Sir Peter MacCallum Department of Oncology - Research Publications
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ItemMolecular comparison of interval and screen-detected breast cancers(WILEY, 2019-06)
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ItemA simple consensus approach improves somatic mutation prediction accuracy(BMC, 2013-09-30)Differentiating true somatic mutations from artifacts in massively parallel sequencing data is an immense challenge. To develop methods for optimal somatic mutation detection and to identify factors influencing somatic mutation prediction accuracy, we validated predictions from three somatic mutation detection algorithms, MuTect, JointSNVMix2 and SomaticSniper, by Sanger sequencing. Full consensus predictions had a validation rate of >98%, but some partial consensus predictions validated too. In cases of partial consensus, read depth and mapping quality data, along with additional prediction methods, aided in removing inaccurate predictions. Our consensus approach is fast, flexible and provides a high-confidence list of putative somatic mutations.
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ItemLoss of heterozygosity: what is it good for?(BMC, 2015-08-01)BACKGROUND: Loss of heterozygosity (LOH) is a common genetic event in cancer development, and is known to be involved in the somatic loss of wild-type alleles in many inherited cancer syndromes. The wider involvement of LOH in cancer is assumed to relate to unmasking a somatically mutated tumour suppressor gene through loss of the wild type allele. METHODS: We analysed 86 ovarian carcinomas for mutations in 980 genes selected on the basis of their location in common regions of LOH. RESULTS: We identified 36 significantly mutated genes, but these could only partly account for the quanta of LOH in the samples. Using our own and TCGA data we then evaluated five possible models to explain the selection for non-random accumulation of LOH in ovarian cancer genomes: 1. Classic two-hit hypothesis: high frequency biallelic genetic inactivation of tumour suppressor genes. 2. Epigenetic two-hit hypothesis: biallelic inactivation through methylation and LOH. 3. Multiple alternate-gene biallelic inactivation: low frequency gene disruption. 4. Haplo-insufficiency: Single copy gene disruption. 5. Modified two-hit hypothesis: reduction to homozygosity of low penetrance germline predisposition alleles. We determined that while high-frequency biallelic gene inactivation under model 1 is rare, regions of LOH (particularly copy-number neutral LOH) are enriched for deleterious mutations and increased promoter methylation, while copy-number loss LOH regions are likely to contain under-expressed genes suggestive of haploinsufficiency. Reduction to homozygosity of cancer predisposition SNPs may also play a minor role. CONCLUSION: It is likely that selection for regions of LOH depends on its effect on multiple genes. Selection for copy number neutral LOH may better fit the classic two-hit model whereas selection for copy number loss may be attributed to its effect on multi-gene haploinsufficiency. LOH mapping alone is unlikely to be successful in identifying novel tumour suppressor genes; a combined approach may be more effective.
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ItemThe genetic architecture of breast papillary lesions as a predictor of progression to carcinoma(NATURE PORTFOLIO, 2020-03-12)Intraductal papillomas (IDP) are challenging breast findings because of their variable risk of progression to malignancy. The molecular events driving IDP development and genomic features of malignant progression are poorly understood. In this study, genome-wide CNA and/or targeted mutation analysis was performed on 44 cases of IDP, of which 20 cases had coexisting ductal carcinoma in situ (DCIS), papillary DCIS or invasive ductal carcinoma (IDC). CNA were rare in pure IDP, but 69% carried an activating PIK3CA mutation. Among the synchronous IDP cases, 55% (11/20) were clonally related to the synchronous DCIS and/or IDC, only one of which had papillary histology. In contrast to pure IDP, PIK3CA mutations were absent from clonal cases. CNAs in any of chromosomes 1, 16 or 11 were significantly enriched in clonal IDP lesions compared to pure and non-clonal IDP. The observation that 55% of IDP are clonal to DCIS/IDC indicates that IDP can be a direct precursor for breast carcinoma, not limited to the papillary type. The absence of PIK3CA mutations and presence of CNAs in IDP could be used clinically to identify patients at high risk of progression to carcinoma.
