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Novel protein kinase inhibitor TT-00420 inhibits gallbladder cancer by inhibiting JNK/JUN-mediated signaling pathway

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Abstract

Purpose

This study aimed to investigate the efficiency of our chemically synthesized TT-00420, a novel spectrum-selective multiple protein kinase inhibitor, in cultured cells and animal models of gallbladder cancer (GBC) and explore its potential mechanism.

Methods

Multiple GBC models were established to assess the anti-tumor efficiency, toxicity, and pharmacokinetics of TT-00420. Integrated transcriptomic, proteomic and phosphoproteomic analysis was conducted to identify potential downstream effectors of TT-00420. Western blotting, qRT-PCR, nuclear-cytoplasm separation, and immunofluorescence were performed to confirm the multi-omic results and explore the molecular mechanism of TT-00420. Immunohistochemistry was used to detect FGFR1 and p-FGFR1 expression levels in GBC samples. Autodock software was utilized to investigate the potential binding mode between the TT-00420 and the human FGFR1.

Results

We found that TT-00420 exerted potent growth inhibition of GBC cell lines and multiple xenograft models. Treatment of mice with 15 mg/kg TT-00420 via gavage displayed a half-life of 1.8 h in the blood and rapid distribution to the liver, kidneys, lungs, spleen, and tumors at 0.25 h, but no toxicity to these organs over 2 weeks. Multi-omic analysis revealed c-Jun as a potential downstream effector after TT-00420 treatment. Mechanistically, TT-00420 showed rigorous ability to block FGFR1 and its downstream JNK-JUN (S63/S73) signaling pathway, and induce c-Jun S243-dependent MEK/ERK reactivation, leading to FASLG-dependent tumor cell death. Finally, we found that FGFR1 and p-FGFR1 expression was elevated in GBC patients and these levels correlated with decreased patient survival.

Conclusions

TT-00420 shows potent antitumor efficacy and may serve as a novel agent to improve GBC prognosis.

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Data availability

All data generated or analyzed during this study are included in this published article (and its supplementary information files).

Abbreviations

GBC:

Gallbladder cancer

PDAC:

Pancreatic Ductal Adenocarcinoma

PDX:

Patient-derived Xenograft

CDX:

Cell-derived Xenograft

FIH:

First-in-human

SAE:

Severe Adverse Effects

References

  1. M. Rakic, L. Patrlj, M. Kopljar, R. Klicek, M. Kolovrat, B. Loncar et al., Gallbladder cancer. Hepatobiliary Surg Nutr. 3, 221–226 (2014)

    PubMed  PubMed Central  Google Scholar 

  2. M.A. Schmidt, L. Marcano-Bonilla, L.R. Roberts, Gallbladder cancer: epidemiology and genetic risk associations. Chin Clin Oncol. 8, 31 (2019)

    Article  PubMed  Google Scholar 

  3. T. Ren, Y. Li, X. Zhang, Y. Geng, Z. Shao, M. Li et al., Protocol for a gallbladder cancer registry study in China: the Chinese Research Group of Gallbladder Cancer (CRGGC) study. BMJ Open 11, e038634 (2021)

    Article  PubMed  PubMed Central  Google Scholar 

  4. T. Ren, Y.S. Li, Y.J. Geng, M.L. Li, X.S. Wu, W.W. Wu et al., Analysis of treatment modalities and prognosis of patients with gallbladder cancer in China from 2010 to 2017. Zhonghua Wai Ke Za Zhi. 58, 697–706 (2020)

    CAS  PubMed  Google Scholar 

  5. T. Ren, Y.S. Li, X.Y. Dang, Y. Li, Z.Y. Shao, R.F. Bao et al., Prognostic significance of regional lymphadenectomy in T1b gallbladder cancer: Results from 24 hospitals in China. World J Gastrointest Surg. 13, 176–186 (2021)

    Article  PubMed  PubMed Central  Google Scholar 

  6. C. Boutros, M. Gary, K. Baldwin, P. Somasundar, Gallbladder cancer: past, present and an uncertain future. Surg Oncol. 21, e183–e191 (2012)

    Article  CAS  PubMed  Google Scholar 

  7. X-Y. Cui, X-C. Li, J-J. Cui, X-S. Wu, L. Zou, X-L. Song et al., Modified FOLFIRINOX for unresectable locally advanced or metastatic gallbladder cancer, a comparison with GEMOX regimen. Hepatobiliary Surg. Nutr. 10, 498–506 (2021)

