Biology subjects

Cadell, A. L.

Publications and source records attributed to Cadell, A. L..

5 recordsLinked to original sources

bioRxiv2025

Inclusion of JNK-independent drugs within multi-agent chemotherapy improves response in relapsed high-risk neuroblastoma

The acquisition of a chemoresistant state underlies poor prognosis in many cancers, including neuroblastoma. We previously demonstrated that heterogeneity in apoptosis induction through c-Jun N-Terminal Kinase (JNK) promotes a form of non-genetic chemoresistance in neuroblastoma observable at both patient and single-cell levels. As the maintenance of this JNK-impaired state in the relapse setting is a significant barrier to the efficacy of many standard-of-care chemotherapy drugs, we combined a mechanistic, mathematical model of JNK activation with a paediatric focused drug screen and identified approved oncology drugs capable of inducing apoptosis in a JNK-independent manner. Functional genomics further revealed that synergy between these JNK-independent drugs and standard-of-care chemotherapies emerged from differential utilisation of apoptotic network components, rather than from their direct mechanistic targets. Efficacy studies with PDX models also confirmed that including a JNK-independent drug within existing chemotherapy backbones significantly improved response in the relapse setting, where new approaches are urgently needed. One Sentence SummaryDrug combinations utilising differential sets of network components can overcome the apoptotic impaired state of relapsed high-risk neuroblastoma.

cancer biology↗

bioRxiv2025

JNK pathway suppression drives resistance to combination endocrine therapy and CDK4/6 inhibition in ER+ breast cancer

PurposeEndocrine therapy in combination with CDK4/6 inhibition doubles the progression-free survival of patients with advanced ER+ breast cancer, but resistance is inevitable, leaving patients with limited treatment options. Experimental DesignWe performed unbiased genome-wide CRISPR/Cas9 knockout screens using ER+ breast cancer cells to identify novel drivers of resistance to combination endocrine therapy (tamoxifen) and CDK4/6 inhibitor (palbociclib) treatment. Screen hits were validated by CRISPR/Cas9 knockout models, mechanistic analyses and evaluation of patient samples. ResultsOur screens identified the inactivation of JNK signalling, including loss of the kinase MAP2K7, as a key driver of combination resistance. We developed multiple CRISPR/Cas9 knockout ER+ breast cancer cell lines (MCF-7 and T-47D) to investigate the effects of MAP2K7, MAPK8 and MAPK9 loss. MAP2K7 knockout increased metastatic burden in vivo and led to impaired JNK-mediated stress responses, as well as promoting cell survival and reducing senescence entry following endocrine therapy and CDK4/6 inhibitor treatment. Mechanistically, this occurred via loss of the AP-1 transcription factor c-JUN, leading to an attenuated response to combination endocrine therapy plus CDK4/6 inhibition. Furthermore, we analysed ER+ advanced breast cancer patient cohorts and found that inactivation of the JNK pathway was associated with increased metastatic burden, and low pJNKT183/Y185 activity correlated with a poorer response to systemic endocrine and CDK4/6 inhibitor therapies. ConclusionsOverall, we demonstrate that suppression of JNK signalling enables persistent growth during combined endocrine therapy and CDK4/6 inhibition. Furthermore, our data provide a pre-clinical rationale to screen patients tumours for JNK signalling deficiency prior to receiving combined endocrine therapy and CDK4/6 inhibition. STATEMENT OF TRANSLATIONAL RELEVANCEResistance to CDK4/6 inhibitors in the context of endocrine therapy resistance presents an urgent clinical challenge for the management of estrogen receptor positive (ER+) breast cancer. However, the mechanisms driving resistance to this therapeutic combination remain poorly understood. Here, we have identified inactivation of JNK signalling, specifically loss of the JNK kinase MAP2K7, as a major determinant of resistance to combined endocrine therapy and CDK4/6 inhibition in ER+ breast cancer. We uncovered that MAP2K7 loss augments tumour survival in vitro and in vivo. This occurs via disruption of activator protein-1 (AP-1) transcription factors, and thus prevents the induction of therapy-induced senescence and JNK-induced stress response. These findings reveal a critical tumour suppressor role for the JNK pathway in ER+ breast cancer, highlighting the importance of identifying patients with deficient JNK signalling and cautioning against the development of JNK inhibitors for this setting.

cancer biology↗

bioRxiv2024

PSKH1 kinase activity is differentially modulated via allosteric binding of Ca2+ sensor proteins

Protein Serine Kinase H1 (PSKH1) was recently identified as a crucial factor in kidney development and is overexpressed in prostate, lung and kidney cancers. However, little is known about PSKH1 regulatory mechanisms, leading to its classification as a "dark" kinase. Here, we used biochemistry and mass spectrometry to define PSKH1s consensus substrate motif, protein interactors, and how interactors, including Ca2+ sensor proteins, promote or suppress activity. Intriguingly, despite the absence of a canonical Calmodulin binding motif, Ca2+-Calmodulin activated PSKH1 while, in contrast, the ER-resident Ca2+ sensor of the CREC family, Reticulocalbin-3, suppressed PSKH1 catalytic activity. In addition to antagonistic regulation of the PSKH1 kinase domain by Ca2+ sensing proteins, we identified UNC119B as a protein interactor that activates PSKH1 via direct engagement of the kinase domain. Our findings identify complementary allosteric mechanisms by which regulatory proteins tune PSKH1s catalytic activity, and raise the possibility that different Ca2+ sensors may act more broadly to tune kinase activities by detecting and decoding extremes of intracellular Ca2+ concentrations.

biochemistry↗

bioRxiv2020

A cryptic tubulin-binding domain links MEKK1 to microtubule remodelling

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-{kappa}B 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. SIGNIFICANCE STATEMENTThe protein kinase MEKK1 activates stress response pathways in response to various cellular stressors, including chemotherapies that disrupt dynamics of the tubulin cytoskeleton. Filip[c]ik 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.

biochemistry↗

bioRxiv2019

Mechanism of preferential complex formation by Apoptosis Signal-regulating Kinases

Apoptosis signal-regulating kinases (ASK1-3) are activators of the P38 and JNK MAP kinase pathways. ASK1-3 form oligomeric complexes known as ASK signalosomes that initiate signalling cascades in response to diverse stress stimuli. Here we demonstrate that oligomerization of ASK proteins is driven by previously uncharacterised sterile-alpha motif (SAM) domains that reside at the C-terminus of each ASK protein. SAM domains from ASK1-3 have distinct behaviours: ASK1 forms unstable oligomers, ASK2 is predominantly monomeric, and the ASK3 SAM domain forms a stable oligomer even at low concentration. In contrast to their isolated behaviour, the ASK1 and ASK2 SAM domains preferentially form a stable heterocomplex. The crystal structure of the ASK3 SAM domain, small-angle X-ray scattering, and mutagenesis suggests that ASK3 oligomers and ASK1-ASK2 complexes form discrete quasi-helical rings, via the mid-loop-end-helix interface. Preferential ASK1-ASK2 binding is consistent with mass spectrometry showing that full-length ASK1 forms heterooligomeric complexes incorporating high levels of ASK2. Accordingly, disruption of SAM domain-association impairs ASK activity in the context of electrophilic stress induced by 4-hydroxy-2-nonenal. These findings provide a structural template for how ASK proteins assemble foci to drive inflammatory signalling, and reinforce that strategies targeting ASK kinases should consider the concerted actions of multiple ASK family members.

biochemistry↗