- The Walter and Eliza Hall Institute
1G Royal Parade
Parkville, Victoria, Austrlia, 3052
Ashley Ng
University of Melbourne, Medical Biology, Post-Doc
- Dr Ashley Ng is a clinical haematologist with expertise in treating a range of benign and malignant haematological co... moreDr Ashley Ng is a clinical haematologist with expertise in treating a range of benign and malignant haematological conditions and bone marrow transplant.
He graduated first in class with honours for his medical degree (MBBS), and completing a Bachelor of Medical Science (BMedSci) during his undergraduate training. He undertook haematology training while conducting clinical research projects in the areas of thrombosis, infectious diseases, lymphoma, positron-emission tomography scanning and haemato-pathology, and participating in the design of the Australia Lymphoma and Leukemia Group Non-Hodgkin Lymphoma NHL21 clinical trial.
Following completion of dual fellowships in haematology (FRACP) and haemato-pathology (FRCPA Haem), he trained in molecular and cellular biology undertaking a PhD with Professor Warren Alexander and Associate Professor Benjamin Kile at WEHI (2007-2010), elucidating the role of the transcription factor Erg in haematopoietic stem cell function.
His research provided the first definitive evidence that trisomy of Erg was required for the malignant myeloproliferation in Down syndrome and provided a mechanism by which ERG trisomy predisposes to abnormal haematopoiesis in human disease. His laboratory established the importance of genetic co-operativity between increased Erg expression with signaling molecules and epigenetic modifiers in leukaemogenesis of erythro-megakaryocytic leukaemia.
He initiated collaborative projects examining megakaryocytic lineage specification from haemopoietic stem cells, identified thrombopoietin dependent bi-potential erythroid-megakaryocytic progenitors and definitively demonstrated the importance of the Mpl receptor on megakaryocytes in mitigating thrombopoietin signaling on haemopoietic stem and progenitor cells to prevent myeloproliferation, a mechanism which may contribute to the pathogenesis of essential thrombocytosis. This research demonstrated a biological mechanism of action for Eltrombopag in refractory aplastic anaemia.
He has published over 27 peer reviewed primary research articles, 14 over the last 5 years, with contributions in high impact journals including Immunity, Blood, Proceedings of the National Academy of Sciences, Oncogene, Haematologica, plus an invited review for Leukemia and Lymphoma. His work has been cited over 900 times, with an H-index of 11 over the last 5 years.
Four publications have received editorial commentaries, and others cited by the British Committee for Standards in Haematology/UK Myeloma Forum Guidelines (2010) and the Infectious Diseases Working Party (AGIHO) of the German Society of Hematology and Oncology (DGHO) (2013) establishing that his research has impacted directly on patient clinical outcomes.edit - Warren Alexanderedit
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Missense mutations in PLCG2 can cause autoinflammation with phospholipase C gamma 2-associated antibody deficiency and immune dysregulation (APLAID). Here, we generated a mouse model carrying an APLAID mutation (p.Ser707Tyr) and found... more
Missense mutations in PLCG2 can cause autoinflammation with phospholipase C gamma 2-associated antibody deficiency and immune dysregulation (APLAID). Here, we generated a mouse model carrying an APLAID mutation (p.Ser707Tyr) and found that inflammatory infiltrates in the skin and lungs were only partially ameliorated by removing inflammasome function via the deletion of caspase-1. Also, deleting interleukin-6 or tumor necrosis factor did not fully prevent APLAID mutant mice from autoinflammation. Overall, these findings are in accordance with the poor response individuals with APLAID have to treatments that block interleukin-1, JAK1/2 or tumor necrosis factor. Cytokine analysis revealed increased granulocyte colony-stimulating factor (G-CSF) levels as the most distinct feature in mice and individuals with APLAID. Remarkably, treatment with a G-CSF antibody completely reversed established disease in APLAID mice. Furthermore, excessive myelopoiesis was normalized and lymphocyte number...
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The importance of c-MYC in regulating lymphopoiesis and promoting lymphomagenesis is well-established. Far less appreciated is the vital supporting role of MYC’s relative MNT. Using Rag1Cre-mediated Mnt deletion in lymphoid progenitor... more
The importance of c-MYC in regulating lymphopoiesis and promoting lymphomagenesis is well-established. Far less appreciated is the vital supporting role of MYC’s relative MNT. Using Rag1Cre-mediated Mnt deletion in lymphoid progenitor cells, we show here that, during normal T cell development, MNT loss enhances apoptosis, at least in part by elevating expression of the pro-apoptotic BH3-only protein BIM. Moreover, using T lymphoma-prone VavP-MYC transgenic mice, we show that Mnt deletion reduces the pool of pre-malignant MYC-driven T lymphoid cells and abrogates thymic T lymphomagenesis. In addition, we establish that Mnt deletion prevents T lymphoma development in γ-irradiated mice, most likely by enhancing apoptosis of T lymphoid cells repopulating the depleted thymus. Taken together with our recent demonstration that MNT is vital for the survival of MYC-driven pre-malignant and malignant B lymphoid cells, these results suggest that MNT represents an important new drug target for ...
