N⁶-Methyladenosine modification mediated by METTL3 promotes DNA-PKcs expression to induce anlotinib resistance in osteosarcoma

Abstract

	•DNA-PKcs knockdown heightens osteosarcoma sensitivity to anlotinib.
	•DNA-PKcs modulates anlotinib-induced protective autophagy through interacts with Beclin-1 and regulates its ubiquitination.
	•m6A modification of OLE_LINK82PRKDC mRNA induced by METTL3 contributes to anlotinib resistance in osteosarcoma.
	•m6A methylation of PRKDC mRNA recognised by YTHDF1 amplifies the expression of DNA-PKcs.

Keywords

anlotinib / DNA-PKcs / m ⁶A / METTL3 / osteosarcoma

References

[1] RitterJ, Bielack SS. Osteosarcoma. Ann Oncol. 2010; 21(7): vii320-5.

[2] ChenC, XieL, RenT, HuangY, XuJ, GuoW. Immunotherapy for osteosarcoma: fundamental mechanism, rationale, and recent breakthroughs. Cancer Lett. 2021; 500: 1-10.

[3] HarrisonDJ, GellerDS, GillJD, Lewis VO, GorlickR. Current and future therapeutic approaches for osteosarcoma. Expert Rev Anticancer Ther. 2018; 18: 39-50.

[4] LilienthalI, HeroldN. Targeting molecular mechanisms underlying treatment efficacy and resistance in osteosarcoma: a review of current and future strategies. Int J Mol Sci. 2020; 21.

[5] WangG, SunM, JiangY, et al. Anlotinib, a novel small molecular tyrosine kinase inhibitor, suppresses growth and metastasis via dual blockade of VEGFR2 and MET in osteosarcoma. Int J Cancer. 2019; 145: 979-993.

[6] LiangL, HuiK, HuC, et al. Autophagy inhibition potentiates the anti-angiogenic property of multikinase inhibitor anlotinib through JAK2/STAT3/VEGFA signaling in non-small cell lung cancer cells. J Exp Clin Cancer Res. 2019; 38: 71.

[7] HanB, LiK, ZhaoY, et al. Anlotinib as a third-line therapy in patients with refractory advanced non-small-cell lung cancer: a multicentre, randomised phase II trial (ALTER0302). Br J Cancer. 2018; 118: 654-661.

[8] HanB, LiK, WangQ, et al. Effect of anlotinib as a third-line or further treatment on overall survival of patients with advanced non-small cell lung cancer: the ALTER 0303 Phase 3 Randomized Clinical Trial. JAMA Oncol. 2018; 4: 1569-1575.

[9] SunY, NiuW, DuF, et al. Safety, pharmacokinetics, and antitumor properties of anlotinib, an oral multi-target tyrosine kinase inhibitor, in patients with advanced refractory solid tumors. J Hematol Oncol. 2016; 9: 105.

[10] LiuZ, GaoS, ZhuL, et al. Efficacy and safety of anlotinib in patients with unresectable or metastatic bone sarcoma: a retrospective multiple institution study. Cancer Med. 2021; 10: 7593-7600.

[11] ChenQ, ZhengK, XuM, et al. Anlotinib combined with radiotherapy and chemotherapy for recurrent pelvic osteosarcoma treatment: a case report and literature review. Front Oncol. 2023; 13: 1283932.

[12] WangG, CaoL, JiangY, et al. Anlotinib reverses multidrug resistance (MDR) in osteosarcoma by inhibiting P-Glycoprotein (PGP1) function in vitro and in vivo. Front Pharmacol. 2021; 12: 798837.

[13] ZhangLL, LuJ, LiuRQ, et al. Chromatin accessibility analysis reveals that TFAP2A promotes angiogenesis in acquired resistance to anlotinib in lung cancer cells. Acta Pharmacol Sin. 2020; 41: 1357-1365.

[14] GuG, HuC, HuiK, et al. Exosomal miR-136-5p derived from anlotinib-resistant NSCLC cells confers anlotinib resistance in non-small cell lung cancer through targeting PPP2R2A. Int J Nanomedicine. 2021; 16: 6329-6343.

[15] JinZ, LuY, WuX, et al. The cross-talk between tumor cells and activated fibroblasts mediated by lactate/BDNF/TrkB signaling promotes acquired resistance to anlotinib in human gastric cancer. Redox Biol. 2021; 46: 102076.

[16] XuJ, ChenC, SunK, et al. Tocilizumab (monoclonal anti-IL-6R antibody) reverses anlotinib resistance in osteosarcoma. Front Oncol. 2023; 13: 1192472.

[17] ChenC, GuoY, HuangQ, et al. PI3K inhibitor impairs tumor progression and enhances sensitivity to anlotinib in anlotinib-resistant osteosarcoma. Cancer Lett. 2022; 536: 215660.

