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Pancreatic cancer is a highly lethal malignancy often presenting with advanced disease and characterized by resistance to standard chemotherapy. Immune-based therapies such checkpoint inhibition have been largely ineffective such that pancreatic cancer is categorized as an immunologically "cold tumor". In the present study, we examine the therapeutic efficacy of a personalized cancer vaccine in which tumor cells are fused with dendritic cells (DC) resulting in the broad induction of antitumor immunity.In the KPC spontaneous pancreatic cancer murine model, we demonstrated that vaccination with DC/KPC fusions led to expansion of pancreatic cancer specific lymphocytes with an activated phenotype. Remarkably, vaccination led to a reduction in tumor bulk and near doubling of median survival in this highly aggressive model. In a second murine pancreatic model (Panc02), vaccination with DC/tumor fusions similarly led to expansion of tumor antigen specific lymphocytes and their infiltration to the tumor site. Having shown efficacy in immunocompetent murine models, we subsequently demonstrated that DC/tumor fusions generated from primary human pancreatic cancer and autologous DCs potently stimulate tumor specific cytotoxic lymphocyte responses.DC/tumor fusions induce the activation and expansion of tumor reactive lymphocytes with the capacity to infiltrate into the pancreatic cancer tumor bed.

Animal models are critical for the preclinical validation of cancer immunotherapies. Unfortunately, mouse breast cancer models do not faithfully reproduce the molecular subtypes and immune environment of the human disease. In particular, there are no good murine models of estrogen receptor–positive (ER+) breast cancer, the predominant subtype in patients. Here, we show that Nitroso-N-methylurea–induced mammary tumors in outbred Sprague-Dawley rats recapitulate the heterogeneity for mutational profiles, ER expression, and immune evasive mechanisms observed in human breast cancer. We demonstrate the utility of this model for preclinical studies by dissecting mechanisms of response to immunotherapy using combination TGFBR inhibition and PD-L1 blockade. Short-term treatment of early-stage tumors induced durable responses. Gene expression profiling and spatial mapping classified tumors as inflammatory and noninflammatory, and identified IFNγ, T-cell receptor (TCR), and B-cell receptor (BCR) signaling, CD74/MHC II, and epithelium-interacting CD8+ T cells as markers of response, whereas the complement system, M2 macrophage phenotype, and translation in mitochondria were associated with resistance. We found that the expression of CD74 correlated with leukocyte fraction and TCR diversity in human breast cancer. We identified a subset of rat ER+ tumors marked by expression of antigen-processing genes that had an active immune environment and responded to treatment. A gene signature characteristic of these tumors predicted disease-free survival in patients with ER+ Luminal A breast cancer and overall survival in patients with metastatic breast cancer receiving anti–PD-L1 therapy. We demonstrate the usefulness of this preclinical model for immunotherapy and suggest examination to expand immunotherapy to a subset of patients with ER+ disease.See related Spotlight by Roussos Torres, p. 672

Supplementary Figure from Insights into Immune Escape During Tumor Evolution and Response to Immunotherapy Using a Rat Model of Breast Cancer

Supplementary Table from Insights into Immune Escape During Tumor Evolution and Response to Immunotherapy Using a Rat Model of Breast Cancer

Supplementary Tables

Supplemental Figure 2. A) Relative expression Left panel: Mean of fluorescence and right panel: qPCR validation of miR-21-5p expression level in the set of cell lines, 3D-models and organoid compared to normal controls. B) qPCR analysis of PDCD4 and PTEN expression levels in the PANC1 stable cell lines (Lenti-21) inhibiting miR- 21 activity (pLenti-III-miR-off). C) qRT-PCR of miR-21-5p expression after lipofectamine transfection of PANC1, BxPC3 and PL-45 cells with anti-miR-21 inhibitor (mirVana ®) at the dose of 50nM after 48h. For qPCR each miRNA sample was normalized on the basis of it's 18s content and on the basis of GAPDH for mRNAs.

Supplemental Figure 3. A) iRGD-TAMRA (red) binding after 15 min at 4 degree in normal cells (Low integrins-NRP1) and in human and mice PDAC cells (High integrins-NRP1). Scale bars 200μm B) qRT-PCR of miR-21-5p expression after treatment of PANC1, BxPC3 and PL-45 cells with TPN-21 at the dose of 100nM after 48h. C) Representation of 3D-model treatment course with TPN-21. For qPCR each miRNA sample was normalized on the basis of its 18s content. Error bars, mean {plus minus} s.d *P = 0.01-0.05; **P = 0.001-0.01; ***P < 0.001; ****P < 0.0001 N.S., not significant, two-tailed t-test; n = 3 biological replicates.