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Genomic markers define molecular subtypes and measurable residual disease (MRD) targets in B-cell acute lymphoblastic leukemia/lymphoma (B-ALL) and are essential determinants of treatment. Current diagnostic approaches typically involve... more
Genomic markers define molecular subtypes and measurable residual disease (MRD) targets in B-cell acute lymphoblastic leukemia/lymphoma (B-ALL) and are essential determinants of treatment. Current diagnostic approaches typically involve serial multi-step testing utilizing conventional cytogenetics (CC)/FISH and molecular genetic (RT-qPCR, MLPA, clonality PCR, NGS panel) techniques which are time and sample consuming and ultimately may not adequately identify genomically complex B-ALL subtypes. In contrast, single-step comprehensive genomic profiling by whole genome and whole transcriptome sequencing (WGS/WTS) may be more efficient for the molecular classification of established and newly described entities which are of increasing therapeutic relevance. We have instituted a multimodal platform for molecular testing in B-ALL performing WGS/WTS in parallel with deep NGS-based immunoglobulin (IG) rearrangement MRD and exploratory DNA-breakpoint based MRD assays. We aimed to determine th...
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We describe one male and one female patient who each developed childhood/adolescent obsessive‐compulsive disorder as a prelude to the development of a typical picture of chorea‐acanthocytosis (ChAc). In each patient, the caudate nucleus... more
We describe one male and one female patient who each developed childhood/adolescent obsessive‐compulsive disorder as a prelude to the development of a typical picture of chorea‐acanthocytosis (ChAc). In each patient, the caudate nucleus showed dramatic atrophy. The role of the caudate in compulsive phenomena, and the predilection for neurological disorders with onset in adolescence to present as major mental illness, is discussed. On the basis of the current evidence and previous findings, we suggest that ChAc can be understood as a disorder whose clinical presentation reflects an interaction between the disease process and the individual's neurodevelopmental stage with both initial interrupted neurodevelopment, and supervening neurodegeneration. © 2007 Movement Disorder Society
Research Interests: Neuroscience, Cognitive Science, Psychiatry, Neurodevelopment, Medicine, and 15 moreMovement disorders, Humans, Caudate Nucleus, Movement, Female, Male, Obsessive Compulsive Disorder, Mental Disorders, Clinical Sciences, Atrophy, Adult, Disease Progression, Chorea, Acanthocytosis, and Obsessive Compulsive
CRISPR technologies have advanced cancer modelling in mice, but CRISPR activation (CRISPRa) methods have not been exploited in this context. We establish a CRISPRa mouse (dCas9a-SAMKI) for inducing gene expression in vivo and in vitro.... more
CRISPR technologies have advanced cancer modelling in mice, but CRISPR activation (CRISPRa) methods have not been exploited in this context. We establish a CRISPRa mouse (dCas9a-SAMKI) for inducing gene expression in vivo and in vitro. Using dCas9a-SAMKI primary lymphocytes, we induce B cell restricted genes in T cells and vice versa, demonstrating the power of this system. There are limited models of aggressive double hit lymphoma. Therefore, we transactivate pro-survival BCL-2 in Eµ-MycT/+;dCas9a-SAMKI/+ haematopoietic stem and progenitor cells. Mice transplanted with these cells rapidly develop lymphomas expressing high BCL-2 and MYC. Unlike standard Eµ-Myc lymphomas, BCL-2 expressing lymphomas are highly sensitive to the BCL-2 inhibitor venetoclax. We perform genome-wide activation screens in these lymphoma cells and find a dominant role for the BCL-2 protein A1 in venetoclax resistance. Here we show the potential of our CRISPRa model for mimicking disease and providing insights...
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The importance of c-MYC in regulating normal lymphopoiesis and promoting lymphomagenesis is well-established, but far less appreciated is the vital supporting role of MYC’s relative MNT. We show here that, during normal T cell... more
The importance of c-MYC in regulating normal lymphopoiesis and promoting lymphomagenesis is well-established, but far less appreciated is the vital supporting role of MYC’s relative MNT. We show here that, during normal T cell development, MNT loss enhances apoptosis and establish the mechanism as elevated expression of the pro-apoptotic BH3-only protein BIM. Using a MYC transgenic mouse model, we also show that MNT loss reduces the pool of premalignant MYC-driven T cells and totally abrogates thymic T lymphomagenesis. Taken together with our recent demonstration that MNT is vital for the survival of MYC-driven premalignant and malignant B lymphoid cells, these results suggest that MNT represents an important new drug target for both T and B lymphoid malignancies.
