ORCID Profile
0000-0003-1128-5315
Current Organisation
Garvan Institute of Medical Research
Does something not look right? The information on this page has been harvested from data sources that may not be up to date. We continue to work with information providers to improve coverage and quality. To report an issue, use the Feedback Form.
Publisher: American Association for the Advancement of Science (AAAS)
Date: 03-03-2023
Abstract: Gene expression noise is known to promote stochastic drug resistance through the elevated expression of in idual genes in rare cancer cells. However, we now demonstrate that chemoresistant neuroblastoma cells emerge at a much higher frequency when the influence of noise is integrated across multiple components of an apoptotic signaling network. Using a JNK activity biosensor with longitudinal high-content and in vivo intravital imaging, we identify a population of stochastic, JNK-impaired, chemoresistant cells that exist because of noise within this signaling network. Furthermore, we reveal that the memory of this initially random state is retained following chemotherapy treatment across a series of in vitro, in vivo, and patient models. Using matched PDX models established at diagnosis and relapse from in idual patients, we show that HDAC inhibitor priming cannot erase the memory of this resistant state within relapsed neuroblastomas but improves response in the first-line setting by restoring drug-induced JNK activity within the chemoresistant population of treatment-naïve tumors.
Publisher: Elsevier BV
Date: 09-2020
Publisher: Springer Science and Business Media LLC
Date: 05-12-2016
DOI: 10.1038/SREP37871
Abstract: Cerebral malaria (CM) is a severe complication of Plasmodium falciparum infection responsible for thousands of deaths in children in sub-Saharan Africa. CM pathogenesis remains incompletely understood but a number of effectors have been proposed, including plasma microparticles (MP). MP numbers are increased in CM patients’ circulation and, in the mouse model, they can be localised within inflamed vessels, suggesting their involvement in vascular damage. In the present work we define, for the first time, the protein cargo of MP during experimental cerebral malaria (ECM) with the overarching hypothesis that this characterisation could help understand CM pathogenesis. Using qualitative and quantitative high-throughput proteomics we compared MP proteins from non-infected and P . berghei ANKA-infected mice. More than 360 proteins were identified, 60 of which were differentially abundant, as determined by quantitative comparison using TMT TM isobaric labelling. Network analyses showed that ECM MP carry proteins implicated in molecular mechanisms relevant to CM pathogenesis, including endothelial activation. Among these proteins, the strict association of carbonic anhydrase I and S100A8 with ECM was verified by western blot on MP from DBA/1 and C57BL/6 mice. These results demonstrate that MP protein cargo represents a novel ECM pathogenic trait to consider in the understanding of CM pathogenesis.
Publisher: Public Library of Science (PLoS)
Date: 13-04-2016
Publisher: Springer Science and Business Media LLC
Date: 10-11-2015
DOI: 10.1038/SREP16314
Abstract: Microparticle (MP) research is clouded by debate regarding the accuracy and validity of flow cytometry (FCM) as an analytical methodology, as it is influenced by many variables including the pre-analytical conditions, instruments physical capabilities and detection parameters. This study utilises a simplistic in vitro system for generating MP and through comparative analysis with immuno-electron microscopy (Immuno-EM) assesses the strengths and limitations of probe selection and high-sensitivity FCM. Of the markers examined, MP were most specifically labelled with phosphatidylserine ligands, annexin V and lactadherin, although only ~60% MP are PS positive. Whilst these two ligands detect comparable absolute MP numbers, they interact with the same population in distinct manners annexin V binding is enhanced on TNF induced MP. CD105 and CD54 expression were, as expected, consistent and enhanced following TNF activation respectively. Their labelling however accounted for as few as 30–40% of MP. The greatest discrepancies between FCM and I-EM were observed in the population solely labelled for the surface antigen. These findings demonstrate that despite significant improvements in resolution, high-sensitivity FCM remains limited in detecting small-size MP expressing low antigen levels. This study highlights factors to consider when selecting endothelial MP probes, as well as interpreting and representing data.
