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Apoptosis induction and cell cycle arrest of pladienolide B in erythroleukemia cell lines

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Summary

Splicing of pre-mRNA into functional mRNA, carried out by the spliceosome, represents a crucial step in eukaryotic gene expression. Mutations and other deregulation in some of the spliceosome components have been identified in multiple pathologies, including hematological malignancies. In this context, we evaluated the therapeutic potential of a splicing inhibitor, Pladienolide B (Pla-B), in two erythroleukemia cell lines. HEL and K562 cell lines were incubated with increasing doses of Pla-B in single and daily administration. Cell viability and density were evaluated using trypan blue assay. Flow cytometry was used to evaluate cell death, cell cycle, and caspase activity. NGS analysis was performed to assess the mutational status of 4 splicing-related genes (SF3B1, U2AF1, ZRSR2 and SRSF2). Expression levels of SF3B1 and unspliced DNAJB1 were evaluated by qPCR. Pla-B significantly decreased the viability and proliferation of both cell lines in time, dose, administration schedule, and cell line-dependent manner. HEL cells were more sensible to Pla-B (IC50 = 1.5 nM) than K562 (IC50 = 25 nM), with an IC50 almost 17 times lower. Pla-B induced cell death, mainly by apoptosis, and cell cycle arrest in G0/G1 phase. No mutations were found in any of the analyzed genes, suggesting that the observed cytotoxic effect is independent of the spliceosome mutations. Splicing modulator Pla-B showed high antitumor activity against HEL and K562 cell lines, inducing apoptosis and cell cycle arrest. These data suggest that Pla-B might represent a new therapeutic approach for erythroleukemia.

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Funding

The present work was supported by CIMAGO - Center of Investigation on Environment, Genetics and Oncobiology, Faculty of Medicine, University of Coimbra, Portugal, by funds from FEDER through the Operational Program Competitiveness Factors - COMPETE, and by Portuguese funds through FCT - Foundation for Science and Technology. Raquel Alves was supported by Portuguese Foundation to Science and Technology (FCT) with a PhD Grant (SFRH/BD/51994/2012).

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Author notes

  1. Joana Jorge and Sara Petronilho contributed equally to this work.

Authors and Affiliations

  1. Laboratory of Oncobiology and Hematology, University Clinic of Hematology, Faculty of Medicine, University of Coimbra, FMUC, Azinhaga de Santa Comba-Celas, 3000-548, Coimbra, Portugal

    Joana Jorge, Sara Petronilho, Raquel Alves, Ana Cristina Gonçalves & Ana Bela Sarmento-Ribeiro

  2. Coimbra Institute for Clinical and Biomedical Research (iCBR) - Group of Environment Genetics and Oncobiology (CIMAGO), FMUC, Coimbra, Portugal

    Joana Jorge, Raquel Alves, Margarida Coucelo, Ana Cristina Gonçalves, José Manuel Nascimento Costa & Ana Bela Sarmento-Ribeiro

  3. Center for Innovative Biomedicine and Biotechnology (CIBB), Coimbra, Portugal

    Raquel Alves, Margarida Coucelo, Ana Cristina Gonçalves, José Manuel Nascimento Costa & Ana Bela Sarmento-Ribeiro

  4. Clinical Hematology Department, Centro Hospitalar Universitário de Coimbra (CHUC), Coimbra, Portugal

    Margarida Coucelo & Ana Bela Sarmento-Ribeiro

  5. University Clinic of Oncology, Faculty of Medicine, University of Coimbra, FMUC, Coimbra, Portugal

    José Manuel Nascimento Costa

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  1. Joana Jorge
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Correspondence to Ana Cristina Gonçalves.

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Jorge, J., Petronilho, S., Alves, R. et al. Apoptosis induction and cell cycle arrest of pladienolide B in erythroleukemia cell lines. Invest New Drugs 38, 369–377 (2020). https://doi.org/10.1007/s10637-019-00796-2

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