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Navitoclax (ABT-263), an inhibitor of the pro-survival BCL-2 family proteins BCL-2, BCL-XL and BCL-W, has shown clinical efficacy in certain BCL-2-dependent haematological cancers, but causes dose-limiting thrombocytopaenia. The latter... more
Navitoclax (ABT-263), an inhibitor of the pro-survival BCL-2 family proteins BCL-2, BCL-XL and BCL-W, has shown clinical efficacy in certain BCL-2-dependent haematological cancers, but causes dose-limiting thrombocytopaenia. The latter effect is caused by Navitoclax directly inducing the apoptotic death of platelets, which are dependent on BCL-XL for survival. Recently, ABT-199, a selective BCL-2 antagonist, was developed. It has shown promising anti-leukaemia activity in patients whilst sparing platelets, suggesting that the megakaryocyte lineage does not require BCL-2. In order to elucidate the role of BCL-2 in megakaryocyte and platelet survival, we generated mice with a lineage-specific deletion of Bcl2, alone or in combination with loss of Mcl1 or Bclx. Platelet production and platelet survival were analysed. Additionally, we made use of BH3 mimetics that selectively inhibit BCL-2 or BCL-XL. We show that the deletion of BCL-2, on its own or in concert with MCL-1, does not affec...
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Many cytokines exert their biological effect through members of the hemopoietin receptor family. Using degenerate oligonucleotides to the common WSXWS motif, we have cloned from human hemopoietic cell cDNA libraries various forms of the... more
Many cytokines exert their biological effect through members of the hemopoietin receptor family. Using degenerate oligonucleotides to the common WSXWS motif, we have cloned from human hemopoietic cell cDNA libraries various forms of the receptor that was recently shown to bind the obesity hormone, leptin. mRNAs encoding long and short forms of the human leptin receptor were found to be coexpressed in a range of human and murine hemopoietic organs, and a subset of cells from these tissues bound leptin at the cell surface. Ectopic expression in murine Ba/F3 and M1 cell lines revealed that the long, but not the short, form of the leptin receptor can signal proliferation and differentiation, respectively. In cultures of murine or human marrow cells, human leptin exhibited no capacity to stimulate cell survival or proliferation, but it enhanced cytokine production and phagocytosis of Leishmania parasites by murine peritoneal macrophages. Our data provide evidence that, in addition to its...
Research Interests: Biology, Cell Biology, Medicine, Gene expression, Cell Division, and 15 moreHematopoietic Stem Cells, Cell line, Cell Differentiation, Humans, Mice, Animals, Leptin, Haematopoiesis, Degeneration, Leptin Receptor, Amino Acid Sequence, Cell Surface Markers, Cell Survival, cDNA library, and biological effect
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Significance Blood platelets, the small circulating cells that coordinate hemostasis, are produced by specialized bone marrow cells called megakaryocytes. The cytokine thrombopoietin (TPO) is a key regulator of platelet production acting... more
Significance Blood platelets, the small circulating cells that coordinate hemostasis, are produced by specialized bone marrow cells called megakaryocytes. The cytokine thrombopoietin (TPO) is a key regulator of platelet production acting via its specific cell receptor, Mpl. Via genetic modification of the Mpl allele in mice, we precisely define the bone marrow cells that express Mpl and, by genetically removing Mpl from megakaryocytes and platelets, we show TPO signaling via Mpl is not required in megakaryocytes for their expansion, maturation, or platelet production. Rather, Mpl expression on megakaryocytes is essential for regulating TPO availability in the bone marrow microenvironment to prevent myeloproliferation, a model we suggest is important for human disease.
Research Interests: Biology, Cell Biology, Medicine, Multidisciplinary, Platelet, and 14 moreHematopoietic Stem Cells, Mice, Animals, Megakaryocytes, Bone marrow, Gene Targeting, Thrombopoietin, Cell Proliferation, Myeloid Cells, Gene Expression Regulation, Gene expression profiling, Blood platelets, Thrombocytosis, and Thrombopoiesis
Two distinct bone marrow-derived blast colony-forming cells can generate colonies of lineage-restricted progenitor cells in agar cultures of murine bone marrow. Both cell types selectively had a Kit + ScaI + phenotype distinguishing them... more
Two distinct bone marrow-derived blast colony-forming cells can generate colonies of lineage-restricted progenitor cells in agar cultures of murine bone marrow. Both cell types selectively had a Kit + ScaI + phenotype distinguishing them from most lineage-restricted progenitor cells. Multicentric blast colony-forming cells stimulated by stem cell factor plus interleukin-6 (IL-6) (BL-CFC-S) were separable from most dispersed blast colony-forming cells stimulated by Flt3 ligand and IL-6 (BL-CFC-F) using CD34 and Flt3R probes. Multicentric BL-CFC-S cofractionated with colony-forming units, spleen (CFU-S) supporting the possibility that the 2 cells may be identical. The colony populations generated by BL-CFC-S were similar in their phenotype and proliferative capacity to progenitor cells in whole bone marrow but the progeny of BL-CFC-F were skewed with an abnormally high proportion of Kit − Flt3R + cells whose clonogenic cells tended to generate only macrophage progeny. Both blast colon...
