ORCID Profile
0000-0002-8792-3021
Current Organisation
University of South Australia
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Publisher: Future Science Ltd
Date: 07-2011
DOI: 10.2144/000113702
Abstract: Neutral lipid droplets (LDs) are dynamic lipid storage organelles found in all eukaryotic cells from yeast to mammals and higher plants. LDs are important to many physiological processes that include basic cellular maintenance, metabolism, and erse medical pathologies. LD accumulation has been studied extensively by a range of methods, but particularly by microscopy with several fluorescent dyes extensively used for qualitative and quantitative imaging. Here, we compared established LD stains Nile Red and BODIPY 493/503 to the 4′, 6-diamidino-2-phenylindole (DAPI)—range dye 1,6-diphenyl-1,3,5-hexatriene (DPH excitation/emission λ max =350 nm/420 nm) using high-content image analysis. HeLa cells treated with oleic acid or vehicle were used to compare staining patterns between DPH and Nile Red as well as DPH and the LD protein adipophilin. DPH, Nile Red, and BODIPY 493/503 were compared as assay reagents in oleic acid dose-response experiments. Treatment of MCF-7 cells with sodium butyrate was used as a second cellular system for high-content analysis of LD formation. In this experimental context, we demonstrate the compatibility of DPH with GFP, a technical limitation of Nile Red and BODIPY 493/503 dyes. These data show that DPH has comparable sensitivity and specificity to that of Nile Red. Z′-factor analysis of dose-response experiments indicated that DPH and BODIPY 493/503 are well suited for quantitative analysis of LDs for high-throughput screening (HTS) applications.
Publisher: Wiley
Date: 18-04-2017
DOI: 10.1002/JCB.25851
Abstract: The importance of Wnt pathway signaling in development of bone has been well established. Here we investigated the role of a known Wnt target, ENC1 (ectodermal-neural cortex 1 NRP/B), in osteoblast differentiation. Enc1 expression was detected in mouse osteoblasts, chondrocytes, and osteocytes by in situ hybridization, and osteoblastic expression was verified in differentiating primary cultures and MC3T3-E1 pre-osteoblast cells, with 57 kDa and 67 kDa ENC1 protein isoforms detected throughout differentiation. Induced knockdown of both ENC1 isoforms reduced alkaline phosphatase staining and virtually abolished MC3T3-E1 mineralization. At culture confluence, Alpl (alkaline phosphatase liver/bone/kidney) expression was markedly reduced compared with control cells, and there was significant and coordinated alteration of other genes involved in cellular phosphate biochemistry. In contrast, with 67 kDa-selective knockdown mineralized nodule formation was enhanced and there was a two-fold increase in Alpl expression at confluence. There was enhanced expression of Wnt/β-catenin target genes with knockdown of both isoforms at this time-point and a five-fold increase in Frzb (Frizzled related protein) with 67 kDa-selective knockdown at mineralization, indicating possible ENC1 interactions with Wnt signaling in osteoblasts. These results are the first to demonstrate a role for ENC1 in the control of osteoblast differentiation. Additionally, the contrasting mineralization phenotypes and transcriptional patterns seen with coordinate knockdown of both ENC1 isoforms vs selective knockdown of 67 kDa ENC1 suggest opposing roles for the isoforms in regulation of osteoblastic differentiation, through effects on Alpl expression and phosphate cellular biochemistry. This study is the first to report differential roles for the ENC1 isoforms in any cell lineage. J. Cell. Biochem. 118: 2141-2150, 2017. © 2016 Wiley Periodicals, Inc.
Publisher: Elsevier BV
Date: 02-2014
DOI: 10.1016/J.CANLET.2013.09.022
Abstract: The effect of combining MYB suppression with the histone deacetylase inhibitor LBH589 was studied in human myeloid leukemia cell lines. MYB knockdown inhibited proliferation and induced apoptosis in U937 and K562 cells in vitro, and also sensitized both to the pro-apoptotic effect of LBH589. This was accompanied by enhanced expression of the pro-apoptotic BCL2 family members BOK and BIM. U937 cells carrying inducible MYB shRNA were also transplanted into NOD/SCID mice. The combination of MYB knockdown and LBH589 prolonged survival compared to either treatment alone, suggesting that further development of such combinations might lead to effective and safe leukemia therapies.