[18] RoundtreeIA, EvansME, PanT, HeC. Dynamic RNA modifications in gene expression regulation. Cell. 2017; 169: 1187-1200.

[19] WangCX, CuiGS, LiuX, et al. METTL3-mediated m6A modification is required for cerebellar development. PLoS Biol. 2018; 16.

[20] PingXL, SunBF, WangL, et al. Mammalian WTAP is a regulatory subunit of the RNA N6-methyladenosine methyltransferase. Cell Res. 2014; 24: 177-189.

[21] YangY, HsuPJ, ChenYS, Yang YG. Dynamic transcriptomic m(6)A decoration: writers, erasers, readers and functions in RNA metabolism. Cell Res. 2018; 28: 616-624.

[22] ZhouL, YangC, ZhangN, Zhang X, ZhaoT, YuJ. Silencing METTL3 inhibits the proliferation and invasion of osteosarcoma by regulating ATAD2. Biomed Pharmacother. 2020; 125: 109964.

[23] HanJ, WangJZ, YangX, et al. METTL3 promote tumor proliferation of bladder cancer by accelerating pri-miR221/222 maturation in m6A-dependent manner. Mol Cancer. 2019; 18: 110.

[24] TassinariV, Cesarini V, TomaselliS, et al. ADAR1 is a new target of METTL3 and plays a pro-oncogenic role in glioblastoma by an editing-independent mechanism. Genome Biol. 2021; 22: 51.

[25] ZhaoW, CuiY, LiuL, et al. METTL3 facilitates oral squamous cell carcinoma tumorigenesis by enhancing c-Myc stability via YTHDF1-mediated m(6)A modification. Mol Ther Nucleic Acids. 2020; 20: 1-12.

[26] LiuL, WuY, LiQ, et al. METTL3 promotes tumorigenesis and metastasis through BMI1 m(6)A methylation in oral squamous cell carcinoma. Mol Ther. 2020; 28: 2177-2190.

[27] LiK, LiX, TianJ, Wang H, PanJ, LiJ. Downregulation of DNA-PKcs suppresses P-gp expression via inhibition of the Akt/NF-κB pathway in CD133-positive osteosarcoma MG-63 cells. Oncol Rep. 2016; 36: 1973-1980.

[28] LiX, TianJ, BoQ, et al. Targeting DNA-PKcs increased anticancer drug sensitivity by suppressing DNA damage repair in osteosarcoma cell line MG63. Tumour Biol. 2015; 36: 9365-9372.

[29] ZhangY, ShenG, MengT, et al. Eicosapentaenoic acid enhances the sensitivity of osteosarcoma to cisplatin by inducing ferroptosis through the DNA-PKcs/AKT/NRF2 pathway and reducing PD-L1 expression to attenuate immune evasion. Int Immunopharmacol. 2023; 125: 111181.

[30] XuJ, WanZ, TangM, et al. N(6)-methyladenosine-modified CircRNA-SORE sustains sorafenib resistance in hepatocellular carcinoma by regulating β-catenin signaling. Mol Cancer. 2020; 19: 163.

[31] FuY, Dominissini D, RechaviG, HeC. Gene expression regulation mediated through reversible m⁶A RNA methylation. Nat Rev Genet. 2014; 15: 293-306.

[32] ShiH, WangX, LuZ, et al. YTHDF3 facilitates translation and decay of N(6)-methyladenosine-modified RNA. Cell Res. 2017; 27: 315-328.

[33] XieL, XuJ, SunX, et al. Apatinib for advanced osteosarcoma after failure of standard multimodal therapy: an open label Phase II clinical trial. Oncologist. 2019; 24: e542-e50.

[34] PascaleRM, JosephC, LatteG, Evert M, FeoF, CalvisiDF. DNA-PKcs: a promising therapeutic target in human hepatocellular carcinoma? DNA Repair (Amst). 2016; 47: 12-20.

[35] LeeYR, KangGS, OhT, JoHJ, ParkHJ, Ahn GO. DNA-dependent protein kinase catalytic subunit (DNA-PKcs): beyond the DNA Double-Strand Break Repair. Mol Cells. 2023; 46: 200-205.

[36] WilloughbyCE, JiangY, ThomasHD, et al. Selective DNA-PKcs inhibition extends the therapeutic index of localized radiotherapy and chemotherapy. J Clin Invest. 2020; 130: 258-271.

[37] GurungRL, LimHK, VenkatesanS, Lee PS, HandeMP. Targeting DNA-PKcs and telomerase in brain tumour cells. Mol Cancer. 2014; 13: 232.