Supplemental Figure 1. A) Left panel: Visualization of the miRNA profile of normal pancreas samples (n=3 control). Right panel: Visualization of the miRNA profile of PDAC PDX tumor samples (n=27 cases). Relative expression is shown as mean of fluorescence (MFI). miRNAs profiling was done through Firefly Circulating miRNA Assay and normalized using two miRNAs that are not significantly deregulated between all samples miR-22-3p and miR-30b-5p defined by the geNorm-like 37 algorithm. B) Visualization of miRNA profiles of PDAC cell lines (n=4), PDAC 3Dmodel (n=2) and Patient-derived-organoid (PDO 286 and PDO 281) compared to normal pancreatic cell and organoid (n=2 control). miRNAs profiling was done via Firefly Circulating miRNA Assay and normalized using miR-181b-5p, miR-103-3p, miR-30b-5p defined by the geNorm-like algorithm. Normalized miRNA signal intensities are presented as fold-change (log10 of the ratio between a probe value to the average of all the other samples for that probe). Green represents highly expressed miRNAs and red represents lowly expressed miRNAs.

Purpose: Since drug responses vary between patients, it is crucial to develop pre-clinical or co-clinical strategies that forecast patient response. In this study, we tested whether RNA-based therapeutics were suitable for personalized medicine by using patient-derived-organoid (PDO) and patient-derived-xenograft (PDX) models.Experimental Design: We performed microRNA (miRNA) profiling of PDX samples to determine the status of miRNA deregulation in individual pancreatic ductal adenocarcinoma (PDAC) patients. To deliver personalized RNA-based-therapy targeting oncogenic miRNAs that form part of this common PDAC miRNA over-expression signature, we packaged antimiR oligonucleotides against one of these miRNAs in tumor-penetrating nanocomplexes (TPN) targeting cell surface proteins on PDAC tumors.Results: As a validation for our pre-clinical strategy, the therapeutic potential of one of our nano-drugs, TPN-21, was first shown to decrease tumor cell growth and survival in PDO avatars for individual patients, then in their PDX avatars.Conclusions: This general approach appears suitable for co-clinical validation of personalized RNA medicine and paves the way to prospectively identify patients with eligible miRNA profiles for personalized RNA-based therapy. Clin Cancer Res; 24(7); 1734–47. ©2018 AACR.

Supplemental legend

Supplemental Figure 4. A) qPCR analysis of miR-21-5p expression level after 4 treatments with TPN-21 in PDO 286. B) Relative tumor burden after (4 I.V injection of PBS (n=6) TPN-control n=6 or TPN-21 n=5 (5mg/kg). For qPCR each miRNA sample was normalized on the basis of its 18s content.

Supplemental table 1A List of miRNAs, miRBase accession number and mature sequence of 46 selected-miRNAs used to build the Firefly 46plex circulating custom panel. There miRNAs have previously been linked to pancreatic cancer or to KRAS.

Supplemental table 1B List of PDX pancreatic samples. Information regarding KRAS-mutation and gemcitabine resistance profile. S for sensitive, R for Resistant.

Supplemental Figure 5. A) Representatives PDO 286 images and metabolic activity measured from MTT assay after repeated TPN-21 or gemcitabine treatments (100nM) or both treatments. Scale bars 1000μm. B) Quantification of organoid size after treatments. Organoids size was measure through ImageJ software on a total of ⩾ 35 spheroids and are presented in arbitrary units (UA). Error bars, mean {plus minus} s.d *P = 0.01- 0.05; **P = 0.001-0.01; ***P < 0.001; ****P < 0.0001 N.S., not significant, two-tailed ttest; n = 3 biological replicates.

Supplementary Data from Organoid Sensitivity Correlates with Therapeutic Response in Patients with Pancreatic Cancer

Purpose:Pancreatic ductal adenocarcinoma (PDAC) remains a significant health issue. For most patients, there are no options for targeted therapy, and existing treatments are limited by toxicity. The HOPE trial (Harnessing Organoids for PErsonalized Therapy) was a pilot feasibility trial aiming to prospectively generate patient-derived organoids (PDO) from patients with PDAC and test their drug sensitivity and correlation with clinical outcomes.Experimental Design:PDOs were established from a heterogeneous population of patients with PDAC including both basal and classical PDAC subtypes.Results:A method for classifying PDOs as sensitive or resistant to chemotherapy regimens was developed to predict the clinical outcome of patients. Drug sensitivity testing on PDOs correlated with clinical responses to treatment in individual patients.Conclusions:These data support the investigation of PDOs to guide treatment in prospective interventional trials in PDAC.

