Skip to main content
SpringerLink
Log in

Nilotinib in combination with sunitinib renders MCL-1 for degradation and activates autophagy that overcomes sunitinib resistance in renal cell carcinoma

  • Research
  • Published:
Cellular Oncology Aims and scope Submit manuscript

Abstract

Purpose

Sunitinib is a recommended drug for metastatic renal cell carcinoma (RCC). However, the therapeutic potential of sunitinib is impaired by toxicity and resistance. Therefore, we seek to explore a combinatorial strategy to improve sunitinib efficacy of low-toxicity dose for better clinical application.

Methods

We screen synergistic reagents of sunitinib from a compound library containing 1374 FDA-approved drugs by in vitro cell viability evaluation. The synergistically antiproliferative and proapoptotic effects were demonstrated on in vitro and in vivo models. The molecular mechanism was investigated by phosphoproteomics, co-immunoprecipitation, immunofluorescence and western-blot assays, etc.

Results

From the four-step screening, nilotinib stood out as a potential synergistic killer combined with sunitinib. Subsequent functional evaluation demonstrated that nilotinib and sunitinib synergistically inhibit RCC cell proliferation and promote apoptosis in vitro and in vivo. Mechanistically, nilotinib activates E3-ligase HUWE1 and in combination with sunitinib renders MCL-1 for degradation via proteasome pathway, resulting in the release of Beclin-1 from MCL-1/Beclin-1 complex. Subsequently, Beclin-1 induces complete autophagy flux to promote antitumor effect.

Conclusion

Our findings revealed that a novel mechanism that nilotinib in combination with sunitinib overcomes sunitinib resistance in RCC. Therefore, this novel rational combination regimen provides a promising therapeutic avenue for metastatic RCC and rationale for evaluating this combination clinically.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

Data availability

All data are available within the article, supplementary information, or available from the corresponding author upon reasonable request.

Abbreviations

AXL:

AXL receptor tyrosine kinase

BCL-2:

BCL2 apoptosis regulator

BCL-XL:

Apoptosis regulator Bcl-X

c-Kit:

Stem cell growth factor receptor Kit

FDA:

Food and drug administration

FLT-3:

Fms related receptor tyrosine kinase 3

HBSS:

Hanks’ balance salt solution

HUWE1:

HECT, UBA and WWE domain containing E3 ubiquitin protein ligase 1

IP:

Immunoprecipitation

IF:

Immunofluorescence

LC3:

Microtubule associated protein 1 light chain 3 alpha

MCL-1:

Myeloid cell leukemia 1

MET:

Tyrosine-protein kinase Met

mTORC1:

Mammalian target of rapamycin complex 1

PDGFR:

Platelet-derived growth factor receptor

PFS:

Progression free survival

RCC:

Renal cell carcinoma

TKI:

Tyrosine kinase inhibitor

USP9X:

Ubiquitin specific peptidase 9 X-linked

VEGFR:

Vascular endothelial growth factor receptor

WB:

Western blot

References

  1. B. Shuch, A. Amin, A.J. Armstrong, J.N. Eble, V. Ficarra, A. Lopez-Beltran et al., Understanding pathologic variants of renal cell carcinoma: distilling therapeutic opportunities from biologic complexity. Europ. Urol. 67, 85–97 (2015). https://doi.org/10.1016/j.eururo.2014.04.029

    Article  Google Scholar 

  2. J.S. Lam, J.T. Leppert, A.S. Belldegrun, R.A. Figlin, Novel approaches in the therapy of metastatic renal cell carcinoma. World J. Urol. 23, 202–212 (2005). https://doi.org/10.1007/s00345-004-0466-0

    Article  CAS  PubMed  Google Scholar 

  3. N.K. Janzen, H.L. Kim, R.A. Figlin, A.S. Belldegrun, Surveillance after radical or partial nephrectomy for localized renal cell carcinoma and management of recurrent disease. Urologic Clini. North Am. 30, 843–852 (2003). https://doi.org/10.1016/s0094-0143(03)00056-9

    Article  Google Scholar 

  4. A.M. Molina, X. Lin, B. Korytowsky, E. Matczak, M.J. Lechuga, R. Wiltshire et al., Sunitinib objective response in metastatic renal cell carcinoma: analysis of 1059 patients treated on clinical trials. Eur. J. Cancer 50, 351–358 (2014). https://doi.org/10.1016/j.ejca.2013.08.021

