RNA seq on primary pancreatic cancer cells growing on collagen 1 homotrimers and heterotrimers
Ontology highlight
ABSTRACT: RNA seq on primary pancreatic cancer (KPPC;Col1pdxKO cancer-collagen1 knockout) cells growing on collagen 1 homotrimers and heterotrimers. In this study, we identify that pancreatic cancer cells produce a unique Collagen1 homotrimer variant, in contrast to the Collagen1 heterotrimer produced by normal cells (such as fibroblasts). The variant forms of Collagen1 (homotrimer and heterotrimer) have distinct effects on cancer cell behaviors. We culture the pancreatic cancer cells on purified Collagen1 homotrimer and heterotrimer (as well as vehicle control) and examine the global changes of gene expression profile in cancer cells by variant Collagen1 subtypes.
Project description:RNA sequencing analysis on two primary pancreatic cancer cell lines from transgenic mice: (1) cancer-collagen1 knockout cell line from KPPC;Col1pdxKO cancer-collagen1 knockout tumor and (2) control pancreatic cancer cell line from KPPC tumor
Project description:Activated pancreatic stellate cells produce the fibrotic matrix in chronic pancreatitis and pancreatic cancer. In vitro protocols examining PSC biology have usually involved PSCs cultured on plastic, a non-physiological surface. However, PSCs cultured on physiological matrices e.g. MatrigelTM (normal basement membrane) and collagen (fibrotic pancreas), may have distinctly different behaviours compared to cells cultured on plastic. Therefore, we aimed to compare PSC gene expression after culture on plastic, MatrigelTM and collagen I.
Project description:Activated pancreatic stellate cells produce the fibrotic matrix in chronic pancreatitis and pancreatic cancer. In vitro protocols examining PSC biology have usually involved PSCs cultured on plastic, a non-physiological surface. However, PSCs cultured on physiological matrices e.g. MatrigelTM (normal basement membrane) and collagen (fibrotic pancreas), may have distinctly different behaviours compared to cells cultured on plastic. Therefore, we aimed to compare PSC gene expression after culture on plastic, MatrigelTM and collagen I. Total RNA from stellate cells in 10 cm Petri dishes was isolated by Qiagen RNeasy Mini Plus kit as per manufacturer’s instructions. The Agilent 2100 Bioanalyzer (Agilent Technologies Inc. Santa Clara, CA) was used for quality control of the isolated total RNA. Gene expression profiles of rat PSCs cultured on MatrigelTM, collagen I and plastic were analysed by whole rat genome microarray purchased from Affymetrix (Rat Gene 1.0 ST Array). This array was able to detect 27,342 rat genes, with approximately 26 probes on average per gene (referred to as a probe set).
Project description:Collagen, a critical component of the tumor microenvironment (TME), significantly influences cancer-associated fibrosis, tissue stiffness, immune response, and metastasis. Pancreatic ductal adenocarcinoma (PDAC) is characterized by excessive collagen deposition, and effective therapies remain limited. Our research challenges conventional understanding by highlighting the remarkable ability of specific tumor epithelial populations, particularly those with low levels of L1 cell adhesion molecule (L1CAM) expression (L1low), to actively modulate collagen dynamics within the TME. Transcriptome analysis of PDAC cells with varying L1 expression reveals shifts in collagen-related pathways, with heightened expression of collagen 17A1 (COL17A1) in L1low cells. Alongside, L1low cells exhibit enhanced migratory capacity. In an orthotopic mouse model, L1low cancer cells demonstrate increased collagen deposition, resistance to gemcitabine treatment, and an elevated propensity for liver metastases. Treatment with Tranilast, an anti-fibrotic drug, significantly reduces tumor volume and collagen deposition while enhancing L1 expression. This, in turn, mitigates tumor invasiveness and suppresses metastases formation. Overall, our study uncovers a novel role for L1CAM in PDAC aggressiveness and fibrosis, revealing a new epithelial population capable of collagen secretion and challenging the traditional notion that this function is solely attributed to stromal cells.