  8. A. Sharma, B. Kalyan Mohanti, S. Pal Chaudhary, V. Sreenivas, R. Kumar Sahoo, N. Kumar Shukla et al., Modified gemcitabine and oxaliplatin or gemcitabine + cisplatin in unresectable gallbladder cancer: Results of a phase III randomised controlled trial. Eur J Cancer. 123, 162–170 (2019)

    Article  CAS  PubMed  Google Scholar 

  9. J.S. Jang, H.Y. Lim, I.G. Hwang, H.S. Song, N. Yoo, S. Yoon et al., Gemcitabine and oxaliplatin in patients with unresectable biliary cancer including gall bladder cancer: a Korean Cancer Study Group phase II trial. Cancer Chemother Pharmacol. 65, 641–647 (2010)

    Article  CAS  PubMed  Google Scholar 

  10. L. Zou, X. Li, X. Wu, J. Cui, X. Cui, X. Song et al., Modified FOLFIRINOX versus gemcitabine plus oxaliplatin as first-line chemotherapy for patients with locally advanced or metastatic cholangiocarcinoma: a retrospective comparative study. BMC Cancer 21, 818 (2021)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. R.T. Shroff, M.M. Javle, L. Xiao, A.O. Kaseb, G.R. Varadhachary, R.A. Wolff et al., Gemcitabine, Cisplatin, and nab-Paclitaxel for the Treatment of Advanced Biliary Tract Cancers: A Phase 2 Clinical Trial. JAMA Oncol. 5, 824–830 (2019)

    Article  PubMed  PubMed Central  Google Scholar 

  12. J. Valle, H. Wasan, D. H. Palmer, D. Cunningham, A. Anthoney, A. Maraveyas, S. Madhusudan, T. Iveson, S. Hughes, S. P. Pereira, M. Roughton, J. Bridgewater, for the ABC-02 Trial Investigators. Cisplatin plus Gemcitabine versus Gemcitabine for Biliary Tract Cancer. N. Engl. J. Med. 362:1273–1281 (2010)

  13. X. Song, Y. Hu, Y. Li, R. Shao, F. Liu, Y. Liu, Overview of current targeted therapy in gallbladder cancer. Signal Transduct Target Ther. 5, 230 (2020)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. B.I. Rini, S.K. Pal, B.J. Escudier, M.B. Atkins, T.E. Hutson, C. Porta et al., Tivozanib versus sorafenib in patients with advanced renal cell carcinoma (TIVO-3): a phase 3, multicentre, randomised, controlled, open-label study. Lancet Oncol. 21, 95–104 (2020)

    Article  CAS  PubMed  Google Scholar 

  15. J. Ricke, H.J. Klumpen, H. Amthauer, I. Bargellini, P. Bartenstein, E.N. de Toni et al., Impact of combined selective internal radiation therapy and sorafenib on survival in advanced hepatocellular carcinoma. J Hepatol. 71, 1164–1174 (2019)

    Article  CAS  PubMed  Google Scholar 

  16. A. Burchert, G. Bug, L.V. Fritz, J. Finke, M. Stelljes, C. Rollig et al., Sorafenib Maintenance After Allogeneic Hematopoietic Stem Cell Transplantation for Acute Myeloid Leukemia With FLT3-Internal Tandem Duplication Mutation (SORMAIN). J Clin Oncol. 38, 2993–3002 (2020)

    Article  PubMed  Google Scholar 

  17. J.C. Yang, S.M. Gadgeel, L.V. Sequist, C.L. Wu, V.A. Papadimitrakopoulou, W.C. Su et al., Pembrolizumab in Combination With Erlotinib or Gefitinib as First-Line Therapy for Advanced NSCLC With Sensitizing EGFR Mutation. J Thorac Oncol. 14, 553–559 (2019)

    Article  CAS  PubMed  Google Scholar 

  18. T. Seto, T. Kato, M. Nishio, K. Goto, S. Atagi, Y. Hosomi et al., Erlotinib alone or with bevacizumab as first-line therapy in patients with advanced non-squamous non-small-cell lung cancer harbouring EGFR mutations (JO25567): an open-label, randomised, multicentre, phase 2 study. Lancet Oncol. 15, 1236–1244 (2014)

