Medical Biology - Research Publications
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ItemNo Preview AvailableTargeting histone acetylation dynamics and oncogenic transcription by catalytic P300/CBP inhibition(CELL PRESS, 2021-05-20)To separate causal effects of histone acetylation on chromatin accessibility and transcriptional output, we used integrated epigenomic and transcriptomic analyses following acute inhibition of major cellular lysine acetyltransferases P300 and CBP in hematological malignancies. We found that catalytic P300/CBP inhibition dynamically perturbs steady-state acetylation kinetics and suppresses oncogenic transcriptional networks in the absence of changes to chromatin accessibility. CRISPR-Cas9 screening identified NCOR1 and HDAC3 transcriptional co-repressors as the principal antagonists of P300/CBP by counteracting acetylation turnover kinetics. Finally, deacetylation of H3K27 provides nucleation sites for reciprocal methylation switching, a feature that can be exploited therapeutically by concomitant KDM6A and P300/CBP inhibition. Overall, this study indicates that the steady-state histone acetylation-methylation equilibrium functions as a molecular rheostat governing cellular transcription that is amenable to therapeutic exploitation as an anti-cancer regimen.
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ItemMelanoma brain metastases that progress on BRAF-MEK inhibitors demonstrate resistance to ipilimumab-nivolumab that is associated with the Innate PD-1 Resistance Signature (IPRES)(BMJ PUBLISHING GROUP, 2021-10)BACKGROUND: Melanoma brain metastases (MBMs) are a challenging clinical problem with high morbidity and mortality. Although first-line dabrafenib-trametinib and ipilimumab-nivolumab have similar intracranial response rates (50%-55%), central nervous system (CNS) resistance to BRAF-MEK inhibitors (BRAF-MEKi) usually occurs around 6 months, and durable responses are only seen with combination immunotherapy. We sought to investigate the utility of ipilimumab-nivolumab after MBM progression on BRAF-MEKi and identify mechanisms of resistance. METHODS: Patients who received first-line ipilimumab-nivolumab for MBMs or second/third line ipilimumab-nivolumab for intracranial metastases with BRAFV600 mutations with prior progression on BRAF-MEKi and MRI brain staging from March 1, 2015 to June 30, 2018 were included. Modified intracranial RECIST was used to assess response. Formalin-fixed paraffin-embedded samples of BRAFV600 mutant MBMs that were naïve to systemic treatment (n=18) or excised after progression on BRAF-MEKi (n=14) underwent whole transcriptome sequencing. Comparative analyses of MBMs naïve to systemic treatment versus BRAF-MEKi progression were performed. RESULTS: Twenty-five and 30 patients who received first and second/third line ipilimumab-nivolumab, were included respectively. Median sum of MBM diameters was 13 and 20.5 mm for the first and second/third line ipilimumab-nivolumab groups, respectively. Intracranial response rate was 75.0% (12/16), and median progression-free survival (PFS) was 41.6 months for first-line ipilimumab-nivolumab. Efficacy of second/third line ipilimumab-nivolumab after BRAF-MEKi progression was poor with an intracranial response rate of 4.8% (1/21) and median PFS of 1.3 months. Given the poor activity of ipilimumab-nivolumab after BRAF-MEKi MBM progression, we performed whole transcriptome sequencing to identify mechanisms of drug resistance. We identified a set of 178 differentially expressed genes (DEGs) between naïve and MBMs with progression on BRAF-MEKi treatment (p value <0.05, false discovery rate (FDR) <0.1). No distinct pathways were identified from gene set enrichment analyses using Kyoto Encyclopedia of Genes and Genomes, Gene Ontogeny or Hallmark libraries; however, enrichment of DEG from the Innate Anti-PD1 Resistance Signature (IPRES) was identified (p value=0.007, FDR=0.03). CONCLUSIONS: Second-line ipilimumab-nivolumab for MBMs after BRAF-MEKi progression has poor activity. MBMs that are resistant to BRAF-MEKi that also conferred resistance to second-line ipilimumab-nivolumab showed enrichment of the IPRES gene signature.
