319 results on '"Sean M Grimmond"'
Search Results
2. Recommendations for Accurate Resolution of Gene and Isoform Allele-Specific Expression in RNA-Seq Data.
- Author
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David L A Wood, Katia Nones, Anita Steptoe, Angelika Christ, Ivon Harliwong, Felicity Newell, Timothy J C Bruxner, David Miller, Nicole Cloonan, and Sean M Grimmond
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Medicine ,Science - Abstract
Genetic variation modulates gene expression transcriptionally or post-transcriptionally, and can profoundly alter an individual's phenotype. Measuring allelic differential expression at heterozygous loci within an individual, a phenomenon called allele-specific expression (ASE), can assist in identifying such factors. Massively parallel DNA and RNA sequencing and advances in bioinformatic methodologies provide an outstanding opportunity to measure ASE genome-wide. In this study, matched DNA and RNA sequencing, genotyping arrays and computationally phased haplotypes were integrated to comprehensively and conservatively quantify ASE in a single human brain and liver tissue sample. We describe a methodological evaluation and assessment of common bioinformatic steps for ASE quantification, and recommend a robust approach to accurately measure SNP, gene and isoform ASE through the use of personalized haplotype genome alignment, strict alignment quality control and intragenic SNP aggregation. Our results indicate that accurate ASE quantification requires careful bioinformatic analyses and is adversely affected by sample specific alignment confounders and random sampling even at moderate sequence depths. We identified multiple known and several novel ASE genes in liver, including WDR72, DSP and UBD, as well as genes that contained ASE SNPs with imbalance direction discordant with haplotype phase, explainable by annotated transcript structure, suggesting isoform derived ASE. The methods evaluated in this study will be of use to researchers performing highly conservative quantification of ASE, and the genes and isoforms identified as ASE of interest to researchers studying those loci.
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- 2015
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3. Towards the systematic mapping and engineering of the protein prenylation machinery in Saccharomyces cerevisiae.
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Viktor Stein, Marta H Kubala, Jason Steen, Sean M Grimmond, and Kirill Alexandrov
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Medicine ,Science - Abstract
Protein prenylation is a widespread and highly conserved eukaryotic post-translational modification that endows proteins with the ability to reversibly attach to intracellular membranes. The dynamic interaction of prenylated proteins with intracellular membranes is essential for their signalling functions and is frequently deregulated in disease processes such as cancer. As a result, protein prenylation has been pharmacologically targeted by numerous drug discovery programs, albeit with limited success. To a large extent, this can be attributed to an insufficient understanding of the interplay of different protein prenyltransferases and the combinatorial diversity of the prenylatable sequence space. Here, we report a high-throughput, growth-based genetic selection assay in Saccharomyces cerevisiae based on the Ras Recruitment System which, for the first time, has allowed us to create a comprehensive map of prenylatable protein sequences in S. cerevisiae. We demonstrate that potential prenylatable space is sparsely (6.2%) occupied leaving room for creation of synthetic orthogonal prenylatable sequences. To experimentally demonstrate that, we used the developed platform to engineer mutant farnesyltransferases that efficiently prenylate substrate motives that are not recognised by endogenous protein prenyltransferases. These uncoupled mutants can now be used as starting points for the systematic engineering of the eukaryotic protein prenylation machinery.
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- 2015
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4. Using the MCF10A/MCF10CA1a Breast Cancer Progression Cell Line Model to Investigate the Effect of Active, Mutant Forms of EGFR in Breast Cancer Development and Treatment Using Gefitinib.
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Darrell C Bessette, Erik Tilch, Tatjana Seidens, Michael C J Quinn, Adrian P Wiegmans, Wei Shi, Sibylle Cocciardi, Amy McCart-Reed, Jodi M Saunus, Peter T Simpson, Sean M Grimmond, Sunil R Lakhani, Kum Kum Khanna, Nic Waddell, Fares Al-Ejeh, and Georgia Chenevix-Trench
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Medicine ,Science - Abstract
Basal-like and triple negative breast cancer (TNBC) share common molecular features, poor prognosis and a propensity for metastasis to the brain. Amplification of epidermal growth factor receptor (EGFR) occurs in ~50% of basal-like breast cancer, and mutations in the epidermal growth factor receptor (EGFR) have been reported in up to ~ 10% of Asian TNBC patients. In non-small cell lung cancer several different mutations in the EGFR tyrosine kinase domain confer sensitivity to receptor tyrosine kinase inhibitors, but the tumourigenic potential of EGFR mutations in breast cells and their potential for targeted therapy is unknown.Constructs containing wild type, G719S or E746-A750 deletion mutant forms of EGFR were transfected into the MCF10A breast cells and their tumorigenic derivative, MCF10CA1a. The effects of EGFR over-expression and mutation on proliferation, migration, invasion, response to gefitinib, and tumour formation in vivo was investigated. Copy number analysis and whole exome sequencing of the MCF10A and MCF10CA1a cell lines were also performed.Mutant EGFR increased MCF10A and MCF10CA1a proliferation and MCF10A gefitinib sensitivity. The EGFR-E746-A750 deletion increased MCF10CA1a cell migration and invasion, and greatly increased MCF10CA1a xenograft tumour formation and growth. Compared to MCF10A cells, MCF10CA1a cells exhibited large regions of gain on chromosomes 3 and 9, deletion on chromosome 7, and mutations in many genes implicated in cancer.Mutant EGFR enhances the oncogenic properties of MCF10A cell line, and increases sensitivity to gefitinib. Although the addition of EGFR E746-A750 renders the MCF10CA1a cells more tumourigenic in vivo it is not accompanied by increased gefitinib sensitivity, perhaps due to additional mutations, including the PIK3CA H1047R mutation, that the MCF10CA1a cell line has acquired. Screening TNBC/basal-like breast cancer for EGFR mutations may prove useful for directing therapy but, as in non-small cell lung cancer, accompanying mutations in PIK3CA may confer gefitinib resistance.
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- 2015
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5. SnapShot-Seq: a method for extracting genome-wide, in vivo mRNA dynamics from a single total RNA sample.
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Jesse M Gray, David A Harmin, Sarah A Boswell, Nicole Cloonan, Thomas E Mullen, Joseph J Ling, Nimrod Miller, Scott Kuersten, Yong-Chao Ma, Steven A McCarroll, Sean M Grimmond, and Michael Springer
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Medicine ,Science - Abstract
mRNA synthesis, processing, and destruction involve a complex series of molecular steps that are incompletely understood. Because the RNA intermediates in each of these steps have finite lifetimes, extensive mechanistic and dynamical information is encoded in total cellular RNA. Here we report the development of SnapShot-Seq, a set of computational methods that allow the determination of in vivo rates of pre-mRNA synthesis, splicing, intron degradation, and mRNA decay from a single RNA-Seq snapshot of total cellular RNA. SnapShot-Seq can detect in vivo changes in the rates of specific steps of splicing, and it provides genome-wide estimates of pre-mRNA synthesis rates comparable to those obtained via labeling of newly synthesized RNA. We used SnapShot-Seq to investigate the origins of the intrinsic bimodality of metazoan gene expression levels, and our results suggest that this bimodality is partly due to spillover of transcriptional activation from highly expressed genes to their poorly expressed neighbors. SnapShot-Seq dramatically expands the information obtainable from a standard RNA-Seq experiment.
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- 2014
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6. Rapid identification of a novel complex I MT-ND3 m.10134C>A mutation in a Leigh syndrome patient.
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David K Miller, Minal J Menezes, Cas Simons, Lisa G Riley, Sandra T Cooper, Sean M Grimmond, David R Thorburn, John Christodoulou, and Ryan J Taft
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Medicine ,Science - Abstract
Leigh syndrome (LS) is a rare progressive multi-system neurodegenerative disorder, the genetics of which is frequently difficult to resolve. Rapid determination of the genetic etiology of LS in a 5-year-old girl facilitated inclusion in Edison Pharmaceutical's phase 2B clinical trial of EPI-743. SNP-arrays and high-coverage whole exome sequencing were performed on the proband, both parents and three unaffected siblings. Subsequent multi-tissue targeted high-depth mitochondrial sequencing was performed using custom long-range PCR amplicons. Tissue-specific mutant load was also assessed by qPCR. Complex I was interrogated by spectrophotometric enzyme assays and Western Blot. No putatively causal mutations were identified in nuclear-encoded genes. Analysis of low-coverage off-target mitochondrial reads revealed a previously unreported mitochondrial mutation in the proband in MT-ND3 (m.10134C>A, p.Q26K), a Complex I mitochondrial gene previously associated with LS. Targeted investigations demonstrated that this mutation was 1% heteroplasmic in the mother's blood and homoplasmic in the proband's blood, fibroblasts, liver and muscle. Enzyme assays revealed decreased Complex I activity. The identification of this novel LS MT-ND3 variant, the genomics of which was accomplished in less than 3.5 weeks, indicates that rapid genomic approaches may prove useful in time-sensitive cases with an unresolved genetic diagnosis.
