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Targeting NAE1-mediated protein hyper-NEDDylation halts cholangiocarcinogenesis and impacts on tumor-stroma crosstalk
Paula Olaizola1, Pui Yuen Lee-Law2, Maite G Fernandez-Barrena3
1Department of Liver and Gastrointestinal Diseases, Biodonostia Health Research Institute - Donostia University Hospital -, University of the Basque Country (UPV/EHU), San Sebastian, Spain.
Background & Aims:
Cholangiocarcinoma (CCA) comprises a heterogeneous group of malignant tumors associated with dismal prognosis. Alterations in post-translational modifications (PTMs), including NEDDylation, result in abnormal protein dynamics, cell disturbances and disease. Herein, we investigate the role of NEDDylation in CCA development and progression.
Methods:
Levels and functions of NEDDylation, together with response to pevonedistat (NEDDylation inhibitor) or CRISPR/Cas9 against NAE1 were evaluated in vitro, in vivo and/or in patients with CCA. The development of preneoplastic lesions in Nae1+/- mice was investigated using an oncogene-driven CCA model. The impact of NEDDylation in CCA cells on tumor-stroma crosstalk was assessed using CCA-derived cancer-associated fibroblasts (CAFs). Proteomic analyses were carried out by mass-spectrometry.
Results:
The NEDDylation machinery was found overexpressed and overactivated in human CCA cells and tumors. Most NEDDylated proteins found upregulated in CCA cells, after NEDD8-immunoprecipitation and further proteomics, participate in the cell cycle, proliferation or survival. Genetic (CRISPR/Cas9-NAE1) and pharmacological (pevonedistat) inhibition of NEDDylation reduced CCA cell proliferation and impeded colony formation in vitro. NEDDylation depletion (pevonedistat or Nae1+/- mice) halted tumorigenesis in subcutaneous, orthotopic, and oncogene-driven models of CCA in vivo. Moreover, pevonedistat potentiated chemotherapy-induced cell death in CCA cells in vitro. Mechanistically, impaired NEDDylation triggered the accumulation of both cullin RING ligase and NEDD8 substrates, inducing DNA damage and cell cycle arrest. Furthermore, impaired NEDDylation in CCA cells reduced the secretion of proteins involved in fibroblast activation, angiogenesis, and oncogenic pathways, ultimately hampering CAF proliferation and migration.
Conclusion:
Aberrant protein NEDDylation contributes to cholangiocarcinogenesis by promoting cell survival and proliferation. Moreover, NEDDylation impacts the CCA-stroma crosstalk. Inhibition of NEDDylation with pevonedistat may represent a potential therapeutic strategy for patients with CCA.
Lay Summary:
Little is known about the role of post-translational modifications of proteins in cholangiocarcinoma development and progression. Herein, we show that protein NEDDylation is upregulated and hyperactivated in cholangiocarcinoma, promoting tumor growth. Pharmacological inhibition of NEDDylation halts cholangiocarcinogenesis and could be an effective therapeutic strategy to tackle these tumors.
Insights
Aberrant protein NEDDylation drives cholangiocarcinoma (CCA) growth by promoting cell survival and proliferation. Inhibiting NEDDylation with pevonedistat halts tumor progression and may offer a new therapeutic strategy for CCA patients.
Area of Science:
- Oncology
- Molecular Biology
- Biochemistry
Background:
- Cholangiocarcinoma (CCA) is a heterogeneous malignancy with poor prognosis.
- Altered post-translational modifications (PTMs), such as NEDDylation, are implicated in disease development.
- The specific role of NEDDylation in CCA pathogenesis remains largely unexplored.
Purpose of the Study:
- To investigate the role and therapeutic potential of NEDDylation in cholangiocarcinoma.
- To evaluate the impact of NEDDylation inhibition on CCA progression and tumor-stroma interactions.
Main Methods:
- Assessed NEDDylation levels and function in CCA cells and patient tumors.
- Utilized in vitro and in vivo models, including genetic (CRISPR/Cas9-NAE1) and pharmacological (pevonedistat) inhibition.
- Investigated oncogene-driven CCA models and tumor-stroma crosstalk with cancer-associated fibroblasts (CAFs).
- Performed proteomic analysis via mass spectrometry.
Main Results:
- NEDDylation machinery is overexpressed and hyperactivated in human CCA.
- Inhibition of NEDDylation reduced CCA cell proliferation, colony formation, and tumorigenesis in vivo.
- NEDDylation inhibition induced DNA damage and cell cycle arrest.
- Impaired NEDDylation modulated tumor-stroma crosstalk by reducing pro-tumorigenic CAF-associated factors.
Conclusions:
- Aberrant protein NEDDylation significantly contributes to cholangiocarcinogenesis.
- NEDDylation inhibition, particularly with pevonedistat, demonstrates therapeutic potential for CCA.
- Targeting NEDDylation may represent a novel strategy for treating cholangiocarcinoma.
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