158 research outputs found
Introme accurately predicts the impact of coding and noncoding variants on gene splicing, with clinical applications
No full textData source: Supplementary information, https://doi.org/10.1186/s13059-023-02936-7Predicting the impact of coding and noncoding variants on splicing is challenging, particularly in non-canonical splice sites, leading to missed diagnoses in patients. Existing splice prediction tools are complementary but knowing which to use for each splicing context remains difficult. Here, we describe Introme, which uses machine learning to integrate predictions from several splice detection tools, additional splicing rules, and gene architecture features to comprehensively evaluate the likelihood of a variant impacting splicing. Through extensive benchmarking across 21,000 splice-altering variants, Introme outperformed all tools (auPRC: 0.98) for the detection of clinically significant splice variants. Introme is available at https://github.com/CCICB/introme .Patricia J. Sullivan, Velimir Gayevskiy, Ryan L. Davis, Marie Wong, Chelsea Mayoh, Amali Mallawaarachchi, Yvonne Hort, Mark J. McCabe, Sarah Beecroft, Matilda R. Jackson, Peer Arts, Andrew Dubowsky, Nigel Laing, Marcel E. Dinger, Hamish S. Scott, Emily Oates, Mark Pinese, and Mark J. Cowle
Germline analysis of an international cohort of pediatric diffuse midline glioma patients
No full textAbstract Background Factors that drive the development of diffuse midline gliomas (DMG) are unknown. Our study aimed to determine the prevalence of pathogenic/likely pathogenic (P/LP) germline variants in pediatric patients with DMG. Methods We assembled an international cohort of 252 pediatric patients with DMG, including diffuse intrinsic pontine glioma (n=153), with germline whole genome or whole exome sequencing. Results We identified P/LP germline variants in cancer predisposition genes in 7.5% (19/252) of patients. Tumor profiles differed, with absence of somatic drivers in the PI3K/mTOR pathway in patients with germline P/LP variants versus those without (P = 0.023). P/LP germline variants were recurrent in homologous recombination (n=9; BRCA1, BRCA2, PALB2) and Fanconi anemia genes (n=4). Somatic findings established that the germline variants definitively contributed to tumorigenesis in at least 1% of cases. One patient with recurrent DMG and pathogenic germline variants (BRCA2, FANCE) showed near-complete radiological response to PARP and immune checkpoint inhibition. Conclusions Our study determined the prevalence of pathogenic germline variants in pediatric DMG, and suggests a role in tumorigenesis for a subset of patients
DIPG-76. PEDIATRIC PATIENTS WITH DIFFUSE MIDLINE GLIOMA DEMONSTRATE AN UNEXPECTED PREVALENCE OF GERMLINE VARIANTS IN HOMOLOGOUS RECOMBINATION GENES
No full textAbstract BACKGROUND Pediatric patients with Diffuse Midline Gliomas (DMG), H3K27M altered have a dismal prognosis and novel therapeutic approaches are urgently needed. Factors that drive development of pediatric DMG are unknown. METHODS To determine the prevalence of germline pathogenic/likely pathogenic variants (P/LPV) in DMG, we assembled an international cohort of 252 patients with germline whole genome or whole exome sequencing data, including diffuse intrinsic pontine glioma (DIPG; n=153), from Australian, European and North American centres. RESULTS We identified germline P/LPV in cancer predisposition genes in 7.5% (19/252) of patients, mainly homologous recombination (n=9; BRCA1, BRCA2, PALB2) and Fanconi anemia genes (n=4). Germline P/LPV in mismatch repair genes (MSH2, PMS2) were found in two patients. Two patients each had two separate germline P/LPV. The prevalence of germline P/LPV was not significantly different according to age, location of DMG nor H3K27M mutational status. Furthermore, tumor profiles differed, with absence of somatic drivers in the PI3K/mTOR pathway in patients with germline P/LPV compared to those without (P = 0.023). Knockdown of BRCA1 in DMG cell cultures sensitized tumor cells to PARP