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ItemBioinformatics Pipelines for Targeted Resequencing and Whole-Exome Sequencing of Human and Mouse Genomes: A Virtual Appliance Approach for Instant Deployment(PUBLIC LIBRARY SCIENCE, 2014-04-21)Targeted resequencing by massively parallel sequencing has become an effective and affordable way to survey small to large portions of the genome for genetic variation. Despite the rapid development in open source software for analysis of such data, the practical implementation of these tools through construction of sequencing analysis pipelines still remains a challenging and laborious activity, and a major hurdle for many small research and clinical laboratories. We developed TREVA (Targeted REsequencing Virtual Appliance), making pre-built pipelines immediately available as a virtual appliance. Based on virtual machine technologies, TREVA is a solution for rapid and efficient deployment of complex bioinformatics pipelines to laboratories of all sizes, enabling reproducible results. The analyses that are supported in TREVA include: somatic and germline single-nucleotide and insertion/deletion variant calling, copy number analysis, and cohort-based analyses such as pathway and significantly mutated genes analyses. TREVA is flexible and easy to use, and can be customised by Linux-based extensions if required. TREVA can also be deployed on the cloud (cloud computing), enabling instant access without investment overheads for additional hardware. TREVA is available at http://bioinformatics.petermac.org/treva/.
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ItemEvaluating the breast cancer predisposition role of rare variants in genes associated with low-penetrance breast cancer risk SNPs(BIOMED CENTRAL LTD, 2018-01-09)BACKGROUND: Genome-wide association studies (GWASs) have identified numerous single-nucleotide polymorphisms (SNPs) associated with small increases in breast cancer risk. Studies to date suggest that some SNPs alter the expression of the associated genes, which potentially mediates risk modification. On this basis, we hypothesised that some of these genes may be enriched for rare coding variants associated with a higher breast cancer risk. METHODS: The coding regions and exon-intron boundaries of 56 genes that have either been proposed by GWASs to be the regulatory targets of the SNPs and/or located < 500 kb from the risk SNPs were sequenced in index cases from 1043 familial breast cancer families that previously had negative test results for BRCA1 and BRCA2 mutations and 944 population-matched cancer-free control participants from an Australian population. Rare (minor allele frequency ≤ 0.001 in the Exome Aggregation Consortium and Exome Variant Server databases) loss-of-function (LoF) and missense variants were studied. RESULTS: LoF variants were rare in both the cases and control participants across all the candidate genes, with only 38 different LoF variants observed in a total of 39 carriers. For the majority of genes (n = 36), no LoF variants were detected in either the case or control cohorts. No individual gene showed a significant excess of LoF or missense variants in the cases compared with control participants. Among all candidate genes as a group, the total number of carriers with LoF variants was higher in the cases than in the control participants (26 cases and 13 control participants), as was the total number of carriers with missense variants (406 versus 353), but neither reached statistical significance (p = 0.077 and p = 0.512, respectively). The genes contributing most of the excess of LoF variants in the cases included TET2, NRIP1, RAD51B and SNX32 (12 cases versus 2 control participants), whereas ZNF283 and CASP8 contributed largely to the excess of missense variants (25 cases versus 8 control participants). CONCLUSIONS: Our data suggest that rare LoF and missense variants in genes associated with low-penetrance breast cancer risk SNPs may contribute some additional risk, but as a group these genes are unlikely to be major contributors to breast cancer heritability.
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ItemCopy number analysis by low coverage whole genome sequencing using ultra low-input DNA from formalin-fixed paraffin embedded tumor tissue(BMC, 2016-11-15)Unlocking clinically translatable genomic information, including copy number alterations (CNA), from formalin-fixed paraffin-embedded (FFPE) tissue is challenging due to low yields and degraded DNA. We describe a robust, cost-effective low-coverage whole genome sequencing (LC WGS) method for CNA detection using 5 ng of FFPE-derived DNA. CN profiles using 100 ng or 5 ng input DNA were highly concordant and comparable with molecular inversion probe (MIP) array profiles. LC WGS improved CN profiles of samples that performed poorly using MIP arrays. Our technique enables identification of driver and prognostic CNAs in archival patient samples previously deemed unsuitable for genomic analysis due to DNA limitations.