Publisher: Springer Science and Business Media LLC
Date: 27-11-2013
DOI: 10.1186/BCR3580
Publisher: Springer Science and Business Media LLC
Date: 07-2008
DOI: 10.1038/NRC2439
Abstract: The transcription factor MYB has a key role as a regulator of stem and progenitor cells in the bone marrow, colonic crypts and a neurogenic region of the adult brain. It is in these compartments that a deficit in MYB activity leads to severe or lethal phenotypes. As was predicted from its leukaemogenicity in several animal species, MYB has now been identified as an oncogene that is involved in some human leukaemias. Moreover, recent evidence has strengthened the case that MYB is activated in colon and breast cancer: a block to MYB expression is overcome by mutation of the regulatory machinery in the former disease and by oestrogen receptor-alpha (ERalpha) in the latter.
Publisher: Springer Science and Business Media LLC
Date: 09-08-2012
DOI: 10.1038/LEU.2012.225
Abstract: The Myb protein was first identified as an oncogene that causes leukemia in chickens. Since then, it has been widely associated with different types of cancers and studied in detail in myeloid leukemias. However, despite these studies, its role in the induction, pathogenesis and maintenance of AML, and other blood disorders, is still not well understood. Recent efforts to uncover its plethora of transcriptional targets have provided key insights into understanding its mechanism of action. This review evaluates our current knowledge of the role of Myb in leukemia, with a particular focus on AML, from the vast literature spanning three decades, highlighting key studies that have influenced our understanding. We discuss recent insights into its role in leukemogenesis and how these could be exploited for the therapeutic targeting of Myb, its associated co-regulators or its target genes, in order to improve outcomes in the treatment of a wide range of hematopoietic malignancies.
Publisher: Public Library of Science (PLoS)
Date: 12-12-2012
Publisher: Springer Science and Business Media LLC
Date: 23-10-2015
DOI: 10.1038/NRC4018
Abstract: Drugs that target intracellular signalling pathways have markedly improved progression-free survival of patients with cancers who were previously regarded as untreatable. However, the rapid emergence of therapeutic resistance, as a result of bypass signalling or downstream mutation within kinase-mediated signalling cascades, has curtailed the benefit gained from these therapies. Such resistance mechanisms are facilitated by the linearity and redundancy of kinase signalling pathways. We argue that, in each cancer, the dysregulation of key transcriptional regulators not only defines the cancer phenotype but is essential for its development and maintenance. Furthermore, we propose that, as therapeutic targets, these transcriptional regulators are less prone to bypass by alternative mutational events or clonal heterogeneity, and therefore we must rekindle our efforts to directly target transcriptional regulation across a broad range of cancers.
Publisher: Elsevier BV
Date: 02-2015
DOI: 10.1016/J.JBIOTEC.2014.12.006
Abstract: Although numerous techniques for protein expression and production are available the pace of genome sequencing outstrips our ability to analyze the encoded proteins. To address this bottleneck, we have established a system for parallelized cloning, DNA production and cell-free expression of large numbers of proteins. This system is based on a suite of pCellFree Gateway destination vectors that utilize a Species Independent Translation Initiation Sequence (SITS) that mediates recombinant protein expression in any in vitro translation system. These vectors introduce C or N terminal EGFP and mCherry fluorescent and affinity tags, enabling direct analysis and purification of the expressed proteins. To maximize throughput and minimize the cost of protein production we combined Gateway cloning with Rolling Circle DNA Amplification. We demonstrate that as little as 0.1 ng of plasmid DNA is sufficient for template lification and production of recombinant human protein in Leishmania tarentolae and Escherichia coli cell-free expression systems. Our experiments indicate that this approach can be applied to large gene libraries as it can be reliably performed in multi-well plates. The resulting protein expression pipeline provides a valuable new tool for applications of the post genomic era.
Publisher: Springer Science and Business Media LLC
Date: 09-04-2010
DOI: 10.1007/S10637-009-9245-5
Abstract: Heparan sulfate mimetics, which we have called the PG500 series, have been developed to target the inhibition of both angiogenesis and heparanase activity. This series extends the technology underpinning PI-88, a mixture of highly sulfated oligosaccharides which reached Phase III clinical development for hepatocellular carcinoma. Advances in the chemistry of the PG500 series provide numerous advantages over PI-88. These new compounds are fully sulfated, single entity oligosaccharides attached to a lipophilic moiety, which have been optimized for drug development. The rational design of these compounds has led to vast improvements in potency compared to PI-88, based on in vitro angiogenesis assays and in vivo tumor models. Based on these and other data, PG545 has been selected as the lead clinical candidate for oncology and is currently undergoing formal preclinical development as a novel treatment for advanced cancer.
