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MECHANISMS OF RESISTANCE

Sustained activation of non-canonical NF-κB signalling drives glycolytic reprogramming in doxorubicin-resistant DLBCL

Leukemia volume 37, pages 441–452 (2023)Cite this article

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Abstract

DLBCL is the most common lymphoma with high tumor heterogeneity. Treatment refractoriness and relapse from R-CHOP therapy in patients remain a clinical problem. Activation of the non-canonical NF-κB pathway is associated with R-CHOP resistance. However, downstream targets of non-canonical NF-κB mediating R-CHOP-induced resistance remains uncharacterized. Here, we identify the common mechanisms underlying both intrinsic and acquired resistance that are induced by doxorubicin, the main cytotoxic component of R-CHOP. We performed global transcriptomic analysis of (1) a panel of resistant versus sensitive and (2) isogenic acquired doxorubicin-resistant DLBCL cell lines following short and chronic exposure to doxorubicin respectively. Doxorubicin-induced stress in resistant cells activates a distinct transcriptional signature that is enriched in metabolic reprogramming and oncogenic signalling. Selective and sustained activation of non-canonical NF-κB signalling in these resistant cells exacerbated their survival by augmenting glycolysis. In response to doxorubicin, p52-RelB complexes transcriptionally activated multiple glycolytic regulators with prognostic significance through increased recruitment at their gene promoters. Targeting p52-RelB and their targets in resistant cells increased doxorubicin sensitivity in vitro and in vivo. Collectively, our study uncovered novel molecular drivers of doxorubicin-induced resistance that are regulated by non-canonical NF-κB pathway. We reveal new avenues of therapeutic targeting for R-CHOP-treated refractory/relapsed DLBCL patients.

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Fig. 1: DLBCL cell lines display differential responsiveness to doxorubicin-induced cytotoxicity which correlates with the apoptotic response to doxorubicin-induced DDR.
Fig. 2: Resistant DLBCL cells display a distinct doxorubicin-induced transcriptomic signature that predicts pan cancer doxorubicin sensitivity.
Fig. 3: Periodic treatment of sensitive DLBCL cells to increasing doses of doxorubicin results in the acquisition of doxorubicin resistance and reduction in doxorubicin-induced DDR.
Fig. 4: Dox-induced transcriptomes of acquired and primary Dox-resistant DLBCL cells are similarly enriched in metabolic reprogramming and oncogenic processes.
Fig. 5: Sustained activation of non-canonical NF-κB pathway promotes survival in Dox-resistant cells via enhanced glycolysis.
Fig. 6: Dox-induced recruitment of p52-RelB to the promoters of clinically important metabolic regulators results in their increased expression in Dox-resistant cells.
Fig. 7: Inhibition of p52-RelB regulated targets reverses doxorubicin resistance and attenuates doxorubicin-induced glycolysis in DLBCL cells.
Fig. 8: Inhibition of non-canonical NF-κB signalling abolishes doxorubicin resistance in DLBCL in vivo.

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Data availability

The RNA-seq datasets generated during and/or analysed during the current study had been deposited in the GEO (Gene Expression Omnibus) database under the accession number GSE215900.

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Acknowledgements

We thank Hui Fen Kerry May Lim and Dachuan Huang from National Cancer Centre, Singapore for establishing the PDX model. This study is funded by the National Research Foundation (NRF) Singapore, under its Singapore NRF Fellowship (NRF-NRFF2018-04). In addition, we thank the Nanyang Assistant Professorship (NAP) Start-up-grant to Y.L. lab and National Medical Research Council (NMRC-OFLCG-18May0028), Tanoto Foundation and Ling Foundation for their support.

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Authors and Affiliations

  1. School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore, 637551, Singapore

    Shen Kiat Lim, Chen Chen Peng, Shannon Low, Varsheni Vijay, Andrea Budiman, Suet-Mien Tan & Yinghui Li

  2. Lymphoma Genomic Translational Research Laboratory, Cellular and Molecular Research, National Cancer Centre Singapore, 11 Hospital Crescent, Singapore, 169610, Singapore

    Beng Hooi Phang, Jing Quan Lim & Choon Kiat Ong

  3. ONCO-ACP, Duke-NUS Medical School, 8 College Road, Singapore, 169857, Singapore

    Jing Quan Lim

  4. Cancer Science Institute of Singapore, National University of Singapore, Singapore, 117599, Singapore

    Anand D. Jeyasekharan

  5. Department of Haematology-Oncology, National University Hospital, Singapore, 119074, Singapore

    Anand D. Jeyasekharan

  6. Director’s office, National Cancer Centre Singapore, 11 Hospital Crescent, Singapore, 169610, Singapore

    Soon Thye Lim

  7. Office of Education, Duke-NUS Medical School, 8 College Road, Singapore, 169857, Singapore

    Soon Thye Lim

  8. Cancer and Stem Cell Biology Program, Duke-NUS Graduate Medical School, 8 College Road, Singapore, 169857, Singapore

    Choon Kiat Ong

  9. Institute of Molecular and Cell Biology (IMCB), Agency for Science Technology and Research, Singapore, 138673, Singapore

    Yinghui Li

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Contributions

YL conceptualized and supervised the study. YL and SKL planned and devised the experiments. SKL, SL and AB performed all molecular and cell biology experiments. CCP and VV performed all bioinformatics analyses and data visualization. STL and CKO contributed the PDX model and BHP performed the PDX ex vivo experiments. ST and SKL conducted the tumor xenograft studies. SKL analysed the data and JQL reviewed all statistical analyses. ADJ provided feedback on the study design. SKL wrote the manuscript and YL edited it.

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Correspondence to Yinghui Li.

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Lim, S.K., Peng, C.C., Low, S. et al. Sustained activation of non-canonical NF-κB signalling drives glycolytic reprogramming in doxorubicin-resistant DLBCL. Leukemia 37, 441–452 (2023). https://doi.org/10.1038/s41375-022-01769-w

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