    Article  CAS  PubMed  Google Scholar 

  19. H. Saito, T. Fukuhara, N. Furuya, K. Watanabe, S. Sugawara, S. Iwasawa et al., Erlotinib plus bevacizumab versus erlotinib alone in patients with EGFR-positive advanced non-squamous non-small-cell lung cancer (NEJ026): interim analysis of an open-label, randomised, multicentre, phase 3 trial. Lancet Oncol. 20, 625–635 (2019)

    Article  CAS  PubMed  Google Scholar 

  20. K. Nakagawa, E.B. Garon, T. Seto, M. Nishio, S. Ponce Aix, L. Paz-Ares et al., Ramucirumab plus erlotinib in patients with untreated, EGFR-mutated, advanced non-small-cell lung cancer (RELAY): a randomised, double-blind, placebo-controlled, phase 3 trial. Lancet Oncol. 20, 1655–1669 (2019)

    Article  CAS  PubMed  Google Scholar 

  21. S.J. Lubner, M.R. Mahoney, J.L. Kolesar, N.K. Loconte, G.P. Kim, H.C. Pitot et al., Report of a multicenter phase II trial testing a combination of biweekly bevacizumab and daily erlotinib in patients with unresectable biliary cancer: a phase II Consortium study. J Clin Oncol. 28, 3491–3497 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. H. Horiuchi, H. Kawamata, T. Fujimori, Y. Kuroda. A MEK inhibitor (U0126) prolongs survival in nude mice bearing human gallbladder cancer cells with K-ras mutation: analysis in a novel orthotopic inoculation model. Int. J. Oncol. 23(4), 957–63 (2003)

  23. K. Kiguchi, L. Ruffino, T. Kawamoto, T. Ajiki, J. DiGiovanni. Chemopreventive and Therapeutic Efficacy of Orally Active Tyrosine Kinase Inhibitors in a Transgenic Mouse Model of Gallbladder Carcinoma. Clin. Cancer Res. 11(15), 5572–80 (2005)

  24. L-X. Fu, Q-W Lian, J-D. Pan, Z-L. Xu, T-M. Zhou, B. Ye. JAK2 tyrosine kinase inhibitor ag490 suppresses cell growth and invasion of gallbladder cancer cells via inhibition of JAK2/STAT3 signaling. J Biol Regul Homeost Agents. 31(1), 51–58 (2017)

  25. Y. Zhang, C. Zuo, L. Liu, Y. Hu, B. Yang, S. Qiu et al., Single-cell RNA-sequencing atlas reveals an MDK-dependent immunosuppressive environment in ErbB pathway-mutated gallbladder cancer. J Hepatol. 75, 1128–1141 (2021)

    Article  CAS  PubMed  Google Scholar 

  26. F. Liu, Y. Li, D. Ying, S. Qiu, Y. He, M. Li et al., Whole-exome mutational landscape of neuroendocrine carcinomas of the gallbladder. Signal Transduct Target Ther. 6, 55 (2021)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. M. Li, Z. Zhang, X. Li, J. Ye, X. Wu, Z. Tan et al., Whole-exome and targeted gene sequencing of gallbladder carcinoma identifies recurrent mutations in the ErbB pathway. Nat Genet. 46, 872–876 (2014)

    Article  CAS  PubMed  Google Scholar 

  28. M. Li, F. Liu, Y. Zhang, X. Wu, W. Wu, X.A. Wang et al., Whole-genome sequencing reveals the mutational landscape of metastatic small-cell gallbladder neuroendocrine carcinoma (GB-SCNEC). Cancer Lett. 391, 20–27 (2017)

    Article  CAS  PubMed  Google Scholar 

  29. M. Li, F. Liu, F. Zhang, W. Zhou, X. Jiang, Y. Yang et al., Genomic ERBB2/ERBB3 mutations promote PD-L1-mediated immune escape in gallbladder cancer: a whole-exome sequencing analysis. Gut 68, 1024–1033 (2019)

    Article  CAS  PubMed  Google Scholar 

  30. P. Peng, G. Li, X. Qiang, Y. Tian, Q. Yu, E. Marangoni, F. Wu. TT-00420, a novel multiple kinase inhibitor to treat TNBC. American Association for Cancer Research, (2019)