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Hematopoietic progenitor cells are the progeny of hematopoietic stem cells that coordinate the production of precise numbers of mature blood cells of diverse functional lineages. Identification of cell-surface antigen expression... more
Hematopoietic progenitor cells are the progeny of hematopoietic stem cells that coordinate the production of precise numbers of mature blood cells of diverse functional lineages. Identification of cell-surface antigen expression associated with hematopoietic lineage restriction has allowed prospective isolation of progenitor cells with defined hematopoietic potential. To clarify further the cellular origins of megakaryocyte commitment, we assessed the in vitro and in vivo megakaryocyte and platelet potential of defined progenitor populations in the adult mouse bone marrow. We show that megakaryocytes arise from CD150 + bipotential progenitors that display both platelet- and erythrocyte-producing potential in vivo and that can develop from the Flt3 − fraction of the pregranulocyte-macrophage population. We define a bipotential erythroid-megakaryocyte progenitor population, the CD150 + CD9 lo endoglin lo fraction of Lin − cKit + IL7 receptor alpha − FcγRII/III lo Sca1 − cells, which c...
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Murine hematopoietic blast colony-forming cells (BL-CFCs) are able to generate up to 30,000 progeny blast cells within 10 d in agar cultures. Contained in these populations are large numbers of lineage-committed progenitor cells in the... more
Murine hematopoietic blast colony-forming cells (BL-CFCs) are able to generate up to 30,000 progeny blast cells within 10 d in agar cultures. Contained in these populations are large numbers of lineage-committed progenitor cells in the granulocytic and macrophage lineages. Sequential analyses of blast colonies revealed that self-generation of BL-CFCs occurs but is surprisingly late in clonal expansion, as is the emergence of progenitor cells committed to megakaryocytic and eosinophil lineages. Self-generating BL-CFCs were highly enriched in lineage − Kit + Sca1 + CD34 − Flt3R − populations, and colonies generated by such cells contained colony-forming units–spleen and formed erythroid and lymphoid progeny in vivo. The data suggest the existence of a hierarchical structure in BL-CFC populations with at least a subset being cells assayable as colony-forming units–spleen. Because BL-CFCs can self-generate and are able to generate lymphoid and myeloid populations, BL-CFCs appear to be i...
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GFI1B is a transcription factor important for erythropoiesis and megakaryocyte development but previously unknown to be associated with human disease. A family with a novel bleeding disorder was identified and characterized. Genetic... more
GFI1B is a transcription factor important for erythropoiesis and megakaryocyte development but previously unknown to be associated with human disease. A family with a novel bleeding disorder was identified and characterized. Genetic linkage analysis and massively parallel sequencing were used to localize the mutation causing the disease phenotype on chromosome 9. Functional studies were then performed in megakaryocytic cell lines to determine the biological effects of the mutant transcript. We have identified a family with an autosomal dominant bleeding disorder associated with macrothrombocytopenia, red cell anisopoikilocytosis, and platelet dysfunction. The severity of bleeding is variable with some affected individuals experiencing spontaneous bleeding while other family members exhibit only abnormal bleeding with surgery. A single nucleotide insertion was identified in GFI1B that predicts a frameshift mutation in the fifth zinc finger DNA-binding domain. This mutation alters the transcriptional activity of the protein, resulting in a reduction in platelet α-granule content and aberrant expression of key platelet proteins. GFI1B mutation represents a novel human bleeding disorder, and the described phenotype identifies GFI1B as a critical regulator of platelet shape, number, and function.
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Research Interests: Genetics, Stem Cells, Biology, Transcription Factors, Medicine, and 15 moreDown Syndrome, Hematopoiesis, Transcriptome, Humans, Mice, Animals, Megakaryocytes, Trisomy 21, Genotype, Microarray Analysis, PLoS Genetics, Cell Proliferation, Gene Expression Regulation, Gene expression profiling, and alleles
Research Interests: Immunology and Biology
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Accurate quantification of the plasmacell burden in bone-marrow is essential for diagnosis and assessment of therapeutic response in plasma-cell dyscrasias. 2 It has particular importance in defining a prognostically favorable group of... more
Accurate quantification of the plasmacell burden in bone-marrow is essential for diagnosis and assessment of therapeutic response in plasma-cell dyscrasias. 2 It has particular importance in defining a prognostically favorable group of patients with multiple myeloma who obtain a complete response following high-dose myeloablative therapy. Most laboratories assess the extent of plasma cell infiltration by morphological examination of Romanowsky-stained bonemarrow aspirate samples and hematoxylin and eosin (H&E)-stained trephine sections. Immunohistological and flow cytometric techniques provide additional diagnostic and prognostic information, although their value in plasma-cell quantification has not been systematically assessed against other methods. We compared four methodologies for plasmacell quantification on bone marrow samples in patients with monoclonal gammopathies of undetermined significance (MGUS) or multiple myeloma to determine i) the degree of correlation between the ...