    Article  CAS  PubMed  Google Scholar 

  5. S. Bracarda, R. Iacovelli, L. Boni, M. Rizzo, L. Derosa, M. Rossi et al., Sunitinib administered on 2/1 schedule in patients with metastatic renal cell carcinoma: the RAINBOW analysis. Ann. Oncol. 26, 2107–2113 (2015). https://doi.org/10.1093/annonc/mdv315

    Article  CAS  PubMed  Google Scholar 

  6. M.E. Gore, C. Szczylik, C. Porta, S. Bracarda, G.A. Bjarnason, S. Oudard et al., Safety and efficacy of sunitinib for metastatic renal-cell carcinoma: an expanded-access trial. Lancet Oncol. 10, 757–763 (2009). https://doi.org/10.1016/s1470-2045(09)70162-7

    Article  CAS  PubMed  Google Scholar 

  7. B.E. Houk, C.L. Bello, B. Poland, L.S. Rosen, G.D. Demetri, R.J. Motzer, Relationship between exposure to sunitinib and efficacy and tolerability endpoints in patients with cancer: results of a pharmacokinetic/pharmacodynamic meta-analysis. Cancer Chemother. Pharmacol. 66, 357–371 (2010). https://doi.org/10.1007/s00280-009-1170-y

    Article  CAS  PubMed  Google Scholar 

  8. D.R. Feldman, M.S. Baum, M.S. Ginsberg, H. Hassoun, C.D. Flombaum, S. Velasco et al., Phase I trial of bevacizumab plus escalated doses of sunitinib in patients with metastatic renal cell carcinoma. J. Clin. Oncol. 27, 1432–1439 (2009). https://doi.org/10.1200/jco.2008.19.0108

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. P.H. Patel, P.L. Senico, R.E. Curiel, R.J. Motzer, Phase I study combining treatment with temsirolimus and sunitinib malate in patients with advanced renal cell carcinoma. Clin. Genitourinary Cancer 7, 24–27 (2009). https://doi.org/10.3816/CGC.2009.n.004

    Article  CAS  Google Scholar 

  10. A. Amin, E.R. Plimack, M.S. Ernstoff, L.D. Lewis, T.M. Bauer, D.F. McDermott et al., Safety and efficacy of nivolumab in combination with sunitinib or pazopanib in advanced or metastatic renal cell carcinoma: the CheckMate 016 study. J. Immunother. Cancer 6 (2018). https://doi.org/10.1186/s40425-018-0420-0

  11. M. Elgendy, A.K. Abdel-Aziz, S.L. Renne, V. Bornaghi, G. Procopio, M. Colecchia et al., Dual modulation of MCL-1 and mTOR determines the response to sunitinib. J. Clin. Investig. 127, 153–168 (2017). https://doi.org/10.1172/jci84386

    Article  PubMed  Google Scholar 

  12. S. Giuliano, Y. Cormerais, M. Dufies, R. Grepin, P. Colosetti, A. Belaid et al., Resistance to sunitinib in renal clear cell carcinoma results from sequestration in lysosomes and inhibition of the autophagic flux. Autophagy 11, 1891–1904 (2015). https://doi.org/10.1080/15548627.2015.1085742

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. L. DeVorkin, M. Hattersley, P. Kim, J. Ries, J. Spowart, M.S. Anglesio et al., Autophagy inhibition enhances sunitinib efficacy in clear cell ovarian carcinoma. Mol. Cancer Res. 15, 250–258 (2017). https://doi.org/10.1158/1541-7786.Mcr-16-0132

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. T. Ikeda, K. Ishii, Y. Saito, M. Miura, A. Otagiri, Y. Kawakami et al., Inhibition of autophagy enhances sunitinib-induced cytotoxicity in rat pheochromocytoma PC12 cells. J. Pharmacol. Sci. 121, 67–73 (2013). https://doi.org/10.1254/jphs.12158FP

    Article  CAS  PubMed  Google Scholar 

  15. T. Wiedmer, A. Blank, S. Pantasis, L. Normand, R. Bill, P. Krebs et al., Autophagy inhibition improves sunitinib efficacy in pancreatic neuroendocrine tumors via a lysosome-dependent mechanism. Mol. Cancer Ther. 16, 2502–2515 (2017). https://doi.org/10.1158/1535-7163.Mct-17-0136