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- 2014
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7. Whole genome sequence analysis of the first Australian OXA-48-producing outbreak-associated Klebsiella pneumoniae isolates: the resistome and in vivo evolution.
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Björn A Espedido, Jason A Steen, Helen Ziochos, Sean M Grimmond, Matthew A Cooper, Iain B Gosbell, Sebastiaan J van Hal, and Slade O Jensen
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Medicine ,Science - Abstract
Whole genome sequencing was used to characterize the resistome of intensive care unit (ICU) outbreak-associated carbapenem-resistant K. pneumoniae isolates. Importantly, and of particular concern, the carbapenem-hydrolyzing β-lactamase gene bla(OXA-48) and the extended-spectrum β-lactamase gene bla(CTX-M-14), were identified on a single broad host-range conjugative plasmid. This represents the first report of bla(OXA-48) in Australia and highlights the importance of resistance gene surveillance, as such plasmids can silently spread amongst enterobacterial populations and have the potential to drastically limit treatment options. Furthermore, the in vivo evolution of these isolates was also examined after 18 months of intra-abdominal carriage in a patient that transited through the ICU during the outbreak period. Reflecting the clonality of K. pneumoniae, only 11 single nucleotide polymorphisms (SNPs) were accumulated during this time-period and many of these were associated with genes involved in tolerance/resistance to antibiotics, metals or organic solvents, and transcriptional regulation. Collectively, these SNPs are likely to be associated with changes in virulence (at least to some extent) that have refined the in vivo colonization capacity of the original outbreak isolate.
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- 2013
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8. Somatic point mutation calling in low cellularity tumors.
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Karin S Kassahn, Oliver Holmes, Katia Nones, Ann-Marie Patch, David K Miller, Angelika N Christ, Ivon Harliwong, Timothy J Bruxner, Qinying Xu, Matthew Anderson, Scott Wood, Conrad Leonard, Darrin Taylor, Felicity Newell, Sarah Song, Senel Idrisoglu, Craig Nourse, Ehsan Nourbakhsh, Suzanne Manning, Shivangi Wani, Anita Steptoe, Marina Pajic, Mark J Cowley, Mark Pinese, David K Chang, Anthony J Gill, Amber L Johns, Jianmin Wu, Peter J Wilson, Lynn Fink, Andrew V Biankin, Nicola Waddell, Sean M Grimmond, and John V Pearson
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Medicine ,Science - Abstract
Somatic mutation calling from next-generation sequencing data remains a challenge due to the difficulties of distinguishing true somatic events from artifacts arising from PCR, sequencing errors or mis-mapping. Tumor cellularity or purity, sub-clonality and copy number changes also confound the identification of true somatic events against a background of germline variants. We have developed a heuristic strategy and software (http://www.qcmg.org/bioinformatics/qsnp/) for somatic mutation calling in samples with low tumor content and we show the superior sensitivity and precision of our approach using a previously sequenced cell line, a series of tumor/normal admixtures, and 3,253 putative somatic SNVs verified on an orthogonal platform.
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- 2013
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9. qpure: A tool to estimate tumor cellularity from genome-wide single-nucleotide polymorphism profiles.
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Sarah Song, Katia Nones, David Miller, Ivon Harliwong, Karin S Kassahn, Mark Pinese, Marina Pajic, Anthony J Gill, Amber L Johns, Matthew Anderson, Oliver Holmes, Conrad Leonard, Darrin Taylor, Scott Wood, Qinying Xu, Felicity Newell, Mark J Cowley, Jianmin Wu, Peter Wilson, Lynn Fink, Andrew V Biankin, Nic Waddell, Sean M Grimmond, and John V Pearson
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Medicine ,Science - Abstract
Tumour cellularity, the relative proportion of tumour and normal cells in a sample, affects the sensitivity of mutation detection, copy number analysis, cancer gene expression and methylation profiling. Tumour cellularity is traditionally estimated by pathological review of sectioned specimens; however this method is both subjective and prone to error due to heterogeneity within lesions and cellularity differences between the sample viewed during pathological review and tissue used for research purposes. In this paper we describe a statistical model to estimate tumour cellularity from SNP array profiles of paired tumour and normal samples using shifts in SNP allele frequency at regions of loss of heterozygosity (LOH) in the tumour. We also provide qpure, a software implementation of the method. Our experiments showed that there is a medium correlation 0.42 ([Formula: see text]-value=0.0001) between tumor cellularity estimated by qpure and pathology review. Interestingly there is a high correlation 0.87 ([Formula: see text]-value [Formula: see text] 2.2e-16) between cellularity estimates by qpure and deep Ion Torrent sequencing of known somatic KRAS mutations; and a weaker correlation 0.32 ([Formula: see text]-value=0.004) between IonTorrent sequencing and pathology review. This suggests that qpure may be a more accurate predictor of tumour cellularity than pathology review. qpure can be downloaded from https://sourceforge.net/projects/qpure/.
- Published
- 2012
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10. Identification of novel markers of mouse fetal ovary development.
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Huijun Chen, James S Palmer, Rathi D Thiagarajan, Marcel E Dinger, Emmanuelle Lesieur, Hansheng Chiu, Alexandra Schulz, Cassy Spiller, Sean M Grimmond, Melissa H Little, Peter Koopman, and Dagmar Wilhelm
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Medicine ,Science - Abstract
In contrast to the developing testis, molecular pathways driving fetal ovarian development have been difficult to characterise. To date no single master regulator of ovarian development has been identified that would be considered the female equivalent of Sry. Using a genomic approach we identified a number of novel protein-coding as well as non-coding genes that were detectable at higher levels in the ovary compared to testis during early mouse gonad development. We were able to cluster these ovarian genes into different temporal expression categories. Of note, Lrrc34 and AK015184 were detected in XX but not XY germ cells before the onset of sex-specific germ cell differentiation marked by entry into meiosis in an ovary and mitotic arrest in a testis. We also defined distinct spatial expression domains of somatic cell genes in the developing ovary. Our data expands the set of markers of early mouse ovary differentiation and identifies a classification of early ovarian genes, thus providing additional avenues with which to dissect this process.
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- 2012
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11. Identification of anchor genes during kidney development defines ontological relationships, molecular subcompartments and regulatory pathways.
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Rathi D Thiagarajan, Kylie M Georgas, Bree A Rumballe, Emmanuelle Lesieur, Han Sheng Chiu, Darrin Taylor, Dave T P Tang, Sean M Grimmond, and Melissa H Little
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Medicine ,Science - Abstract
The development of the mammalian kidney is well conserved from mouse to man. Despite considerable temporal and spatial data on gene expression in mammalian kidney development, primarily in rodent species, there is a paucity of genes whose expression is absolutely specific to a given anatomical compartment and/or developmental stage, defined here as 'anchor' genes. We previously generated an atlas of gene expression in the developing mouse kidney using microarray analysis of anatomical compartments collected via laser capture microdissection. Here, this data is further analysed to identify anchor genes via stringent bioinformatic filtering followed by high resolution section in situ hybridisation performed on 200 transcripts selected as specific to one of 11 anatomical compartments within the midgestation mouse kidney. A total of 37 anchor genes were identified across 6 compartments with the early proximal tubule being the compartment richest in anchor genes. Analysis of minimal and evolutionarily conserved promoter regions of this set of 25 anchor genes identified enrichment of transcription factor binding sites for Hnf4a and Hnf1b, RbpJ (Notch signalling), PPARγ:RxRA and COUP-TF family transcription factors. This was reinforced by GO analyses which also identified these anchor genes as targets in processes including epithelial proliferation and proximal tubular function. As well as defining anchor genes, this large scale validation of gene expression identified a further 92 compartment-enriched genes able to subcompartmentalise key processes during murine renal organogenesis spatially or ontologically. This included a cohort of 13 ureteric epithelial genes revealing previously unappreciated compartmentalisation of the collecting duct system and a series of early tubule genes suggesting that segmentation into proximal tubule, loop of Henle and distal tubule does not occur until the onset of glomerular vascularisation. Overall, this study serves to illuminate previously ill-defined stages of patterning and will enable further refinement of the lineage relationships within mammalian kidney development.
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- 2011
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12. Phasevarion mediated epigenetic gene regulation in Helicobacter pylori.
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Yogitha N Srikhanta, Rebecca J Gorrell, Jason A Steen, Jayde A Gawthorne, Terry Kwok, Sean M Grimmond, Roy M Robins-Browne, and Michael P Jennings
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Medicine ,Science - Abstract
Many host-adapted bacterial pathogens contain DNA methyltransferases (mod genes) that are subject to phase-variable expression (high-frequency reversible ON/OFF switching of gene expression). In Haemophilus influenzae and pathogenic Neisseria, the random switching of the modA gene, associated with a phase-variable type III restriction modification (R-M) system, controls expression of a phase-variable regulon of genes (a "phasevarion"), via differential methylation of the genome in the modA ON and OFF states. Phase-variable type III R-M systems are also found in Helicobacter pylori, suggesting that phasevarions may also exist in this key human pathogen. Phylogenetic studies on the phase-variable type III modH gene revealed that there are 17 distinct alleles in H. pylori, which differ only in their DNA recognition domain. One of the most commonly found alleles was modH5 (16% of isolates). Microarray analysis comparing the wild-type P12modH5 ON strain to a P12ΔmodH5 mutant revealed that six genes were either up- or down-regulated, and some were virulence-associated. These included flaA, which encodes a flagella protein important in motility and hopG, an outer membrane protein essential for colonization and associated with gastric cancer. This study provides the first evidence of this epigenetic mechanism of gene expression in H. pylori. Characterisation of H. pylori modH phasevarions to define stable immunological targets will be essential for vaccine development and may also contribute to understanding H. pylori pathogenesis.