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Elevated platelet count is associated with poor survival in certain solid cancers, including lung cancer. In addition, experimental transplantation of cancer cell lines has uncovered a role for platelets in blood-borne metastasis. These... more
Elevated platelet count is associated with poor survival in certain solid cancers, including lung cancer. In addition, experimental transplantation of cancer cell lines has uncovered a role for platelets in blood-borne metastasis. These studies, however, do not account for heterogeneity between lung cancer subtypes. Subsequently, the role of platelets in the major subtypes of non-small cell lung cancer (adenocarcinoma (ADC) and squamous cell carcinoma (SqCC)) is not fully understood. We utilised an autochthonous KrasLSL-G12D/+;p53flox/flox mouse model of lung ADC together with genetic models of thrombocytopenia to interrogate the role of platelets in lung cancer growth and progression. While thrombocytopenia failed to impact primary tumour growth, in experimental metastatic models however, thrombocytopenic mice displayed significantly extended survival. Utilising a novel thrombocytopenic immunocompromised mouse, the importance of platelets in metastatic dissemination was confirmed with human KRAS-mutant ADC cell lines. Finally, retrospective analysis of a NSCLC patient cohort revealed thrombocytosis was predictive of poor survival in ADC patients with metastatic disease. Interestingly, this association was not apparent in SqCC patients. Overall, these data highlight the possibility of patient stratification using thrombocytosis as a biomarker, and indicates opportunities for potential novel treatment strategies that combine anti-platelet and lung cancer therapies.
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We hypothesize that thrombosis with thrombocytopenia syndrome recently described after administration of adenovirus-vectored vaccines for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) occurs as a result of the unique... more
We hypothesize that thrombosis with thrombocytopenia syndrome recently described after administration of adenovirus-vectored vaccines for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) occurs as a result of the unique properties of the adenovirus vectors, which can have widespread biodistribution throughout the body. The antigen is delivered to megakaryocyte cells, which act as part of the primary immune system and distribute the antigen within progeny platelets, also a key component of the immune system. The interaction of the antigen induces preformed antiplatelet factor 4 (PF4) antibodies to bind to PF4-heparan sulfate complexes in the absence of exogenous heparin, at sites where the heparan sulfate concentration in the vascular glycocalyx is optimal for complex formation, causing thrombosis and thrombocytopenia as observed clinically. This hypothesis is testable in cell culture and animal models, and potentially in vivo, and if proven correct has significant implications for vaccine development and our understanding of the links between the coagulation and immune systems.
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Hairy cell leukaemia (HCL) is a rare CD20+ B cell malignancy characterised by rare “hairy” B cells and extensive bone marrow (BM) infiltration. Frontline treatment with the purine analogue cladribine (CDA) results in a highly variable... more
Hairy cell leukaemia (HCL) is a rare CD20+ B cell malignancy characterised by rare “hairy” B cells and extensive bone marrow (BM) infiltration. Frontline treatment with the purine analogue cladribine (CDA) results in a highly variable response duration. We hypothesised that analysis of the BM tumour microenvironment would identify prognostic biomarkers of response to CDA. HCL BM immunology pre and post CDA treatment and healthy controls were analysed using Digital Spatial Profiling to assess the expression of 57 proteins using an immunology panel. A bioinformatics pipeline was developed to accommodate the more complex experimental design of a spatially resolved study. Treatment with CDA was associated with the reduction in expression of HCL tumour markers (CD20, CD11c) and increased expression of myeloid markers (CD14, CD68, CD66b, ARG1). Expression of HLA-DR, STING, CTLA4, VISTA, OX40L were dysregulated pre- and post-CDA. Duration of response to treatment was associated with greate...
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Regulation of haematopoietic stem and progenitor cell (HSPC) fate is crucial during homeostasis and under stress conditions. Here we examine the aetiology of the Flt3 ligand (Flt3L)-mediated increase of type 1 conventional dendritic cells... more
Regulation of haematopoietic stem and progenitor cell (HSPC) fate is crucial during homeostasis and under stress conditions. Here we examine the aetiology of the Flt3 ligand (Flt3L)-mediated increase of type 1 conventional dendritic cells (cDC1s). Using cellular barcoding we demonstrate this occurs through selective clonal expansion of HSPCs that are primed to produce cDC1s and not through activation of cDC1 fate by other HSPCs. In particular, multi/oligo-potent clones selectively amplify their cDC1 output, without compromising the production of other lineages, via a process we term tuning. We then develop Divi-Seq to simultaneously profile the division history, surface phenotype and transcriptome of individual HSPCs. We discover that Flt3L-responsive HSPCs maintain a proliferative 'early progenitor'-like state, leading to the selective expansion of multiple transitional cDC1-primed progenitor stages that are marked by Irf8 expression. These findings define the mechanistic action of Flt3L through clonal tuning, which has important implications for other models of 'emergency' haematopoiesis.