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ItemSafety, infectivity and immunogenicity of a Chick for genetically attenuated blood-stage malaria updates vaccine(BMC, 2021-11-22)BACKGROUND: There is a clear need for novel approaches to malaria vaccine development. We aimed to develop a genetically attenuated blood-stage vaccine and test its safety, infectivity, and immunogenicity in healthy volunteers. Our approach was to target the gene encoding the knob-associated histidine-rich protein (KAHRP), which is responsible for the assembly of knob structures at the infected erythrocyte surface. Knobs are required for correct display of the polymorphic adhesion ligand P. falciparum erythrocyte membrane protein 1 (PfEMP1), a key virulence determinant encoded by a repertoire of var genes. METHODS: The gene encoding KAHRP was deleted from P. falciparum 3D7 and a master cell bank was produced in accordance with Good Manufacturing Practice. Eight malaria naïve males were intravenously inoculated (day 0) with 1800 (2 subjects), 1.8 × 105 (2 subjects), or 3 × 106 viable parasites (4 subjects). Parasitemia was measured using qPCR; immunogenicity was determined using standard assays. Parasites were rescued into culture for in vitro analyses (genome sequencing, cytoadhesion assays, scanning electron microscopy, var gene expression). RESULTS: None of the subjects who were administered with 1800 or 1.8 × 105 parasites developed parasitemia; 3/4 subjects administered 3× 106 parasites developed significant parasitemia, first detected on days 13, 18, and 22. One of these three subjects developed symptoms of malaria simultaneously with influenza B (day 17; 14,022 parasites/mL); one subject developed mild symptoms on day 28 (19,956 parasites/mL); and one subject remained asymptomatic up to day 35 (5046 parasites/mL). Parasitemia rapidly cleared with artemether/lumefantrine. Parasitemia induced a parasite-specific antibody and cell-mediated immune response. Parasites cultured ex vivo exhibited genotypic and phenotypic properties similar to inoculated parasites, although the var gene expression profile changed during growth in vivo. CONCLUSIONS: This study represents the first clinical investigation of a genetically attenuated blood-stage human malaria vaccine. A P. falciparum 3D7 kahrp- strain was tested in vivo and found to be immunogenic but can lead to patent parasitemia at high doses. TRIAL REGISTRATION: Australian New Zealand Clinical Trials Registry (number: ACTRN12617000824369 ; date: 06 June 2017).
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ItemType 1 diabetes in pregnancy is associated with distinct changes in the composition and function of the gut microbiome(BMC, 2021-08-06)BACKGROUND: The gut microbiome changes in response to a range of environmental conditions, life events and disease states. Pregnancy is a natural life event that involves major physiological adaptation yet studies of the microbiome in pregnancy are limited and their findings inconsistent. Pregnancy with type 1 diabetes (T1D) is associated with increased maternal and fetal risks but the gut microbiome in this context has not been characterized. By whole metagenome sequencing (WMS), we defined the taxonomic composition and function of the gut bacterial microbiome across 70 pregnancies, 36 in women with T1D. RESULTS: Women with and without T1D exhibited compositional and functional changes in the gut microbiome across pregnancy. Profiles in women with T1D were distinct, with an increase in bacteria that produce lipopolysaccharides and a decrease in those that produce short-chain fatty acids, especially in the third trimester. In addition, women with T1D had elevated concentrations of fecal calprotectin, a marker of intestinal inflammation, and serum intestinal fatty acid-binding protein (I-FABP), a marker of intestinal epithelial damage. CONCLUSIONS: Women with T1D exhibit a shift towards a more pro-inflammatory gut microbiome during pregnancy, associated with evidence of intestinal inflammation. These changes could contribute to the increased risk of pregnancy complications in women with T1D and are potentially modifiable by dietary means. Video abstract.
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ItemTranscriptome sequencing and multi-plex imaging of prostate cancer microenvironment reveals a dominant role for monocytic cells in progression(BMC, 2021-07-22)BACKGROUND: Prostate cancer is caused by genomic aberrations in normal epithelial cells, however clinical translation of findings from analyses of cancer cells alone has been very limited. A deeper understanding of the tumour microenvironment is needed to identify the key drivers of disease progression and reveal novel therapeutic opportunities. RESULTS: In this study, the experimental enrichment of selected cell-types, the development of a Bayesian inference model for continuous differential transcript abundance, and multiplex immunohistochemistry permitted us to define the transcriptional landscape of the prostate cancer microenvironment along the disease progression axis. An important role of monocytes and macrophages in prostate cancer progression and disease recurrence was uncovered, supported by both transcriptional landscape findings and by differential tissue composition analyses. These findings were corroborated and validated by spatial analyses at the single-cell level using multiplex immunohistochemistry. CONCLUSIONS: This study advances our knowledge concerning the role of monocyte-derived recruitment in primary prostate cancer, and supports their key role in disease progression, patient survival and prostate microenvironment immune modulation.