  31. M.E. Ritchie, B. Phipson, D. Wu, Y. Hu, C.W. Law, W. Shi et al., limma powers differential expression analyses for RNA-sequencing and microarray studies. Nucleic Acids Res. 43, e47 (2015)

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  32. S. Tyanova, T. Temu, J. Cox, The MaxQuant computational platform for mass spectrometry-based shotgun proteomics. Nat Protoc. 11, 2301–2319 (2016)

    Article  CAS  PubMed  Google Scholar 

  33. S. Tyanova, J. Cox, Perseus: A Bioinformatics Platform for Integrative Analysis of Proteomics Data in Cancer Research. Methods Mol Biol. 1711, 133–148 (2018)

    Article  CAS  PubMed  Google Scholar 

  34. M.V. Kuleshov, M.R. Jones, A.D. Rouillard, N.F. Fernandez, Q. Duan, Z. Wang et al., Enrichr: a comprehensive gene set enrichment analysis web server 2016 update. Nucleic Acids Res. 44, W90–W97 (2016)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. D. Seeliger, B.L. de Groot, Ligand docking and binding site analysis with PyMOL and Autodock/Vina. J Comput Aided Mol Des. 24, 417–422 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. E. Shaulian, AP-1–The Jun proteins: Oncogenes or tumor suppressors in disguise? Cell Signal. 22, 894–899 (2010)

    Article  CAS  PubMed  Google Scholar 

  37. G. Raivich, c-Jun expression, activation and function in neural cell death, inflammation and repair. J Neurochem. 107, 898–906 (2008)

    CAS  PubMed  Google Scholar 

  38. Q. Meng, Y. Xia, c-Jun, at the crossroad of the signaling network. Protein Cell. 2, 889–898 (2011)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. E. Shaulian, M. Karin. AP-1 in cell proliferation and survival. Oncogene. 20(19), 2390–400 (2001)

  40. E. Shaulian, M. Karin. AP-1 as a regulator of cell life and death. Nat. Cell. Biol. 4(5), E131–6 (2002)

  41. L. Gorecki, M. Andrs, M. Rezacova, J. Korabecny, Discovery of ATR kinase inhibitor berzosertib (VX-970, M6620): Clinical candidate for cancer therapy. Pharmacol Ther. 210, 107518 (2020)

    Article  CAS  PubMed  Google Scholar 

  42. J.J. Lin, A.T. Shaw, Resisting Resistance: Targeted Therapies in Lung Cancer. Trends Cancer. 2, 350–364 (2016)

    Article  PubMed  PubMed Central  Google Scholar 

  43. B.C. Medeiros, J. Possick, M. Fradley, Cardiovascular, pulmonary, and metabolic toxicities complicating tyrosine kinase inhibitor therapy in chronic myeloid leukemia: Strategies for monitoring, detecting, and managing. Blood Rev. 32, 289–299 (2018)

    Article  CAS  PubMed  Google Scholar 

  44. G. Shen, F. Zheng, D. Ren, F. Du, Q. Dong, Z. Wang et al., Anlotinib: a novel multi-targeting tyrosine kinase inhibitor in clinical development. J Hematol Oncol. 11, 120 (2018)

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  45. L. Huang, S. Jiang, Y. Shi, Tyrosine kinase inhibitors for solid tumors in the past 20 years (2001–2020). J Hematol Oncol. 13, 143 (2020)

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  46. S. Qin, A. Li, M. Yi, S. Yu, M. Zhang, K. Wu, Recent advances on anti-angiogenesis receptor tyrosine kinase inhibitors in cancer therapy. J Hematol Oncol. 12, 27 (2019)

    Article  PubMed  PubMed Central  Google Scholar 

  47. E. Rassy, R. Flippot, L. Albiges, Tyrosine kinase inhibitors and immunotherapy combinations in renal cell carcinoma. Ther Adv Med Oncol. 12, 1758835920907504 (2020)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. A. Spallanzani, G. Orsi, K. Andrikou, F. Gelsomino, M. Rimini, L. Riggi et al., Lenvatinib as a therapy for unresectable hepatocellular carcinoma. Expert Rev Anticancer Ther. 18, 1069–1076 (2018)