- Published
- 2011
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13. A high-throughput platform for lentiviral overexpression screening of the human ORFeome.
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Dubravka Škalamera, Max V Ranall, Benjamin M Wilson, Paul Leo, Amy S Purdon, Carolyn Hyde, Ehsan Nourbakhsh, Sean M Grimmond, Simon C Barry, Brian Gabrielli, and Thomas J Gonda
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Medicine ,Science - Abstract
In response to the growing need for functional analysis of the human genome, we have developed a platform for high-throughput functional screening of genes overexpressed from lentiviral vectors. Protein-coding human open reading frames (ORFs) from the Mammalian Gene Collection were transferred into lentiviral expression vector using the highly efficient Gateway recombination cloning. Target ORFs were inserted into the vector downstream of a constitutive promoter and upstream of an IRES controlled GFP reporter, so that their transfection, transduction and expression could be monitored by fluorescence. The expression plasmids and viral packaging plasmids were combined and transfected into 293T cells to produce virus, which was then used to transduce the screening cell line. We have optimised the transfection and transduction procedures so that they can be performed using robotic liquid handling systems in arrayed 96-well microplate, one-gene-per-well format, without the need to concentrate the viral supernatant. Since lentiviruses can infect both dividing and non-dividing cells, this system can be used to overexpress human ORFs in a broad spectrum of experimental contexts. We tested the platform in a 1990 gene pilot screen for genes that can increase proliferation of the non-tumorigenic mammary epithelial cell line MCF-10A after removal of growth factors. Transduced cells were labelled with the nucleoside analogue 5-ethynyl-2'-deoxyuridine (EdU) to detect cells progressing through S phase. Hits were identified using high-content imaging and statistical analysis and confirmed with vectors using two different promoters (CMV and EF1α). The screen demonstrates the reliability, versatility and utility of our screening platform, and identifies novel cell cycle/proliferative activities for a number of genes.
- Published
- 2011
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14. Use of DNA-damaging agents and RNA pooling to assess expression profiles associated with BRCA1 and BRCA2 mutation status in familial breast cancer patients.
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Logan C Walker, Bryony A Thompson, Nic Waddell, kConFab Investigators, Sean M Grimmond, and Amanda B Spurdle
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Genetics ,QH426-470 - Abstract
A large number of rare sequence variants of unknown clinical significance have been identified in the breast cancer susceptibility genes, BRCA1 and BRCA2. Laboratory-based methods that can distinguish between carriers of pathogenic mutations and non-carriers are likely to have utility for the classification of these sequence variants. To identify predictors of pathogenic mutation status in familial breast cancer patients, we explored the use of gene expression arrays to assess the effect of two DNA-damaging agents (irradiation and mitomycin C) on cellular response in relation to BRCA1 and BRCA2 mutation status. A range of regimes was used to treat 27 lymphoblastoid cell-lines (LCLs) derived from affected women in high-risk breast cancer families (nine BRCA1, nine BRCA2, and nine non-BRCA1/2 or BRCAX individuals) and nine LCLs from healthy individuals. Using an RNA-pooling strategy, we found that treating LCLs with 1.2 microM mitomycin C and measuring the gene expression profiles 1 hour post-treatment had the greatest potential to discriminate BRCA1, BRCA2, and BRCAX mutation status. A classifier was built using the expression profile of nine QRT-PCR validated genes that were associated with BRCA1, BRCA2, and BRCAX status in RNA pools. These nine genes could distinguish BRCA1 from BRCA2 carriers with 83% accuracy in individual samples, but three-way analysis for BRCA1, BRCA2, and BRCAX had a maximum of 59% prediction accuracy. Our results suggest that, compared to BRCA1 and BRCA2 mutation carriers, non-BRCA1/2 (BRCAX) individuals are genetically heterogeneous. This study also demonstrates the effectiveness of RNA pools to compare the expression profiles of cell-lines from BRCA1, BRCA2, and BRCAX cases after treatment with irradiation and mitomycin C as a method to prioritize treatment regimes for detailed downstream expression analysis.
- Published
- 2010
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15. Transcriptome-wide prediction of miRNA targets in human and mouse using FASTH.
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Chikako Ragan, Nicole Cloonan, Sean M Grimmond, Michael Zuker, and Mark A Ragan
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Medicine ,Science - Abstract
Transcriptional regulation by microRNAs (miRNAs) involves complementary base-pairing at target sites on mRNAs, yielding complex secondary structures. Here we introduce an efficient computational approach and software (FASTH) for genome-scale prediction of miRNA target sites based on minimizing the free energy of duplex structure. We apply our approach to identify miRNA target sites in the human and mouse transcriptomes. Our results show that short sequence motifs in the 5' end of miRNAs frequently match mRNAs perfectly, not only at validated target sites but additionally at many other, energetically favourable sites. High-quality matching regions are abundant and occur at similar frequencies in all mRNA regions, not only the 3'UTR. About one-third of potential miRNA target sites are reassigned to different mRNA regions, or gained or lost altogether, among different transcript isoforms from the same gene. Many potential miRNA target sites predicted in human are not found in mouse, and vice-versa, but among those that do occur in orthologous human and mouse mRNAs most are situated in corresponding mRNA regions, i.e. these sites are themselves orthologous. Using a luciferase assay in HEK293 cells, we validate four of six predicted miRNA-mRNA interactions, with the mRNA level reduced by an average of 73%. We demonstrate that a thermodynamically based computational approach to prediction of miRNA binding sites on mRNAs can be scaled to analyse complete mammalian transcriptome datasets. These results confirm and extend the scope of miRNA-mediated species- and transcript-specific regulation in different cell types, tissues and developmental conditions.
- Published
- 2009
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16. Phasevarions mediate random switching of gene expression in pathogenic Neisseria.
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Yogitha N Srikhanta, Stefanie J Dowideit, Jennifer L Edwards, Megan L Falsetta, Hsing-Ju Wu, Odile B Harrison, Kate L Fox, Kate L Seib, Tina L Maguire, Andrew H-J Wang, Martin C Maiden, Sean M Grimmond, Michael A Apicella, and Michael P Jennings
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Immunologic diseases. Allergy ,RC581-607 ,Biology (General) ,QH301-705.5 - Abstract
Many host-adapted bacterial pathogens contain DNA methyltransferases (mod genes) that are subject to phase-variable expression (high-frequency reversible ON/OFF switching of gene expression). In Haemophilus influenzae, the random switching of the modA gene controls expression of a phase-variable regulon of genes (a "phasevarion"), via differential methylation of the genome in the modA ON and OFF states. Phase-variable mod genes are also present in Neisseria meningitidis and Neisseria gonorrhoeae, suggesting that phasevarions may occur in these important human pathogens. Phylogenetic studies on phase-variable mod genes associated with type III restriction modification (R-M) systems revealed that these organisms have two distinct mod genes--modA and modB. There are also distinct alleles of modA (abundant: modA11, 12, 13; minor: modA4, 15, 18) and modB (modB1, 2). These alleles differ only in their DNA recognition domain. ModA11 was only found in N. meningitidis and modA13 only in N. gonorrhoeae. The recognition site for the modA13 methyltransferase in N. gonorrhoeae strain FA1090 was identified as 5'-AGAAA-3'. Mutant strains lacking the modA11, 12 or 13 genes were made in N. meningitidis and N. gonorrhoeae and their phenotype analyzed in comparison to a corresponding mod ON wild-type strain. Microarray analysis revealed that in all three modA alleles multiple genes were either upregulated or downregulated, some of which were virulence-associated. For example, in N. meningitidis MC58 (modA11), differentially expressed genes included those encoding the candidate vaccine antigens lactoferrin binding proteins A and B. Functional studies using N. gonorrhoeae FA1090 and the clinical isolate O1G1370 confirmed that modA13 ON and OFF strains have distinct phenotypes in antimicrobial resistance, in a primary human cervical epithelial cell model of infection, and in biofilm formation. This study, in conjunction with our previous work in H. influenzae, indicates that phasevarions may be a common strategy used by host-adapted bacterial pathogens to randomly switch between "differentiated" cell types.
- Published
- 2009
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17. The abundance of short proteins in the mammalian proteome.
