Biology subjects

Clemons, N. J.

Publications and source records attributed to Clemons, N. J..

8 recordsLinked to original sources

bioRxiv2026

Mapping Functional Tumor Suppressor Networks in Esophageal Adenocarcinoma Using In Vivo CRISPR Screening and Perturb-sequencing

Esophageal adenocarcinoma (EAC) is a genetically heterogeneous malignancy with few recurrent drivers, limiting effective targeted therapies. Although EAC arises from Barretts esophagus (BE), mechanisms driving progression from this premalignant state to invasive cancer remain unclear. We combined pooled CRISPR-Cas9 loss-of-function screening, in vivo tumorigenicity assays, and Perturb-seq profiling to define functional drivers of BE transformation. We identified 37 tumor suppressors whose loss promotes progression to EAC, defining a functional landscape of tumor initiation. Despite genetic diversity, these losses converged on four transcriptional programs involving metabolic reprogramming, cell cycle progression, RNA processing, and cellular motility. Furthermore, we identify loss of NIPBL, TGFBR2, and RPL22 as key mediators of resistance to platinum- and taxane-based chemotherapy. Collectively, these findings provide a unifying framework for genomic heterogeneity in EAC, uncover underappreciated tumor suppressor pathways, and establish a resource to guide mechanistic and translational studies aimed at improving treatment strategies in this aggressive cancer.

cancer biology↗

bioRxiv2026

SMAD4 loss drives chromosomal instability during tumourigenesis via translational reprogramming

Chromosomal instability (CIN), arising from errors in chromosome segregation during cell division, is a hallmark of cancer. Whilst CIN can result from several mitotic defects, the mechanisms that initiate CIN to drive tumourigenesis remain incompletely understood. Here, we show that loss of SMAD4 reprograms translation to induce CIN, resulting in tumour formation. Multi-omics analysis of tumourigenesis models driven by loss of SMAD4 complemented by functional studies demonstrate that loss of SMAD4 in pre-neoplastic cells leads to dysfunctional mitosis and an altered global translation landscape. We show that CDK11B is translationally downregulated in SMAD4-\- cells, and re-expression of the mitosis-specific isoform of this protein (CDK11B-p58) rescues the mitotic defects. Analysis of patient tumours reveals a strong correlation between markers of CIN and SMAD4 status, indicating the clinical relevance of this phenotype. Collectively, we reveal a previously unrecognised role for SMAD4 as a gatekeeper for CIN-mediated tumourigenesis via regulation of translation.

cancer biology↗

bioRxiv2026

Systematic Evaluation Defines the Limits of Ferroptosis in Cancer Therapy

Ferroptosis is a cell death mechanism characterized by the accumulation of iron-catalyzed lipid peroxides in membrane lipid acyl chains and subsequent loss of membrane integrity.1 Despite thorough investigation of its mechanisms in cultured cells, induction of ferroptosis has unresolved clinical utility in cancer therapy. Here, we systematically evaluate ferroptosis induction via multiple mechanisms, in both cell and tumor models, using focused genetic screens, genetic loss-of-function systems, and pharmacological perturbations. Through this analysis we identify cancer cell line subsets with distinct responses to canonical ferroptosis inducers and suppressors and define the underpinnings of each. Inhibition of central in vitro ferroptosis suppressors GPX4, GCLC, or SLC7A11 across these multiple models fails to impact established tumor growth. In contrast, deficiency in the cytosolic thioredoxin reductase and pharmacologic GCLC inhibition potently induces tumor regression and triggers a form of non-ferroptotic cell death regulated by cystine availability and translation. These analyses further reveal that the principal essential function of environmental cystine in cultured cells is to support selenoprotein function, identified through investigating our finding that {beta}-mercaptoethanol supports exponential growth in cystine-free conditions. Thus, while ferroptosis activation may be efficacious alone or in combination with other therapies in specific tumor contexts, cell culture systems greatly overestimate the potential anti-cancer effects of ferroptosis induction via the GPX4 axis.