    Article  CAS  PubMed  Google Scholar 

  49. Z. Xue, D.J. Vis, A. Bruna, T. Sustic, S. van Wageningen, A.S. Batra et al., MAP3K1 and MAP2K4 mutations are associated with sensitivity to MEK inhibitors in multiple cancer models. Cell Res. 28, 719–729 (2018)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. A. Prahallad, C. Sun, S. Huang, F. Di Nicolantonio, R. Salazar, D. Zecchin et al., Unresponsiveness of colon cancer to BRAF(V600E) inhibition through feedback activation of EGFR. Nature 483, 100–103 (2012)

    Article  CAS  PubMed  Google Scholar 

  51. C. Sun, S. Hobor, A. Bertotti, D. Zecchin, S. Huang, F. Galimi et al., Intrinsic resistance to MEK inhibition in KRAS mutant lung and colon cancer through transcriptional induction of ERBB3. Cell Rep. 7, 86–93 (2014)

    Article  CAS  PubMed  Google Scholar 

  52. R.B. Corcoran, H. Ebi, A.B. Turke, E.M. Coffee, M. Nishino, A.P. Cogdill et al., EGFR-mediated re-activation of MAPK signaling contributes to insensitivity of BRAF mutant colorectal cancers to RAF inhibition with vemurafenib. Cancer Discov. 2, 227–235 (2012)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. S. Karaman, V. M. Leppanen, K. Alitalo. Vascular endothelial growth factor signaling in development and disease. Development. 145(14), dev151019 (2018)

  54. M. Simons, E. Gordon, L. Claesson-Welsh, Mechanisms and regulation of endothelial VEGF receptor signalling. Nat Rev Mol Cell Biol. 17, 611–625 (2016)

    Article  CAS  PubMed  Google Scholar 

  55. W.H. Toh, M.M. Siddique, L. Boominathan, K.W. Lin, K. Sabapathy, c-Jun regulates the stability and activity of the p53 homologue, p73. J Biol Chem. 279, 44713–44722 (2004)

    Article  CAS  PubMed  Google Scholar 

  56. A. Skopelitou, M. Hadjiyannakis, D. Dimopoulos, S. Kamina, O. Krikoni, V. Alexopoulou, C. Rigas, N. J. Agnantis. p53 and c-jun Expression in Urinary Bladder Transitional Cell Carcinoma: Correlation with Proliferating Cell Nuclear Antigen (PCNA) Histological Grade and Clinical Stage. Eur Urol. 31(4), 464–71 (1997)

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Acknowledgements

We really appreciate the Prof. Rong Shao from Shanghai Jiao Tong University School of Medicine for substantially editing the manuscript.

Funding

This project was supported in part by grants from the National Natural Science Foundation of China (31620103910, 81874181, 81903035, 82103308, 3213000192), National Science and Technology Major Project for “Major New Drug Innovation and Development” (2019ZX09301-158), Shanghai Municipal Science and Technology Major Project (20JC1419101).

Author information

Author notes

  1. Huijie Miao, Yajun Geng, Yang Li, Shijie Tang and Feiling Feng contributed equally to this work.

Authors and Affiliations

  1. Department of Biliary-Pancreatic Surgery, Ren Ji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200127, China

    Huijie Miao, Yajun Geng, Yang Li, Weijian Li, Yongsheng Li, Liguo Liu, Zeyu Wang, Ziyi Wang, Ke Liu, Lu Zou, Mao Yang, Yuhao Zhao, Maolan Li & Yingbin Liu

  2. State Key Laboratory of Cell Biology, Shanghai Institute of Biochemistry and Cell Biology, CAS Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, Shanghai, 200031, China

    Shijie Tang & Luonan Chen

  3. University of Chinese Academy of Sciences, Beijing, 100049, China

    Shijie Tang

  4. Department of Biliary Surgery, Eastern Hepatobiliary Surgery Hospital, Shanghai, 200433, China

    Feiling Feng, Zhizhen Li & Xiaoqing Jiang

  5. Department of Hepatobiliary Surgery, the First Affiliated Hospital of Xi’an Jiao Tong University, Xi’an, 710061, China