    Article  CAS  PubMed  Google Scholar 

  16. K.J. Gotink, H.J. Broxterman, M. Labots, R.R. de Haas, H. Dekker, R.J. Honeywell et al., Lysosomal sequestration of sunitinib: a novel mechanism of drug resistance. Clin. Cancer Res. 17, 7337–7346 (2011). https://doi.org/10.1158/1078-0432.ccr-11-1667

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. M. Elgendy, C. Sheridan, G. Brumatti, S.J. Martin, Oncogenic ras-induced expression of noxa and beclin-1 promotes autophagic cell death and limits clonogenic survival. Molecular Cell 42, 23–35 (2011). https://doi.org/10.1016/j.molcel.2011.02.009

    Article  CAS  PubMed  Google Scholar 

  18. M.C. Maiuri, G. Le Toumelin, A. Criollo, J.C. Rain, F. Gautier, P. Juin et al., Functional and physical interaction between Bcl-X-L and a BH3-like domain in Beclin-1. EMBO J. 26, 2527–2539 (2007). https://doi.org/10.1038/sj.emboj.7601689

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. M. Elgendy, M. Ciro, A.K. Abdel-Aziz, G. Belmonte, R. Dal Zuffo, C. Mercurio et al., Beclin 1 restrains tumorigenesis through Mcl-1 destabilization in an autophagy-independent reciprocal manner. Nat. Commun. 5 (2014). https://doi.org/10.1038/ncomms6637

  20. J.R. Wisniewski, A. Zougman, N. Nagaraj, M. Mann, Universal sample preparation method for proteome analysis. Nature Methods 6, 359–U360 (2009). https://doi.org/10.1038/nmeth.1322

    Article  CAS  PubMed  Google Scholar 

  21. M. Zarei, A. Sprenger, M. Rackiewicz, J. Dengjel, Fast and easy phosphopeptide fractionation by combinatorial ERLIC-SCX solid-phase extraction for in-depth phosphoproteome analysis. Nature Protocols 11, 37–45 (2016). https://doi.org/10.1038/nprot.2015.134

    Article  CAS  PubMed  Google Scholar 

  22. T.C. Chou, P. Talalay, Quantitative analysis of dose-effect relationships: the combined effects of multiple drugs or enzyme inhibitors. Adv. Enzyme Regul. 22, 27–55 (1984). https://doi.org/10.1016/0065-2571(84)90007-4

    Article  CAS  PubMed  Google Scholar 

  23. Z.J. Jin, About the evaluation of drug combination. Acta Pharmacol. Sin. 25, 146–147 (2004)

    CAS  PubMed  Google Scholar 

  24. D. Huang, Y. Ding, Y. Li, W.M. Luo, Z.F. Zhang, J. Snider et al., Sunitinib acts primarily on tumor endothelium rather than tumor cells to inhibit the growth of renal cell carcinoma. Cancer Res. 70, 1053–1062 (2010). https://doi.org/10.1158/0008-5472.Can-09-3722

    Article  CAS  PubMed  Google Scholar 

  25. Q. Ding, X. He, J.M. Hsu, W. Xia, C.T. Chen, L.Y. Li et al., Degradation of Mcl-1 by beta-TrCP mediates glycogen synthase kinase 3-induced tumor suppression and chemosensitization. Mol. Cell Biol. 27, 4006–4017 (2007). https://doi.org/10.1128/mcb.00620-06

    Article  CAS  PubMed  Google Scholar 

  26. A.M. Domina, J.A. Vrana, M.A. Gregory, S.R. Hann, R.W. Craig, MCL1 is phosphorylated in the PEST region and stabilized upon ERK activation in viable cells, and at additional sites with cytotoxic okadaic acid or taxol. Oncogene 23, 5301–5315 (2004). https://doi.org/10.1038/sj.onc.1207692

    Article  CAS  PubMed  Google Scholar 

  27. W.T. Tai, C.W. Shiau, H.L. Chen, C.Y. Liu, C.S. Lin, A.L. Cheng et al., Mcl-1-dependent activation of Beclin 1 mediates autophagic cell death induced by sorafenib and SC-59 in hepatocellular carcinoma cells. Cell Death Dis. 4, e485 (2013). https://doi.org/10.1038/cddis.2013.18