Supplementary Figure 3 from Epithelial Cell Organization Suppresses Myc Function by Attenuating Myc Expression

PDF file - 103K, Mislocalization of SCRIB enhances EGF-induced AKT activation.

PDF file - 187K, H&E staining of hyperplasia and tumors from hSCRIBP305L mice.

Supplementary Figure 1 from Epithelial Cell Organization Suppresses Myc Function by Attenuating Myc Expression

Supplementary Figure 6 from Epithelial Cell Organization Suppresses Myc Function by Attenuating Myc Expression

PDF file - 68K

c-Myc is an oncogene transcription factor that causes cancer in many settings, including solid tumors that arise in the context of organized tissue structures. Given that disruption of tissue architecture frequently occurs in cancer, there is considerable interest in how cell organization impacts oncogene function. A previous report found that organization of mammary epithelial cells into defined 3-dimensional structures renders them insensitive to the effects of retrovirus-mediated overexpression of Myc, leading to the notion that organization tempers the sensitivity of individual cells to Myc activity. In this article, we report that epithelial cell organization does not profoundly alter Myc activity but, instead, suppresses Myc by modulating its expression. We show that the morphogenesis of mammary epithelial cells into organized acinar structures in vitro is accompanied by widespread changes in gene expression patterns, including a substantial decrease in the expression of Myc. Concomitant with the decrease in endogenous Myc expression, we observe a decrease in transcription from retroviral vectors during morphogenesis and find that Myc transgene expression in acini is much lower than in unorganized cells. This decrease in Myc transgene activity is responsible for the apparent recalcitrance of organized cells to ectopic Myc, as adenovirus-mediated expression of Myc in organized structures potently induces apoptosis. These observations reveal that organization does not alter the inherent response of epithelial cells to Myc and suggest that other tumor suppression mechanisms, apart from structure, antagonize Myc in the development of solid tumors. Cancer Res; 71(11); 3822–30. ©2011 AACR.

Supplementary Figure 4 from Epithelial Cell Organization Suppresses Myc Function by Attenuating Myc Expression

Supplementary Figure 5 from Epithelial Cell Organization Suppresses Myc Function by Attenuating Myc Expression

Supplementary Tables 1-5 from Epithelial Cell Organization Suppresses Myc Function by Attenuating Myc Expression

The polarity protein complex Par6/atypical protein kinase (aPKC)/Cdc42 regulates polarization processes during epithelial morphogenesis, astrocyte migration, and axon specification. We, as well as others, have shown that this complex is also required for disruption of apical-basal polarity during the oncogene ErbB2-induced transformation and transforming growth factor β–induced epithelial-mesenchymal transition of mammary epithelial cells. Here, we report that expression of Par6 by itself in mammary epithelial cells induces epidermal growth factor–independent cell proliferation and development of hyperplastic three-dimensional acini without affecting apical-basal polarity. This is dependent on the ability of Par6 to interact with aPKC and Cdc42, but not Lgl and Par3, and its ability to promote sustained activation of MEK/ERK signaling. Down-regulation of Cdc42 or aPKC expression suppresses the ability of Par6 to induce proliferation, demonstrating that Par6 promotes cell proliferation by interacting with aPKC and Cdc42. We also show that Par6 is overexpressed in breast cancer–derived cell lines and in both precancerous breast lesions and advanced primary human breast cancers, suggesting that Par6 overexpression regulates tumor initiation and progression. Thus, in addition to regulating cell polarization processes, Par6 is an inducer of cell proliferation in breast epithelial cells. [Cancer Res 2008;68(20):8201–9]

Supplementary Methods, Figure and Table Legends from Epithelial Cell Organization Suppresses Myc Function by Attenuating Myc Expression

Supplementary Figure 2 from Epithelial Cell Organization Suppresses Myc Function by Attenuating Myc Expression

Supplementary Figures 1-6 from The Polarity Protein Par6 Induces Cell Proliferation and Is Overexpressed in Breast Cancer

PDF file - 99K, Expression of SCRIB mRNA in human breast cancer.

PDF file - 204K, Characterization of hSCRIBP305L tumors.

