Titolo della tesi: Jagged1, a novel non-canonical player in Pancreatic Ductal Adenocarcinoma (PDAC) and Colorectal Cancer (CRC)
Pancreatic Ductal Adenocarcinoma (PDAC) and Colorectal Cancer (CRC) are two of the leading causes of cancer death worldwide. Since chemotherapeutic treatments have limited success, it is imperative to understand the molecular mechanisms involved in tumour pathology to improve current therapies and increase patient survival. Both PDAC and CRC are characterised by numerous mutations in specific oncogenes and tumour suppressor genes. However, a notable common alteration between them is the presence of activating mutations in the KRAS proto-oncogene, which is observed in approximately 90% of PDAC cases and 50% of CRC cases.
Notch signalling is a highly evolutionarily conserved pathway that mediates cell-to-cell communication and consists of transmembrane receptors and ligands. Ligand binding renders the transmembrane Notch receptors susceptible to proteolytic processing involving ADAM metalloproteinases and the PS/γ-secretase complex, resulting in the release of the Notch intracellular domain, Notch-IC, which translocates to the nucleus and modulates gene expression involved in various cellular processes such as proliferation, cell differentiation, angiogenesis, apoptosis, and others. Interestingly, Notch signalling has been implicated in various aspects of PDAC and CRC biology, but the role of Notch receptors is controversial.
The Notch ligand, Jagged1, is a single-pass transmembrane protein that has been reported to be overexpressed in both PDAC and CRC, where it is associated with tumour size, metastasis, high TNM stage and poor prognosis. Notably, recent works have shown that Jagged1 itself is processed by sequential proteolytic cleavages involving ADAM-17/TACE and the PS/γ-secretase complex, releasing the Jagged1 intracellular domain, Jag1-ICD, which can translocate to the nucleus and induce reverse signalling. In CRC, Jagged1 is constitutively processed via K-Ras/MEK/Erk axis and Jag1-ICD has been shown to contribute to tumour biology, aggressiveness, and drug resistance. Based on these reports, we aimed to investigate the role of Jagged1 in PDAC tumours and the chemoresistance mechanisms of Jag1-ICD in CRC.
Firstly, our preliminary results showed that there is a direct relationship between K-Ras, Notch receptors and Jagged1 in PDAC. In fact, K-Ras controls Notch receptor expression through K-Ras/MEK/Erk signalling. Notch, in turn, can sustain Jagged1 expression. In addition, Jagged1 is not only overexpressed but is also proteolytically processed, resulting in the release of Jag1-ICD, which is mainly localised to the nuclear compartment in several PDAC cell lines, triggering Jag1-ICD-associated reverse signalling. To study the role of Jagged1, we generated Jagged1-depleted cell lines using Crispr-Cas9 technology and Jag1-ICD-overexpressing cell lines. Our results show modulations of proliferation, EMT and apoptosis-related genes, suggesting a tumorigenic role of Jagged1 in PDAC.
Chemotherapy resistance in CRC patients remains a clinical challenge as it allows the development of metastasis and disease recurrence. However, the molecular mechanisms that characterise drug resistance in CRC are not well understood. It has been previously reported that 5-FU and Oxaliplatin contribute to Jagged1 processing in CRC via Erk1/2, suggesting chemoresistance effect. In the present study, we investigated the effects of OXP and 5-FU chemotherapeutic agents, alone or in combination with GSIs, on Jagged1 processing in CRC cells. Here we show that GSIs are able to inhibit Notch signalling, but, surprisingly, lead to a strong activation of Jagged1 reverse signalling through Erk1/2 activation. Furthermore, both OXP and 5-FU treatment drive mechanisms of intrinsic chemoresistance and protects CRC cells from apoptosis by triggering Jagged1 reverse signalling. Taken together, OXP/5FU and GSIs synergistically promote the selection and growth of a Jag1-ICD positive population that is resistant to chemotherapy.
Collectively, our data implicate Jagged1 as a novel oncogenic player in PDAC and CRC, contributing to the maintenance of carcinogenesis in a non-canonical manner.