Marc Giry-Laterriere

Whole genomes redefine the mutational landscape of pancreatic cancer

Pancreatic cancer remains one of the most lethal of malignancies and a major health burden. We performed whole-genome sequencing and copy number variation (CNV) analysis of 100 pancreatic ductal adenocarcinomas (PDACs). Chromosomal rearrangements leading to gene disruption were prevalent, affecting genes known to be important in pancreatic cancer (TP53, SMAD4, CDKN2A, ARID1A and ROBO2) and new candidate drivers of pancreatic carcinogenesis (KDM6A and PREX2). Patterns of structural variation (variation in chromosomal structure) classified PDACs into 4 subtypes with potential clinical utility: the subtypes were termed stable, locally rearranged, scattered and unstable. A significant proportion harboured focal amplifications, many of which contained druggable oncogenes (ERBB2, MET, FGFR1, CDK6, PIK3R3 and PIK3CA), but at low individual patient prevalence. Genomic instability co-segregated with inactivation of DNA maintenance genes (BRCA1, BRCA2 or PALB2) and a mutational signature of DNA damage repair deficiency. Of 8 patients who received platinum therapy, 4 of 5 individuals with these measures of defective DNA maintenance responded.

Transient tissue priming via ROCK inhibition uncouples pancreatic cancer progression, sensitivity to chemotherapy, and metastasis

Fine-tuned manipulation of tumor tension and vasculature enhances response to chemotherapy and impairs metastatic spread in pancreatic cancer. ROCK-ing pancreatic cancer to the core Pancreatic cancer, one of the most deadly and difficult-to-treat tumor types in patients, usually has a dense stroma that can be difficult for drugs to penetrate. Stromal characteristics can also affect multiple other aspects of tumor biology, including metastatic spread, vascular supply, and immune response. Vennin et al. used Fasudil, a drug that inhibits a protein called ROCK and is already used for some conditions in people, to demonstrate the feasibility including short-term tumor stroma remodeling as part of cancer treatment. In genetically engineered and patient-derived mouse models of pancreatic cancer, priming with Fasudil disrupted the tumors’ extracellular matrix and improved the effectiveness of subsequent treatment with standard-of-care chemotherapy for this disease. The emerging standard of care for patients with inoperable pancreatic cancer is a combination of cytotoxic drugs gemcitabine and Abraxane, but patient response remains moderate. Pancreatic cancer development and metastasis occur in complex settings, with reciprocal feedback from microenvironmental cues influencing both disease progression and drug response. Little is known about how sequential dual targeting of tumor tissue tension and vasculature before chemotherapy can affect tumor response. We used intravital imaging to assess how transient manipulation of the tumor tissue, or “priming,” using the pharmaceutical Rho kinase inhibitor Fasudil affects response to chemotherapy. Intravital Förster resonance energy transfer imaging of a cyclin-dependent kinase 1 biosensor to monitor the efficacy of cytotoxic drugs revealed that priming improves pancreatic cancer response to gemcitabine/Abraxane at both primary and secondary sites. Transient priming also sensitized cells to shear stress and impaired colonization efficiency and fibrotic niche remodeling within the liver, three important features of cancer spread. Last, we demonstrate a graded response to priming in stratified patient-derived tumors, indicating that fine-tuned tissue manipulation before chemotherapy may offer opportunities in both primary and metastatic targeting of pancreatic cancer.

Hypermutation In Pancreatic Cancer

Pancreatic cancer is molecularly diverse, with few effective therapies. Increased mutation burden and defective DNA repair are associated with response to immune checkpoint inhibitors in several other cancer types. We interrogated 385 pancreatic cancer genomes to define hypermutation and its causes. Mutational signatures inferring defects in DNA repair were enriched in those with the highest mutation burdens. Mismatch repair deficiency was identified in 1% of tumors harboring different mechanisms of somatic inactivation of MLH1 and MSH2. Defining mutation load in individual pancreatic cancers and the optimal assay for patient selection may inform clinical trial design for immunotherapy in pancreatic cancer.