Publisher: The American Association of Immunologists
Date: 10-2014
Abstract: Endothelial cells closely interact with circulating lymphocytes. Aggression or activation of the endothelium leads to an increased shedding of endothelial cell microparticles (MP). Endothelial MP (EMP) are found in high plasma levels in numerous immunoinflammatory diseases, such as atherosclerosis, sepsis, multiple sclerosis, and cerebral malaria, supporting their role as effectors and markers of vascular dysfunction. Given our recently described role for human brain microvascular endothelial cells (HBEC) in modulating immune responses, we investigated how HBEC-derived MP could interact with and support the proliferation of T cells. Like their mother cells, EMP expressed molecules important for Ag presentation and T cell costimulation, that is, β2-microglobulin, MHC II, CD40, and ICOSL. HBEC were able to take up fluorescently labeled Ags with EMP also containing fluorescent Ags, suggestive of Ag carryover from HBEC to EMP. In cocultures, fluorescently labeled EMP from resting or cytokine-stimulated HBEC formed conjugates with both CD4+ and CD8+ subsets, with higher proportions of T cells binding EMP from cytokine-stimulated cells. The increased binding of EMP from cytokinestimulated HBEC to T cells was VCAM-1 and ICAM-1 dependent. Finally, in CFSE T cell proliferation assays using anti-CD3 mAb or T cell mitogens, EMP promoted the proliferation of CD4+ T cells and that of CD8+ T cells in the absence of exogenous stimuli and in the T cell mitogenic stimulation. Our findings provide novel evidence that EMP can enhance T cell activation and potentially ensuing Ag presentation, thereby pointing toward a novel role for MP in neuroimmunological complications of infectious diseases.
Publisher: Springer Science and Business Media LLC
Date: 28-08-2023
DOI: 10.1038/S43018-023-00614-Y
Abstract: The lysyl oxidase family represents a promising target in stromal targeting of solid tumors due to the importance of this family in crosslinking and stabilizing fibrillar collagens and its known role in tumor desmoplasia. Using small-molecule drug-design approaches, we generated and validated PXS-5505, a first-in-class highly selective and potent pan-lysyl oxidase inhibitor. We demonstrate in vitro and in vivo that pan-lysyl oxidase inhibition decreases chemotherapy-induced pancreatic tumor desmoplasia and stiffness, reduces cancer cell invasion and metastasis, improves tumor perfusion and enhances the efficacy of chemotherapy in the autochthonous genetically engineered KPC model, while also demonstrating antifibrotic effects in human patient-derived xenograft models of pancreatic cancer. PXS-5505 is orally bioavailable, safe and effective at inhibiting lysyl oxidase activity in tissues. Our findings present the rationale for progression of a pan-lysyl oxidase inhibitor aimed at eliciting a reduction in stromal matrix to potentiate chemotherapy in pancreatic ductal adenocarcinoma.
Publisher: Springer Science and Business Media LLC
Date: 19-11-2018
DOI: 10.1038/S41467-018-07404-6
Abstract: The original version of this Article contained an error in Figure 4. In panel i, the lower CYA and α-SMA images were inadvertently inverted. This has been corrected in both the PDF and HTML versions of the Article.
Publisher: Wiley
Date: 17-03-2017
DOI: 10.1002/CM.21356
Publisher: American Association for the Advancement of Science (AAAS)
Date: 28-04-2023
Abstract: Aberrant AKT activation occurs in a number of cancers, metabolic syndrome, and immune disorders, making it an important target for the treatment of many diseases. To monitor spatial and temporal AKT activity in a live setting, we generated an Akt-FRET biosensor mouse that allows longitudinal assessment of AKT activity using intravital imaging in conjunction with image stabilization and optical window technology. We demonstrate the sensitivity of the Akt-FRET biosensor mouse using various cancer models and verify its suitability to monitor response to drug targeting in spheroid and organotypic models. We also show that the dynamics of AKT activation can be monitored in real time in erse tissues, including in in idual islets of the pancreas, in the brown and white adipose tissue, and in the skeletal muscle. Thus, the Akt-FRET biosensor mouse provides an important tool to study AKT dynamics in live tissue contexts and has broad preclinical applications.