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Martin C Frith, Alistair R Forrest, Ehsan Nourbakhsh, Ken C Pang, Chikatoshi Kai, Jun Kawai, Piero Carninci, Yoshihide Hayashizaki, Timothy L Bailey, and Sean M Grimmond
- Subjects
Genetics ,QH426-470 - Abstract
Short proteins play key roles in cell signalling and other processes, but their abundance in the mammalian proteome is unknown. Current catalogues of mammalian proteins exhibit an artefactual discontinuity at a length of 100 aa, so that protein abundance peaks just above this length and falls off sharply below it. To clarify the abundance of short proteins, we identify proteins in the FANTOM collection of mouse cDNAs by analysing synonymous and non-synonymous substitutions with the computer program CRITICA. This analysis confirms that there is no real discontinuity at length 100. Roughly 10% of mouse proteins are shorter than 100 aa, although the majority of these are variants of proteins longer than 100 aa. We identify many novel short proteins, including a "dark matter" subset containing ones that lack detectable homology to other known proteins. Translation assays confirm that some of these novel proteins can be translated and localised to the secretory pathway.
- Published
- 2006
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18. A rare diagnostically challenging case of CIC-DUX4 sarcoma arising in the neck
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Stefanie Aranza, Christina Roydhouse, Catherine Mitchell, Joseph H.A. Vissers, Wing-Yee Lo, Sean M. Grimmond, and Jane Kho
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Pathology and Forensic Medicine - Published
- 2023
19. Australian Genomics: Outcomes of a 5-year national program to accelerate the integration of genomics in healthcare
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Zornitza Stark, Tiffany Boughtwood, Matilda Haas, Jeffrey Braithwaite, Clara L. Gaff, Ilias Goranitis, Amanda B. Spurdle, David P. Hansen, Oliver Hofmann, Nigel Laing, Sylvia Metcalfe, Ainsley J. Newson, Hamish S. Scott, Natalie Thorne, Robyn L. Ward, Marcel E. Dinger, Stephanie Best, Janet C. Long, Sean M. Grimmond, John Pearson, Nicola Waddell, Christopher P. Barnett, Matthew Cook, Michael Field, David Fielding, Stephen B. Fox, Jozef Gecz, Adam Jaffe, Richard J. Leventer, Paul J. Lockhart, Sebastian Lunke, Andrew J. Mallett, Julie McGaughran, Linda Mileshkin, Katia Nones, Tony Roscioli, Ingrid E. Scheffer, Christopher Semsarian, Cas Simons, David M. Thomas, David R. Thorburn, Richard Tothill, Deborah White, Sally Dunwoodie, Peter T. Simpson, Peta Phillips, Marie-Jo Brion, Keri Finlay, Michael CJ. Quinn, Tessa Mattiske, Emma Tudini, Kirsten Boggs, Sean Murray, Kathy Wells, John Cannings, Andrew H. Sinclair, John Christodoulou, Kathryn N. North, Stark, Zornitza, Boughtwood, Tiffany, Haas, Matilda, Braithwaite, Jeffrey, Scott, Hamish S, and North, Kathryn N
- Subjects
Genetics ,Genetics (clinical) - Abstract
Refereed/Peer-reviewed Australian Genomics is a national collaborative partnership of more than 100 organizations piloting a whole-of-system approach to integrating genomics into healthcare, based on federation principles. In the first five years of operation, Australian Genomics has evaluated the outcomes of genomic testing in more than 5,200 individuals across 19 rare disease and cancer flagship studies. Comprehensive analyses of the health economic, policy, ethical, legal, implementation and workforce implications of incorporating genomics in the Australian context have informed evidence-based change in policy and practice, resulting in national government funding and equity of access for a range of genomic tests. Simultaneously, Australian Genomics has built national skills, infrastructure, policy, and data resources to enable effective data sharing to drive discovery research and support improvements in clinical genomic delivery.
- Published
- 2023
20. S100 family proteins are linked to organoid morphology and EMT in pancreatic cancer
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Ronnie Ren Jie Low, Ka Yee Fung, Hugh Gao, Adele Preaudet, Laura F. Dagley, Jumana Yousef, Belinda Lee, Samantha J. Emery-Corbin, Paul M. Nguyen, Rune H. Larsen, Nadia J. Kershaw, Antony W. Burgess, Peter Gibbs, Frédéric Hollande, Michael D. W. Griffin, Sean M. Grimmond, and Tracy L. Putoczki
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Cell Biology ,Molecular Biology - Abstract
Epithelial-mesenchymal transition (EMT) is a continuum that includes epithelial, partial EMT, and mesenchymal states, each of which is associated with cancer progression, invasive capabilities, and ultimately, metastasis. We used a lineage-traced sporadic model of pancreatic cancer to generate a murine organoid biobank from primary and secondary tumors, including sublines that underwent partial EMT and complete EMT. Using an unbiased proteomics approach, we found that organoid morphology predicts the EMT state, and the solid organoids are associated with a partial EMT signature. We also observed that exogenous TGFβ1 induces solid organoid morphology that is associated with changes in the S100 family, complete EMT, and the formation of high-grade tumors. S100A4 may be a useful biomarker for predicting EMT state, disease progression, and outcome in patients with pancreatic cancer.
- Published
- 2023
21. Description of a novel subtype of acute myeloid leukemia defined by recurrent CBFB insertions
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Georgina L. Ryland, Masayuki Umeda, Linda Holmfeldt, Sören Lehmann, Morten Krogh Herlin, Jing Ma, Mahsa Khanlari, Jeffrey E. Rubnitz, Rhonda E. Ries, Hansen J. Kosasih, Paul G. Ekert, Hwee Ngee Goh, Ing S. Tiong, Sean M. Grimmond, Claudia Haferlach, Ryan B. Day, Timothy J. Ley, Soheil Meshinchi, Xiaotu Ma, Piers Blombery, and Jeffery M. Klco
- Subjects
Oncogene Proteins, Fusion ,Immunology ,Humans ,Leukemia, Myeloid, Acute/genetics ,Cell Biology ,Hematology ,Biochemistry ,Core Binding Factor beta Subunit - Published
- 2023
22. Enhancer retargeting of CDX2 and UBTF::ATXN7L3 define a subtype of high-risk B-progenitor acute lymphoblastic leukemia
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Shunsuke Kimura, Lindsey Montefiori, Ilaria Iacobucci, Yaqi Zhao, Qingsong Gao, Elisabeth M. Paietta, Claudia Haferlach, A. Douglas Laird, Paul E. Mead, Zhaohui Gu, Wendy Stock, Mark Litzow, Jacob M. Rowe, Selina M. Luger, Stephen P. Hunger, Georgina L. Ryland, Breon Schmidt, Paul G. Ekert, Alicia Oshlack, Sean M. Grimmond, Jacqueline Rehn, James Breen, David Yeung, Deborah L. White, Ibrahim Aldoss, Elias J. Jabbour, Ching-Hon Pui, Manja Meggendorfer, Wencke Walter, Wolfgang Kern, Torsten Haferlach, Samuel Brady, Jinghui Zhang, Kathryn G. Roberts, Piers Blombery, and Charles G. Mullighan
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Adult ,Male ,Lymphoid Neoplasia ,Adolescent ,Immunology ,Genomics ,Cell Biology ,Hematology ,Middle Aged ,Precursor Cell Lymphoblastic Leukemia-Lymphoma ,Prognosis ,Biochemistry ,Chromatin ,Young Adult ,Precursor B-Cell Lymphoblastic Leukemia-Lymphoma ,Humans ,CDX2 Transcription Factor ,Female ,Child ,Transcriptome ,Pol1 Transcription Initiation Complex Proteins ,Aged ,Transcription Factors - Abstract
Transcriptome sequencing has identified multiple subtypes of B-progenitor acute lymphoblastic leukemia (B-ALL) of prognostic significance, but a minority of cases lack a known genetic driver. Here, we used integrated whole-genome (WGS) and -transcriptome sequencing (RNA-seq), enhancer mapping, and chromatin topology analysis to identify previously unrecognized genomic drivers in B-ALL. Newly diagnosed (n = 3221) and relapsed (n = 177) B-ALL cases with tumor RNA-seq were studied. WGS was performed to detect mutations, structural variants, and copy number alterations. Integrated analysis of histone 3 lysine 27 acetylation and chromatin looping was performed using HiChIP. We identified a subset of 17 newly diagnosed and 5 relapsed B-ALL cases with a distinct gene expression profile and 2 universal and unique genomic alterations resulting from aberrant recombination-activating gene activation: a focal deletion downstream of PAN3 at 13q12.2 resulting in CDX2 deregulation by the PAN3 enhancer and a focal deletion of exons 18-21 of UBTF at 17q21.31 resulting in a chimeric fusion, UBTF::ATXN7L3. A subset of cases also had rearrangement and increased expression of the PAX5 gene, which is otherwise uncommon in B-ALL. Patients were more commonly female and young adult with median age 35 (range,12-70 years). The immunophenotype was characterized by CD10 negativity and immunoglobulin M positivity. Among 16 patients with known clinical response, 9 (56.3%) had high-risk features including relapse (n = 4) or minimal residual disease >1% at the end of remission induction (n = 5). CDX2-deregulated, UBTF::ATXN7L3 rearranged (CDX2/UBTF) B-ALL is a high-risk subtype of leukemia in young adults for which novel therapeutic approaches are required.