Publisher: Impact Journals, LLC
Date: 16-08-2016
Publisher: Elsevier BV
Date: 05-2008
DOI: 10.1016/J.BBRC.2008.03.032
Abstract: Although the first members of the Schlafen gene family were first described almost 10 years ago, the precise molecular/biochemical functions of the proteins they encode still remain largely unknown. Roles in cell growth, haematopoietic cell differentiation, and T cell development/maturation have, with some experimental support, been postulated, but none have been conclusively verified. Here, we have determined the subcellular localization of Schlafens 1, 2, 4, 5, 8, and 9, representing all three of the murine subgroups. We show that the proteins from subgroups I and II localize to the cytoplasm, while the longer forms in subgroup III localize exclusively to the nuclear compartment. We also demonstrate upregulation of Schlafen2 upon differentiation of haematopoietic cells and show this endogenous protein localizes to the cytoplasm. Thus, we propose the different subgroups of Schlafen proteins are likely to have functionally distinct roles, reflecting their differing localizations within the cell.
Publisher: Springer Science and Business Media LLC
Date: 30-08-2010
DOI: 10.1038/ONC.2010.397
Abstract: c-Myb is an essential hematopoietic transcription factor that controls proliferation and differentiation of progenitors during blood cell development. Whereas sumoylation of the C-terminal regulatory domain (CRD) is known to have a major impact on the activity of c-Myb, no role for noncovalent binding of small ubiquitin-like modifier (SUMO) to c-Myb has been described. Based on the consensus SUMO-interacting motif (SIM), we identified and examined putative SIMs in human c-Myb. Interaction and reporter assays showed that the SIM in the in the transactivation domain of c-Myb (V(267)NIV) is functional. This motif is necessary for c-Myb to be able to interact noncovalently with SUMO, preferentially SUMO2/3. Destroying the SUMO-binding properties by mutation resulted in a large increase in the transactivation potential of c-Myb. Mutational analysis and overexpression of conjugation-defective SUMO argued against intramolecular repression caused by sumoylated CRD and in favor of SUMO-dependent repression in trans. Using both a myeloid cell line-based assay and a primary hematopoietic cell assay, we addressed the transforming abilities of SUMO binding and conjugation mutants. Interestingly, only loss of SUMO binding, and not SUMO conjugation, enhanced the myeloid transformational potential of c-Myb. c-Myb with the SIM mutated conferred a higher proliferative ability than the wild-type and caused an effective differentiation block. This establishes SUMO binding as a mechanism involved in modulating the transactivation activity of c-Myb, and responsible for keeping the transforming potential of the oncoprotein in check.
Publisher: Elsevier BV
Date: 05-2013
DOI: 10.1016/J.LEUKRES.2013.01.012
Abstract: RNA interference-mediated suppression of MYB expression promoted apoptosis in the AML cell line U937, without affecting expression of the anti-apoptotic MYB target BCL2. This was accompanied by up-regulation of the pro-apoptotic gene DRAK2 and stimulation of caspase-9 activity. Moreover, RNA interference-mediated suppression of DRAK2 in U937 cells alleviated apoptosis induced by MYB down-regulation. Finally ChIP assays showed that in U937 cells MYB binds to a conserved element upstream of the DRAK2 transcription start site. Together, these findings identify a novel mechanism by which MYB suppresses apoptosis in an AML model cell line.
Publisher: Elsevier BV
Date: 09-2014
Publisher: Cold Spring Harbor Laboratory
Date: 21-01-2021
DOI: 10.1101/2021.01.21.427535
Abstract: The nucleolar surveillance pathway (NSP) monitors nucleolar fidelity and responds to nucleolar stresses (i.e., inactivation of ribosome biogenesis) by mediating the inhibitory binding of ribosomal proteins (RPs) to mouse double minute 2 homolog (MDM2), a nuclear-localised E3 ubiquitin ligase, which results in p53 accumulation. Inappropriate activation of the NSP has been implicated in the pathogenesis of collection of human diseases termed “ribosomopathies”, while drugs that selectively activate the NSP are now in trials for cancer. Despite the clinical significance, the precise molecular mechanism(s) regulating the NSP remain poorly understood. Using genome-wide loss of function screens, we demonstrate the ribosome biogenesis (RiBi) axis as the most potent class of genes whose disruption stabilises p53. Furthermore, we identified a novel suite of genes critical for the NSP, including a novel mammalian protein implicated in 5S ribonucleoprotein particle (5S-RNP) biogenesis, HEATR3. By selectively disabling the NSP, we unexpectedly demonstrate that a functional NSP is required for the ability of all nuclear acting stresses tested, including DNA damage, to robustly induce p53 accumulation. Together, our data demonstrates that the NSP has evolved as the dominant central integrator of stresses that regulate nuclear p53 abundance, thus ensuring RiBi is hardwired to cellular proliferative capacity.