Publisher: Proceedings of the National Academy of Sciences
Date: 17-08-2020
Abstract: The protein kinase MEKK1 activates stress-signaling pathways in response to various cellular stressors, including chemotherapies that disrupt dynamics of the tubulin cytoskeleton. We show that MEKK1 contains a previously uncharacterized domain that can preferentially bind to the curved tubulin heterodimer—which is found in soluble tubulin and at sites of microtubule assembly and disassembly. Mutations that interfere with MEKK1−tubulin binding disrupt microtubule networks in migrating cells and are enriched in patient-derived tumor sequences. These results suggest that MEKK1−tubulin binding may be relevant to cancer progression, and the efficacy of microtubule-disrupting chemotherapies that require the activity of MEKK1.
Publisher: Springer International Publishing
Date: 2020
Publisher: Cold Spring Harbor Laboratory
Date: 08-04-2020
DOI: 10.1101/2020.04.07.030676
Abstract: The MEKK1 protein is a pivotal kinase activator of responses to cellular stress. Activation of MEKK1 can trigger various responses, including mitogen activated protein (MAP) kinases, NF-κB signalling, or cell migration. Notably, MEKK1 activity is triggered by microtubule-targeting chemotherapies, amongst other stressors. Here we show that MEKK1 contains a previously unidentified tumour overexpressed gene (TOG) domain. The MEKK1 TOG domain binds to tubulin heterodimers—a canonical function of TOG domains—but is unusual in that it appears alone rather than as part of a multi-TOG array, and has structural features distinct from previously characterised TOG domains. MEKK1 TOG demonstrates a clear preference for binding curved tubulin heterodimers, which exist in soluble tubulin and at sites of microtubule polymerisation and depolymerisation. Mutations disrupting tubulin-binding lead to destabilisation of the MEKK1 protein in cells, and ultimately a decrease in microtubule density at the leading edge of polarised cells. We also show that MEKK1 mutations at the tubulin-binding interface of the TOG domain recur in patient derived tumour sequences, suggesting selective enrichment of tumour cells with disrupted MEKK1–microtubule association. Together, these findings provide a direct link between the MEKK1 protein and tubulin, which is likely to be relevant to cancer cell migration and response to microtubule-modulating therapies. The protein kinase MEKK1 activates stress response pathways in response to various cellular stressors, including chemotherapies that disrupt dynamics of the tubulin cytoskeleton. Filipčík et al., show that MEKK1 contains a previously uncharacterised domain that can preferentially bind to the curved tubulin heterodimer—which is found at sites of microtubule assembly and disassembly. Mutations that interfere with MEKK1-tubulin binding disrupt microtubule networks in migrating cells and are enriched in patient-derived tumour sequences. These results suggest that MEKK1-tubulin binding may be relevant to cancer progression, and the efficacy of microtubule-disrupting chemotherapies that require the activity of MEKK1.
Publisher: Springer Science and Business Media LLC
Date: 12-10-2018
DOI: 10.1038/S41467-018-06713-0
Abstract: Germline mutations in the ubiquitously expressed ACTB , which encodes β-cytoplasmic actin (CYA), are almost exclusively associated with Baraitser-Winter Cerebrofrontofacial syndrome (BWCFF). Here, we report six patients with previously undescribed heterozygous variants clustered in the 3′-coding region of ACTB . Patients present with clinical features distinct from BWCFF, including mild developmental disability, microcephaly, and thrombocytopenia with platelet anisotropy. Using patient-derived fibroblasts, we demonstrate cohort specific changes to β-CYA filament populations, which include the enhanced recruitment of thrombocytopenia-associated actin binding proteins (ABPs). These perturbed interactions are supported by in silico modeling and are validated in disease-relevant thrombocytes. Co-examination of actin and microtubule cytoskeleton constituents in patient-derived megakaryocytes and thrombocytes indicates that these β-CYA mutations inhibit the final stages of platelet maturation by compromising microtubule organization. Our results define an ACTB -associated clinical syndrome with a distinct genotype-phenotype correlation and delineate molecular mechanisms underlying thrombocytopenia in this patient cohort.
