Opendata, web and dolomites

Report

Teaser, summary, work performed and final results

Periodic Reporting for period 1 - PDA_TGFb (TGF-beta mediated crosstalk between cancer (stem) cells and tumor microenvironment in pancreatic cancer metastasis)

Teaser

Pancreatic ductal adenocarcinoma (PDAC) is the fourth-leading cause of cancer-related death in the world, with a 5-year survival rate of less than 5%. Chemotherapy resistance and tumour relapse are two unresolved problems in PDAC treatment. Cancer stem cells (CSCs) are key...

Summary

Pancreatic ductal adenocarcinoma (PDAC) is the fourth-leading cause of cancer-related death in the world, with a 5-year survival rate of less than 5%. Chemotherapy resistance and tumour relapse are two unresolved problems in PDAC treatment. Cancer stem cells (CSCs) are key drivers in tumour progression, resistance and relapse and studying CSC biology may provide novel insights that could help to overcome these problems. Indeed, up-regulation of detoxifying enzymes and drug transporters in pancreatic CSCs have already been identified as important mechanisms for chemo-resistance in PDAC. Hence, targeting the CSC niche and their plasticity could be a complementary therapeutic strategy against cancer. Transforming growth factor beta (TGF-β) signalling is one of the most important pathways supporting the CSC niche and plasticity in PDAC. High levels of TGF-β1 in patients with PDAC are associated with poor prognosis in the clinical setting. It has been reported that the pancreatic stellate cells (PSC) within the tumour microenvironment represent the principal source of TGF-β1, but very little is known about the TGF-β1 mediated crosstalk between PSC and PDAC cells. The main goal of this project was to increase survival of pancreatic cancer patients by exploring the contribution of the tumour microenvironment to the failure of presently available oncological treatments. Specifically, we aimed to: 1) Obtain pure subpopulations from epithelium and stroma of PDAC human samples; 2) Define the role of TGFB_SLs and the signaling pathways activated during the tumor-stroma crosstalk; and 3) Determine whether the metastatic cells activate the TGFB_SLs-driven stromal response at the metastatic site.

Work performed

During the last two years I demonstrated that the lack of L1CAM is a hallmark of tumour differentiation, thus providing strong evidence that L1CAM regulates the cancer stem cells phenotype and bears potential as a biomarker for patient stratification.
I found that L1CAM is downregulated in the majority of pancreas carcinomas, which is associated with poor outcome. L1CAMnegative PDAC cells (either L1CAM L1- sorted or shL1CAM knockdown) were strongly enriched for CSC phenotypes, including enhanced self-renewal, tumour initiation, migration, invasion and chemoresistance. The lack of L1CAM is functionally linked to stemness and aggressiveness in PDAC and therefore may represent a more suitable functional CSC biomarker. pancreatic cancer, the combination of CD44 and CD133 has emerged as reliable biomarkers for the identification, tracking and isolation of CSC. I studied the expression of CD44 and CD133 in relation to the L1CAM status using flow cytometry. Diminished levels of L1CAM correlated with a higher proportion of CD44 and CD133 cells in pancreatic tumours, respectively. The ectopic downregulation of L1CAM increased the proportion of CD44 or CD133 cells. This effect was accompanied by increased expression of stemness-associated genes such as SOX2. Conversely, L1CAM overexpression reduced the expression of these CSCs genes. Furthermore, L1CAMnegative cells were strongly enriched for CSC-like properties such as tumour sphere and organoids formation and are resistant to gemcitabine treatment. Therefore, loss of L1CAM increases the CSC population and/or features, suggesting that L1CAM promotes epithelial (cancer) cell differentiation. I found that L1CAM expression diminishes stem cell properties, along with the percentage of these cells, but does not seem to functionally alter non-CSC cells (i.e., CD44- and CD133-). My data are conceivable with the notion that L1CAM loss results in a partial and/or preferential expansion of certain progenitor pools. Specifically, I noted a fractional increase in CSC-like cells as opposed to a complete conversion of the entire population towards CSC upon L1CAM inhibition. My data are in line with the notion that a predetermined subpopulation of PDAC bears the potential to convert into CSC upon L1CAM downregulation. Consistent data were obtained for invasiveness as another hallmark of CSCs. Culturing PDAC cells as organoids in Matrigel revealed that L1CAMnegative cells bear enhanced invasive capacity and EMT signature as compared to control cells. Importantly the L1- cells were able to recapitulate the tumour heterogeneity. Conversely, ectopic overexpression of L1CAM cDNA did not only reduce their stem-like properties and subsequently tumourigenicity, but also resulted in diminished invasiveness. I propose that L1CAM belongs to a new type of cancer genes that can act both as an oncogene and as a tumour suppressor, depending on the cellular/genomic context. I provided a thorough mechanistic analysis of the function of L1CAM in PDAC cells. I showed that L1CAM decreases stemness, EMT and subsequently their dissemination, supporting its functional role as tumour suppressor in fully established PDAC. The PDAC microenvironment is characterised by an extensive desmoplastic response, including influx of inflammatory cells, extensive deposition of collagen-rich extracellular matrix as well as activation and expansion of PSC. PSC are the predominant source of TGF-β1, which plays a critical role in tumour initiation and progression, at least in part by modulating the interactions between pancreatic CSCs and PSC. I found that TGF-β/SMAD signalling is involved in the downregulation of L1CAM and concomitant maintenance of pancreatic CSC-like properties. I demonstrated that TGF-β1 secreted by PSCs strongly decreases L1CAM in PDAC cells both in vitro and in vivo. These data provide the mechanistic basis for PSCs counteracting L1CAM as a suppressor of CSC-like characteristics in PDAC cells. Intriguingly, PDAC

Final results

A large number of deaths by cancer are the consequence of recurrent tumors that develop months or years after therapy. Although effective therapies continue to evolve and improve for several types of cancer (e.g., breast and colorectal cancer), for pancreatic cancer we are far away from having successful therapies and the current ones only confer a marginal survival advantage. Within this project I identified a novel mechanism of tumor formation. Additionally:
▸The discovery of the essential role of the TGF-β1-L1CAM axis in CSCs represents an important progression in our thoughtful of CSC biology
▸ Restoring the L1CAM expression by targeting the TGF-β1 pathway could be a highly specific therapeutic approach against pancreatic CSCs
Overall, thanks to the Marie Curie IF, I became independent scientist.

Website & more info

More info: http://www.igb.cnr.it/staff/people/lonardoe.