SOX9 regulates ERBB signalling in pancreatic cancer development

Objective The transcription factor SOX9 was recently shown to stimulate ductal gene expression in pancreatic acinar-to-ductal metaplasia and to accelerate development of premalignant lesions preceding pancreatic ductal adenocarcinoma (PDAC). Here, we investigate how SOX9 operates in pancreatic tumourigenesis. Design We analysed genomic and transcriptomic data from surgically resected PDAC and extended the expression analysis to xenografts from PDAC samples and to PDAC cell lines. SOX9 expression was manipulated in human cell lines and mouse models developing PDAC. Results We found genetic aberrations in the SOX9 gene in about 15% of patient tumours. Most PDAC samples strongly express SOX9 protein, and SOX9 levels are higher in classical PDAC. This tumour subtype is associated with better patient outcome, and cell lines of this subtype respond to therapy targeting epidermal growth factor receptor (EGFR/ERBB1) signalling, a pathway essential for pancreatic tumourigenesis. In human PDAC, high expression of SOX9 correlates with expression of genes belonging to the ERBB pathway. In particular, ERBB2 expression in PDAC cell lines is stimulated by SOX9. Inactivating Sox9 expression in mice confirmed its role in PDAC initiation; it demonstrated that Sox9 stimulates expression of several members of the ERBB pathway and is required for ERBB signalling activity. Conclusions By integrating data from patient samples and mouse models, we found that SOX9 regulates the ERBB pathway throughout pancreatic tumourigenesis. Our work opens perspectives for therapy targeting tumourigenic mechanisms.

JRK is a positive regulator of β-catenin transcriptional activity commonly overexpressed in colon, breast and ovarian cancer

The loss of β-catenin inhibitory components is a well-established mechanism of carcinogenesis but β-catenin hyperactivity can also be enhanced through its coactivators. Here we first interrogated a highly validated genomic screen and the largest repository of cancer genomics data and identified JRK as a potential new oncogene and therapeutic target of the β-catenin pathway. We proceeded to validate the oncogenic role of JRK in colon cancer cells and primary tumors. Consistent with a β-catenin activator function, depletion of JRK in several cancer cell lines repressed β-catenin transcriptional activity and reduced cell proliferation. Importantly, JRK expression was aberrantly elevated in 21% of colorectal cancers, 15% of breast and ovarian cancers and was associated with increased expression of β-catenin target genes and increased cell proliferation. This study shows that JRK is required for β-catenin hyperactivity regardless of the adenomatous polyposis coli/β-catenin mutation status and targeting JRK presents new opportunities for therapeutic intervention in cancer.

Sirtuin1 stimulates the proliferation and the expression of glycolysis genes in pancreatic neoplastic lesions.

Metabolic reprogramming is a feature of neoplasia and tumor growth. Sirtuin 1 (SIRT1) is a lysine deacetylase of multiple targets including metabolic regulators such as p53. SIRT1 regulates metaplasia in the pancreas. Nevertheless, it is unclear if SIRT1 affects the development of neoplastic lesions and whether metabolic gene expression is altered. To assess neoplastic lesion development, mice with a pancreas-specific loss of Sirt1 (Pdx1-Cre;Sirt1-lox) were bred into a KrasG12D mutant background (KC) that predisposes to the development of pancreatic intra-epithelial neoplasia (PanIN) and ductal adenocarcinoma (PDAC). Similar grade PanIN lesions developed in KC and KC;Sirt1-lox mice but specifically early mucinous PanINs occupied 40% less area in the KC;Sirt1-lox line, attributed to reduced proliferation. This was accompanied by reduced expression of proteins in the glycolysis pathway, such as GLUT1 and GAPDH. The stimulatory effect of SIRT1 on proliferation and glycolysis gene expression was confirmed in a human PDAC cell line. In resected PDAC samples, higher proliferation and expression of glycolysis genes correlated with poor patient survival. SIRT1 expression per se was not prognostic but low expression of Cell Cycle and Apoptosis Regulator 2 (CCAR2), a reported SIRT1 inhibitor, corresponded to poor patient survival. These findings open perspectives for novel targeted therapies in pancreatic cancer.