inhibition. Reflecting the potential therapeutic relevance of these findings, we describe one H3.3 K27M-mutant DMG patient with a pathogenic germline BRCA2 and FANCE variant and multiple recurrences, who was treated with a PARP inhibitor (olaparib) and immune checkpoint inhibitor, leading to a near complete radiological response after 4 months. CONCLUSION Our study is the largest series to date investigating germline P/LPV in cancer predisposition genes in DMG and provides new therapeutic insights. It is expected that these germline findings will also guide cascade testing for a proband’s relatives. Our data highlight the importance of germline testing in H3K27-altered DMG patients at diagnosis
Abstract A123: Reversal of glucocorticoid resistance in pediatric acute lymphoblastic leukemia is dependent on restoring BIM expression
No full textAbstract
Introduction: Acute lymphoblastic leukemia (ALL) is the most common pediatric malignancy. Glucocorticoids (e.g. dexamethasone, prednisolone) form a critical component of chemotherapy regimens for pediatric ALL and initial resistance to glucocorticoid therapy is predictive of poor outcome. Acquired resistance to glucocorticoids is common at relapse and is disparate compared with other chemotherapeutic drugs used in the treatment of pediatric ALL. Glucocorticoids induce apoptosis in lymphoid cells which is mediated through the glucocorticoid receptor (GR). Resistance to glucocorticoids in pediatric ALL is associated with upregulation of the oncogene C-MYC and failure to induce the proapoptotic gene BIM. The purpose of this study was to identify small molecules that are able to reverse glucocorticoid resistance in pediatric ALL patient-derived xenografts (PDXs). Methods: A 40,000 compound high-throughput screening (HTS) campaign was carried out to identify compounds that potentiated the cytotoxicity of dexamethasone against a pediatric ALL PDX derived from a patient with aggressive and fatal relapse. Cytotoxicity assays were carried out by alamarBlue assay. Gene expression analysis was carried out using Illumina microarrays. mRNA and protein analysis was carried out by RT-PCR and immunoblotting, respectively. Results: The HTS campaign identified a novel glucocorticoid sensitiser, 2-((4,5-dihydro-1H-imidazol-2-yl)thio)-N-isopropyl-N-phenylacetamide (GCS-3). The sensitizing effect was specific to glucocorticoids, with synergy observed when GCS-3 was combined with dexamethasone or prednisolone, but not with other chemotherapeutic drugs. Synergy was observed in a range of dexamethasone-resistant and -sensitive xenografts representative of B-ALL, T-ALL, early T-cell precursor ALL and Philadelphia chromosome positive ALL. GCS-3 required a functional GR to sensitise ALL xenografts to dexamethasone. Gene expression analysis showed that GCS-3 in the presence of dexamethasone upregulated a previously published gene expression signature induced by glucocorticoid treatment of B-ALL patients (Rhein et al. Leukemia, 2007, 21:897-905). Moreover, GCS-3 in combination with dexamethasone significantly downregulated C-MYC and upregulated BIM expression in a glucocorticoid-resistant ALL xenograft. Treatment with the GR antagonist, RU486, significantly abrogated the cytotoxic effects of the GCS-3/dexamethasone combination. Although RU486 increased C-MYC expression in the dexamethasone treated cells, there was no increase in C-MYC expression in the GCS-3/dexamethasone combination. RU486 suppressed the marked induction of BIM following treatment with the GCS-3/dexamethasone combination, indicating that BIM upregulation is critical for the cytotoxicity of the GCS-3/dexamethasone combination. The GCS-3/dexamethasone combination significantly increased binding of the GR to an intronic binding site at the BIM locus which is associated with BIM transcription and glucocorticoid sensitivity in pediatric ALL. Conclusion: This study describes the potential of the novel glucocorticoid sensitizing agent, GCS-3, as a biological tool to understand glucocorticoid action and resistance.