Publisher: Elsevier BV
Date: 2011
Publisher: Springer Science and Business Media LLC
Date: 26-07-2010
DOI: 10.1186/BCR2614
Publisher: American Society of Hematology
Date: 24-04-2014
DOI: 10.1182/BLOOD-2012-02-413187
Abstract: A mutation preventing interaction between c-Myb and p300 prevents transformation and leukemia induction by MLL-AF9 and AML1-ETO9a oncogenes. Identifying agents that block the c-Myb-p300 interaction may be a valuable approach to developing a therapy for acute myeloid leukemia.
Publisher: Mary Ann Liebert Inc
Date: 08-2010
DOI: 10.1089/HUM.2009.107
Abstract: Manipulation of gene expression is an invaluable tool to study gene function in vitro and in vivo. The application of small inhibitory RNAs to knock down gene expression provides a relatively simple, elegant, but transient approach to study gene function in many cell types as well as in whole animals. Short hairpin structures (shRNAs) are a logical advance as they can be expressed continuously and are hence suitable for stable gene knockdown. Drug-inducible systems have now been developed however, application of the technology has been h ered by persistent problems with low or transient expression, leakiness or poor inducibility of the short hairpin, and lack of reversibility. We have developed a robust, versatile, single lentiviral vector tool that delivers tightly regulated, fully reversible, doxycycline-responsive knockdown of target genes (FOXP3 and MYB), using single short hairpin RNAs. To demonstrate the capabilities of the vector we targeted FOXP3 because it plays a critical role in the development and function of regulatory T cells. We also targeted MYB because of its essential role in hematopoiesis and implication in breast cancer progression. The versatility of this vector is hence demonstrated by knockdown of distinct genes in two biologically separate systems.
Publisher: Impact Journals, LLC
Date: 24-01-2016
Publisher: Springer Science and Business Media LLC
Date: 27-12-2019
DOI: 10.1038/S41598-019-56426-7
Abstract: Acute leukaemias express high levels of MYB which are required for the initiation and maintenance of the disease. Inhibition of MYB expression or activity has been shown to suppress MLL-fusion oncoprotein-induced acute myeloid leukaemias (AML), which are among the most aggressive forms of AML, and indeed MYB transcription has been reported to be regulated by the MLL-AF9 oncoprotein. This highlights the importance of understanding the mechanism of MYB transcriptional regulation in these leukaemias. Here we have demonstrated that the MLL-AF9 fusion protein regulates MYB transcription directly at the promoter region, in part by recruiting the transcriptional regulator kinase CDK9, and CDK9 inhibition effectively suppresses MYB expression as well as cell proliferation. However, MYB regulation by MLL-AF9 does not require H3K79 methylation mediated by the methyltransferase DOT1L, which has also been shown to be a key mediator of MLL-AF9 leukemogenicity. The identification of specific, essential and druggable transcriptional regulators may enable effective targeting of MYB expression, which in turn could potentially lead to new therapeutic approaches for acute myeloid leukaemia with MLL-AF9.
Publisher: American Society of Hematology
Date: 22-04-2010
DOI: 10.1182/BLOOD-2009-08-235846
Abstract: Granulocyte/macrophage colony-stimulating factor promotes growth, survival, differentiation, and activation of normal myeloid cells and plays an important role in myeloid leukemias. The GM-CSF receptor (GMR) shares a signaling subunit, βc, with interleukin-3 and interleukin-5 receptors and has recently been shown to induce activation of Janus kinase 2 (JAK2) and downstream signaling via formation of a unique dodecameric receptor complex. In this study we use 2 activated βc mutants that display distinct signaling capacity and have differential requirements for the GMR α-subunit (GMR-α) to dissect the signaling pathways associated with the GM-CSF response. The V449E transmembrane mutant selectively activates JAK2/signal transducer and activator of transcription 5 and extracellular signal-regulated kinase (ERK) pathways, resulting in a high level of sensitivity to JAK and ERK inhibitors, whereas the extracellular mutant (FIΔ) selectively activates the phosphoinositide 3-kinase/Akt and IκKβ/nuclear factorκB pathways. We also demonstrate a novel and direct interaction between the SH3 domains of Lyn and Src with a conserved proline-rich motif in GMR-α and show a selective requirement for Src family kinases by the FIΔ mutant. We relate the nonoverlapping nature of signaling by the activated mutants to the structure of the unique GMR complex and propose alternative modes of receptor activation acting synergistically in the mature liganded receptor complex.