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There is little information regarding CMV reactivation in the non-allogeneic transplantation setting. We report the incidence of CMV-DNAemia and CMV-associated disease in a cohort of patients treated for haematological malignancies at the... more
There is little information regarding CMV reactivation in the non-allogeneic transplantation setting. We report the incidence of CMV-DNAemia and CMV-associated disease in a cohort of patients treated for haematological malignancies at the Peter MacCallum Cancer Center from 6/99-6/04. Among the 36 patients identified, potential predictive factors were analysed, including underlying malignancy, previous therapy and therapy immediately preceding CMV-DNAemia/disease. Patients received a median of 2 prior therapies (range 1–9), age ranged from 35–77 years, with 19 males. Diseases were: acute leukaemia(6), CLL(6), NHL(21), HD(1), myeloma(2). Therapies immediately preceding CMV-DNAemia/disease were: “conventional chemotherapy” (39%, including HyperCVAD-14%), fludarabine-based (19%), autologous BMT (22%), and antibody therapy (20%, including denileukin diftitox, 6%, alemtuzumab, 14%). Risk of CMV-DNAemia/disease was calculated from the total number of patients treated over the period of analysis: alemtuzumab, (6/12; 50%), denileukin (2/33; 6%), autologous BMT (9/223; 4%), fludarabine-based (8/177; 4.5%), HyperCVAD (5/77; 6.5%). 94% were symptomatic on investigation (fever 92%, CXR changes 25%, abnormal LFT’s 3%, GI symptoms 8%, rash 6%). 7 had CMV-disease (3 pneumonitis, 2 oesophagitis, 2 enteritis/colitis). 11 had isolated fever. 14/19 treated with ganciclovir had possible/proven CMV-related illness; in 12, fever, CMV-DNAemia or symptoms resolved. 2 died from underlying disease or treatment complications. 3 asymptomatic patients with CMV-DNAemia on >1 occasion were observed until DNA negativity. 17/19 with CMV-DNAemia on only 1 occasion did not develop CMV-related illness. We recommend that patients receiving alemtuzumab, should have CMV surveillance to facilitate early therapy, though the role of ongoing surveillance/prophylaxis is unknown. Patients with persistent/rising CMV-DNA titres or evidence of CMV-disease, should be treated.
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Research Interests: Stem Cells and Biology
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Improving survival outcomes in adult B-cell acute lymphoblastic leukemia (B-ALL) remains a clinical challenge. Relapsed disease has a poor prognosis despite the use of tyrosine kinase inhibitors (TKIs) for Philadelphia chromosome positive... more
Improving survival outcomes in adult B-cell acute lymphoblastic leukemia (B-ALL) remains a clinical challenge. Relapsed disease has a poor prognosis despite the use of tyrosine kinase inhibitors (TKIs) for Philadelphia chromosome positive (Ph+ ALL) cases and immunotherapeutic approaches, including blinatumomab and chimeric antigen receptor T cells. Targeting aberrant cell survival pathways with selective small molecule BH3-mimetic inhibitors of BCL-2 (venetoclax, S55746), BCL-XL (A1331852), or MCL1 (S63845) is an emerging therapeutic option. We report that combined targeting of BCL-2 and MCL1 is synergistic in B-ALL in vitro. The combination demonstrated greater efficacy than standard chemotherapeutics and TKIs in primary samples from adult B-ALL with Ph+ ALL, Ph-like ALL, and other B-ALL. Moreover, combined BCL-2 or MCL1 inhibition with dasatinib showed potent killing in primary Ph+ B-ALL cases, but the BH3-mimetic combination appeared superior in vitro in a variety of Ph-like ALL ...
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SummaryExtrinsic regulation of single haematopoietic stem and progenitor cell (HSPC) fate is crucial for immune cell development. Here, we examine the aetiology of Flt3 ligand (Flt3L)-mediated emergency development of type 1 conventional... more
SummaryExtrinsic regulation of single haematopoietic stem and progenitor cell (HSPC) fate is crucial for immune cell development. Here, we examine the aetiology of Flt3 ligand (Flt3L)-mediated emergency development of type 1 conventional dendritic cells (cDC1s), which results in enhanced immunity against infections and cancer. Using cellular barcoding, we demonstrate a predominant role of enhanced clonal expansion and moderate contribution via recruitment of additional cDC1-generating HSPCs. The selective cDC1 expansion occurs primarily via multi-/oligo-potent clones, without compromising output to other lineages. To understand the molecular hallmarks early during a Flt3L response, we develop Divi-Seq to simultaneously profile cell division history, surface phenotype and transcriptional state of single HSPCs. We discover that Flt3L-responsive HSPCs maintain a proliferative ‘early progenitor’-like state, which leads to selective emergence of CD11c+cKit+ transitional precursors with h...