Pancreas-Specific Sirt1-Deficiency in Mice Compromises Beta-Cell Function without Development of Hyperglycemia

Aims/Hypothesis Sirtuin 1 (Sirt1) has been reported to be a critical positive regulator of glucose-stimulated insulin secretion in pancreatic beta-cells. The effects on islet cells and blood glucose levels when Sirt1 is deleted specifically in the pancreas are still unclear. Methods This study examined islet glucose responsiveness, blood glucose levels, pancreatic islet histology and gene expression in Pdx1Cre; Sirt1ex4F/F mice that have loss of function and loss of expression of Sirt1 specifically in the pancreas. Results We found that in the Pdx1Cre; Sirt1ex4F/F mice, the relative insulin positive area and the islet size distribution were unchanged. However, beta-cells were functionally impaired, presenting with lower glucose-stimulated insulin secretion. This defect was not due to a reduced expression of insulin but was associated with a decreased expression of the glucose transporter Slc2a2/Glut2 and of the Glucagon like peptide-1 receptor (Glp1r) as well as a marked down regulation of endoplasmic reticulum (ER) chaperones that participate in the Unfolded Protein Response (UPR) pathway. Counter intuitively, the Sirt1-deficient mice did not develop hyperglycemia. Pancreatic polypeptide (PP) cells were the only other islet cells affected, with reduced numbers in the Sirt1-deficient pancreas. Conclusions/Interpretation This study provides new mechanistic insights showing that beta-cell function in Sirt1-deficient pancreas is affected due to altered glucose sensing and deregulation of the UPR pathway. Interestingly, we uncovered a context in which impaired beta-cell function is not accompanied by increased glycemia. This points to a unique compensatory mechanism. Given the reduction in PP, investigation of its role in the control of blood glucose is warranted.

Emerging Drug Target In Pancreatic Cancer: Placing Sirtuin 1 on the Canvas

Sirtuin 1 is a protein deacetylase that regulates a large number of proteins often functionally implicated in tumor development and progression. Its pleiotropic function has turned SIRT1 into an attractive chemotherapeutic target, underscored by very promising preclinical results with SIRT1 inhibitors in the treatment of chronic myeloid leukemia. Here, we revisit the studies on SIRT1 as an emerging target for therapy in pancreatic cancer, a tumor with dismal outcomes for which currently few therapeutic options are available. We highlight those potential SIRT1 target genes that are commonly affected in pancreatic cancer according to recent genomic analyses.

'MCC' protein interacts with E-cadherin and β-catenin strengthening cell-cell adhesion of HCT116 colon cancer cells.

E-cadherin and β-catenin are key proteins that are essential in the formation of the epithelial cell layer in the colon but their regulatory pathways that are disrupted in cancer metastasis are not completely understood. Mutated in colorectal cancer (MCC) is a tumour suppressor gene that is silenced by promoter methylation in colorectal cancer and particularly in patients with increased lymph node metastasis. Here, we show that MCC methylation is found in 45% of colon and 24% of rectal cancers and is associated with proximal colon, poorly differentiated, circumferential and mucinous tumours as well as increasing T stage and larger tumour size. Knockdown of MCC in HCT116 colon cancer cells caused a reduction in E-cadherin protein level, which is a hallmark of epithelial–mesenchymal transition in cancer, and consequently diminished the E-cadherin/β-catenin complex. MCC knockdown disrupted cell–cell adhesive strength and integrity in the dispase and transepithelial electrical resistance assays, enhanced hepatocyte growth factor-induced cell scatter and increased tumour cell invasiveness in an organotypic assay. The Src/Abl inhibitor dasatinib, a candidate anti-invasive drug, abrogated the invasive properties induced by MCC deficiency. Mechanistically, we establish that MCC interacts with the E-cadherin/β-catenin complex. These data provide a significant advance in the current understanding of cell–cell adhesion in colon cancer cells.

Proteogenomic Analysis Identifies a Novel Human SHANK3 Isoform.

Mutations of the SHANK3 gene have been associated with autism spectrum disorder. Individuals harboring different SHANK3 mutations display considerable heterogeneity in their cognitive impairment, likely due to the high SHANK3 transcriptional diversity. In this study, we report a novel interaction between the Mutated in colorectal cancer (MCC) protein and a newly identified SHANK3 protein isoform in human colon cancer cells and mouse brain tissue. Hence, our proteogenomic analysis identifies a new human long isoform of the key synaptic protein SHANK3 that was not predicted by the human reference genome. Taken together, our findings describe a potential new role for MCC in neurons, a new human SHANK3 long isoform and, importantly, highlight the use of proteomic data towards the re-annotation of GC-rich genomic regions.