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- 2022
23. Table S5 from Homologous Recombination DNA Repair Pathway Disruption and Retinoblastoma Protein Loss Are Associated with Exceptional Survival in High-Grade Serous Ovarian Cancer
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Anna deFazio, David D. L. Bowtell, Brad H. Nelson, Paul Harnett, Martin Köbel, Alexander Dobrovic, Alison Brand, Michael Friedlander, Penny Blomfield, Philip Beale, Robert M. Rome, Yee C. Leung, Paul A. Cohen, Marisa Grossi, Sumitra Ananda, Anne Hamilton, Linda Mileshkin, Orla McNally, Peter F. Rambau, Prue E. Allan, Raghwa Sharma, Colin J.R. Stewart, Jillian Hung, Nicola Waddell, John V. Pearson, Sean M. Grimmond, Kaushalya Amarasinghe, Giada V. Zapparoli, Thomas Mikeska, Joy Hendley, Yoke-Eng Chiew, Sreeja R. Gadipally, Timothy Semple, Gisela Mir Arnau, Jason Li, Ann-Marie Patch, Joshy George, Katy Milne, Maartje C.A. Wouters, Elizabeth L. Christie, Valérie Garès, Val Gebski, Bo Gao, Dariush Etemadmoghadam, Catherine J. Kennedy, Catherine Emmanuel, Sian Fereday, Kathryn Alsop, and Dale W. Garsed
- Abstract
HR pathway mutations
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- 2023
24. Data from Gemcitabine and CHK1 Inhibition Potentiate EGFR-Directed Radioimmunotherapy against Pancreatic Ductal Adenocarcinoma
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Kum Kum Khanna, Michael P. Brown, Sean M. Grimmond, Andrew V. Biankin, Angela Chou, Adnan M. Nagrial, Mariska Miranda, Murugan Kalimutho, Wei Shi, Marina Pajic, and Fares Al-Ejeh
- Abstract
Purpose: To develop effective combination therapy against pancreatic ductal adenocarcinoma (PDAC) with a combination of chemotherapy, CHK1 inhibition, and EGFR-targeted radioimmunotherapy.Experimental Design: Maximum tolerated doses were determined for the combination of gemcitabine, the CHK1 inhibitor PF-477736, and Lutetium-177 (177Lu)–labeled anti-EGFR antibody. This triple combination therapy was investigated using PDAC models from well-established cell lines, recently established patient-derived cell lines, and fresh patient-derived xenografts. Tumors were investigated for the accumulation of 177Lu-anti-EGFR antibody, survival of tumor-initiating cells, induction of DNA damage, cell death, and tumor tissue degeneration.Results: The combination of gemcitabine and CHK1 inhibitor PF-477736 with 177Lu-anti-EGFR antibody was tolerated in mice. This triplet was effective in established tumors and prevented the recurrence of PDAC in four cell line–derived and one patient-derived xenograft model. This exquisite response was associated with the loss of tumor-initiating cells as measured by flow cytometric analysis and secondary implantation of tumors from treated mice into treatment-naïve mice. Extensive DNA damage, apoptosis, and tumor degeneration were detected in the patient-derived xenograft. Mechanistically, we observed CDC25A stabilization as a result of CHK1 inhibition with consequent inhibition of gemcitabine-induced S-phase arrest as well as a decrease in canonical (ERK1/2 phosphorylation) and noncanonical EGFR signaling (RAD51 degradation) as a result of EGFR inhibition.Conclusions: Our study developed an effective combination therapy against PDAC that has potential in the treatment of PDAC. Clin Cancer Res; 20(12); 3187–97. ©2014 AACR.
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- 2023
25. Data from Homologous Recombination DNA Repair Pathway Disruption and Retinoblastoma Protein Loss Are Associated with Exceptional Survival in High-Grade Serous Ovarian Cancer
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Anna deFazio, David D. L. Bowtell, Brad H. Nelson, Paul Harnett, Martin Köbel, Alexander Dobrovic, Alison Brand, Michael Friedlander, Penny Blomfield, Philip Beale, Robert M. Rome, Yee C. Leung, Paul A. Cohen, Marisa Grossi, Sumitra Ananda, Anne Hamilton, Linda Mileshkin, Orla McNally, Peter F. Rambau, Prue E. Allan, Raghwa Sharma, Colin J.R. Stewart, Jillian Hung, Nicola Waddell, John V. Pearson, Sean M. Grimmond, Kaushalya Amarasinghe, Giada V. Zapparoli, Thomas Mikeska, Joy Hendley, Yoke-Eng Chiew, Sreeja R. Gadipally, Timothy Semple, Gisela Mir Arnau, Jason Li, Ann-Marie Patch, Joshy George, Katy Milne, Maartje C.A. Wouters, Elizabeth L. Christie, Valérie Garès, Val Gebski, Bo Gao, Dariush Etemadmoghadam, Catherine J. Kennedy, Catherine Emmanuel, Sian Fereday, Kathryn Alsop, and Dale W. Garsed
- Abstract
Purpose: Women with epithelial ovarian cancer generally have a poor prognosis; however, a subset of patients has an unexpected dramatic and durable response to treatment. We sought to identify clinical, pathological, and molecular determinants of exceptional survival in women with high-grade serous cancer (HGSC), a disease associated with the majority of ovarian cancer deaths.Experimental Design: We evaluated the histories of 2,283 ovarian cancer patients and, after applying stringent clinical and pathological selection criteria, identified 96 with HGSC that represented significant outliers in terms of treatment response and overall survival. Patient samples were characterized immunohistochemically and by genome sequencing.Results: Different patterns of clinical response were seen: long progression-free survival (Long-PFS), multiple objective responses to chemotherapy (Multiple Responder), and/or greater than 10-year overall survival (Long-Term Survivors). Pathogenic germline and somatic mutations in genes involved in homologous recombination (HR) repair were enriched in all three groups relative to a population-based series. However, 29% of 10-year survivors lacked an identifiable HR pathway alteration, and tumors from these patients had increased Ki-67 staining. CD8+ tumor-infiltrating lymphocytes were more commonly present in Long-Term Survivors. RB1 loss was associated with long progression-free and overall survival. HR deficiency and RB1 loss were correlated, and co-occurrence was significantly associated with prolonged survival.Conclusions: There was diversity in the clinical trajectory of exceptional survivors associated with multiple molecular determinants of exceptional outcome in HGSC patients. Concurrent HR deficiency and RB1 loss were associated with favorable outcomes, suggesting that co-occurrence of specific mutations might mediate durable responses in such patients. Clin Cancer Res; 24(3); 569–80. ©2017 AACR.See related commentary by Peng and Mills, p. 508
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- 2023
26. Supplementary Figure 1 from Gemcitabine and CHK1 Inhibition Potentiate EGFR-Directed Radioimmunotherapy against Pancreatic Ductal Adenocarcinoma
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Kum Kum Khanna, Michael P. Brown, Sean M. Grimmond, Andrew V. Biankin, Angela Chou, Adnan M. Nagrial, Mariska Miranda, Murugan Kalimutho, Wei Shi, Marina Pajic, and Fares Al-Ejeh
- Abstract
Supplementary Figure 1: Sensitization of PANC-1 cells to gemcitabine and EGFR-directed RIT by Chk1 inhibition. (A) Adherent cultures of the PDAC PANC-1 cell line were treated in the absence or presence of escalating doses of gemcitabine, in the absence of presence of Chk1i alone (PF-477736, 180 nM) or the combinations. Chk1i was added either 16 hours after gemcitabine or concurrently. Cells were collected 24-96 hours after treatment for standard cell cycle analysis by DNA content using FACS. (B) PANC-1 cells were treated with escalating concentrations of 177Lu-anti-EGFR mAb (RIT) to achieve the specified radiation doses (0-4 Gy) over 72 hours of incubation. RIT was performed alone or in combination with Chk1i (180 nM) and then clonogenic survival was determined by standard assays. Chk1i alone at 180 nM did not have any effect on clonogenic survival (data not shown). (C) PANC-1 cells were left untreated (vehicle control, data not shown) or incubated with anti-EGFR mAb (unlabeled, control which was not different from the vehicle control), 177Lu-DOTA, 177Lu-labelled mAb with irrelevant specificity (Sal5, raised against Salmonella antigen) or 177Lu-anti-EGFR mAb (2 Gy over 72 hours). Cells were washed 3 hours after incubation to remove unbound material and left untreated or treated with Chk1i alone (180 nM), gemcitabine alone (40 ng/mL) or the combination (Chk1i+gemcitabine) before standard clonogenic survival assays.
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- 2023
27. Supplementary Figure 3 from Gemcitabine and CHK1 Inhibition Potentiate EGFR-Directed Radioimmunotherapy against Pancreatic Ductal Adenocarcinoma
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Kum Kum Khanna, Michael P. Brown, Sean M. Grimmond, Andrew V. Biankin, Angela Chou, Adnan M. Nagrial, Mariska Miranda, Murugan Kalimutho, Wei Shi, Marina Pajic, and Fares Al-Ejeh
- Abstract
Supplementary Figure 3: EGFR expression in the PANC-1 and BxPC-3. Cell cultures were used for standard immunoblot and flow cytometry analysis for EGFR expression. The anti-EGFR mAb clone 225 was used for staining and anti-mouse IgG antibody conjugated with HRP was used for immunoblot or conjugated with Alexa488 for flow cytometry. Anti-tubulin was used to confirm equal loading for immunoblot. Both PDAC cell lines express EGFR.