Publisher: Frontiers Media SA
Date: 07-10-2020
Publisher: Springer Science and Business Media LLC
Date: 05-07-2018
DOI: 10.1038/S41598-018-28459-X
Abstract: Megakaryocytes (MKs) are the precursors of platelets (PLTs) and may be used for PLT production in vivo or in vitro , as well as a source for PLT-derived growth factors. Induced pluripotent stem cells represent an unlimited cell source for the in vitro production of MKs. This study aimed at developing an effective, xeno-free and scalable system to produce high numbers of MKs. In particular, microcarrier beads-assisted stirred bioreactors were evaluated as a means of improving MK yields. This method resulted in the production of 18.7 × 10 7 MKs per 50 ml medium. Laminin-coated microcarriers increased MK production per iPSC by up to 10-fold. MKs obtained in this system showed typical features of mature MKs and were able to produce PLTs in vitro and in vivo . To increase safety, MKs produced in the bioreactors were irradiated a procedure that did not affect their capability to form proPLTs and PTLs after transfusion. In vitro generated MKs represent a promising alternative to donor PLTs and open the possibility for the development of innovative MK-based cell therapies.
Publisher: Springer Science and Business Media LLC
Date: 09-08-2017
DOI: 10.1038/S41598-017-07933-Y
Abstract: Myosin motor proteins convert chemical energy into force and movement through their interactions with nucleotide and filamentous actin (F-actin). The evolutionarily conserved lysine-265 (K265) of the myosin-2 motor from Dictyostelium discoideum ( Dd ) is proposed to be a key residue in an allosteric communication pathway that mediates actin-nucleotide coupling. To better understand the role of K265, point mutations were introduced within the Dd myosin-2 M765-2R framework, replacing this lysine with alanine (K265A), glutamic acid (K265E) or glutamine (K265Q), and the functional and kinetic properties of the resulting myosin motors were assessed. The alanine and glutamic acid substitutions reduced actin-activated ATPase activity, slowed the in vitro sliding velocity and attenuated the inhibitory potential of the allosteric myosin inhibitor pentabromopseudilin (PBP). However, glutamine substitution did not substantially change these parameters. Structural modelling suggests that K265 interacts with D590 and Q633 to establish a pivotal allosteric branching point. Based on our results, we propose: (1) that the K265-D590 interaction functions to reduce myosins basal ATPase activity in the absence of F-actin, and (2) that the dynamic formation of the K265-Q633 salt bridge upon actin cleft closure regulates the activation of product release by actin filaments.
Publisher: Elsevier BV
Date: 07-2022
Publisher: Portland Press Ltd.
Date: 12-2022
DOI: 10.1042/BST20220808
Abstract: c-Jun N-terminal Kinases (JNKs) have been identified as key disease drivers in a number of pathophysiological settings and central oncogenic signaling nodes in various cancers. Their roles in driving primary tumor growth, positively regulating cancer stem cell populations, promoting invasion and facilitating metastatic outgrowth have led JNKs to be considered attractive targets for anti-cancer therapies. However, the homeostatic, apoptotic and tumor-suppressive activities of JNK proteins limit the use of direct JNK inhibitors in a clinical setting. In this review, we will provide an overview of the different JNK targeting strategies developed to date, which include various ATP-competitive, non-kinase and substrate-competitive inhibitors. We aim to summarize their distinct mechanisms of action, review some of the insights they have provided regarding JNK-targeting in cancer, and outline the limitations as well as challenges of all strategies that target JNKs directly. Furthermore, we will highlight alternate drug targets within JNK signaling complexes, including recently identified scaffold proteins, and discuss how these findings may open up novel therapeutic options for targeting discrete oncogenic JNK signaling complexes in specific cancer settings.