Pancreatic cancer has the worst prognosis of all common tumors. Earlier cancer diagnosis could increase survival rates and better assessment of metastatic disease could improve patient care. As such, there is an urgent need to develop biomarkers to diagnose this deadly malignancy earlier. Analyzing circulating extracellular vesicles (cEVs) using ‘liquid biopsies’ offers an attractive approach to diagnose and monitor disease status. However, it is important to differentiate EV-associated proteins enriched in patients with pancreatic ductal adenocarcinoma (PDAC) from those with benign pancreatic diseases such as chronic pancreatitis and intraductal papillary mucinous neoplasm (IPMN). To meet this need, we combined the novel EVtrap method for highly efficient isolation of EVs from plasma and conducted proteomics analysis of samples from 124 individuals, including patients with PDAC, benign pancreatic diseases and controls. On average, 912 EV proteins were identified per 100µL of plasma. EVs containing high levels of PDCD6IP, SERPINA12 and RUVBL2 were associated with PDAC compared to the benign diseases in both discovery and validation cohorts. EVs with PSMB4, RUVBL2 and ANKAR were associated with metastasis, and those with CRP, RALB and CD55 correlated with poor clinical prognosis. Finally, we validated a 7-EV protein PDAC signature against a background of benign pancreatic diseases that yielded an 89% prediction accuracy for the diagnosis of PDAC. To our knowledge, our study represents the largest proteomics profiling of circulating EVs ever conducted in pancreatic cancer and provides a valuable open-source atlas to the scientific community with a comprehensive catalogue of novel cEVs that may assist in the development of biomarkers and improve the outcomes of patients with PDAC.

Purpose: Pancreatic ductal adenocarcinoma (PDAC) remains a significant health issue. For most patients, there are no options for targeted therapy, and existing treatments are limited by toxicity. The HOPE trial (Harnessing Organoids for PErsonalized Therapy) was a pilot feasibility trial aiming to prospectively generate patient-derived organoids (PDO) from patients with PDAC and test their drug sensitivity and correlation with clinical outcomes. Experimental Design: PDOs were established from a heterogeneous population of patients with PDAC including both basal and classical PDAC subtypes. Results: A method for classifying PDOs as sensitive or resistant to chemotherapy regimens was developed to predict the clinical outcome of patients. Drug sensitivity testing on PDOs correlated with clinical responses to treatment in individual patients. Conclusions: These data support the investigation of PDOs to guide treatment in prospective interventional trials in PDAC.

Animal models are critical for the preclinical validation of cancer immunotherapies. Unfortunately, mouse breast cancer models do not faithfully reproduce the molecular subtypes and immune environment of the human disease. In particular, there are no good murine models of estrogen receptor–positive (ER+) breast cancer, the predominant subtype in patients. Here, we show that Nitroso-N-methylurea–induced mammary tumors in outbred Sprague-Dawley rats recapitulate the heterogeneity for mutational profiles, ER expression, and immune evasive mechanisms observed in human breast cancer. We demonstrate the utility of this model for preclinical studies by dissecting mechanisms of response to immunotherapy using combination TGFBR inhibition and PD-L1 blockade. Short-term treatment of early-stage tumors induced durable responses. Gene expression profiling and spatial mapping classified tumors as inflammatory and noninflammatory, and identified IFNγ, T-cell receptor (TCR), and B-cell receptor (BCR) signaling, CD74/MHC II, and epithelium-interacting CD8+ T cells as markers of response, whereas the complement system, M2 macrophage phenotype, and translation in mitochondria were associated with resistance. We found that the expression of CD74 correlated with leukocyte fraction and TCR diversity in human breast cancer. We identified a subset of rat ER+ tumors marked by expression of antigen-processing genes that had an active immune environment and responded to treatment. A gene signature characteristic of these tumors predicted disease-free survival in patients with ER+ Luminal A breast cancer and overall survival in patients with metastatic breast cancer receiving anti–PD-L1 therapy. We demonstrate the usefulness of this preclinical model for immunotherapy and suggest examination to expand immunotherapy to a subset of patients with ER+ disease. See related Spotlight by Roussos Torres, p. 672