    Rui Zhang

  6. State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Shanghai, 200127, China

    Shimei Qiu, Ying Wu, Chen Chen, Yongning He, Dongxi Xiang, Yun Liu & Yingbin Liu

  7. Shanghai Key Laboratory of Biliary Tract Disease Research, Shanghai, 200127, China

    Shimei Qiu, Ying Wu, Chen Chen, Maolan Li & Yingbin Liu

  8. Department of General Surgery, Xinhua Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, 200092, China

    Ziyu Shao

  9. 3D Medicines Lnc, Shanghai, 201114, China

    Dadong Zhang

  10. Transthera Sciences (Nanjing), lnc, Nanjing, 210032, Jiangsu, China

    Peng Peng, Xiaoyan Qiang & Frank Wu

  11. Center for Excellence in Animal Evolution and Genetics, Chinese Academy of Sciences, Kunming, 650223, China

    Luonan Chen

  12. Key Laboratory of Systems Health Science of Zhejiang Province, Hangzhou Institute for Advanced Study, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Hangzhou, 310024, China

    Luonan Chen

Authors
  1. Huijie Miao
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  2. Yajun Geng
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  3. Yang Li
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  4. Shijie Tang
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  5. Feiling Feng
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  6. Weijian Li
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  7. Yongsheng Li
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  8. Liguo Liu
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  9. Rui Zhang
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  15. Ke Liu
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Contributions

HM, YG, YL, ST and FF performed most of the experiments with assistance from WL, YL, LL, RZ, SQ, YW, ZW, ZW and ZS. LZ, CC and KL helped to perform the animal experiments. MY, YZ, and ZL helped to perform the bioinformatic analysis and statistical analysis. PP and XQ helped to do the pharmacokinetic analysis. YH, DX, LC, JQ, ML, YL and YL conceived and supervised the study. HM, YG, YL, ST and FF wrote most of the manuscript. All authors commented on the manuscript.

Corresponding authors

Correspondence to Dongxi Xiang, Xiaoqing Jiang, Maolan Li, Yun Liu or Yingbin Liu.

Ethics declarations

Ethical approval and consent to participate

All patient samples and clinical data were obtained with informed consent from the Shanghai Jiao Tong University School of Medicine Affiliated Renji Hospital, Shanghai, China. The study was approved by the Ethics Committee of Shanghai Jiao Tong University School of Medicine Affiliated Renji Hospital (RA-2021–442).

Consent for publication

Not applicable.

Competing interest

The authors declare that they have no competing interests.

Additional information

Publisher's Note

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

Supplementary Information

Below is the link to the electronic supplementary material.

13402_2022_692_MOESM1_ESM.jpg

Supplementary file1 (JPG 900 KB) Figure S1. (a) The chemical structure of TT-00420. (b) The kinase panel and potential kinase targets of TT-00420.

13402_2022_692_MOESM2_ESM.jpg

Supplementary file2 (JPG 1132 KB) Figure S2, related to Fig.1 (a-b) The IC50 analysis of TT-00420 in cholangiocarcinoma cell lines and pancreatic cancer cell lines by using CCK-8 assay. (c-f) The IC50 analysis of Infigratinib, Tofacitinib, Sorafenib and Salirasib in GBC commercial cell lines by using CCK-8 assay. (g) The proliferation ability of GBC cells was measured by CCK-8 assay. The cells were treated TT-00420 with different concentration.

13402_2022_692_MOESM3_ESM.jpg

Supplementary file3 (JPG 5267 KB) Figure S3, related to Fig.2 (a) The food consumption observation and body weight change in acute toxicity analysis. The male nude mice were administered TT-00420 with different concentration for consecutive 7 days following observation for additional 21 days. (b) HE staining of heart, liver, spleen, lung, kidney in acute toxicity analysis. The mice were administered TT-00420 with different concentration. (c) Tissue distribution analysis (left) and drug plasma concentration analysis (right) at different time points (n=3 for each time point) of TT-00420 in male nude mice after intragastric gavage at 15mg/kg loaded with 0.5%MC. The organs and serum were collected and performed LC-MS analysis after the mice were euthanized.

13402_2022_692_MOESM4_ESM.jpg

Supplementary file4 (JPG 5739 KB) Figure S4, related to Fig.2 GBC-SD cells (a) and SW1990 cells (b) were subcutaneously transplanted into the flanks of male nude mice to develop tumors (n=6 for each group). After the tumor was formed, TT-00420 was administered into mice with 15mg/kg by IG gavage once a day. Mouse weight was measured every day before drug administration. Tumor size was measured once every three days and tumor weight was measured after mice were euthanized. (c) HE staining of heart, liver, spleen, lung, kidney in NOZ CDX model after administered with different concentration of TT-00420. (d) The ki67 IHC staining in GBC-SD CDX model sections.

