CDK4/6 and BET inhibitors synergistically suppress pancreatic tumor growth and epithelial-to-mesenchymal transition by regulating the GSK3β-mediated Wnt/β-catenin pathway

Abstract

Aim: Cyclin-dependent kinases 4 and 6 (CDK4/6) are frequently upregulated in pancreatic ductal adenocarcinoma (PDAC) and are associated with poor overall survival. Although CDK4/6 inhibition suppresses tumor cell proliferation, it paradoxically promotes metastasis and invasion, and the mechanisms underlying this effect remain unclear.

Methods: We evaluated the effects of the CDK4/6 inhibitor palbociclib (PD-0332991) and the bromodomain and extra-terminal (BET) inhibitor JQ1, administered individually and in combination, on human PDAC cell lines in vitro and on tumor growth in an orthotopic mouse model.

Results: Palbociclib modestly inhibited pancreatic tumor growth but significantly enhanced tumor cell migration, invasion, and epithelial-to-mesenchymal transition (EMT). In contrast, co-treatment with JQ1 potentiated palbociclib’s anti-proliferative effects and reversed EMT. Mechanistically, CDK4/6 inhibition activated the canonical Wnt/β-catenin pathway via Ser9 phosphorylation of GSK3β, whereas BET inhibition disrupted the cross-talk between Wnt/β-catenin and TGF-β/Smad signaling. Combined inhibition of CDK4/6 and BET produced a synergistic antitumor effect in vitro and in vivo.

Conclusion: Our findings support a combined therapeutic strategy targeting CDK4/6 and BET proteins to achieve synergistic inhibition of PDAC progression.

Keywords

Pancreatic cancer / pancreatic ductal adenocarcinoma / epithelial-to-mesenchymal transition / CDK4/6 inhibitor PD-0332991 / BET inhibitor JQ1 / Wnt/β-catenin pathway

References

[1] Siegel RL,Fuchs HE.Cancer statistics, 2022.CA Cancer J Clin2022;72:7-33

[2] Khalaf N,Abrams HR.Burden of pancreatic cancer: from epidemiology to practice.Clin Gastroenterol Hepatol2021;19:876-84 PMCID:PMC8559554

[3] Cai J,Lu M.Advances in the epidemiology of pancreatic cancer: trends, risk factors, screening, and prognosis.Cancer Lett2021;520:1-11

[4] Buscail L,Cordelier P.Role of oncogenic KRAS in the diagnosis, prognosis and treatment of pancreatic cancer.Nat Rev Gastroenterol Hepatol2020;17:153-68

[5] De Luca A, Maiello MR, D’Alessio A, Pergameno M, Normanno N. The RAS/RAF/MEK/ERK and the PI3K/AKT signalling pathways: role in cancer pathogenesis and implications for therapeutic approaches.Expert Opin Ther Targets2012;16 Suppl 2:S17-27

[6] Bertrand FE.The cross-talk of NOTCH and GSK-3 signaling in colon and other cancers.Biochim Biophys Acta Mol Cell Res2020;1867:118738

[7] Hong DS,Strickler JH.KRAS^G12C inhibition with sotorasib in advanced solid tumors.N Engl J Med2020;383:1207-17 PMCID:PMC7571518

[8] Cicenas J,Meskinyte I,Skeberdyte A.KRAS, TP53, CDKN2A, SMAD4, BRCA1, and BRCA2 mutations in pancreatic cancer.Cancers2017;9:42 PMCID:PMC5447952

[9] Rice A.The mutational landscape of pancreatic and liver cancers, as represented by circulating tumor DNA.Front Oncol2019;9:952 PMCID:PMC6769086

[10] Chiorean EG.Pancreatic cancer: optimizing treatment options, new, and emerging targeted therapies.Drug Des Devel Ther2015;9:3529-45 PMCID:PMC4500614

[11] Bailey P, Chang DK, Nones K, et al; Australian Pancreatic Cancer Genome Initiative. Genomic analyses identify molecular subtypes of pancreatic cancer. Nature. 2016;531:47-52.