BackgroundTumor-specific cytotoxic T cells and T cell receptors are effective tools for cancer immunotherapy. Most efforts to identify them rely on known antigens or lymphocytes that have infiltrated into the tumor bed. Approaches to empirically identify tumor-targeting T cells and T cell receptors by exploiting all antigens expressed on tumor cell surfaces are not well developed for most carcinomas, including pancreatic cancer.MethodsAutologous tumor organoids were stimulated with T cells from the patients’ peripheral blood for 2 weeks to generate the organoid-primed T (opT) cells. opT cell phenotype was analyzed by monitoring changes in the expression levels of 28 cell surface and checkpoint proteins. Expression of ligands of the immune checkpoints was investigated by immunohistochemistry staining. T cells were labeled with carboxyfluorescein succinimidyl ester (CFSE) and assayed by flow cytometry to monitor tumor-induced T cell proliferation changes. opT cell-mediated killing of three-dimensional organoids was measured using an M30 ELISA kit. T cell receptors (TCRs) were identified by deep sequencing of gDNA isolated from T cells, and the TCR specificity was confirmed by transferring TCRs to the T cell line SKW-3 or donor T cells.ResultsThe co-culture was effective in the generation of CD8 + or CD4+opT cells. The opT cells killed autologous tumors in a granzyme B or Fas-Fas ligand-dependent manner and expressed markers of tissue-resident memory phenotype. Each patient-derived opT cell culture displayed a unique complement of checkpoint proteins. Interestingly, only NKG2A blockade showed a potent increase in the interferon-γ production compared with blocking programmed cell death protein 1 (PD-1) or programmed cell death ligand 1 (PD-L1) or TIM3 or TIGIT or LAG3. Importantly, TCR sequencing demonstrated a dramatic clonal expansion of T cells with a restricted subset of TCRs. Cloning and transferring the TCRs to heterologous T cells was sufficient to confer tumor cell recognition and cytotoxic properties in a patient-specific manner.ConclusionWe report a platform for expanding tumor-targeting T cells from the peripheral blood of patients with pancreatic cancer. We identify the NKG2A-HLA-E axis as a potentially important checkpoint for CD8 +T cells for pancreatic cancer. Lastly, we demonstrate empirical identification of tumor-targeting TCRs that can be used for TCR-therapeutics.

Targeting carbonic anhydrase IX acidifies intracellular pH, disrupts redox homeostasis, and creates vulnerability to ferroptosis., The metabolic mechanisms involved in the survival of tumor cells within the hypoxic niche remain unclear. We carried out a synthetic lethal CRISPR screen to identify survival mechanisms governed by the tumor hypoxia–induced pH regulator carbonic anhydrase IX (CAIX). We identified a redox homeostasis network containing the iron-sulfur cluster enzyme, NFS1. Depletion of NFS1 or blocking cyst(e)ine availability by inhibiting xCT, while targeting CAIX, enhanced ferroptosis and significantly inhibited tumor growth. Suppression of CAIX activity acidified intracellular pH, increased cellular reactive oxygen species accumulation, and induced susceptibility to alterations in iron homeostasis. Mechanistically, inhibiting bicarbonate production by CAIX or sodium-driven bicarbonate transport, while targeting xCT, decreased adenosine 5′-monophosphate–activated protein kinase activation and increased acetyl–coenzyme A carboxylase 1 activation. Thus, an alkaline intracellular pH plays a critical role in suppressing ferroptosis, a finding that may lead to the development of innovative therapeutic strategies for solid tumors to overcome hypoxia- and acidosis-mediated tumor progression and therapeutic resistance.

Estrogen receptor (ER) positive breast cancer represents 75% of all breast cancers in women. Although patients with ER+ cancers receive endocrine therapies, more than 30% develop resistance and succumb to the disease, highlighting the need to understand endocrine resistance. Here we show an unexpected role for the cell polarity protein SCRIB as a tumor-promoter and a regulator of endocrine resistance in ER-positive breast cancer cells. SCRIB expression is induced by estrogen signaling in a MYC-dependent manner. SCRIB interacts with SLC3A2, a heteromeric component of leucine amino acid transporter SLC7A5. SLC3A2 binds to the N-terminus of SCRIB to facilitate the formation of SCRIB/SLC3A2/LLGL2/SLC7A5 quaternary complex required for membrane localization of the amino acid transporter complex. Both SCRIB and SLC3A2 are required for cell proliferation and tamoxifen resistance in ER+ cells identifying a new role for the SCRIB/SLC3A2 complex in ER+ breast cancer.