13402_2022_692_MOESM5_ESM.jpg

Supplementary file5 (JPG 953 KB) Figure S5, related to Fig.2 (a) Tumor pictures of NOZ CDX model. (b)Tumor pictures of GBC-SD CDX model. (c) Tumor picture of GBC patient-derived xenograft (PDX) model.

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Supplementary file6 (JPG 1554 KB) Figure S6, related to Fig.3 (a) Experimental workflow to analyze the proteomic and phosphoproteomic data of NOZ cells after treated with TT-00420 at 1μM for 48hr. (b) GBC cells were performed RNA-SEQ analysis after treated with TT-00420 (NOZ-1μM, GBC-SD-0.4μM, EHGB-1-0.1μM) for 24hr. Heatmap shows the Pearson Correlation Coefficients (PCC) between the different biological replicates in the transcriptome data. Statistically significantly up- and down-regulated genes were identified by limma (p-value<0.05; FoldChange≥1.2) and presented as volcano plot. Each dot represented one gene.

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Supplementary file7 (JPG 1059 KB) Figure S7, related to Fig.3 (a) Number of quantified proteins and phosphosites in 3 biological replicates of NOZ cells with different experimental conditions. (b) Principal component analysis (PCA) of proteome and phosphoproteome data. (c) Heatmap shows the Pearson Correlation Coefficients (PCC) between the different biological replicates in the proteomic and phosphoproteomic data.

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Supplementary file8 (JPG 1087 KB) Figure S8, related to Fig.3 (a) Distribution of serine, threonine and tyrosine phosphorylation sites in phosphoproteomic data. (b) Venn diagram of common downregulated proteins in both proteomic and phosphoproteomic data. (c) STRING-based analysis of potential kinase targets and common downregulated proteins. (d) Upstream transcription factors of significantly differently expressed proteins were estimated with Ingenuity Pathway Analysis (IPA). Activation z-score ≥1.5 means the TF is in activation condition. (e) Log2FoldChange of TT-00420 compared to NC on proteomic and phosphoproteomic data.

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Supplementary file9 (JPG 761 KB) Figure S9, related to Fig.4 The protein-protein interaction (PPI) network analysis of JUN with its repressed downstream target genes. The genes were imported into STRING database, and then the correlation results were exported to Cytoscape Software to draw the diagram based on the correlation degree.

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Supplementary file10 (JPG 1357 KB) Figure S10, related to Fig.5(a) The western blot analysis of GBC cells after treated TT-00420 with different concentration for 48hr. (b) The quantification analysis of western blot in Fig.5A. (c) Colony formation assay of GBC cells after treated with vehicle control, TT-00420 (0.2μM), Trametinib (0.1μM), or TT-00420/Trametinib combined (TT-00420 0.1μM, Trametinib 0.05μM). (d) Colony formation assay of GBC cells after treated with vehicle control, TT-00420 (0.2μM), SCH772984 (0.1μM), or TT-00420/SCH772984 combined (TT-00420 0.1μM, SCH772984 0.05μM).

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Supplementary file11 (JPG 1324 KB) Figure S11, related to Fig.6 Kaplan-Meier analysis of disease-free survival (DFS) and overall survival (OS) of different gastrointestinal carcinoma according to different FGFR1 mRNA level. The sequencing data was obtained from TCGA database.

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Supplementary file12 (JPG 753 KB) Figure S12 Molecular docking of FGFR1 and TT-00420 using Autodock Tools software. The predicted potential residues were TYR-654 and VAL704. The length of the possible hydrogen bonds was 3.1Å and 3.4Å, respectively.

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Supplementary file13 (JPG 1463 KB) Figure S13 (a) The GSEA analysis of RNA-SEQ data using different gene sets in NOZ cells. (b) The GSEA analysis of proteomics data using different gene sets in NOZ cells.

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Miao, H., Geng, Y., Li, Y. et al. Novel protein kinase inhibitor TT-00420 inhibits gallbladder cancer by inhibiting JNK/JUN-mediated signaling pathway. Cell Oncol. 45, 689–708 (2022). https://doi.org/10.1007/s13402-022-00692-7

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