Citation Format: Richard B Lock, Cara E Toscan, Duohui Jing, Chelsea Mayoh. Reversal of glucocorticoid resistance in pediatric acute lymphoblastic leukemia is dependent on restoring BIM expression [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr A123. doi:10.1158/1535-7163.TARG-19-A12
Rascall: Rapid (Ra) screening (Sc) of RNA-seq data for prognostically significant genomic alterations in acute lymphoblastic leukaemia (ALL)
No full textData source: Accessible data, https://github.com/j-rehn/RaScALLRNA-sequencing (RNA-seq) efforts in acute lymphoblastic leukaemia (ALL) have identified numerous prognostically significant genomic alterations which can guide diagnostic risk stratification and treatment choices when detected early. However, integrating RNA-seq in a clinical setting requires rapid detection and accurate reporting of clinically relevant alterations. Here we present RaScALL, an implementation of the k-mer based variant detection tool km, capable of identifying more than 100 prognostically significant lesions observed in ALL, including gene fusions, single nucleotide variants and focal gene deletions. We compared genomic alterations detected by RaScALL and those reported by alignment-based de novo variant detection tools in a study cohort of 180 Australian patient samples. Results were validated using 100 patient samples from a published North American cohort. RaScALL demonstrated a high degree of accuracy for reporting subtype defining genomic alterations. Gene fusions, including difficult to detect fusions involving EPOR and DUX4, were accurately identified in 98% of reported cases in the study cohort (n = 164) and 95% of samples (n = 63) in the validation cohort. Pathogenic sequence variants were correctly identified in 75% of tested samples, including all cases involving subtype defining variants PAX5 p.P80R (n = 12) and IKZF1 p.N159Y (n = 4). Intragenic IKZF1 deletions resulting in aberrant transcript isoforms were also detectable with 98% accuracy. Importantly, the median analysis time for detection of all targeted alterations averaged 22 minutes per sample, significantly shorter than standard alignment-based approaches. The application of RaScALL enables rapid identification and reporting of previously identified genomic alterations of known clinical relevance.Jacqueline Rehn, Chelsea Mayoh, Susan L Heatley, Barbara J McClure, Laura N Eadie, Caitlin Schutz, David T Yeung, Mark J Cowley, James Breen, Deborah L Whit
Methotrexate-related central neurotoxicity: clinical characteristics, risk factors and genome-wide association study in children treated for acute lymphoblastic leukemia
No full textSymptomatic methotrexate-related central neurotoxicity (MTX neurotoxicity) is a severe toxicity experienced during acute lymphoblastic leukemia (ALL) therapy with potential long-term neurologic complications. Risk factors and long-term outcomes require further study. We conducted a systematic, retrospective review of 1,251 consecutive Australian children enrolled on Berlin-Frankfurt-Münster or Children's Oncology Group-based protocols between 1998-2013. Clinical risk predictors for MTX neurotoxicity were analyzed using regression. A genome-wide association study (GWAS) was performed on 48 cases and 537 controls. The incidence of MTX neurotoxicity was 7.6% (n=95 of 1,251), at a median of 4 months from ALL diagnosis and 8 days after intravenous or intrathecal MTX. Grade 3 elevation of serum aspartate aminotransferase (P=0.005, odds ratio 2.31 [range, 1.28–4.16]) in induction/consolidation was associated with MTX neurotoxicity, after accounting for the only established risk factor, age ≥10 years. Cumulative incidence of CNS relapse was increased in children where intrathecal MTX was omitted following symptomatic MTX neurotoxicity (n=48) compared to where intrathecal MTX was continued throughout therapy (n=1,174) (P=0.047). Five-year central nervous system relapse-free survival was 89.2±4.6% when intrathecal MTX was ceased compared to 95.4±0.6% when intrathecal MTX was continued. Recurrence of MTX neurotoxicity was low (12.9%) for patients whose intrathecal MTX was continued after their first episode. The GWAS identified single-nucletide polymorphism associated with MTX neurotoxicity near genes regulating neuronal growth, neuronal differentiation and cytoskeletal organization (P<1x10-6). In conclusion, increased serum aspartate aminotransferase and age ≥10 years at diagnosis were independent risk factors for MTX neurotoxicity. Our data do not support cessation of intrathecal MTX after a first MTX neurotoxicity event.Marion K. Mateos, Glenn M Marshall, Pasquale M. Barbaro, Michael C.J. Quinn, Carly George, Chelsea Mayoh, Rosemary Sutton, Tamas Revesz, Jodie E Giles, Draga Barbaric, Frank Alvaro, Françoise Mechinaud, Daniel Catchpoole, John A. Lawson, Georgia Chenevix-Trench, Stuart MacGregor, Rishi S.Kotecha, Luciano Dalla-Pozza, and Toby N. Traha
Unfoldment of self-reference in logic and in computer science