[12] Serrano M.The tumor suppressor protein p16INK4a.Exp Cell Res1997;237:7-13

[13] Cheung EC.The role of ROS in tumour development and progression.Nat Rev Cancer2022;22:280-97

[14] Goel S,Zhao JJ.Targeting CDK4 and CDK6 in cancer.Nat Rev Cancer2022;22:356-72 PMCID:PMC9149100

[15] Du Q,Wang M,Sun X.The application and prospect of CDK4/6 inhibitors in malignant solid tumors.J Hematol Oncol2020;13:41 PMCID:PMC7195725

[16] Goel S,McAllister SS.CDK4/6 inhibition in cancer: beyond cell cycle arrest.Trends Cell Biol2018;28:911-25 PMCID:PMC6689321

[17] Chi S,Li Y.BET inhibitor and CDK4/6 inhibitor synergistically inhibit breast cancer by suppressing BRD4 stability and DNA damage repair.Transl Oncol2025;51:102212 PMCID:PMC11629338

[18] Rampioni Vinciguerra GL,Segatto I.CDK4/6 inhibitors in combination therapies: better in company than alone: a mini review.Front Oncol2022;12:891580 PMCID:PMC9197541

[19] Toogood PL,Repine JT.Discovery of a potent and selective inhibitor of cyclin-dependent kinase 4/6.J Med Chem2005;48:2388-406

[20] Gelbert LM,Lin X.Preclinical characterization of the CDK4/6 inhibitor LY2835219: in-vivo cell cycle-dependent/independent anti-tumor activities alone/in combination with gemcitabine.Invest New Drugs2014;32:825-37 PMCID:PMC4169866

[21] Rader J,Hart LS.Dual CDK4/CDK6 inhibition induces cell-cycle arrest and senescence in neuroblastoma.Clin Cancer Res2013;19:6173-82 PMCID:PMC3844928

[22] Palafox M,Bellet M.High p16 expression and heterozygous RB1 loss are biomarkers for CDK4/6 inhibitor resistance in ER⁺ breast cancer.Nat Commun2022;13:5258 PMCID:PMC9452562

[23] Wang L,Kao CY,Tsai MJ.Small molecule JQ1 promotes prostate cancer invasion via BET-independent inactivation of FOXA1.J Clin Invest2020;130:1782-92 PMCID:PMC7108891

[24] Pagliarini R,Sellers WR.Oncogene addiction: pathways of therapeutic response, resistance, and road maps toward a cure.EMBO Rep2015;16:280-96 PMCID:PMC4364868

[25] Wu C,Pilié PG.PARP and CDK4/6 inhibitor combination therapy induces apoptosis and suppresses neuroendocrine differentiation in prostate cancer.Mol Cancer Ther2021;20:1680-91 PMCID:PMC8456452

[26] Malumbres M.CDK4/6 inhibitors: what is the best cocktail?.Clin Cancer Res2019;25:6-8

[27] Wang M,Liao X.The SGK3-Catalase antioxidant signaling axis drives cervical cancer growth and therapy resistance.Redox Biol2023;67:102931 PMCID:PMC10623367

[28] Tang H,You T.A real-world analysis of trametinib in combination with hydroxychloroquine or CDK4/6 inhibitor as third- or later-line therapy in metastatic pancreatic adenocarcinoma.BMC Cancer2023;23:958 PMCID:PMC10563303

[29] Alqahtani A,Ashraf M.Bromodomain and extra-terminal motif inhibitors: a review of preclinical and clinical advances in cancer therapy.Future Sci OA2019;5:FSO372 PMCID:PMC6426170

[30] Shi C,Liu Y,Ren G.Bromodomain and extra-terminal motif (BET) inhibition is synthetic lethal with loss of SMAD4 in colorectal cancer cells via restoring the loss of MYC repression.Oncogene2021;40:937-50

[31] Duan Q,Anand P.BET bromodomain inhibition suppresses innate inflammatory and profibrotic transcriptional networks in heart failure.Sci Transl Med2017;9:eaah5084 PMCID:PMC5544253