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ItemGRIDSS2: comprehensive characterisation of somatic structural variation using single breakend variants and structural variant phasing(BMC, 2021-07-12)GRIDSS2 is the first structural variant caller to explicitly report single breakends-breakpoints in which only one side can be unambiguously determined. By treating single breakends as a fundamental genomic rearrangement signal on par with breakpoints, GRIDSS2 can explain 47% of somatic centromere copy number changes using single breakends to non-centromere sequence. On a cohort of 3782 deeply sequenced metastatic cancers, GRIDSS2 achieves an unprecedented 3.1% false negative rate and 3.3% false discovery rate and identifies a novel 32-100 bp duplication signature. GRIDSS2 simplifies complex rearrangement interpretation through phasing of structural variants with 16% of somatic calls phasable using paired-end sequencing.
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ItemThe site of breast cancer metastases dictates their clonal composition and reversible transcriptomic profile(AMER ASSOC ADVANCEMENT SCIENCE, 2021-07)Intratumoral heterogeneity is a driver of breast cancer progression, but the nature of the clonal interactive network involved in this process remains unclear. Here, we optimized the use of optical barcoding to visualize and characterize 31 cancer subclones in vivo. By mapping the clonal composition of thousands of metastases in two clinically relevant sites, the lungs and liver, we found that metastases were highly polyclonal in lungs but not in the liver. Furthermore, the transcriptome of the subclones varied according to their metastatic niche. We also identified a reversible niche-driven signature that was conserved in lung and liver metastases collected during patient autopsies. Among this signature, we found that the tumor necrosis factor-α pathway was up-regulated in lung compared to liver metastases, and inhibition of this pathway affected metastasis diversity. These results highlight that the cellular and molecular heterogeneity observed in metastases is largely dictated by the tumor microenvironment.
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ItemHigher frequency of vertebrate-infecting viruses in the gut of infants born to mothers with type 1 diabetes(WILEY, 2020-02-05)Background: Microbial exposures in utero and early life shape the infant microbiome, which can profoundly impact on health. Compared to the bacterial microbiome, very little is known about the virome. We set out to characterize longitudinal changes in the gut virome of healthy infants born to mothers with or without type 1 diabetes using comprehensive virome capture sequencing. Methods: Healthy infants were selected from Environmental Determinants of Islet Autoimmunity (ENDIA), a prospective cohort of Australian children with a first‐degree relative with type 1 diabetes, followed from pregnancy. Fecal specimens were collected three‐monthly in the first year of life. Results: Among 25 infants (44% born to mothers with type 1 diabetes) at least one virus was detected in 65% (65/100) of samples and 96% (24/25) of infants during the first year of life. In total, 26 genera of viruses were identified and >150 viruses were differentially abundant between the gut of infants with a mother with type 1 diabetes vs without. Positivity for any virus was associated with maternal type 1 diabetes and older infant age. Enterovirus was associated with older infant age and maternal smoking. Conclusions: We demonstrate a distinct gut virome profile in infants of mothers with type 1 diabetes, which may influence health outcomes later in life. Higher prevalence and greater number of viruses observed compared to previous studies suggests significant underrepresentation in existing virome datasets, arising most likely from less sensitive techniques used in data acquisition.
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ItemDetection of cell-free microbial DNA using a contaminant-controlled analysis framework(BMC, 2021-06-23)BACKGROUND: The human microbiome plays an important role in cancer. Accumulating evidence indicates that commensal microbiome-derived DNA may be represented in minute quantities in the cell-free DNA of human blood and could possibly be harnessed as a new cancer biomarker. However, there has been limited use of rigorous experimental controls to account for contamination, which invariably affects low-biomass microbiome studies. RESULTS: We apply a combination of 16S-rRNA-gene sequencing and droplet digital PCR to determine if the specific detection of cell-free microbial DNA (cfmDNA) is possible in metastatic melanoma patients. Compared to matched stool and saliva samples, the absolute concentration of cfmDNA is low but significantly above the levels detected from negative controls. The microbial community of plasma is strongly influenced by laboratory and reagent contaminants introduced during the DNA extraction and sequencing processes. Through the application of an in silico decontamination strategy including the filtering of amplicon sequence variants (ASVs) with batch dependent abundances and those with a higher prevalence in negative controls, we identify known gut commensal bacteria, such as Faecalibacterium, Bacteroides and Ruminococcus, and also other uncharacterised ASVs. We analyse additional plasma samples, highlighting the potential of this framework to identify differences in cfmDNA between healthy and cancer patients. CONCLUSIONS: Together, these observations indicate that plasma can harbour a low yet detectable level of cfmDNA. The results highlight the importance of accounting for contamination and provide an analytical decontamination framework to allow the accurate detection of cfmDNA for future biomarker studies in cancer and other diseases.
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ItemtidyHeatmap: an R package for modular heatmap production based on tidy principles(The Open Journal, 2020-08-03)