Publisher: Informa Healthcare
Date: 14-05-2008
Abstract: MYB is highly expressed in almost all estrogen receptor (ER)-positive breast tumours and is a direct target of estrogen/ER signalling. Our recent studies have shown that MYB is also required for the proliferation of ER-positive breast tumour cell lines, and have shed further light on the mechanism of ER regulation of MYB expression. Here we discuss the rationale for therapeutic targeting of MYB in breast cancer and consider a number of approaches to developing an anti-MYB therapeutic.
Publisher: Elsevier BV
Date: 07-2012
Publisher: American Chemical Society (ACS)
Date: 03-02-2010
DOI: 10.1021/JM901449M
Abstract: A series of polysulfated penta- and tetrasaccharide glycosides containing alpha(1-->3)/alpha(1-->2)-linked mannose residues were synthesized as heparan sulfate (HS) mimetics and evaluated for their ability to inhibit angiogenesis. The compounds bound tightly to angiogenic growth factors (FGF-1, FGF-2, and VEGF) and strongly inhibited heparanase activity. In addition, the compounds exhibited potent activity in cell-based and ex vivo assays indicative of angiogenesis, with tetrasaccharides exhibiting activity comparable to that of pentasaccharides. Selected compounds also showed good antitumor activity in vivo in a mouse melanoma (solid tumor) model resistant to the phase III HS mimetic 1 (muparfostat, formerly known as PI-88). The lipophilic modifications also resulted in reduced anticoagulant activity, a common side effect of HS mimetics, and conferred a reasonable pharmacokinetic profile in the rat, as exemplified by the sulfated octyl tetrasaccharide 5. The data support the further investigation of this class of compounds as potential antiangiogenic, anticancer therapeutics.
Publisher: Springer Science and Business Media LLC
Date: 06-1201
Publisher: American Society of Hematology
Date: 02-12-2021
Publisher: American Association for Cancer Research (AACR)
Date: 13-11-2011
DOI: 10.1158/0008-5472.CAN-11-1015
Abstract: MYB oncogene upregulation is associated with estrogen receptor (ER)-positive breast cancer, but disease requirements for MYB function in vivo have not been explored. In this study, we provide evidence of a critical requirement for MYB functions in models of human and murine breast cancer. In human breast cancer, we found that MYB expression was critical for tumor cell growth both in vitro and in vivo in xenograft settings. In transgenic knockout mice, tissue-specific deletion of the murine MYB gene caused a transient defect in mammary gland development that was reflected in delayed ductal branching and defective apical bud formation. In mouse mammary tumor virus (MMTV)-NEU mice where tumors are initiated by activation of HER2, MYB deletion was sufficient to abolish tumor formation. In the more aggressive MMTV-PyMT model system, MYB deletion delayed tumorigenesis significantly. Together, the findings in these transgenic knockout models implied that MYB was critical during an early window in mammary development when it was essential for tumor initiation, even though MYB loss did not exert a lasting impact upon normal mammary function. Two important MYB-target genes that promote cell survival, BCL2 and GRP78/BIP, were each elevated compared with nontransformed mammary epithelial cells, thereby promoting survival as confirmed in colony formation assays in vitro. Taken together, our findings establish a role for MYB at the hub of ER- and HER2-dependent pathways in mammary carcinogenesis. Cancer Res 71(22) 7029–37. ©2011 AACR.
Publisher: American Society of Hematology
Date: 13-04-2023
Publisher: Public Library of Science (PLoS)
Date: 08-04-2015
Publisher: Public Library of Science (PLoS)
Date: 24-05-2011
Publisher: Oxford University Press (OUP)
Date: 05-04-2012
DOI: 10.1093/NAR/GKS286
Publisher: Springer Science and Business Media LLC
Date: 08-01-2009
DOI: 10.1038/LEU.2008.349
Abstract: The tumor suppressor Gadd45alpha was earlier shown to be a repressed target of sustained receptor-mediated ERK1/2 signaling. We have identified Gadd45alpha as a downregulated gene in response to constitutive signaling from two FLT3 mutants (FLT3-ITD and FLT3-TKD) commonly found in AML, and a leukemogenic GM-CSF receptor trans-membrane mutant (GMR-V449E). GADD45A mRNA downregulation is also associated with FLT3-ITD(+) AML. Sustained ERK1/2 signaling contributes significantly to receptor-mediated downregulation of Gadd45alpha mRNA in FDB1 cells expressing activated receptor mutants, and in the FLT3-ITD(+) cell line MV4 . Knockdown of Gadd45alpha with shRNA led to increased growth and survival of FDB1 cells and enforced expression of Gadd45alpha in FDB1 cells expressing FLT3-ITD or GMR-V449E resulted in reduced growth and viability. Gadd45alpha overexpression in FLT3-ITD(+) AML cell lines also resulted in reduced growth associated with increased apoptosis and G(1)/S cell cycle arrest. Overexpression of Gadd45alpha in FDB1 cells expressing GMR-V449E was sufficient to induce changes associated with myeloid differentiation suggesting Gadd45alpha downregulation contributes to the maintenance of receptor-induced myeloid differentiation block. Thus, we show that ERK1/2-mediated downregulation of Gadd45alpha by sustained receptor signaling contributes to growth, survival and arrested differentiation in AML.