Project description:RNA-sequencing analyses were performed on cancer cell lines from KPPC and PPSSC pancreatic tumors, so as to identify the changes of tumor immune microenvironment upon Trp53 and Smad4 deletion without KRAS mutation.
Project description:Here, we sought to evaluate the impact of KrasG12D and KrasG12C inhibitors on pancreatic cancer cells. A murine pancreatic cancer cell line with KrasG12D mutation was treated with MRTX1133 (KrasG12D inhibitor) and MRTX849 (KrasG12C inhibitor) and scRNA-seq performed to evaluate transcriptional changes. We further evaluated transcriptional changes in cancer cells, as well as in the tumor microenvironment of immunodeficient (NSG) and immunocompetent (C57BL6/J) mice orthotopically implanted with KPC689 cells and spontaneous KPPC tumors in response to MRTX1133 and MRTX849.
Project description:Metabolic syndrome represents a risk factor of pancreatic ductal adenocarcinoma (PDAC), the gastrointestinal cancer with the lowest survival rate and therapeutic outcome. PDAC results bidirectional interconnected with metabolic alteration, which favors its onset and occurs during early phases of its development. Pancreatic neoplastic lesions evolve within a highly dense desmoplastic stroma, characterized by abundant extracellular matrix deposition consequence of a prominent cancer associated fibroblast (CAF) activation. Here, the complex association between dysmetabolism and PDAC has been analyzed with a specific focus on CAFs to unveil novel targets exploitable for diagnostic and therapeutic purposes. Methods: PDAC development upon dysmetabolic conditions was investigated in: 1) wild type immunocompetent syngeneic mice by orthotopic transplantation of PanIN-bearing Pdx1-Cre; Kras+/LSL-G12D; Trp53+/LSL-R172H organoids (KPC mice) fed with high fat diet to recapitulate dysmetabolism-dependent alterations; and 2) primary pancreatic CAFs isolated from chemotherapy naïve PDAC patients with or without an history of metabolic syndrome. Results: The dysmetabolic-associated higher PDAC aggressiveness was paralleled by an enrichment of collagen fibrils due to an increased function of the prolyl 4-hydroxylase subunit alpha 1 (P4HA1). Upon dysmetabolic conditions, P4HA1 boosts collagen hydroxy-prolination, intensifying collagen contraction strength and thus preventing CD3 positive lymphocyte PDAC infiltration. Interestingly, semaglutide, an incretin agonist with anti-diabetic and anti-obesity properties, is able to prevent the higher stromal deposition of PDAC observed in dysmetabolic conditions, preventing CAF contraction, allowing immune infiltration and reducing tumor development. Conclusion: These results shed light on novel therapeutic options for PDAC patients affected by metabolic syndrome aimed at pancreatic cancer stroma reshape
Project description:RNA sequencing analysis on tumor samples from two groups of mice (1) KPPC;Col1pdxKO cancer-collagen1 knockout group and (2) KPPC control group
Project description:Collagen linearization leads to cancer metastasis and poor prognosis but there are no therapies to target it. In addition to cell-generated mechanical forces, collagen linearization is triggered by the interaction of collagen with WISP1 (CCN4), a TGFb1-induced factor secreted by cancer cells. However, it remains unknown how WISP1 is physiologically regulated and whether it could be targeted to prevent collagen linearization. Here, we show that WISP2 (CCN5), a secreted factor downregulated in tumors, is an endogenous inhibitor of type I collagen (Col I) linearization. Mechanistically, WISP2 prevents WISP1 binding to Col I, thereby inhibiting WISP1-induced Col I linearization. Consequently, restoration of WISP2 via overexpression in tumor cells or via treatments with recombinant WISP2 impairs WISP1-induced or TGFb1-induced cancer cell invasion and spontaneous metastasis in vivo. Overall, this study uncovers WISP2 as an endogenous competitive inhibitor of WISP1 and reveals that collagen linearization is controlled by secreted factors with antagonistic functions.