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. G. Marino, M. Niso-Santano, E.H. Baehrecke, G. Kroemer, Self-consumption: the interplay of autophagy and apoptosis. Nat. Rev. Mol. Cell Biol. 15, 81–94 (2014). https://doi.org/10.1038/nrm3735

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. S. Giuliano, Y. Cormerais, M. Dufies, R. Grépin, P. Colosetti, A. Belaid et al., Resistance to sunitinib in renal clear cell carcinoma results from sequestration in lysosomes and inhibition of the autophagic flux. Autophagy 11, 1891–1904 (2015). https://doi.org/10.1080/15548627.2015.1085742

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. D.J. Klionsky, A.K. Abdel-Aziz, S. Abdelfatah, M. Abdellatif, A. Abdoli, S. Abel et al., Guidelines for the use and interpretation of assays for monitoring autophagy (4th edn). Autophagy 17, 1–382 (2021). https://doi.org/10.1080/15548627.2020.1797280

    Article  PubMed  PubMed Central  Google Scholar 

  31. R.J. Motzer, E. Jonasch, N. Agarwal, A. Alva, M. Baine, K. Beckermann et al., Kidney cancer, version 3.2022. J. Natl. Compr. Cancer Network 20, 71–89 (2022). https://doi.org/10.6004/jnccn.2022.0001

    Article  CAS  Google Scholar 

  32. L. Qu, J. Ding, C. Chen, Z.J. Wu, B. Liu, Y. Gao et al., Exosome-transmitted lncARSR promotes sunitinib resistance in renal cancer by acting as a competing endogenous RNA. Cancer Cell 29, 653–668 (2016). https://doi.org/10.1016/j.ccell.2016.03.004

    Article  CAS  PubMed  Google Scholar 

  33. F. Shojaei, J.H. Lee, B.H. Simmons, A. Wong, C.O. Esparza, P.A. Plumlee et al., HGF/c-met acts as an alternative angiogenic pathway in sunitinib-resistant tumors. Cancer Res. 70, 10090–10100 (2010). https://doi.org/10.1158/0008-5472.Can-10-0489

    Article  CAS  PubMed  Google Scholar 

  34. L. Zhou, X.D. Liu, M. Sun, X. Zhang, P. German, S. Bai et al., Targeting MET and AXL overcomes resistance to sunitinib therapy in renal cell carcinoma. Oncogene 35, 2687–2697 (2016). https://doi.org/10.1038/onc.2015.343

    Article  CAS  PubMed  Google Scholar 

  35. T.K. Choueiri, S. Halabi, B.L. Sanford, O. Hahn, M.D. Michaelson, M.K. Walsh et al., Cabozantinib versus sunitinib as initial targeted therapy for patients with metastatic renal cell carcinoma of poor or intermediate risk: the alliance A031203 CABOSUN trial. J. Clin. Oncol. 35, 591 (2017). https://doi.org/10.1200/jco.2016.70.7398

  36. J. Deng, Q. Zhang, L.P. Lv, P. Ma, Y.Y. Zhang, N. Zhao et al., Identification of an autophagy-related gene signature for predicting prognosis and immune activity in pancreatic adenocarcinoma. Sci. Rep. 12 (2022). https://doi.org/10.1038/s41598-022-11050-w

  37. Y. Ekim, S. Kara, B. Gencer, T. Karaca, Efficacy of sunitinib, sunitinib-hesperetin, and sunitinib-doxycycline combinations on experimentally-induced corneal neovascularization. Curr. Eye Res. 44, 590–598 (2019). https://doi.org/10.1080/02713683.2019.1584320

    Article  CAS  PubMed  Google Scholar 

  38. F. Rossari, F. Minutolo, E. Orciuolo, Past, present, and future of Bcr-Abl inhibitors: from chemical development to clinical efficacy. J. Hematol. Oncol. 11 (2018). https://doi.org/10.1186/s13045-018-0624-2