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- 2023
28. Supplementary Data from Homologous Recombination DNA Repair Pathway Disruption and Retinoblastoma Protein Loss Are Associated with Exceptional Survival in High-Grade Serous Ovarian Cancer
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Anna deFazio, David D. L. Bowtell, Brad H. Nelson, Paul Harnett, Martin Köbel, Alexander Dobrovic, Alison Brand, Michael Friedlander, Penny Blomfield, Philip Beale, Robert M. Rome, Yee C. Leung, Paul A. Cohen, Marisa Grossi, Sumitra Ananda, Anne Hamilton, Linda Mileshkin, Orla McNally, Peter F. Rambau, Prue E. Allan, Raghwa Sharma, Colin J.R. Stewart, Jillian Hung, Nicola Waddell, John V. Pearson, Sean M. Grimmond, Kaushalya Amarasinghe, Giada V. Zapparoli, Thomas Mikeska, Joy Hendley, Yoke-Eng Chiew, Sreeja R. Gadipally, Timothy Semple, Gisela Mir Arnau, Jason Li, Ann-Marie Patch, Joshy George, Katy Milne, Maartje C.A. Wouters, Elizabeth L. Christie, Valérie Garès, Val Gebski, Bo Gao, Dariush Etemadmoghadam, Catherine J. Kennedy, Catherine Emmanuel, Sian Fereday, Kathryn Alsop, and Dale W. Garsed
- Abstract
Revised Supplementary Data, containing Supplementary Methods, Supplementary Figures, Supplementary Tables Supplementary Figure S1. Outline of cohort selection and analyses. Supplementary Figure S2. Clinical response and therapy course of 96 patients with exceptional responses to chemotherapy. Supplementary Figure S3. Distribution and type of TP53 mutations. Supplementary Figure S4. RB1 protein expression altered by genomic inactivation. Supplementary Figure S5. Characterization of CD8 and Ki-67 in tumors according to homologous recombination mutation status. Supplementary Table S2 Immunohistochemical analysis: primary antibodies and staining conditions Supplementary Table S3 Homologous recombination and DNA repair panel Supplementary Table S6 Comparison of molecular alteration prevalence between clinical subgroups Supplementary Table S7 Patient characteristics of tissue microarray cohort
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- 2023
29. Data from EIF1AX and NRAS Mutations Co-occur and Cooperate in Low-Grade Serous Ovarian Carcinomas
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Anna deFazio, David D.L. Bowtell, Ivan B. Lomakin, Helen Rizos, Martin Köbel, Sean M. Grimmond, Nicola Waddell, John V. Pearson, Timothy Semple, Timothy P. Holloway, Gisela Mir Arnau, George Au-Yeung, Joshy George, Ann-Marie Patch, Elizabeth L. Christie, Jason Li, Paul R. Harnett, Raghwa Sharma, Joy Hendley, Yoke-Eng Chiew, Chris Mitchell, Sian Fereday, Catherine J. Kennedy, Dale W. Garsed, Tania Moujaber, Ying Lei, Walid J. Azar, and Dariush Etemadmoghadam
- Abstract
Low-grade serous ovarian carcinomas (LGSC) are associated with a poor response to chemotherapy and are molecularly characterized by RAS pathway activation. Using exome and whole genome sequencing, we identified recurrent mutations in the protein translational regulator EIF1AX and in NF1, USP9X, KRAS, BRAF, and NRAS. RAS pathway mutations were mutually exclusive; however, we found significant co-occurrence of mutations in NRAS and EIF1AX. Missense EIF1AX mutations were clustered at the N-terminus of the protein in a region associated with its role in ensuring translational initiation fidelity. Coexpression of mutant NRAS and EIF1AX proteins promoted proliferation and clonogenic survival in LGSC cells, providing the first example of co-occurring, growth-promoting mutational events in ovarian cancer. Cancer Res; 77(16); 4268–78. ©2017 AACR.
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- 2023
30. Supplementary Figure 2 from Gemcitabine and CHK1 Inhibition Potentiate EGFR-Directed Radioimmunotherapy against Pancreatic Ductal Adenocarcinoma
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Kum Kum Khanna, Michael P. Brown, Sean M. Grimmond, Andrew V. Biankin, Angela Chou, Adnan M. Nagrial, Mariska Miranda, Murugan Kalimutho, Wei Shi, Marina Pajic, and Fares Al-Ejeh
- Abstract
Supplementary Figure 2: Maximum tolerated doses of gemcitabine, Chk1i and EGFR-directed RIT in vivo. (A) Balb/c nude mice bearing PANC-1 subcutaneous xenografts (60 mm3 in volume) were treated with 50 mg/kg or 100 mg/kg of gemcitabine on days 1, 4, 7 and 10 administered intravenously combined with 15 mg/kg Chk1i, administered as two doses per day (7.5 mg/kg per dose) on days 1, 4, 7 and 10 at 3 hours before and after gemcitabine administration. Mice were treated with gemcitabine and Chk1i combinations (gem + Chk1i) alone or in combination with 6 or 9 MBq/20g (300 or 450 MBq/kg) of 177Lu-anti-EGFR mAb. Mice were monitored for 14 days to determine acute toxicities as judged by weight, posture, movement, eating and drinking. Tumor volume was also monitored during this short time as an indication of efficacy. (B) The tumor growth curves were used to measure the growth rate (k = day-1) from exponential growth equations and shown in the bar graph in panel (5 mice per group, error bar is the standard error of the mean, SEM). *** p < 0.001 in One-way ANOVA in GraphPad® Prism comparing treatments involving anti-EGFR RIT and those not involving this RIT (p
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- 2023
31. Supplemental Material - Group Authorship from EIF1AX and NRAS Mutations Co-occur and Cooperate in Low-Grade Serous Ovarian Carcinomas
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Anna deFazio, David D.L. Bowtell, Ivan B. Lomakin, Helen Rizos, Martin Köbel, Sean M. Grimmond, Nicola Waddell, John V. Pearson, Timothy Semple, Timothy P. Holloway, Gisela Mir Arnau, George Au-Yeung, Joshy George, Ann-Marie Patch, Elizabeth L. Christie, Jason Li, Paul R. Harnett, Raghwa Sharma, Joy Hendley, Yoke-Eng Chiew, Chris Mitchell, Sian Fereday, Catherine J. Kennedy, Dale W. Garsed, Tania Moujaber, Ying Lei, Walid J. Azar, and Dariush Etemadmoghadam
- Abstract
Supplemental Material - Group Authorship from EIF1AX and NRAS Mutations Co-occur and Cooperate in Low-Grade Serous Ovarian Carcinomas
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- 2023
32. Supplemental Materials and Methods from EIF1AX and NRAS Mutations Co-occur and Cooperate in Low-Grade Serous Ovarian Carcinomas
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Anna deFazio, David D.L. Bowtell, Ivan B. Lomakin, Helen Rizos, Martin Köbel, Sean M. Grimmond, Nicola Waddell, John V. Pearson, Timothy Semple, Timothy P. Holloway, Gisela Mir Arnau, George Au-Yeung, Joshy George, Ann-Marie Patch, Elizabeth L. Christie, Jason Li, Paul R. Harnett, Raghwa Sharma, Joy Hendley, Yoke-Eng Chiew, Chris Mitchell, Sian Fereday, Catherine J. Kennedy, Dale W. Garsed, Tania Moujaber, Ying Lei, Walid J. Azar, and Dariush Etemadmoghadam
- Abstract
Supplemental materials and methods
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- 2023
33. Supplementary Tables from EIF1AX and NRAS Mutations Co-occur and Cooperate in Low-Grade Serous Ovarian Carcinomas
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Anna deFazio, David D.L. Bowtell, Ivan B. Lomakin, Helen Rizos, Martin Köbel, Sean M. Grimmond, Nicola Waddell, John V. Pearson, Timothy Semple, Timothy P. Holloway, Gisela Mir Arnau, George Au-Yeung, Joshy George, Ann-Marie Patch, Elizabeth L. Christie, Jason Li, Paul R. Harnett, Raghwa Sharma, Joy Hendley, Yoke-Eng Chiew, Chris Mitchell, Sian Fereday, Catherine J. Kennedy, Dale W. Garsed, Tania Moujaber, Ying Lei, Walid J. Azar, and Dariush Etemadmoghadam
- Abstract
Supplementary Table S1. Patient characteristics; Supplementary Table S2. Ki67 proliferation index; Supplementary Table S3. WES and WGS performance statistics; Supplementary Table S4. Amplicon sequencing primers; Supplementary Table S5. High confidence variants from exome and whole genome sequencing; Supplementary Table S6. Summary of SNVs and indels by sample; Supplementary Table S7. Significantly mutated genes (MuSiC output); Supplementary Table S8. Somatic variants in recurrently mutated genes; Supplementary Table S9. Validation cohort variants; Supplementary Table S10. TP53 variants in validation cohort; Supplementary Table S11. EIF1AX mutations reported in cancer
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- 2023
34. Supplementary Methods from Gemcitabine and CHK1 Inhibition Potentiate EGFR-Directed Radioimmunotherapy against Pancreatic Ductal Adenocarcinoma