No full textThe paradoxes induced by unrestricted self-reference have been of interest since antiquity. In this paper the author approaches the interpretation of self-reference in natural languages from the perspective of linguist - using as insight the treatments of self-reference by Russel, Tarski, Kleene and Scott. The subject of self-reference is approached historically. The works of Russell are first discussed and then compared with the more general concepts introduced by Tarski. These are then discussed within the framework of Kleene's recursion (fixed point) theorem. Finally, the pioneering work of Scott is discussed and comparisons made with earlier work. A final section extends the notions discussed earlier into a linguistic/philosophical framework
The transcriptional co-repressor Runx1t1 is essential for MYCN-driven neuroblastoma tumorigenesis
No full textMYCN oncogene amplification is frequently observed in aggressive childhood neuroblastoma. Using an unbiased large-scale mutagenesis screen in neuroblastoma-prone transgenic mice, we identify a single germline point mutation in the transcriptional corepressor Runx1t1, which abolishes MYCN-driven tumorigenesis. This loss-of-function mutation disrupts a highly conserved zinc finger domain within Runx1t1. Deletion of one Runx1t1 allele in an independent Runx1t1 knockout mouse model is also sufficient to prevent MYCN-driven neuroblastoma development, and reverse ganglia hyperplasia, a known pre-requisite for tumorigenesis. Silencing RUNX1T1 in human neuroblastoma cells decreases colony formation in vitro, and inhibits tumor growth in vivo. Moreover, RUNX1T1 knockdown inhibits the viability of PAX3-FOXO1 fusion-driven rhabdomyosarcoma and MYC-driven small cell lung cancer cells. Despite the role of Runx1t1 in MYCN-driven tumorigenesis neither gene directly regulates the other. We show RUNX1T1 forms part of a transcriptional LSD1-CoREST3-HDAC repressive complex recruited by HAND2 to enhancer regions to regulate chromatin accessibility and cell-fate pathway genes.Jayne E. Murray, Emanuele Valli, Giorgio Milazzo, Chelsea Mayoh, Andrew J. Gifford, Jamie I. Fletcher, Chengyuan Xue, Nisitha Jayatilleke, Firoozeh Salehzadeh, Laura D. Gamble, Jourdin R. C. Rouaen, Daniel R. Carter, Helen Forgham, Eric O. Sekyere, Joanna Keating, Georgina Eden, Sophie Allan, Stephanie Alfred, Frances K. Kusuma, Ashleigh Clark, Hannah Webber, Amanda J. Russell, Antoine de Weck, Benjamin T. Kile, Martina Santulli, Piergiuseppe De Rosa, Emmy D. G. Fleuren, Weiman Gao, Lorna Wilkinson-White, Jason K. K. Low, Joel P. Mackay, Glenn M. Marshall, Douglas J. Hilton, Federico M. Giorgi, Jan Koster, Giovanni Perini, Michelle Haber, Murray D. Norri
International experience in the development of patient-derived xenograft models of diffuse intrinsic pontine glioma
No full textPurpose Diffuse intrinsic pontine glioma is the most aggressive form of high grade glioma in children with no effective therapies. There have been no improvements in survival in part due poor understanding of underlying biology, and lack of representative in vitro and in vivo models. Recently, it has been found feasible to use both biopsy and autopsy tumors to generate cultures and xenograft models. Methods To further model development, we evaluated the collective international experience from 8 collaborating centers to develop DIPG pre-clinical models from patient-derived autopsies and biopsies. Univariate and multivariate analysis was performed to determine key factors associated with the success of in vitro and in vivo PDX development. Results In vitro cultures were successfully established from 57% of samples (84.2% of biopsies and 38.2% of autopsies). Samples transferred in DMEM media were more likely to establish successful culture than those transported in Hibernate A. In vitro cultures were more successful from biopsies (84.2%) compared with autopsies (38.2%) and as monolayer on laminin-coated plates than as neurospheres. Primary cultures successfully established from autopsy samples were more likely to engraft in animal models than cultures established from biopsies (86.7% vs. 47.4%). Collectively, tumor engraftment was more successful when DIPG samples were directly implanted in mice (68%), rather than after culturing (40.7%). Conclusion This multi-center study provides valuable information on the success rate of establishing patient-derived preclinical models of DIPG. The results can lead to further optimization of DIPG model development and ultimately assist in the investigation of new therapies for this aggressive pediatric brain tumor.Maria Tsoli, Han Shen, Chelsea Mayoh, Laura Franshaw, Anahid Ehteda, Danielle Upton, Diana Carvalho, Maria Vinci, Michael H. Meel, Dannis van Vuurden, Alexander Plessier, David Castel, Rachid Drissi, Michael Farrell, Jane Cryan, Darach Crimmins, John Caird, Jane Pears, Stephanie Francis, Louise E. A. Ludlow, Andrea Carai, Angela Mastronuzzi, Bing Liu, Jordan Hansford, Nick Gottardo, Tim Hassall, Maria Kirby, Maryam Fouladi, Cynthia Hawkins, Michelle Monje, Jacques Grill, Chris Jones, Esther Hulleman, David S. Ziegle
Reversal of glucocorticoid resistance in paediatric acute lymphoblastic leukaemia is dependent on restoring BIM expression
No full text- …