Introduction Thrombotic Thrombocytopenic Purpura (TTP) is a cause of microangiopathic-haemolytic anaemia and thrombocytopenia, associated with renal and neurological dysfunction with thrombotic complications causing significant morbidity... more
Introduction Thrombotic Thrombocytopenic Purpura (TTP) is a cause of microangiopathic-haemolytic anaemia and thrombocytopenia, associated with renal and neurological dysfunction with thrombotic complications causing significant morbidity and mortality. Methods A restrospective single-institution analysis of patients with TTP treated between 1990–2005. Renal or bone marrow transplantation patients were excluded. Results Forty patients were identified. Aetiology was idiopathic 75% (n=30), connective tissue disease-related 12.5% (n=5), malignancy-related 5% (n=2) and pregnancy-related 7.5% (n=3). Presenting features: neurological 62.5% (n=25), renal impairment (creatinine>0.11 mmol/L) 76% (n=28), microangiopathic-haemolytic anaemia 97.5% (n=39) and thrombocytopenia 100% (mean platelet-count 42x10^9/L). Mean Hb 93 g/L and mean Lactose dehydrogenase (LD) 2517 U/L (<420). 38 patients received up-front single plasma-volume plasma-exchange using fresh-frozen plasma (median 11 exchange...
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Deregulated over-expression of MYC is implicated in the development and malignant progression of most (~70%) human tumors. MYC drives cell growth and proliferation but also, at high levels, promotes apoptosis. Here, we report that the... more
Deregulated over-expression of MYC is implicated in the development and malignant progression of most (~70%) human tumors. MYC drives cell growth and proliferation but also, at high levels, promotes apoptosis. Here, we report that the proliferative capacity of MYC-driven normal and neoplastic B lymphoid cells depends on MNT, a MYC-related transcriptional repressor. Our genetic data establish that MNT synergises with MYC by suppressing MYC-driven apoptosis and that it does so primarily by reducing the level of pro-apoptotic BIM. In Em-Myc mice, which model the MYC/IGH chromosome translocation in Burkitt's lymphoma, homozygous Mnt deletion greatly reduced lymphoma incidence by enhancing apoptosis and markedly decreasing premalignant B lymphoid cell populations. Strikingly, by inducing Mnt deletion within transplanted fully-malignant Em-Myc lymphoma cells, we significantly extended transplant recipient survival. The dependency of lymphomas on MNT for survival suggests that drugs in...
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Summary/B-cell development is initiated by the stepwise differentiation of hematopoietic stem cells into lineage committed progenitors, ultimately generating the mature B-cells that mediate protective immunity. This highly regulated... more
Summary/B-cell development is initiated by the stepwise differentiation of hematopoietic stem cells into lineage committed progenitors, ultimately generating the mature B-cells that mediate protective immunity. This highly regulated process also generates clonal immunological diversity via recombination of immunoglobulin genes. While several transcription factors that control B-cell development and V(D)J recombination have been defined, how these processes are initiated and coordinated into a precise regulatory network remains poorly understood. Here, we show that the transcription factor ETS Related Gene (Erg) is essential for the earliest steps in B-cell differentiation. Erg initiates a transcriptional network involving the B-cell lineage defining genes, Ebf1 and Pax5, that directly promotes the expression of key genes involved in V(D)J recombination and formation of the B-cell receptor. Complementation of the Erg-deficiency with a productively rearranged immunoglobulin gene rescu...
Background Precursor-B acute lymphoblastic leukemia (B-ALL) is an aggressive hematological malignancy. Relapsed disease has a poor prognosis, despite improved outcomes with tyrosine kinase inhibitors for Ph+ cases and immunotherapeutic... more
Background Precursor-B acute lymphoblastic leukemia (B-ALL) is an aggressive hematological malignancy. Relapsed disease has a poor prognosis, despite improved outcomes with tyrosine kinase inhibitors for Ph+ cases and immunotherapeutic approaches, such as blinotumomab and CAR-T cells. Targeting cell survival with novel small molecule BH3-mimetic inhibitors of BCL-2 (e.g. Souers et al Nat Med 2013, Roberts et al, NEJM 2016 and Casara et al, Oncotarget 2018), BCL-XL (Lessene et al, Nat Chem Biol, 2013) or MCL1 (Kotschy et al, Nature 2016) is an emerging therapeutic option. BCL-2 is reported to have a pro-survival role in BCR-ABL1, JAK2 fusion, ETV6-RUNX1 and MLL-r driven ALL (Brown et al., Journal Biological Chemistry 2017). BH3-mimetics targeting BCL-2 and BCL-XL has efficacy in paediatric ALL xenografts (Khaw et al., Blood 2016), while ruxolitinib combined with ABT-737 is synergistic in JAK2-mutant pre-B-ALL (Waibel et al., Cell Reports 2013). We now report that combined targeting o...
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GFI1B is a transcription factor important for erythropoiesis and megakaryocyte development, but previously unknown to be associated with human disease. We have identified a family with an autosomal dominant bleeding disorder associated... more
GFI1B is a transcription factor important for erythropoiesis and megakaryocyte development, but previously unknown to be associated with human disease. We have identified a family with an autosomal dominant bleeding disorder associated with thrombocytopenia, red cell anisopoikilocytosis and platelet dysfunction. The severity of bleeding is variable with some affected individuals experiencing spontaneous bleeding while other family members only exhibit abnormal bleeding with surgery. All affected individuals had a moderate thrombocytopenia ranging from 72-149 x109/L with an elevated mean platelet volume. Platelet function was perturbed with prolonged collagen/epinephrine closure times on the PFA-100 (294 vs 125 sec, P <0.001). Platelet aggregation studies were abnormal, with an absent collagen response and primary aggregation only with ADP, arachidonic acid and adrenaline. Genetic linkage analysis and massively parallel sequencing were performed on 16 family members to localize th...
