Figure S1. Landscape of somatic mutations and cytogenetics in Moffitt MF cohort. Figure S2. Effects of enforced MYC expression in HSCs in vivo. Figure S3. MYC-induced changes in hematopoietic sub-populations and colony forming potential. Figure S4. The JAK/STAT, PI3K/AKT, MEK/ERK, and alarmin pathways in MYC-driven MF.

Background Although inhibitors of mRNA translation are being evaluated as anti-cancer agents, the dynamics of protein synthesis throughout tumour progression are still poorly understood. Here we assess how alterations in mRNA translation during early tumorigenesis affect tumour development in KRAS-driven lung adenocarcinoma (LuAd).Methods We deployed autochthonous mouse models of LuAd driven by oncogenic KRASG12D combined with moderate overexpression of MYC and simultaneously manipulated mRNA translation by deleting the mRNA helicases eIF4A1 and eIF4A2 or by administering pharmacological inhibitors of protein synthesis, such as rapamycin. This permits synchronous assessment of LuAd initiation and progression in vivo and is amenable to parallel ex vivo culture of tumour-derived cells for detailed analysis of protein synthesis (using ribosome footprinting) and metabolic landscapes. These approaches also allowed us to perform multiplex imaging and spatial transcriptomics to characterise tumour formation in altered mRNA translation conditions and to compare results obtained in mice against the Lattice-A cohort of non-small cell lung cancer (NSCLC) patients.Results Deletion of the mRNA-translation repressor, eIF4A2 in KRAS-driven LuAd leads to a dysregulated protein synthesis landscape characterised by a strongly upregulated secretome, enlarged secretory compartments, increased oxidative metabolism and acquisition of senescence-like characteristics. Paradoxically, this overdriven secretory protein synthesis landscape delays tumorigenesis and leads to the appearance of clusters of non-proliferative, p21-positive KRASG12D-expressing cells in the lung. Consistently, reduction of mRNA translation with rapamycin in Eif4a2-deleted tumours suppresses senescence and restores tumorigenesis. Importantly, some Eif4a2 knockout cells overcome senescence to form tumours that exhibit enhanced MAP-kinase signalling and, in contrast to eIF4A2+/+ lesions, these were eradicated by administration of a MEK inhibitor. Consistently, MAP-kinase signalling was significantly increased in human NSCLC expressing low levels of eIF4A2.Conclusions Our study highlights that restraint of mRNA translation by eIF4A2 is critical in the early-stages of KRAS-driven LuAd to allow bypass of oncogene-induced senescence and tumour progression. Importantly, because tumours with dysregulated mRNA translation rely heavily on MAP-kinase signalling they are exquisitely sensitive to MEK inhibition, and this indicates the possibility that low expression of eIF4A2 could be used to identify potential responders to MEK inhibitors in clinical trials.

Comparison of gene expression profiles of Mx1-Cre+/-;Rosa26LSL-MYC/LSL-MYC vs. Mx1-Cre+/-;Rosa26+/+ mouse.

γδ T cells are important for host defense at the respiratory mucosa, acting directly or through interactions with other cells. However, how γδ T cells influence other immune cells in the lung remains unclear. Using a genetically engineered mouse model of lung cancer, we show that tumors drive expansion of both CD27+ and CD27- γδ T cells. Advanced microscopy techniques indicated that CD27- γδ T cells are enriched in tumors, whereas CD27+ γδ T cells are more prone to interact with macrophages in tumor-associated adventitial cuffs. SiglecFlow profibrotic airway macrophages were more prevalent in lung tumor-bearing mice than tumor-free mice. This profibrotic subset was reduced in lungs when the cancer model was crossed to Tcrd knockout mice or treated with Vγ1-depleting antibodies but not in TcrgV4/6 knockout mice. Thus, our findings implicate Vγ1 γδ T cells in driving tumor-associated airway macrophage functional imprinting. Determining the translatability to human health may offer new avenues for refining patient management and immunotherapeutic strategies.

Mosaic chromosomal alterations (mCAs) are a prevalent but poorly understood form of clonal hematopoiesis (CH). Whether mCAs contribute to disease independently of CHIP, and whether their large-scale genomic effects can be resolved to actionable targets, remain unknown. In 452,594 UK Biobank participants, we show that mCAs confer multimorbidity and mortality risk independent of CHIP. Notably, mCA-CHIP co-occurrence defines a very high-risk clonal state with synergistically elevated mortality, identifying a population not captured by CHIP screening alone. To resolve large mCAs to specific disease mechanisms, a cytoband-level mapping framework was developed that links mCAs to discrete genomic loci and candidate effector genes. Functional validation using single-cell transcriptomics and mouse models prioritized MYC (chr8 gain) and S100A9 (chr1 gain) as key drivers of systemic inflammation and multiorgan pathology. These findings establish mCAs as independent, synergistic, and genetically-resolvable drivers of age-related disease, with immediate implications for screening, risk stratification, and therapeutic development.