Publisher: Elsevier BV
Date: 03-2017
Abstract: Malignant melanomas often arise from nevi, which result from initial oncogene-induced hyperproliferation of melanocytes that are maintained in a CDKN2A 16-mediated senescent state. Thus, genes that can bypass this senescence barrier are likely to contribute to melanoma development. We have performed a gain-of-function screen of 17,030 lentivirally expressed human open reading frames (ORFs) in a melanoma cell line containing an inducible p16 construct to identify such genes. Genes known to bypass p16-induced senescence arrest, including the human papilloma virus 18 E7 gene ( HPV18E7), and genes such as the p16-binding CDK6 with expected functions, as well as panel of novel genes, were identified, including high-mobility group box (HMGB) proteins. A number of these were further validated in two other models of p16-induced senescence. Tissue immunohistochemistry demonstrated higher levels of CDK6 in primary melanomas compared with normal skin and nevi. Reduction of CDK6 levels drove melanoma cells expressing functional p16 into senescence, demonstrating its contribution to bypass senescence.
Publisher: Elsevier BV
Date: 09-2008
DOI: 10.1016/J.BCMD.2008.03.006
Abstract: The Schlafen gene family has been implicated in lymphoid and myeloid maturation and differentiation as well as inflammation. However, little is known about the functions of this gene family except that anti-proliferative activities, particularly for Schlafen1, the prototype member of the family, have been reported. This was shown mainly by ectopic expression of Schlafen1 in murine fibroblasts resulting in growth inhibition and a G1 cell cycle arrest apparently via repression of Cyclin D1 expression. However, we have been unable to reproduce these findings. Schlafen1 and Schlafen2 failed to inhibit cell proliferation, cause G1 cell cycle arrest, or affect Cyclin D1 level in murine fibroblasts. This was regardless of whether overexpression was constitutive, induced or from transient transfections. Moreover, in our hands, Schlafen1 and -2 do not appear to regulate the activity of Cyclin D1 promoter. Importantly, we also showed that Schlafen1 and -2 do not play anti-proliferative roles in more physiologically-relevant myeloid cell lines. We therefore suggest that Schlafen1 and Schlafen2 might not have obligatory anti-proliferative activities, at least in vitro, and that efforts to explore their functions should be directed to other aspects, such as haemopoietic development and immune response.
Publisher: American Association for Cancer Research (AACR)
Date: 12-2015
DOI: 10.1158/1535-7163.MCT-15-0506
Abstract: Human papillomavirus (HPV) is the causative agent in cervical cancer. HPV oncogenes are major drivers of the transformed phenotype, and the cancers remain addicted to these oncogenes. A screen of the human kinome has identified inhibition of Aurora kinase A (AURKA) as being synthetically lethal on the background of HPV E7 expression. The investigational AURKA inhibitor MLN8237/Alisertib selectively promoted apoptosis in the HPV cancers. The apoptosis was driven by an extended mitotic delay in the Alisertib-treated HPV E7–expressing cells. This had the effect of reducing Mcl-1 levels, which is destabilized in mitosis, and increasing BIM levels, normally destabilized by Aurora A in mitosis. Overexpression of Mcl-1 reduced sensitivity to the drug. The level of HPV E7 expression influenced the extent of Alisertib-induced mitotic delay and Mcl-1 reduction. Xenograft experiments with three cervical cancer cell lines showed Alisertib inhibited growth of HPV and non-HPV xenografts during treatment. Growth of non-HPV tumors was delayed, but in two separate HPV cancer cell lines, regression with no resumption of growth was detected, even at 50 days after treatment. A transgenic model of premalignant disease driven solely by HPV E7 also demonstrated sensitivity to drug treatment. Here, we show for the first time that targeting of the Aurora A kinase in mice using drugs such as Alisertib results in a curative sterilizing therapy that may be useful in treating HPV-driven cancers. Mol Cancer Ther 14(12) 2753–61. ©2015 AACR.