cancer biology↗

bioRxiv2023

Caspase-2 protects against ferroptotic cell death

Caspase-2, one of the most evolutionarily conserved member of the caspase family, is an important regulator of the cellular response to oxidative stress. Given that ferroptosis is suppressed by antioxidant defense pathways, such as that involving selenoenzyme glutathione peroxidase 4 (GPX4), we hypothesised that caspase-2 may play a role in regulating ferroptosis. This study provides the first demonstration of an important and unprecedented function of caspase-2 in protecting cancer cells from undergoing ferroptotic cell death. Specifically, we show that depletion of caspase-2 leads to downregulation of stress response genes including SESN2, HMOX1, SLC7A11 and sensitises mutant-p53 cancer cells to cell death induced by various ferroptosis inducing compounds. Importantly, the canonical catalytic activity of caspase-2 is not required for its role and suggests that caspase-2 regulates ferroptosis via non-proteolytic interaction with other proteins. Using an unbiased BioID proteomics screen, we identified novel caspase-2 interacting proteins (including heat shock proteins and co-chaperones) that regulate cellular responses to stress. Finally, we demonstrate that caspase-2 limits chaperone mediated autophagic degradation of GPX4 to promote survival of mutant-p53 cancer cells. In conclusion, we document a novel role for caspase-2 as a negative regulator of ferroptosis in cells with mutant-p53. Our results provide evidence for a novel function of caspase-2 functions in cell death regulation and open potential new avenues to exploit ferroptosis in cancer therapy.

cancer biology↗

bioRxiv2021

High-content profiling reveals a unified model of copper ionophore dependent cell death in oesophageal adenocarcinoma

Background and AimsOesophageal adenocarcinoma (OAC) is of increasing global concern due to increasing incidence, a lack of effective treatments, and poor prognosis. Therapeutic target discovery and clinical trials have been hindered by the heterogeneity of the disease, lack of driver mutations, and the dominance of large-scale genomic rearrangements. In this work we have characterised three potent and selective hit compounds identified in an innovative high-content phenotypic screening assay. The three hits include two approved drugs; elesclomol and disulfiram, and another small molecule compound, ammonium pyrrolidinedithiocarbamate. We uncover their mechanism of action, discover a targetable vulnerability, and gain insight into drug sensitivity for biomarker-based clinical trials in OAC. MethodsElesclomol, disulfiram, and ammonium pyrrolidinedithiocarbamate were systematically characterised across panels of oesophageal cell lines and patient-derived organoids. Drug treated oesophageal cell lines were morphologically profiled using a high-content, imaging platform. Compounds were assessed for efficacy across patient-derived organoids. Metabolomics and transcriptomics were assessed for the identification of oesophageal-cancer specific drug mechanisms and patient stratification hypotheses. ResultsHigh-content profiling revealed that all three compounds were highly selective for OAC over tissue-matched controls. Comparison of gene expression and morphological signatures unveiled a unified mechanism of action involving the accumulation of copper selectively in cancer cells, leading to dysregulation of proteostasis and cancer cell death. Basal omic analyses revealed proteasome and metabolic markers of drug sensitivity, forming the basis for biomarker-based clinical trials in OAC. ConclusionsIntegrated analysis of high-content imaging, transcriptomic and metabolomic data has revealed a new therapeutic mechanism for the treatment of OAC and represents an alternative target-agnostic drug discovery strategy.

cancer biology↗

bioRxiv2020

Genome-wide CRISPR screens reveal APR-246 (Eprenetapopt) triggers ferroptosis and inhibits iron-sulfur cluster biogenesis