[32] Mazur PK,Mello SS.Combined inhibition of BET family proteins and histone deacetylases as a potential epigenetics-based therapy for pancreatic ductal adenocarcinoma.Nat Med2015;21:1163-71 PMCID:PMC4959788

[33] Miller AL,Vance RB.BET inhibition decreases HMGCS2 and sensitizes resistant pancreatic tumors to gemcitabine.Cancer Lett2024;592:216919 PMCID:PMC11309032

[34] Zhang Y,Zhao C.CAFs homologous biomimetic liposome bearing BET inhibitor and pirfenidone synergistically promoting antitumor efficacy in pancreatic ductal adenocarcinoma.Adv Sci2024;11:e2305279 PMCID:PMC10767438

[35] Yamamoto K,Kudo Y.Stromal remodeling by the BET bromodomain inhibitor JQ1 suppresses the progression of human pancreatic cancer.Oncotarget2016;7:61469-84 PMCID:PMC5308665

[36] Mueller S,Maresch R.Evolutionary routes and KRAS dosage define pancreatic cancer phenotypes.Nature2018;554:62-8 PMCID:PMC6097607

[37] Ritchie ME,Wu D.limma powers differential expression analyses for RNA-sequencing and microarray studies.Nucleic Acids Res2015;43:e47 PMCID:PMC4402510

[38] Silvis MR,Rohweder R.MYC-mediated resistance to trametinib and HCQ in PDAC is overcome by CDK4/6 and lysosomal inhibition.J Exp Med2023;220:e20221524 PMCID:PMC9930170

[39] Wang Y,Liu G.Disruption of super-enhancers in activated pancreatic stellate cells facilitates chemotherapy and immunotherapy in pancreatic cancer.Adv Sci2024;11:e2308637 PMCID:PMC11040371

[40] DiRenzo DM,Shi X.A crosstalk between TGF-β/Smad3 and Wnt/β-catenin pathways promotes vascular smooth muscle cell proliferation.Cell Signal2016;28:498-505 PMCID:PMC4788971

[41] Loganathan SN,Holler AE,Wang J.Targeting the IGF1R/PI3K/AKT pathway sensitizes ewing sarcoma to BET bromodomain inhibitors.Mol Cancer Ther2019;18:929-36 PMCID:PMC6497538

[42] Franco J,Knudsen ES.CDK4/6 inhibitors have potent activity in combination with pathway selective therapeutic agents in models of pancreatic cancer.Oncotarget2014;5:6512-25 PMCID:PMC4171647

[43] Al Baghdadi T,Garrett-Mayer E.Palbociclib in patients with pancreatic and biliary cancer with CDKN2A alterations: results from the targeted agent and profiling utilization registry study.JCO Precis Oncol2019;3:1-8

[44] Ogata R,Saitoh W,Kurebayashi J.Resistance to cyclin-dependent kinase (CDK) 4/6 inhibitors confers cross-resistance to other CDK inhibitors but not to chemotherapeutic agents in breast cancer cells.Breast Cancer2021;28:206-15 PMCID:PMC7796879

[45] Salvador-Barbero B,Zapatero-Solana E.CDK4/6 inhibitors impair recovery from cytotoxic chemotherapy in pancreatic adenocarcinoma.Cancer Cell2020;37:340-53.e6

[46] Jacobs KM,Ferraro DJ,Hallahan DE.GSK-3β: a bifunctional role in cell death pathways.Int J Cell Biol2012;2012:930710 PMCID:PMC3364548

[47] Armanious H,Gelebart P,Mackey J.Clinical and biological significance of GSK-3β inactivation in breast cancer-an immunohistochemical study.Hum Pathol2010;41:1657-63

[48] Kisoh K,Itoh T.Involvement of GSK-3β phosphorylation through PI3-K/Akt in cerebral ischemia-induced neurogenesis in rats.Mol Neurobiol2017;54:7917-27 PMCID:PMC5684253

[49] Hermida MA,Leslie NR.GSK3 and its interactions with the PI3K/AKT/mTOR signalling network.Adv Biol Regul2017;65:5-15