Figure S5. Changes in signaling in BM of trisomy 8+ TN-MF patient and MYC MF mice. Figure S6. Effects of silencing S100a9 in MYC-driven MF or overexpression of S100a9. Figure S7. Effects of inhibition of S100a9 or MYC in MYC-driven MF. Figure S8. Synthesis and Characterization of Tasquinimod. Figure S9. Synthesis and Characterization of MYCi975.

Table S1. Demographics of Moffitt Total Cancer Care MF patients. Table S2. Demographic Profile of trisomy 8+ TN-MF patients and HDs used in scRNA-seq analysis and PDX studies. Table S3. List of genes used for PROGENy analysis of human BM cells. Table S4. Demographics of TN-MF patients. Table S5. Clinical parameters of Mx1-Cre+/-;Rosa26LSL-MYC/LSL-MYC studies. Table S6. Clinical parameters of Scl-CreERT+/-;Rosa26LSL-MYC/LSL-MYC in vivo studies. Table S7. Clinical parameters in competitive transplant studies. Table S8. Comparison of gene expression profiles of Mx1-Cre+/-;Rosa26LSL-MYC/LSL-MYC vs. Mx1-Cre+/-;Rosa26+/+ mouse. Table S9. List of genes used for PROGENy analysis of mouse BM cells. Table S10. Cell-cell interaction analyses. Table S11. Clinical parameters of Mx1-Cre+/-;Rosa26LSL-MYC/LSL-MYC;S100a9-/- studies. Table S12. Clinical parameters of S100a9 transgenic mouse studies. Table S13. Clinical parameters of Tasquinimod efficacy studies in MYC-driven MF. Table S14. Clinical parameters of MYCi975 efficacy studies in MYC-driven MF. Table S15: Key Resources Table. Table S16: Sequence Information.

Abstract Background Inhalation of asbestos is the primary cause of Pleural Mesothelioma and is an acknowledged risk factor for all types of Lung Cancer. Persistence of asbestos fibres in the lungs and pleural cavity drives chronic inflammation, leading to the accumulation of driver mutations in exposed tissues and the eventual emergence of invasive malignant disease. Chronic inflammation additionally drives local immune suppression, likely contributing to poor responses to immunotherapy. The risk of Lung Cancer and Mesothelioma varies by type of asbestos, with Amphiboles (eg Amosite, Crocidolite) linked to greater risk of Mesothelioma than Serpentine (ie Chrysotile), whereas Chrysotile links to greater risk of Lung Cancer than Amphiboles. The vast majority of Asbestos imported into the UK since 1950 was Chrysotile, however, the contribution of asbestos to Lung Cancer in the UK has not been directly investigated. Methods We analysed data from Public Health Scotland (years 1998 to 2022) to determine if the geographic distribution of Lung Cancer incidence across Scotland is related to that of Mesothelioma, and used dose-controlled intrapleural or intratracheal delivery of asbestos fibres into genetically susceptible mice to determine the impact of defined asbestos exposures on Mesothelioma and Lung tumour development, respectively. Results Across the 17 Scottish Health Boards, standardised incidence rates for these cancers, averaged over 1998 to 2022, were directly correlated, with a Pearson Coefficient R = 0.848. By comparison, Mesothelioma showed no correlation with Pancreatic or Prostate Cancer incidence. Intrapleural injection of 25 ug Chrysotile or Amosite fibres resulted in similarly accelerated onset of morbidity in a genetically predisposed mouse model of Mesothelioma. Repeated oral delivery (5 × 10 ug) of Chrysotile fibres additionally accelerated tumour initiation and onset of morbidity in a mouse model of KRas-driven Lung Adenocarcinoma. Conclusions We conclude from these analyses that the contribution of asbestos to Lung Cancer in Scotland may be greatly underestimated.

Mesothelioma is a highly lethal and poorly biologically understood disease which presents diagnostic challenges due to its morphological complexity. This study uses self-supervised AI (Artificial Intelligence) to map the histomorphological landscape of the disease. The resulting atlas consists of recurrent patterns identified from 3446 Hematoxylin and Eosin (H&E) stained images scanned from resected tumour slides. These patterns generate highly interpretable predictions, achieving state-of-the-art performance with 0.65 concordance index (c-index) for outcomes and 88% AUC in subtyping. Their clinical relevance is endorsed by comprehensive human pathological assessment. Furthermore, we characterise the molecular underpinnings of these diverse, meaningful, predictive patterns. Our approach both improves diagnosis and deepens our understanding of mesothelioma biology, highlighting the power of this self-learning method in clinical applications and scientific discovery.