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Kum Kum Khanna, Michael P. Brown, Sean M. Grimmond, Andrew V. Biankin, Angela Chou, Adnan M. Nagrial, Mariska Miranda, Murugan Kalimutho, Wei Shi, Marina Pajic, and Fares Al-Ejeh
- Abstract
Detailed methods of: EGFR immunohistochemistry of PDAC tissue microarrays Patient derived xenografts (PDXs) and cell lines (PDCLs) generation Cell culture Treatments in vitro Histological and immunoblotting analyses of PDXs post-treatment
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- 2023
35. Supplementary Figure 4 from Gemcitabine and CHK1 Inhibition Potentiate EGFR-Directed Radioimmunotherapy against Pancreatic Ductal Adenocarcinoma
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Kum Kum Khanna, Michael P. Brown, Sean M. Grimmond, Andrew V. Biankin, Angela Chou, Adnan M. Nagrial, Mariska Miranda, Murugan Kalimutho, Wei Shi, Marina Pajic, and Fares Al-Ejeh
- Abstract
Supplementary Figure 4: Efficacy of anti-EGFR directed RIT in combination therapy in vivo against PDAC model from the BxPC-3 ATCCTM cell lines. BxPC-3 cells expressing luciferase were used to inoculate balb/c nude mice subcutaneously at 4 weeks of age. Treatments were initiated when tumors reached 60 mm3 as outlined in diagram in A and in Figure 2. Single agent treatments included: EGFR control: 50 μg of unlabeled anti-EGFR mAb; Chk1i: 15 mg/kg of the Chk1 inhibitor PF-477736 administered as two 7.5 mg/kg injections per day subcutaneously on days 1, 4, 7 and 10; Gem: 50 mg/kg of gemcitabine administered intravenously on days 1, 4, 7 and 10; RIT: 177Lu-anti-EGFR mAb (50 μg with 6 MBq radioactivity per 20 g mouse) injected intravenously on day 2 only. For combinations, treatments were performed as described for the single agents where Chk1i was administered 3 hours before and after gemcitabine on days 1, 4, 7 and 10 and RIT was administered on day 2 only. (B) Tumor growth curves presented as the change in tumor volume compared to day 0 (% change +SEM, n = 5 mice per treatment group). (C) Representative images of live bioluminescence imaging using luciferin performed on day 1 (prior to treatment) and day 14 (96 hours after treatments). (D) Quantification and representative images of live bioluminescence imaging using caspase-3/7 substrate (Z-DEVD-Luciferase) performed on day 7 after treatment initiation. Data shown is the average of caspase-3/7 activation (+SEM, n = 5 per treatment group). *** p < 0.001 in One-way ANOVA in GraphPad® Prism.
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- 2023
36. Supplementary Figures from EIF1AX and NRAS Mutations Co-occur and Cooperate in Low-Grade Serous Ovarian Carcinomas
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Anna deFazio, David D.L. Bowtell, Ivan B. Lomakin, Helen Rizos, Martin Köbel, Sean M. Grimmond, Nicola Waddell, John V. Pearson, Timothy Semple, Timothy P. Holloway, Gisela Mir Arnau, George Au-Yeung, Joshy George, Ann-Marie Patch, Elizabeth L. Christie, Jason Li, Paul R. Harnett, Raghwa Sharma, Joy Hendley, Yoke-Eng Chiew, Chris Mitchell, Sian Fereday, Catherine J. Kennedy, Dale W. Garsed, Tania Moujaber, Ying Lei, Walid J. Azar, and Dariush Etemadmoghadam
- Abstract
Supplementary Figure S1. Representative hematoxylin and eosin stained sections from sequenced LGSC; Supplementary Figure S2. Age distribution analysis of patients from the Australian Ovarian Cancer Study. Includes patients with advanced stage, serous epithelial ovarian cancer (n = 684); Supplementary Figure S3. RAS pathway mutations in LGSC; Supplementary Figure S4. Verification of EIF1AX mutations; Supplementary Figure S5. Characterization of the AOCS2 cell line; Supplementary Figure S6. Functional effect of EIF1AX mutations and gene suppression in a LGSC cell line; Supplementary Figure S7. (a) EIF1AX and NRAS function in the mTOR and RAS/ERK signaling pathways to regulate protein translation, cell proliferation and cell survival
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- 2023
37. Supplementary Figure 2 from Neuropilin-2 Promotes Extravasation and Metastasis by Interacting with Endothelial α5 Integrin
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Debabrata Mukhopadhyay, Xiaohui Zhang, Roger J. Daly, Andrew V. Biankin, Sean M. Grimmond, Nicola Waddell, David K. Chang, Mark J. Cowley, Jianmin Wu, Enfeng Wang, Yan Guo, Guangqi E, Steven Bach, Luke H. Hoeppner, and Ying Cao
- Abstract
PDF file - 239K, Supplemental Fig.2. NRP-2 knockdown in 786-O cells didn't significantly change the blood vessels and lymphatic vessels densities in the primary tumors. (A). Representative images of vWF stained blood vessels. (B). Quantification of vWF staining. One typical area from the edge of the tumor (to avoid the necrotic area) was taken and vWF vessel density was quantified. n=5. (C). Representative images of LYVE stained lymphatic vessels. (D). Quantification of LYVE staining. One typical area from the edge of the tumor (to avoid the necrotic area) was taken and LYVE vessel density was quantified. n=5.
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- 2023
38. Supplementary Figure 6 from Neuropilin-2 Promotes Extravasation and Metastasis by Interacting with Endothelial α5 Integrin
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Debabrata Mukhopadhyay, Xiaohui Zhang, Roger J. Daly, Andrew V. Biankin, Sean M. Grimmond, Nicola Waddell, David K. Chang, Mark J. Cowley, Jianmin Wu, Enfeng Wang, Yan Guo, Guangqi E, Steven Bach, Luke H. Hoeppner, and Ying Cao
- Abstract
PDF file - 59K, Supplemental Fig.6. NRP-2 did not co-immunoprecipitate with Integrin a2, a3, a6 or aV.
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- 2023
39. Supplementary Figure 5 from Neuropilin-2 Promotes Extravasation and Metastasis by Interacting with Endothelial α5 Integrin
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Debabrata Mukhopadhyay, Xiaohui Zhang, Roger J. Daly, Andrew V. Biankin, Sean M. Grimmond, Nicola Waddell, David K. Chang, Mark J. Cowley, Jianmin Wu, Enfeng Wang, Yan Guo, Guangqi E, Steven Bach, Luke H. Hoeppner, and Ying Cao
- Abstract
PDF file - 164K, Supplemental Fig. 5. The effect of NRP-2 knock down on ASPC-1 cell primary tumor growth and liver metastasis. A. NRP-2 knockdown did not significantly reduce ASPC-1 primary tumor growth. 2 x 106 control or NRP-2 shRNA ASPC-1 cells were injected into SCID mice pancreas, and after 15 days, tumor weight was analyzed. B. Total images of livers from control and NRP-2 knockdown ASPC-1 tumor burden mice. The ASPC-1 cells contain GFP and the GFP fluoresce signal in the liver was examined by IVIS Imaging System 200.
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- 2023
40. Supplementary Figure 3 from Neuropilin-2 Promotes Extravasation and Metastasis by Interacting with Endothelial α5 Integrin
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Debabrata Mukhopadhyay, Xiaohui Zhang, Roger J. Daly, Andrew V. Biankin, Sean M. Grimmond, Nicola Waddell, David K. Chang, Mark J. Cowley, Jianmin Wu, Enfeng Wang, Yan Guo, Guangqi E, Steven Bach, Luke H. Hoeppner, and Ying Cao
- Abstract
PDF file - 104K, Supplemental Fig.3. (A). NRP-2 knockdown in 786-O didn't significantly change cell proliferation, migration and invasion. (B). NRP-2 knockdown in 786-O and A-498 did not significantly change p-AKT, p-ERK, p-p38, p53 and MDM-2 levels.
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- 2023
41. Supplementary Figure Legend from Neuropilin-2 Promotes Extravasation and Metastasis by Interacting with Endothelial α5 Integrin
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Debabrata Mukhopadhyay, Xiaohui Zhang, Roger J. Daly, Andrew V. Biankin, Sean M. Grimmond, Nicola Waddell, David K. Chang, Mark J. Cowley, Jianmin Wu, Enfeng Wang, Yan Guo, Guangqi E, Steven Bach, Luke H. Hoeppner, and Ying Cao
- Abstract
PDF file - 49K
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- 2023
42. Supplementary Figure 4 from Neuropilin-2 Promotes Extravasation and Metastasis by Interacting with Endothelial α5 Integrin
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Debabrata Mukhopadhyay, Xiaohui Zhang, Roger J. Daly, Andrew V. Biankin, Sean M. Grimmond, Nicola Waddell, David K. Chang, Mark J. Cowley, Jianmin Wu, Enfeng Wang, Yan Guo, Guangqi E, Steven Bach, Luke H. Hoeppner, and Ying Cao
- Abstract
PDF file - 86K, Supplemental Fig.4. (A). NRP-2 levels in 786-O cells after overexpression or knockdown. (B). NRP-2 overexpression or knockdown didn't significantly change the abilities to adhere to blank plate and Collagen I coated plate.
