Targeting ribosome biogenesis reinforces ERK-dependent senescence in pancreatic cancer

ABSTRACT

Pancreatic adenocarcinomas (PDAC) often possess mutations in K-Ras that stimulate the ERK pathway. Aberrantly high ERK activation triggers oncogene-induced senescence, which halts tumor progression. Here we report that low-grade pancreatic intraepithelial neoplasia displays very high levels of phospho-ERK consistent with a senescence response. However, advanced lesions that have circumvented the senescence barrier exhibit lower phospho-ERK levels. Restoring ERK hyperactivation in PDAC using activated RAF leads to ERK-dependent growth arrest with senescence biomarkers. ERK-dependent senescence in PDAC was characterized by a nucleolar stress response including a selective depletion of nucleolar phosphoproteins and intranucleolar foci containing RNA polymerase I designated as senescence-associated nucleolar foci (SANF). Accordingly, combining ribosome biogenesis inhibitors with ERK hyperactivation reinforced the senescence response in PDAC cells. Notably, comparable mechanisms were observed upon treatment with the platinum-based chemotherapy regimen FOLFIRINOX, currently a first-line treatment option for PDAC. We thus suggest that drugs targeting ribosome biogenesis can improve the senescence anticancer response in pancreatic cancer.

Keywords:

• ribosome biogenesis
• nucleolus
• therapy-induced senescence
• pancreatic cancer
• folfirinox
• MAP kinase

Acknowledgements

For phosphoproteomics analyses, we thank Dr. Bonneil and the Center for Advanced Proteomics Analyses. We thank Drs Roy, Desgroseillers, Lowe, McMahon, Nolan, Oeffinger, Ouelette, and Weinberg for access to equipment, reagents, and/or advice. Thanks to M. Vasseur and Dr. Stifani for help with the Microscopy Platform of the University of Montreal’s Biochemistry Department. This work was supported by Canada Graduate Scholarships Michael Smith Foreign Study Supplements to M.C. Rowell and X. Deschênes-Simard and by FRQS (Fonds de recherche Québec - Santé) fellowships to M.C. Rowell, S. Lopes-Paciencia and P. Kalegari. We thank the Montreal Cancer Institute’s Canderel funds and the Pierre-Saul Funds for pancreatic cancer research for financial support. V. Krizhanovsky is supported by the Israel Science Foundation-Canada program (2633/17) and the European Research Council (856487). Nabeel Bardeesy was supported by NIH/NCI P01CA117969. G. Ferbeyre is supported by the Joint Canada-Israel Research Program of IDRC, CIHR PJT-153217, and the CIBC chair for breast cancer research.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Author contributions:

M.C.R performed most replicates of experiments for Figures 3a-i, 4 –6, 7a-c, S2c, S3, S4a, S5, S6a-f, S8a-c, reproduced data from Figures 1a and 2d, did tissue culture and Congo Red staining for Figure 7d-e and S8d-e, and Western-blots for Fig. S9a and S10a. M.C.R analyzed phosphoproteomics data for Figure 4. X.D.S performed experiments for Figures 1–2, S1, S2a-b, and quantified data for Figure 7b and S8b. S.L.P performed experiments for Figure 5e, quantified Figure 5f, optimized conditions for Fig. S5, did tissue culture for Fig. S6d, performed qPCR experiments for Figure 7f and S8f and immunofluorescences for Fig. S6g, S7, S9f-g, S10b. S.L.P also gave conceptual and technical input and performed many exploratory experiments in the course of project development. B.L.C co-performed experiments for Figure 3a-e, i, Figure 4a, Fig. S3 a-e, S4a, performed IHC and quantification for Figure 3j-l and S4b-d and performed tumor sphere assay for Fig S2. P.K. reproduced data from Figure 3f-g, and optimized drug dosage for Figures 6–7 and S7. L.M. and A.F.R. reproduced experiments for Fig. S5. J.G. reproduced experiments in Figure 7a-c, and f and Fig. S8a-c and f. P.K. and J.G. did tissue culture for Fig. S6g, S7, S9–10. F.L. gave conceptual and technical input during the project. V.B. designed the qPCR primers for rRNA qPCR, did qPCR for Fig. S6h and S9d-e, reproduced qPCR experiments for Figure 7f and S8f, and helped in the design of experiments. S.I. did the image acquisition and quantification for Figure 7d-e and S8d-e. A.M.D. performed crystal violet assays for Fig. S9b-c. Y.S. reproduced data from Fig.S5a-c. F.K. helped in IHC scoring for Figures 1–2, and S1. V.D. reviewed histology for Figures 1–2 and Fig.S1. V.K. supervised the project and provided conceptual input regarding the use of senolytics and the analysis of senescence in PanIN and PDAC samples. N.B. provided patient samples and mouse samples of different stages of progression of PDAC for Figures 1–2 and S2, pancreatic cancer cell lines, and conceptual input regarding pancreatic cancer biology. G.F. supervised the project, provided conceptual input, and wrote the manuscript, except for Figures legends and Materials and methods that were written by M.C.R. Figures were prepared by M.C.R. with help from X.D.S., S.L.P, B.L.C., and S.I. All authors participated in reviewing the manuscript.

Supplementary material

Supplemental data for this article can be accessed online at https://doi.org/10.1080/15384101.2023.2278945

Additional information

Funding

The work was supported by the H2020 European Research Council [856487]; Institute of Cancer Research [PJT-153217]; IDRC [Canada-Israel Research Program]; NIH-NCI [NIH/NCI P01CA117969].