Although protein synthesis inhibitors are being evaluated as anti-cancer agents, the dynamics of mRNA translation in early tumorigenesis are still poorly understood. We report that deletion of the mRNA-translation repressor, eIF4A2 in early KRAS-driven lung adenocarcinoma leads to a dysregulated protein synthesis landscape characterised by a strongly upregulated secretome, enlarged secretory compartments, increased oxidative metabolism and acquisition of senescence-like characteristics. Paradoxically, this overdriven protein synthesis landscape delays tumorigenesis and leads to appearance of clusters of non-proliferative, p21-positive KRAS-expressing cells in the lung. Administration of rapamycin to reduce mRNA translation suppresses senescence and restores tumorigenesis following eIF4A2 deletion. Importantly, some eIF4A2 knockout cells overcome senescence to form tumours that exhibit enhanced MAP-kinase signalling and, in contrast to eIF4A2+/+ lesions, these may be eradicated by administration of a MEK inhibitor. Thus, dysregulated mRNA translation exposes a potential therapeutic vulnerability in KRAS-driven lung adenocarcinoma by forcing cancer cells to rely on MEK signalling. ### Competing Interest Statement The authors have declared no competing interest.

The development of standardised, reproducible preclinical models is essential for advancing pleural mesothelioma (PM) research. Here, we present a simple and reliable minimally invasive transthoracic intrapleural injection technique that could improve the efficiency of orthotopic PM model generation. By incorporating a simple needle sleeve to control the injection depth, this method eliminates the need for surgery or general anaesthesia, reducing technical complexity and animal stress while ensuring precise delivery into the pleural cavity. We demonstrate the effectiveness of this approach by achieving a 100% tumour engraftment rate following the injection of AE17 tumour cells. Additionally, this technique has been successfully used for asbestos fibre injection in mesothelioma models, highlighting its versatility. By providing a more accessible, standardised alternative to existing methods, this protocol improves the reliability of PM models and facilitates broader adoption by researchers, including those with limited experience in invasive procedures.

In recent years, alternative splicing emerged as a major mechanism controlling gene-regulatory networks during brain development, yet how alternative splicing is tuned to the dynamic alterations underlying neuronal maturation remains poorly understood. Here, we identified that NUAK1, an AMPK-related kinase linked to neurodevelopmental disorders, is a key regulator of alternative splicing in developing cortical neurons. Mechanistically, NUAK1 exerts its function through phosphorylation of the splicing co-factor SON, regulating a group of highly conserved splicing events in genes crucial for neurodevelopment. We demonstrate that SON plays an important role in cortical neuron development, which is consistent with the neurodevelopmental phenotypes observed in Zhu-Tokita-Takenouchi-Kim (ZTTK) syndrome, a genetic disorder caused by SON haploinsufficiency. Together, our findings uncover a novel pathway involving NUAK1 and SON, which orchestrate a splicing program required for proper neuronal development. ### Competing Interest Statement The authors have declared no competing interest. Association Francaise contre les Myopathies, https://ror.org/0162y2387, Myoneuralp2 European Research Council, https://ror.org/0472cxd90, 678302-NEUROMET European Commission, Marie Sklodowska Curie Action fellowship 101110819 Fondation pour la Recherche Médicale, https://ror.org/04w6kn183, SPF202110014126 Agence Nationale de la Recherche, https://ror.org/00rbzpz17, ANR-11-LABX-0042, ANR-11-IDEX-0007

Hepatocellular carcinoma (HCC), the most common form of primary liver cancer, is a leading cause of cancer-related mortality worldwide1,2. HCC occurs typically from a background of chronic liver disease, caused by a spectrum of predisposing conditions. Tumour development is driven by the expansion of clones that accumulate progressive driver mutations3, with hepatocytes the most likely cell of origin2. However, the landscape of driver mutations in HCC is broadly independent of the underlying aetiologies4. Despite an increasing range of systemic treatment options for advanced HCC, outcomes remain heterogeneous and typically poor. Emerging data suggest that drug efficacies depend on disease aetiology and genetic alterations5,6. Exploring subtypes in preclinical models with human relevance will therefore be essential to advance precision medicine in HCC7. Here we generated a suite of genetically driven immunocompetent in vivo and matched in vitro HCC models. Our models represent multiple features of human HCC, including clonal origin, histopathological appearance and metastasis. We integrated transcriptomic data from the mouse models with human HCC data and identified four common human-mouse subtype clusters. The subtype clusters had distinct transcriptomic characteristics that aligned with the human histopathology. In a proof-of-principle analysis, we verified response to standard-of-care treatment and used a linked in vitro-in vivo pipeline to identify a promising therapeutic candidate, cladribine, that has not previously been linked to HCC treatment. Cladribine acts in a highly effective subtype-specific manner in combination with standard-of-care therapy.

