Pyrroline-5-Carboxylate Reductase 1: a novel target for sensitizing multiple myeloma cells to bortezomib by inhibition of PRAS40-mediated protein synthesis

Journal of Experimental & Clinical Cancer Research

Kim De Veirman¹  Hatice Satilmis¹  Karin Vanderkerken¹  Anke Maes¹  Elke De Bruyne¹  Inge Oudaert¹  Dirk Hose¹  Eline Menu¹  Philip Vlummens²  Wouter De Brouwer³  Bart Ghesquière⁴ 

[1] Department of Hematology and Immunology-Myeloma Center Brussels, Vrije Universiteit Brussel, Laarbeeklaan 103, 1090, Brussels, Jette, Belgium;Department of Hematology and Immunology-Myeloma Center Brussels, Vrije Universiteit Brussel, Laarbeeklaan 103, 1090, Brussels, Jette, Belgium;Department of Clinical Hematology, Ghent University Hospital, Gent, Belgium;Department of Hematology, Vrije Universiteit Brussel, Universitair Ziekenhuis Brussel, Laarbeeklaan 101, 1090, Brussels, Jette, Belgium;Metabolomics Expertise Center, VIB Center for Cancer Biology (VIB-CCB) - Department of Oncology, KU Leuven, Leuven, Belgium;

关键词: Multiple myeloma;    Proline;    PYCR1;    Hypoxia;    Protein synthesis;   

DOI  :  10.1186/s13046-022-02250-3

来源: Springer

【 摘 要 】

BackgroundMultiple myeloma (MM) remains an incurable cancer despite advances in therapy. Therefore, the search for new targets is still essential to uncover potential treatment strategies. Metabolic changes, induced by the hypoxic bone marrow, contribute to both MM cell survival and drug resistance. Pyrroline-5-carboxylate reductase 1 and 2 (PYCR1 and PYCR2) are two mitochondrial enzymes that facilitate the last step in the glutamine-to-proline conversion. Overexpression of PYCR1 is involved in progression of several cancers, however, its’ role in hematological cancers is unknown. In this study, we investigated whether PYCR affects MM viability, proliferation and response to bortezomib.MethodsCorrelation of PYCR1/2 with overall survival was investigated in the MMRF CoMMpass trial (653 patients). OPM-2 and RPMI-8226 MM cell lines were used to perform in vitro experiments. RPMI-8226 cells were supplemented with 13C-glutamine for 48 h in both normoxia and hypoxia (< 1% O2, by chamber) to perform a tracer study. PYCR1 was inhibited by siRNA or the small molecule inhibitor pargyline. Apoptosis was measured using Annexin V and 7-AAD staining, viability by CellTiterGlo assay and proliferation by BrdU incorporation. Differential protein expression was evaluated using Western Blot. The SUnSET method was used to measure protein synthesis. All in vitro experiments were performed in hypoxic conditions.ResultsWe found that PYCR1 and PYCR2 mRNA expression correlated with an inferior overall survival. MM cells from relapsed/refractory patients express significantly higher levels of PYCR1 mRNA. In line with the strong expression of PYCR1, we performed a tracer study in RPMI-8226 cells, which revealed an increased conversion of 13C-glutamine to proline in hypoxia. PYCR1 inhibition reduced MM viability and proliferation and increased apoptosis. Mechanistically, we found that PYCR1 silencing reduced protein levels of p-PRAS40, p-mTOR, p-p70, p-S6, p-4EBP1 and p-eIF4E levels, suggesting a decrease in protein synthesis, which we also confirmed in vitro. Pargyline and siPYCR1 increased bortezomib-mediated apoptosis. Finally, combination therapy of pargyline with bortezomib reduced viability in CD138+ MM cells and reduced tumor burden in the murine 5TGM1 model compared to single agents.Conclusions This study identifies PYCR1 as a novel target in bortezomib-based combination therapies for MM.

【 授权许可】

CC BY