Summary: Misregulation of alternative splicing is a hallmark of human tumors, yet to what extent and how it contributes to malignancy are only beginning to be unraveled. Here, we define which members of the splicing factor SR and SR-like families contribute to breast cancer and uncover differences and redundancies in their targets and biological functions. We identify splicing factors frequently altered in human breast tumors and assay their oncogenic functions using breast organoid models. We demonstrate that not all splicing factors affect mammary tumorigenesis in MCF-10A cells. Specifically, the upregulation of SRSF4, SRSF6, or TRA2β disrupts acinar morphogenesis and promotes cell proliferation and invasion in MCF-10A cells. By characterizing the targets of these oncogenic splicing factors, we identify shared spliced isoforms associated with well-established cancer hallmarks. Finally, we demonstrate that TRA2β is regulated by the MYC oncogene, plays a role in metastasis maintenance in vivo, and its levels correlate with breast cancer patient survival. : Park et al. demonstrate that >50% of human breast tumors exhibit an alteration in one of the splicing factors from the SR protein family. Using in vitro and in vivo breast cancer models, they identify three splicing factors that promote cell proliferation and invasion by regulating isoforms associated with cancer hallmarks. Keywords: alternative RNA splicing, breast cancer, splicing factor, SR protein, metastasis, MYC, TRA2-beta, triple negative breast cancer

Drosophila Lgl and its mammalian homologues, LLGL1 and LLGL2, are scaffolding proteins that regulate the establishment of apical-basal polarity in epithelial cells1,2. Whereas Lgl functions as a tumour suppressor in Drosophila1, the roles of mammalian LLGL1 and LLGL2 in cancer are unclear. The majority (about 75%) of breast cancers express oestrogen receptors (ERs)3, and patients with these tumours receive endocrine treatment4. However, the development of resistance to endocrine therapy and metastatic progression are leading causes of death for patients with ER+ disease4. Here we report that, unlike LLGL1, LLGL2 is overexpressed in ER+ breast cancer and promotes cell proliferation under nutrient stress. LLGL2 regulates cell surface levels of a leucine transporter, SLC7A5, by forming a trimeric complex with SLC7A5 and a regulator of membrane fusion, YKT6, to promote leucine uptake and cell proliferation. The oestrogen receptor targets LLGL2 expression. Resistance to endocrine treatment in breast cancer cells was associated with SLC7A5- and LLGL2-dependent adaption to nutrient stress. SLC7A5 was necessary and sufficient to confer resistance to tamoxifen treatment, identifying SLC7A5 as a potential therapeutic target for overcoming resistance to endocrine treatments in breast cancer. Thus, LLGL2 functions as a promoter of tumour growth and not as a tumour suppressor in ER+ breast cancer. Beyond breast cancer, adaptation to nutrient stress is critically important5, and our findings identify an unexpected role for LLGL2 in this process.

SummaryEpithelial to mesenchymal transition (EMT) is a complex cellular program proceeding through a hybrid E/M state linked to cancer-associated stemness, migration and chemoresistance. Deeper molecular understanding of this dynamic physiological landscape is needed to define events which regulate the transition and entry into and exit from the E/M state. Here, we quantified >60,000 molecules across ten time points and twelve omic layers in human mammary epithelial cells undergoing TGFβ-induced EMT. Deep proteomic profiles of whole cells, nuclei, extracellular vesicles, secretome, membrane and phosphoproteome defined state-specific signatures and major transition points. Parallel metabolomics showed metabolic reprogramming preceded changes in other layers, while single-cell RNA sequencing identified transcription factors controlling entry into E/M. Covariance analysis exposed unexpected discordance between the molecular layers. Integrative causal modeling revealed co-dependencies governing entry into E/M that were verified experimentally using combinatorial inhibition. Overall, this dataset provides an unprecedented resource on TGFβ signaling, EMT and cancer.

Reactive oxygen species (ROS) are implicated in triggering cell signalling events and pathways to promote and maintain tumorigenicity. Chemotherapy and radiation can induce ROS to elicit cell death allows for targeting ROS pathways for effective anti-cancer therapeutics. Coenzyme Q10 is a critical cofactor in the electron transport chain with complex biological functions that extend beyond mitochondrial respiration. This study demonstrates that delivery of oxidized Coenzyme Q10 (ubidecarenone) to increase mitochondrial Q-pool is associated with an increase in ROS generation, effectuating anti-cancer effects in a pancreatic cancer model. Consequent activation of cell death was observed in vitro in pancreatic cancer cells, and both human patient-derived organoids and tumour xenografts. The study is a first to demonstrate the effectiveness of oxidized ubidecarenone in targeting mitochondrial function resulting in an anti-cancer effect. Furthermore, these findings support the clinical development of proprietary formulation, BPM31510, for treatment of cancers with high ROS burden with potential sensitivity to ubidecarenone.