  39. D.B. Mendel, A.D. Laird, X.H. Xin, S.G. Louie, J.G. Christensen, G.M. Li et al., In vivo antitumor activity of SU11248, a novel tyrosine kinase inhibitor targeting vascular endothelial growth factor and platelet-derived growth factor receptors: determination of a pharmacokinetic/pharmacodynamic relationship. Clin. Cancer Res. 9, 327–337 (2003)

    CAS  PubMed  Google Scholar 

  40. S. Takasaki, Y. Kawasaki, M. Kikuchi, M. Tanaka, M. Suzuka, A. Noda et al., Relationships between sunitinib plasma concentration and clinical outcomes in Japanese patients with metastatic renal cell carcinoma. Int. J. Clin. Oncol. 23, 936–943 (2018). https://doi.org/10.1007/s10147-018-1302-7

    Article  CAS  PubMed  Google Scholar 

  41. M. Golemovic, S. Verstovsek, F. Giles, J. Cortes, T. Manshouri, P.W. Manley et al., AMN107, a novel aminopyrimidine inhibitor of Bcr-Abl, has in vitro activity against imatinib-resistant chronic myeloid leukemia. Clin. Cancer Res. 11, 4941–4947 (2005). https://doi.org/10.1158/1078-0432.Ccr-04-2601

    Article  CAS  PubMed  Google Scholar 

  42. F. Strappazzon, A. Di Rita, A. Peschiaroli, P.P. Leoncini, F. Locatelli, G. Melino et al., HUWE1 controls MCL1 stability to unleash AMBRA1-induced mitophagy. Cell Death Differ. 27, 1155–1168 (2020). https://doi.org/10.1038/s41418-019-0404-8

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

Not applicable.

Funding

This project was supported by the National Natural Science Foundation of China (NSFC) (No. 82172812 to R.L. and No. 82272743 to X.Y.), The Natural Science Foundation of Guangdong Province (No. 2022A1515012321 to H.H. and No. 2021A1515012496 to X.Y.), The Enterprise United Foundation of Guangdong Province (No. 2021A1515220182 to H.H.).

Author information

Author notes

  1. Tingyu Liu, Xin Yue and Xue Chen have contributed equally to this study.

Authors and Affiliations

  1. State Key Laboratory of Oncology in South China, Guangdong Provincial Clinical Research Center for Cancer, Sun Yat-sen University Cancer Center, Guangzhou, 510060, China

    Tingyu Liu, Xue Chen, Ru Yan, Chong Wu, Hui Han & Ran-Yi Liu

  2. Institute of Precision Medicine, The First Affiliated Hospital, Sun Yat-Sen University, Guangzhou, 510080, China

    Xin Yue

  3. School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou, 510006, China

    Yunzhi Li & Xianzhang Bu

Authors
  1. Tingyu Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Xin Yue
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Xue Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ru Yan
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Chong Wu
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Yunzhi Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Xianzhang Bu
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Hui Han
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Ran-Yi Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Study concept and design: X.Y., R.L. and H.H.; Performed in vitro functional studies and analyzed data: T.L., X.C. and X.Y.; Performed animal experiments and analyzed data: T.L., R.Y. and C.W.; Generated critical cellular tools: T.L. and X.C.; Performed mechanistical studies and analyzed data: T.L., X.Y., X.C. and Y.L.; Statistic analysis: T.L. and X.Y.; Supervision of data: X.B.; Analysis and interpretation of data and writing of the manuscript: T.L., X.Y., R.L, and H.H. All authors have read and approved the manuscript.

Corresponding authors

Correspondence to Xin Yue, Hui Han or Ran-Yi Liu.

Ethics declarations

Ethics approval and consent to participate

Animal experiments were approved by the Sun Yat-Sen University Cancer Center Institutional Animal Care and Usage Committee following the Animal Welfare and Rights in China (Approval number: L102012019040I). All the experiments were conducted in the Laboratory Animal Center of Sun Yat-sen University.

Consent for publication

Not applicable.

Conflict of interest

None of the authors have any potential conflicts to disclose.

Additional information

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Liu, T., Yue, X., Chen, X. et al. Nilotinib in combination with sunitinib renders MCL-1 for degradation and activates autophagy that overcomes sunitinib resistance in renal cell carcinoma. Cell Oncol. (2024). https://doi.org/10.1007/s13402-024-00927-9

Download citation

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1007/s13402-024-00927-9

Keywords

Search

Navigation