Publisher: Elsevier BV
Date: 2018
Publisher: eLife Sciences Publications, Ltd
Date: 09-06-2020
DOI: 10.7554/ELIFE.53367
Abstract: The identification of clinically viable strategies for overcoming resistance to platinum chemotherapy in lung adenocarcinoma has previously been h ered by inappropriately tailored in vitro assays of drug response. Therefore, using a pulse model that closely mimics the in vivo pharmacokinetics of platinum therapy, we profiled cisplatin-induced signalling, DNA-damage and apoptotic responses across a panel of human lung adenocarcinoma cell lines. By coupling this data to real-time, single-cell imaging of cell cycle and apoptosis we provide a fine-grained stratification of response, where a P70S6K-mediated signalling axis promotes resistance on a TP53 wildtype or null background, but not a mutant TP53 background. This finding highlights the value of in vitro models that match the physiological pharmacokinetics of drug exposure. Furthermore, it also demonstrates the importance of a mechanistic understanding of the interplay between somatic mutations and the signalling networks that govern drug response for the implementation of any consistently effective, patient-specific therapy.
Publisher: Elsevier BV
Date: 09-2021
DOI: 10.1016/J.CELREP.2021.109689
Abstract: Assessing drug response within live native tissue provides increased fidelity with regards to optimizing efficacy while minimizing off-target effects. Here, using longitudinal intravital imaging of a Rac1-Förster resonance energy transfer (FRET) biosensor mouse coupled with in vivo photoswitching to track intratumoral movement, we help guide treatment scheduling in a live breast cancer setting to impair metastatic progression. We uncover altered Rac1 activity at the center versus invasive border of tumors and demonstrate enhanced Rac1 activity of cells in close proximity to live tumor vasculature using optical window imaging. We further reveal that Rac1 inhibition can enhance tumor cell vulnerability to fluid-flow-induced shear stress and therefore improves overall anti-metastatic response to therapy during transit to secondary sites such as the lung. Collectively, this study demonstrates the utility of single-cell intravital imaging in vivo to demonstrate that Rac1 inhibition can reduce tumor progression and metastases in an autochthonous setting to improve overall survival.
Publisher: Springer Science and Business Media LLC
Date: 21-03-2019
Publisher: Wiley
Date: 16-11-2012
DOI: 10.1096/FJ.12-216531
Abstract: Elevated endothelial microparticle (MP) levels are observed in numerous diseases, increasingly supporting roles as effectors and valuable markers of vascular dysfunction. While a contractile role for the actin cytoskeleton has been implicated in vesiculation, i.e., MP production, the precise interactions and mechanisms of its constituents, β- and γ-cytoplasmic actins, is unknown. Human cerebral microvascular endothelial cells were stimulated with known agonists, and vesiculation development was monitored by scanning electron microscopy (SEM) and flow cytometry. These data in combination provide new insight into the kinetics, patterns of vesiculating cell recruitment, and degrees of response specific to stimuli. Reorganization of β- and γ-actins, F-actin, vinculin, and talin accompanied significant MP release. β-Actin redistribution into basal stress fibers following stimulation was associated with increased apically situated actin-rich particulate structures, which in turn directly correlated with electron-lucent membrane protrusions observed by SEM. Y-27632 Rho-kinase inhibition abolished basal β-actin fiber formation, minimizing apically associated actin-rich structures, significantly reducing membrane protrusions and MP release to near basal levels. Cytoskeletal protein expression and distribution varied between MPs and mother cells, as determined by Western blot. These data strongly suggest that β-actin plays an active facilitative role in agonist-induced protuberance formation, through mechanical interactions with newly described actin-rich structures.
Location: Australia
No related grants have been discovered for Sharissa Latham.