Lymphoma

Blood ◽

10.1182/blood-2021-150284 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 3358-3358

Author(s):

Anne Schedel ◽

Ulrike Anne Friedrich ◽

Rabea Wagener ◽

Juha Mehtonen ◽

Claudia Saitta ◽

...

Keyword(s):

Cell Cycle ◽

Acute Lymphoblastic Leukemia ◽

B Cell ◽

Pediatric Cancer ◽

Potential Role ◽

Lymphoblastic Leukemia ◽

Cohesin Complex ◽

Germline Variant ◽

Student’S T ◽

Student’S T Test

Abstract Introduction: Cohesin complex genes are commonly mutated in cancer particularly in myeloid malignancies. Yet patients with germline mutations in cohesin genes, leading to cohesinopathies like Cornelia-de-Lange syndrome (CdLS) are generally not known to be tumor-prone. The complex plays a major role in chromosome alignment and segregation (Uhlmann, Nature Reviews Molecular Cell Biology, 2016), homologous recombination-driven DNA repair (Ström et al., Molecular Cell, 2004) and regulation of gene expression (Busslinger et al., Nature, 2017). To deepen the understanding of cohesin variants in cancer predisposition, we performed TRIO Sequencing in two independent pediatric cancer cohorts. Thereby, we identified a novel recurrent heterozygous germline variant in the cohesin gene RAD21 not described in CdLS patients , located in the binding domain of the cofactors WAPL and PDS5B . Methods: Whole exome sequencing (WES) in a TRIO (child-parent datasets) setting was carried out in two independent, unselected cancer cohorts (TRIO-D, n=158 (Wagener et al., European Journal of Human Genetics, 2021) and TRIO-DD, n=60). To investigate the oncogenic potential of the novel RAD21 variant molecular and functional assessment was performed focusing on potential implications on the complex. Results: The newly identified RAD21 variant at amino acid position 298 resulting in a Proline to Serine (p.P298S) and a Proline to Alanine exchange, respectively, (p.P298A) is only rarely mutated in the general population (gnomAD database n=118,479; RAD21 p.P298S MAF <10 -6 and RAD21 p.P298A MAF <10 -5). While both patients did not show any signs of CdLS, they both have a remarkable family history of cancer. Patient 1 (13y) was diagnosed with T-cell acute lymphoblastic leukemia (T-ALL) whose father had died from breast cancer (41y), while patient 2 (2y) presented with precursor B-cell lymphoblastic lymphoma (pB-LBL) whose uncle had died from pediatric cancer of unknown subtype (8y). To assess the influence of RAD21 p.P298S/A on the binding capacity of the complex, RAD21 variants and the wildtype (WT) were cloned and transfected into HEK293T cells, respectively. Immunoprecipitation analysis of RAD21 with the cofactors WAPL and PDS5B showed no differential binding between the WT and the variants, suggesting that RAD21 p.P298S/A does not impact the formation of the complex. Nevertheless, on a transcriptional level 83 genes were significantly differentially expressed in RAD21 p.P298S and p.P298A compared to the wildtype (fc>1.5, adj. p-value <0.05) with enrichment of genes in p53 signaling pathways. We further observed an increased number of γH2AX and 53BP1 co-localized foci compared to the WT (p≤0.01; Student's t-test). In line, following ionizing radiation, primary patients' samples showed increased cell cycle arrest at G2/M cell-cycle stage compared to a healthy control (p.P298S: p=0.0049 [6Gy]; p=0.0026 [10Gy]; p.P298A: p=0.0054 [6Gy]; p=0.0006 [10Gy]; Student's t-test). For cross-validation of the germline variant RAD21 p.P298S/A and its potential role in pediatric lymphoblastic malignancies, we analysed a third cohort of 150 children with relapsed ALL (IntReALL) for RAD21 p.P298S/A. We again identified RAD21 p.P298A in a boy (12y) with B-cell precursor acute lymphoblastic leukemia. To compare our data to a non-pediatric cancer setting, a cohort of 2300 young adults (<51 years) with cancer was mined (MASTER program). Here, one patient carrying RAD21 p.P298A with a solid tumor was identified. Therefore, amongst all cohorts, RAD21 p.P298S/A was found to be enriched in pediatric vs. adult cancers (3/479 vs. 1/2299; Fisher's exact test; p=0.018). Conclusion: Taken together, we present for the first time the potential role of RAD21 germline variants in pediatric lymphoblastic malignancies. This may shed new light on the many roles of the cohesin complex and its implication outside the typical syndromal presentation. Disclosures No relevant conflicts of interest to declare.

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