Publisher: American Association for Cancer Research (AACR)
Date: 02-2016
DOI: 10.1158/2159-8290.CD-15-1470
Abstract: Summary: A majority of adenoid cystic carcinomas (AdCC)—rare tumors of the salivary gland and some other organs—have recently been found to be driven by chromosomal translocations resulting in MYB–NFIB fusions. Brayer and colleagues and Mitani and colleagues have now reported that AdCCs can alternatively be driven by similar rearrangements involving a second MYB family gene, MYBL1, and that these two drivers act in remarkably similar ways. Cancer Discov 6(2) 125–7. ©2016 AACR. See related article by Brayer et al., p. 176.
Publisher: Springer Science and Business Media LLC
Date: 02-2011
DOI: 10.1038/BJC.2011.11
Publisher: Springer Science and Business Media LLC
Date: 12-05-2022
DOI: 10.1038/S41467-022-30223-9
Abstract: The interaction of germline variation and somatic cancer driver mutations is under-investigated. Here we describe the genomic mitochondrial landscape in adult acute myeloid leukaemia (AML) and show that rare variants affecting the nuclear- and mitochondrially-encoded complex I genes show near-mutual exclusivity with somatic driver mutations affecting isocitrate dehydrogenase 1 ( IDH1 ), but not IDH2 suggesting a unique epistatic relationship. Whereas AML cells with rare complex I variants or mutations in IDH1 or IDH2 all display attenuated mitochondrial respiration, heightened sensitivity to complex I inhibitors including the clinical-grade inhibitor, IACS-010759, is observed only for IDH1 -mutant AML. Furthermore, IDH1 mutant blasts that are resistant to the IDH1-mutant inhibitor, ivosidenib, retain sensitivity to complex I inhibition. We propose that the IDH1 mutation limits the flexibility for citrate utilization in the presence of impaired complex I activity to a degree that is not apparent in IDH2 mutant cells, exposing a mutation-specific metabolic vulnerability. This reduced metabolic plasticity explains the epistatic relationship between the germline complex I variants and oncogenic IDH1 mutation underscoring the utility of genomic data in revealing metabolic vulnerabilities with implications for therapy.
Publisher: Springer Science and Business Media LLC
Date: 14-10-2013
DOI: 10.1038/ONC.2013.419
Abstract: The MYB gene is a master regulator of hematopoiesis and contributes to leukemogenesis in several species including humans. Although it is clear that MYB can promote proliferation, suppress apoptosis and block differentiation, the identities of the MYB target genes that mediate these effects have only been partially elucidated. Several studies, including our own, have collectively identified substantial numbers of MYB target genes, including candidates for each of these activities however, functional validation, particularly in the case of differentiation suppression, has lagged well behind. Here we show that GFI1, which encodes an important regulator of hematopoietic stem cell (HSC) function and granulocytic differentiation, is a direct target of MYB in myeloid leukemia cells. Chromatin immunoprecipitation and reporter studies identified a functional MYB-binding site in the promoter region of GFI, whereas ectopic expression and small hairpin RNA-mediated knockdown of MYB resulted in concomitant increases and decreases, respectively, in GFI1 expression. We also demonstrate that GFI1, like MYB, can block the induced monocytic differentiation of a human acute myeloid leukemia cell line, and most importantly, that GFI1 is essential for MYB's ability to block monocytic differentiation. Thus, we have identified a target of MYB that is a likely mediator of its myeloid differentiation-blocking activity, and which may also be involved in MYB's activities in regulating normal HSC function and myeloid differentiation.
Publisher: Wiley
Date: 23-06-2014
DOI: 10.1111/PCMR.12268
Abstract: Melanoma cell lines are commonly defective for the G2-phase cell cycle checkpoint that responds to incomplete catenation of the replicated chromosomes. Here, we demonstrate that melanomas defective for this checkpoint response are less sensitive to genotoxic stress, suggesting that the defective cell lines compensated for the checkpoint loss by increasing their ability to cope with DNA damage. We performed an siRNA kinome screen to identify kinases responsible and identified PI3K pathway components. Checkpoint-defective cell lines were three-fold more sensitive to small molecule inhibitors of PI3K. The PI3K inhibitor PF-05212384 promoted apoptosis in the checkpoint-defective lines, and the increased sensitivity to PI3K inhibition correlated with increased levels of activated Akt. This work demonstrates that increased PI3K pathway activation is a necessary adaption for the continued viability of melanomas with a defective decatenation checkpoint.