The mechanisms by which cells respond and adapt to oxidative stress are largely unknown but are key to developing a rationale for cancer therapies that target antioxidant pathways. APR-246 is a mutant-p53 targeted therapeutic currently under clinical investigation in myeloid dysplastic syndrome (MDS) and acute myeloid leukemia1. Whilst the mechanism of action of APR-246 is thought to be reactivation of wild-type p53 activity through covalent modification of cysteine residues in the core domain of mutant-p53 protein2,3, here we report that the anti-neoplastic capacity of APR-246 lies predominantly in the conjugation of free cysteine. Genome-wide CRISPR perturbation screening, metabolite profiling and proteomics in response to APR-246 treatment in mutant-p53 cancer cells highlighted the role of GSH and mitochondrial metabolism in determining APR-246 efficacy. APR-246 sensitivity was increased through loss of key enzymes in mitochondrial one-carbon metabolism, SHMT2 and MTHFD1L, due to diminished glycine supply for de novo GSH synthesis. Critically, we show that APR-246 induces iron-dependent, apoptotic machinery-independent cell death, ferroptosis. Whole-cell proteomics analyses indicated an upregulation of proteins involved in iron-sulfur cluster biogenesis (eg. FDX1). GSH, acetyl-CoA and NADH levels were also depleted in APR-246 treated cells. Importantly, we found that APR-246 inhibits iron-sulfur cluster biogenesis in the mitochondria of cancer cells through cysteine conjugation. This work not only details novel determinants of APR-246 activity in cancer cells, but also provides a clinical roadmap for targeting antioxidant pathways in tumours - beyond targeting mutant-p53 tumours. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=104 SRC="FIGDIR/small/398867v1_ufig1.gif" ALT="Figure 1"> View larger version (16K): org.highwire.dtl.DTLVardef@11155f8org.highwire.dtl.DTLVardef@1231cb7org.highwire.dtl.DTLVardef@324881org.highwire.dtl.DTLVardef@136146f_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2020

SLC7A11 is a superior determinant of APR-246 (Eprenetapopt) response than TP53 mutation status

PurposeAPR-246 (Eprenetapopt) is in clinical development with a focus on haematological malignancies and is marketed as a mutant-p53 reactivation therapy. Currently, the detection of at least one TP53 mutation is an inclusion criterion for patient selection into most clinical trials. Preliminary results from our phase Ib/II clinical trial investigating APR-246 combined with combination chemotherapy (cisplatin and 5-Fluorouracil) in metastatic oesophageal cancer, together with previous pre-clinical studies, indicate that TP53 mutation status alone may not be a sufficient biomarker for response to APR-246. This study aimed to identify a robust biomarker for response to APR-246. MethodsCorrelation analysis of the PRIMA-1 activity (lead compound to APR-246) with mutational status, gene expression, protein expression and metabolite abundance across over 800 cancer cell lines was performed. Functional validation and a boutique siRNA screen of over 750 redox-related genes were also conducted. ResultsTP53 mutation status was not predictive of response to APR-246. The expression of SLC7A11, the cystine/glutamate transporter, was identified as a superior determinant of response to APR-246. Genetic regulators of SLC7A11, including ATF4, MDM2, wild-type p53 and c-Myc were confirmed to also regulate cancer cell sensitivity to APR-246. ConclusionsSLC7A11 expression is the major determinant of sensitivity to APR-246 and should be utilised as a predictive biomarker in future clinical investigation of APR-246.

cancer biology↗

bioRxiv2019

Neuronal-specific function of hTim8a in Complex IV assembly provides insight into the molecular mechanism underlying Mohr-Tranebjaerg syndrome

Human Tim8a is a member of an intermembrane space chaperone network, known as the small TIM family, which transport hydrophobic membrane proteins through this compartment. Mutations in TIMM8A cause a neurodegenerative disease, Mohr-Tranebjaerg syndrome (MTS), which is characterised by sensorineural hearing loss, dystonia and blindness. Nothing is known about the function of hTim8a in neuronal cells and consequently how lack of hTim8a leads to a neurodegenerative disease. We identified a novel cell-specific function of hTim8a in the assembly of Complex IV, which is mediated through a transient interaction with the copper chaperone COX17. Complex IV assembly defects in cells lacking hTim8a leads to oxidative stress and changes to key apoptotic regulators, including cytochrome c and Bax, which primes cells for cell death. Alleviation of oxidative stress using Vitamin E rescues cells from apoptotic vulnerability. We hypothesis that enhanced sensitivity of neuronal cells to apoptosis is the underlying mechanism of MTS.

cell biology↗