Research Interests:
Research Interests:
RNA-seq datasets can contain millions of intron reads per sequenced library that are typically removed from downstream analysis. Only reads overlapping annotated exons are considered to be informative since mature mRNA is assumed to be... more
RNA-seq datasets can contain millions of intron reads per sequenced library that are typically removed from downstream analysis. Only reads overlapping annotated exons are considered to be informative since mature mRNA is assumed to be the major component sequenced, especially when examining poly(A) RNA samples. By examining multiple datasets, we demonstrate that intron reads are informative and that signal is shared between exon and intron counts. The majority of expressed genes contain reads in both exon and intron regions, where exon and intron log-counts are positively correlated. On a per gene basis intron counts are larger in Total RNA libraries than the same biological sample sequenced under a poly(A) RNA protocol. This is due to 3’ coverage bias in poly(A) RNA libraries affecting medium to long intron regions, where a considerable drop in read coverage for introns residing at the 5’ end of genes is observed. Looking across thousands of genes simultaneously we characterise co...
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Research Interests:
The hematopoietically expressed homeobox (Hhex) transcription factor is overexpressed in human myeloid leukemias. Conditional knockout models of murine acute myeloid leukemia indicate that maintains leukemia stem cell self-renewal by... more
The hematopoietically expressed homeobox (Hhex) transcription factor is overexpressed in human myeloid leukemias. Conditional knockout models of murine acute myeloid leukemia indicate that maintains leukemia stem cell self-renewal by enabling Polycomb-mediated epigenetic repression of the Cdkn2a tumor suppressor locus, encoding p16 and p19 However, whether overexpression also affects hematopoietic differentiation is unknown. To study this, we retrovirally overexpressed in hematopoietic progenitors. This enabled serial replating of myeloid progenitors, leading to the rapid establishment of interleukin-3 (IL-3)-dependent promyelocytic cell lines. Use of a Hhex-ERT2 fusion protein demonstrated that continuous nuclear Hhex is required for transformation, and structure function analysis demonstrated a requirement of the DNA-binding and N-terminal-repressive domains of Hhex for promyelocytic transformation. This included the N-terminal promyelocytic leukemia protein (Pml) interaction doma...
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Activating JAK2 point mutations are implicated in the pathogenesis of myeloid and lymphoid malignancies, including high-risk B-cell acute lymphoblastic leukemia (B-ALL). In preclinical studies, treatment of JAK2 mutant leukemias with type... more
Activating JAK2 point mutations are implicated in the pathogenesis of myeloid and lymphoid malignancies, including high-risk B-cell acute lymphoblastic leukemia (B-ALL). In preclinical studies, treatment of JAK2 mutant leukemias with type I JAK2 inhibitors (e.g., Food and Drug Administration [FDA]-approved ruxolitinib) provided limited single-agent responses, possibly due to paradoxical JAK2 hyperphosphorylation induced by these agents. To determine the importance of mutant JAK2 in B-ALL initiation and maintenance, we developed unique genetically engineered mouse models of B-ALL driven by overexpressed Crlf2 and mutant Jak2, recapitulating the genetic aberrations found in human B-ALL. While expression of mutant Jak2 was necessary for leukemia induction, neither its continued expression nor enzymatic activity was required to maintain leukemia survival and rapid proliferation. CRLF2/JAK2 mutant B-ALLs with sustained depletion or pharmacological inhibition of JAK2 exhibited enhanced ex...
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RIPK1 is an essential downstream component of many pattern recognition and death receptors. RIPK1 can promote the activation of caspase-8 induced apoptosis and RIPK3-MLKL-mediated necroptosis, however, during development RIPK1 limits both... more
RIPK1 is an essential downstream component of many pattern recognition and death receptors. RIPK1 can promote the activation of caspase-8 induced apoptosis and RIPK3-MLKL-mediated necroptosis, however, during development RIPK1 limits both forms of cell death. Accordingly, Ripk1 mice present with systemic cell death and consequent multi-organ inflammation, which is driven through the activation of both FADD-caspase-8 and RIPK3-MLKL signaling pathways causing perinatal lethality. TRADD is a death domain (DD) containing molecule that mediates signaling downstream of TNFR1 and the TLRs. Following the disassembly of the upstream receptor complexes either RIPK1 or TRADD can form a complex with FADD-caspase-8-cFLIP, via DD-DD interactions with FADD, facilitating the activation of caspase-8. We show that genetic deletion of Ripk1 licenses TRADD to complex with FADD-caspase-8 and activates caspase-8 during development. Deletion of Tradd provided no survival advantage to Ripk1 animals and yet...