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- 2023
43. Supplementary Figure 1 from Neuropilin-2 Promotes Extravasation and Metastasis by Interacting with Endothelial α5 Integrin
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Debabrata Mukhopadhyay, Xiaohui Zhang, Roger J. Daly, Andrew V. Biankin, Sean M. Grimmond, Nicola Waddell, David K. Chang, Mark J. Cowley, Jianmin Wu, Enfeng Wang, Yan Guo, Guangqi E, Steven Bach, Luke H. Hoeppner, and Ying Cao
- Abstract
PDF file - 101K, Supplemental Fig.1. The impact of NRP-2 knockdown on primary tumor growth. (A) Knockdown of NRP-2 by shRNA in 786O cells caused upregulation of NRP-1. (B & C) 2 x 106 control and NRP-2 knockdown 786-O (B) and A-498 cells (C) were subcutaneously injected into nude mice. There was no significant different in the tumor volume between the control and NRP-2 knockdown groups in both 786-O and A-498 cells.
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- 2023
44. Novel RET Fusion RET-SEPTIN9 Predicts Response to Selective RET Inhibition With Selpercatinib in Malignant Pheochromocytoma
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Andrew D Pattison, Angela Mweempwa, Huiling Xu, Sean M. Grimmond, Stephen B. Fox, Benjamin Solomon, Rodney J. Hicks, Richard W. Tothill, Stephen J Luen, David Thomas, Gary Richardson, Joseph H.A. Vissers, Jayesh Desai, and Andrew Fellowes
- Subjects
Malignant Pheochromocytoma ,Cancer Research ,Oncology ,business.industry ,Cancer research ,Medicine ,RET Fusion ,business - Published
- 2021
45. Comprehensive genomic and tumour immune profiling reveals potential therapeutic targets in malignant pleural mesothelioma
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Jenette Creaney, Ann-Marie Patch, Venkateswar Addala, Sophie A. Sneddon, Katia Nones, Ian M. Dick, Y. C. Gary Lee, Felicity Newell, Ebony J. Rouse, Marjan M. Naeini, Olga Kondrashova, Vanessa Lakis, Apostolos Nakas, David Waller, Annabel Sharkey, Pamela Mukhopadhyay, Stephen H. Kazakoff, Lambros T. Koufariotis, Aimee L. Davidson, Priya Ramarao-Milne, Oliver Holmes, Qinying Xu, Conrad Leonard, Scott Wood, Sean M. Grimmond, Raphael Bueno, Dean A. Fennell, John V. Pearson, Bruce W. Robinson, and Nicola Waddell
- Subjects
Mesothelioma ,Lung Neoplasms ,Pleural Neoplasms ,Mesothelioma, Malignant ,Tumor Microenvironment ,Genetics ,Humans ,Molecular Medicine ,Genomics ,Molecular Biology ,Genetics (clinical) - Abstract
Background Malignant pleural mesothelioma (MPM) has a poor overall survival with few treatment options. Whole genome sequencing (WGS) combined with the immune features of MPM offers the prospect of identifying changes that could inform future clinical trials. Methods We analysed somatic mutations from 229 MPM samples, including previously published data and 58 samples that had undergone WGS within this study. This was combined with RNA-seq analysis to characterize the tumour immune environment. Results The comprehensive genome analysis identified 12 driver genes, including new candidate genes. Whole genome doubling was a frequent event that correlated with shorter survival. Mutational signature analysis revealed SBS5/40 were dominant in 93% of samples, and defects in homologous recombination repair were infrequent in our cohort. The tumour immune environment contained high M2 macrophage infiltrate linked with MMP2, MMP14, TGFB1 and CCL2 expression, representing an immune suppressive environment. The expression of TGFB1 was associated with overall survival. A small subset of samples (less than 10%) had a higher proportion of CD8 T cells and a high cytolytic score, suggesting a ‘hot’ immune environment independent of the somatic mutations. Conclusions We propose accounting for genomic and immune microenvironment status may influence therapeutic planning in the future.
- Published
- 2022
46. Proliferation drives quorum sensing of microbial products in human macrophage populations
- Author
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Nadia Rajab, Linden J. Gearing, Ruqian Lyu, Yair D.J. Prawer, Paul W. Angel, Sean M. Grimmond, Andrew L. Laslett, Davis J. McCarthy, and Christine A. Wells
- Abstract
Macrophages coordinate the initial host inflammatory response to tissue infection, as well as mediating the reparative phase, by producing growth factors that promote tissue repair. One model of this functional dichotomy is that peripherally recruited monocyte-derived macrophages drive acute inflammatory responses to infection, whereas tissue-resident macrophages are responsible for tissue repair. Alternatively, inflammation and repair may be inter-dependent molecular programs, such that both recruited and resident cells have equivalent capacity to contribute. Repeated exposure to pathogenic challenge results in innate tolerance, which may also alter the contributions of discrete macrophage populations to inflammation or repair. In this study a village model of tissue resident and recruited macrophages was created using induced pluripotent stem cell-derived macrophages and peripheral blood monocyte-derived macrophages, respectively. Population responses to repeated exposure to lipopolysaccharide were assessed with single-cell RNA sequencing and donors demultiplexed with Vireo. A subset of genes escaped classical tolerance programs in the iPSC, but not monocyte-derived macrophages, and this was associated with differences in their proliferative capacity. This suggests that targeting the proliferative resident macrophages would be most effective to limit inflammatory signaling.
- Published
- 2022
47. Correction to: S100 family proteins are linked to organoid morphology and EMT in pancreatic cancer
- Author
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Ronnie Ren Jie Low, Ka Yee Fung, Hugh Gao, Adele Preaudet, Laura F. Dagley, Jumana Yousef, Belinda Lee, Samantha J. Emery-Corbin, Paul M. Nguyen, Rune H. Larsen, Nadia J. Kershaw, Antony W. Burgess, Peter Gibbs, Frédéric Hollande, Michael D. W. Griffin, Sean M. Grimmond, and Tracy L. Putoczki
- Subjects
Cell Biology ,Molecular Biology - Published
- 2023
48. Methyl-CpG binding domain 4, DNA glycosylase (MBD4)-associated neoplasia syndrome associated with a homozygous missense variant in MBD4: Expansion of an emerging phenotype
- Author
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Piers Blombery, Georgina L. Ryland, Lucy C. Fox, Zornitza Stark, Meg Wall, Anna Jarmolowicz, Ain Roesley, Ella R. Thompson, Sean M. Grimmond, Shyam Panicker, and Fiona Kwok
- Subjects
Endodeoxyribonucleases ,Phenotype ,DNA Repair ,Neoplasms ,Humans ,Hematology ,Amino Acid Sequence ,DNA Methylation ,DNA Glycosylases - Published
- 2022
49. TGF-β1 induced S100 family protein expression is associated with epithelial to mesenchymal transition states and poor survival in pancreatic cancer
- Author
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Ronnie Ren Jie Low, Ka Yee Fung, Hugh Gao, Adele Preaudet, Laura F. Dagley, Jumana Yousef, Belinda Lee, Rune H. Larsen, Nadia J. Kershaw, Antony W. Burgess, Peter Gibbs, Frédéric Hollande, Michael D.W. Griffin, Sean M. Grimmond, and Tracy L. Putoczki
- Subjects
embryonic structures - Abstract
SUMMARYEpithelial-mesenchymal transition (EMT) is a continuum that includes epithelial, partial EMT (P-EMT) and mesenchymal states, each of which are associated with cancer progression, invasive capabilities and ultimately metastasis. We have employed a lineage traced sporadic model of pancreatic cancer to generate a murine organoid biobank from primary and secondary tumors, including sublines that have undergone P-EMT and complete EMT (C-EMT). Using an unbiased proteomics approach, we found that the morphology of the organoids predicts the EMT state, with solid organoids associated with a P-EMT signature. We also observed that exogenous TGFβ1 induces a solid organoid morphology that is associated with changes in the S100 family, C-EMT and the formation of high-grade tumors. S100A4 may represent a useful biomarker to predict EMT state, disease progression and outcome for pancreatic cancer patients.
- Published
- 2022
50. PINA v2.0: mining interactome modules.
- Author
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Mark J. Cowley, Mark Pinese, Karin S. Kassahn, Nic Waddell, John V. Pearson, Sean M. Grimmond, Andrew V. Biankin, Sampsa Hautaniemi, and Jianmin Wu
- Published
- 2012
- Full Text
- View/download PDF
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