[38] LiuL, DengY, ZhengZ, et al. Hsp90 inhibitor STA9090 sensitizes hepatocellular carcinoma to hyperthermia-induced DNA damage by suppressing DNA-PKcs protein stability and mRNA transcription. Mol Cancer Ther. 2021; 20: 1880-1892.

[39] FangX, HuangZ, ZhaiK, et al. Inhibiting DNA-PK induces glioma stem cell differentiation and sensitizes glioblastoma to radiation in mice. Sci Transl Med. 2021; 13.

[40] LiuJJ, LinM, YuJY, LiuB, BaoJK. Targeting apoptotic and autophagic pathways for cancer therapeutics. Cancer Lett. 2011; 300: 105-114.

[41] SuM, MeiY, SinhaS. Role of the crosstalk between autophagy and apoptosis in cancer. J Oncol. 2013; 2013: 102735.

[42] FengH, ChengX, KuangJ, et al. Apatinib-induced protective autophagy and apoptosis through the AKT-mTOR pathway in anaplastic thyroid cancer. Cell Death Dis. 2018; 9: 1030.

[43] HouW, HanJ, LuC, Goldstein LA, RabinowichH. Autophagic degradation of active caspase-8: a crosstalk mechanism between autophagy and apoptosis. Autophagy. 2010; 6: 891-900.

[44] LiuK, RenT, HuangY, et al. Apatinib promotes autophagy and apoptosis through VEGFR2/STAT3/BCL-2 signaling in osteosarcoma. Cell Death Dis. 2017; 8: e3015.

[45] LevyJMM, TowersCG, ThorburnA. Targeting autophagy in cancer. Nat Rev Cancer. 2017; 17: 528-542.

[46] MaL, WangZ, XieM, et al. Silencing of circRACGAP1 sensitizes gastric cancer cells to apatinib via modulating autophagy by targeting miR-3657 and ATG7. Cell Death Dis. 2020; 11: 169.

[47] WuJ, LiangJ, LiuR, et al. Autophagic blockade potentiates anlotinib-mediated ferroptosis in anaplastic thyroid cancer. Endocr Relat Cancer. 2023; 30.

[48] ZhengB, SunX, ZhangL, et al. Inhibition of anlotinib-induced autophagy attenuates invasion and migration by regulating epithelial-mesenchymal transition and cytoskeletal rearrangement through ATG5 in human osteosarcoma cells. Braz J Med Biol Res. 2024; 57: e13152.

[49] YoonJH, AhnSG, LeeBH, Jung SH, OhSH. Role of autophagy in chemoresistance: regulation of the ATM-mediated DNA-damage signaling pathway through activation of DNA-PKcs and PARP-1. Biochem Pharmacol. 2012; 83: 747-757.

[50] PuustinenP, Keldsbo A, Corcelle-TermeauE, et al. DNA-dependent protein kinase regulates lysosomal AMP-dependent protein kinase activation and autophagy. Autophagy. 2020; 16: 1871-1888.

[51] DaidoS, Yamamoto A, FujiwaraK, SawayaR, KondoS, KondoY. Inhibition of the DNA-dependent protein kinase catalytic subunit radiosensitizes malignant glioma cells by inducing autophagy. Cancer Res. 2005; 65: 4368-4375.

[52] ZhuangW, LiB, LongL, Chen L, HuangQ, LiangZQ. Knockdown of the DNA-dependent protein kinase catalytic subunit radiosensitizes glioma-initiating cells by inducing autophagy. Brain Res. 2011; 1371: 7-15.

[53] ZhouC, ZhangZ, ZhuX, et al. N6-Methyladenosine modification of the TRIM7 positively regulates tumorigenesis and chemoresistance in osteosarcoma through ubiquitination of BRMS1. EBioMedicine. 2020; 59: 102955.

[54] MiaoW, ChenJ, JiaL, MaJ, SongD. The m6A methyltransferase METTL3 promotes osteosarcoma progression by regulating the m6A level of LEF1. Biochem Biophys Res Commun. 2019; 516: 719-725.

[55] YuanY, YanG, HeM, et al. ALKBH5 suppresses tumor progression via an m(6)A-dependent epigenetic silencing of pre-miR-181b-1/YAP signaling axis in osteosarcoma. Cell Death Dis. 2021; 12: 60.

[56] WangX, ZhaoBS, RoundtreeIA, et al. N(6)-methyladenosine modulates messenger RNA translation efficiency. Cell. 2015; 161: 1388-1399.

[57] BaiY, YangC, WuR, et al. YTHDF1 regulates tumorigenicity and cancer stem cell-like activity in human colorectal carcinoma. Front Oncol. 2019; 9: 332.

[58] WangX, LuZ, GomezA, et al. N6-methyladenosine-dependent regulation of messenger RNA stability. Nature. 2014; 505: 117-120.

RIGHTS & PERMISSIONS

2025 The Author(s). Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.