Mitochondrial outer membrane permeabilisation (MOMP) is often essential for apoptosis, by enabling cytochrome c release that leads to caspase activation and rapid cell death. Recently, MOMP has been shown to be inherently pro-inflammatory with emerging cellular roles, including its ability to elicit anti-tumour immunity. Nonetheless, how MOMP triggers inflammation and how the cell regulates this remains poorly defined. We find that upon MOMP, many proteins localised either to inner or outer mitochondrial membranes are ubiquitylated in a promiscuous manner. This extensive ubiquitylation serves to recruit the essential adaptor molecule NEMO, leading to the activation of pro-inflammatory NF-κB signalling. We show that disruption of mitochondrial outer membrane integrity through different means leads to the engagement of a similar pro-inflammatory signalling platform. Therefore, mitochondrial integrity directly controls inflammation, such that permeabilised mitochondria initiate NF-κB signalling.

Abstract Despite advances in understanding the genetic abnormalities in myeloproliferative neoplasms (MPN) and the development of JAK2 inhibitors, there is an urgent need to devise new treatment strategies, particularly for patients with triple-negative (TN) myelofibrosis (MF) who lack mutations in the JAK2 kinase pathway and have very poor clinical outcomes. Here we report that MYC copy number gain and increased MYC expression frequently occur in TN-MF and that MYC-directed activation of S100A9, an alarmin protein that plays pivotal roles in inflammation and innate immunity, is necessary and sufficient to drive development and progression of MF. Notably, the MYC-S100A9 circuit provokes a complex network of inflammatory signaling that involves numerous hematopoietic cell types in the bone marrow microenvironment. Accordingly, genetic ablation of S100A9 or treatment with small molecules targeting the MYC-S100A9 pathway effectively ameliorates MF phenotypes, highlighting the MYC–alarmin axis as a novel therapeutic vulnerability for this subgroup of MPNs. Significance: This study establishes that MYC expression is increased in TN-MPNs via trisomy 8, that a MYC-S100A9 circuit manifest in these cases is sufficient to provoke myelofibrosis and inflammation in diverse hematopoietic cell types in the BM niche, and that the MYC-S100A9 circuit is targetable in TN-MPNs.

NUAK1 and NUAK2 belong to a family of kinases related to the catalytic α-subunits of the AMP-activated protein kinase (AMPK) complexes. Despite canonical activation by the tumour suppressor kinase LKB1, both NUAKs exhibit a spectrum of activities that favour tumour development and progression. Here, we review similarities in structure and function of the NUAKs, their regulation at gene, transcript and protein level, and discuss their phosphorylation of specific downstream targets in the context of the signal transduction pathways and biological activities regulated by each or both NUAKs.

Investigating lung cancer using genetically engineered mouse models (GEMMS) Daniel J. Murphy, Professor of Lung Cancer & Mesothelioma at the University of Glasgow, School of Cancer Sciences, discusses opportunities for improving cancer research and care through the use of genetically engineered mouse models. Despite changes in smoking habits, cancers of the lung remain the most lethal types of cancer in the UK and worldwide. In the UK, lung cancer accounts for over 20% of all deaths from cancer, claiming almost 35,000 victims annually (2017-2019 CRUK Lung Cancer Statistics). Until very recently, the vast majority of lung cancers were diagnosed at a late stage – meaning the cancers were very well established and at their most aggressive. Few of these cases respond effectively to treatment; when they do, responses are often short-lived.

Mitochondria form a critical control nexus which are essential for maintaining correct tissue homeostasis. An increasing number of studies have identified dysregulation of mitochondria as a driver in cancer. However, which pathways support and promote this adapted mitochondrial function? A key hallmark of cancer is perturbation of kinase signalling pathways. These pathways include mitogen activated protein kinases (MAPK), lipid secondary messenger networks, cyclic-AMP-activated (cAMP)/AMP-activated kinases (AMPK), and Ca2+/calmodulin-dependent protein kinase (CaMK) networks. These signalling pathways have multiple substrates which support initiation and persistence of cancer. Many of these are involved in the regulation of mitochondrial morphology, mitochondrial apoptosis, mitochondrial calcium homeostasis, mitochondrial associated membranes (MAMs), and retrograde ROS signalling. This review will aim to both explore how kinase signalling integrates with these critical mitochondrial pathways and highlight how these systems can be usurped to support the development of disease. In addition, we will identify areas which require further investigation to fully understand the complexities of these regulatory interactions. Overall, this review will emphasize how studying the interaction between kinase signalling and mitochondria improves our understanding of mitochondrial homeostasis and can yield novel therapeutic targets to treat disease.