Publisher: Elsevier BV
Date: 12-2018
Publisher: Elsevier BV
Date: 08-2012
Publisher: Oxford University Press (OUP)
Date: 11-02-2011
DOI: 10.1093/NAR/GKR024
Publisher: Springer Science and Business Media LLC
Date: 06-05-2021
DOI: 10.1038/S41388-021-01800-X
Abstract: Transcription factor MYB has recently emerged as a promising drug target for the treatment of acute myeloid leukemia (AML). Here, we have characterized a group of natural sesquiterpene lactones (STLs), previously shown to suppress MYB activity, for their potential to decrease AML cell proliferation. Unlike what was initially thought, these compounds inhibit MYB indirectly via its cooperation partner C/EBPβ. C/EBPβ-inhibitory STLs affect the expression of a large number of MYB-regulated genes, suggesting that the cooperation of MYB and C/EBPβ broadly shapes the transcriptional program of AML cells. We show that expression of GFI1 , a direct MYB target gene, is controlled cooperatively by MYB, C/EBPβ, and co-activator p300, and is down-regulated by C/EBPβ-inhibitory STLs, exemplifying that they target the activity of composite MYB-C/EBPβ-p300 transcriptional modules. Ectopic expression of GFI1, a zinc-finger protein that is required for the maintenance of hematopoietic stem and progenitor cells, partially abrogated STL-induced myelomonocytic differentiation, implicating GFI1 as a relevant target of C/EBPβ-inhibitory STLs. Overall, our data identify C/EBPβ as a pro-leukemogenic factor in AML and suggest that targeting of C/EBPβ may have therapeutic potential against AML.
Publisher: INIST-CNRS
Date: 11-2010
DOI: 10.4267/2042/44658
Publisher: Springer Science and Business Media LLC
Date: 10-04-2019
DOI: 10.1038/S41388-019-0789-3
Abstract: Over 70% of human breast cancers are estrogen receptor-positive (ER
Publisher: American Chemical Society (ACS)
Date: 09-12-2009
DOI: 10.1021/PR900884G
Abstract: Understanding networks of interacting proteins is a major goal in cell biology. The survival of motor neurons protein (SMN) interacts, directly or indirectly, with a large number of other proteins and reduced levels of SMN cause the inherited disorder spinal muscular atrophy (SMA). Some SMN interactions are stable and stoichiometric, such as those with gemins, while others are expected to be transient and substoichiometric, such as the functional interaction of SMN with coilin in Cajal bodies. This study set out to determine whether novel components of the extensive SMN interactome can be identified by a proteomic approach. SMN complexes were immuno-precipitated from HeLa nuclear extracts, using anti-SMN monoclonal antibody attached to magnetic beads, digested with trypsin, separated by capillary-liquid chromatography and analyzed by MALDI TOF/TOF mass spectrometry. One-hundred and one proteins were detected with a p value of <0.05, SMN, gemins and U snRNPs being the dominant "hits". Sixty-nine of these were rejected after MALDI analysis of two control pull-downs using antibodies against unrelated nuclear proteins. The proteins found only in anti-SMN pulldowns were either known SMN partners, and/or contained dimethylated RG domains involved in direct interaction with the SMN tudor domain, or they were known binding partners of such direct SMN interactors. Myb-binding protein 1a, identified as a novel candidate, is a mainly nucleolar protein of unknown function but it partially colocalized with SMN in Cajal bodies in HeLa cell nucleoplasm and, like SMN, was reduced in cells from an SMA patient.
Publisher: Future Science Ltd
Date: 05-2010
DOI: 10.2144/000113410
Abstract: Cellular proliferation is fundamental to organism development, tissue renewal, and erse disease states such as cancer. In vitro measurement of proliferation by high-throughput screening allows rapid characterization of the effects of small-molecule or genetic treatments on primary and established cell lines. Current assays that directly measure the cell cycle are not amenable to high-throughput processing and analysis. Here we report the adaptation of the chemical method for detecting DNA synthesis by 5-ethynyl-2′-deoxyuridine (EdU) incorporation into both high-throughput liquid handling and high-content imaging analysis. We demonstrate that chemical detection of EdU incorporation is effective for high-resolution analysis and quantitation of DNA synthesis by high-content imaging. To validate this assay platform we used treatments of MCF10A cells with media supplements and pharmacological inhibitors that are known to affect cell proliferation. Treatments with specific kinase inhibitors indicate that EGF and serum stimulation employs both the mitogen extracellular kinase (MEK)/extracellular-regulated kinase (ERK) and phosphoinositol-3 kinase (PI3K)/AKT signaling networks. As described here, this method is fast, reliable, and inexpensive and yields robust data that can be easily interpreted.
Publisher: Spandidos Publications
Date: 19-01-2018
DOI: 10.3892/OL.2018.7827
No related grants have been discovered for Thomas Gonda.