Abstract
The efficacy of systemic therapy for hepatocellular carcinoma (HCC) related to non-alcoholic steatohepatitis (NASH) is poorly understood. In this study we evaluated the effects of sorafenib based on the expression of molecular markers related to major hepatocarcinogenesis pathways and angiogenesis in a NASH-related HCC model. Forty male rats were submitted to NASH-HCC induction through the combination of a high-fat and choline deficient diet and diethylnitrosamine (100 mg/L) administration in the drinking water for 13 and 16 weeks. After the induction period, the rats received daily gavage administration of saline solution (control) or Sorafenib (5 mg/kg/day) for 3 weeks. Thereafter, the animals were euthanized and samples from liver nodules were collected for histopathological analysis and immunohistochemical assessment of HEP-PAR-1, glutamine-synthetase, VEGF, survivin, β-catenin and p53. A semi-quantitative score was used for VEGF, survivin and β-catenin analysis. For p53, the percentage of positive cells was determined. Results were processed by Wilcoxon’s test or Student’s t-test. Both protocols efficiently induced HCC, most of them being moderately to poorly differentiated. Sorafenib-treated animals showed a decreased expression of VEGF and p53 in HCCs generated at 13 weeks when compared to control animals (p = 0.03; p = 0.04, respectively). No significant difference in β-catenin and survivin were observed. There was a significant decrease in VEGF and p53 expression when comparing the two control groups (13 vs. 16 weeks, p < 0.01). p53 and VEGF are promising biomarkers for assessment of efficacy of Sorafenib, whereas survivin and β-catenin were not found useful. Decreased immunohistochemical expression of p53 and VEGF in the 16 week control group may indicate a different metabolic status of HCC.
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References
Abdelgalil AA, Alkahtani HM, Al-Jenoobi FI (2019) Sorafenib. Profiles of drug substances, excipients, and related methodology, 1st edn. Elsevier, Cambridge, pp 239–266. https://doi.org/10.1016/bs.podrm.2018.11.003
Anstee QM, Reeves HL, Kotsiliti E, Govaere O, Heikenwalder M (2019) From NASH to HCC: current concepts and future challenges. Nat Rev Gastroenterol Hepatol 16:411–428. https://doi.org/10.1038/s41575-019-0145-7
Baugh EH, Ke H, Levine AJ, Bonneau RA, Chan CS (2018) Why are there hotspot mutations in the TP53 gene in human cancers? Cell Death Differ 25:154–160. https://doi.org/10.1038/cdd.2017.180
Bosco J, Zhou Z, Gabriëls S, Verma M, Liu N, Miller BK, Gu S, Lundberg DM, Huang Y, Brown E, Josiah S, Meiyappan M, Traylor MJ, Chen N, Asakura A, De Jonge N, Blanchetot C, de Haard H, Duffy HS, Keefe D (2021) VEGFR-1/Flt-1 inhibition increases angiogenesis and improves muscle function in a mouse model of Duchenne muscular dystrophy. Mol Ther Methods Clin Dev 21:369–381. https://doi.org/10.1016/j.omtm.2021.03.013
Bray F, Ferlay J, Soerjomataram I, Siegel RL, Torre LA, Jemal A (2018) Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin 68:394–424. https://doi.org/10.3322/caac.21492
Cai Y, Ma W, Huang X, Cao L, Li H, Jiang Y, Lu N, Yin Y (2015) Effect of survivin on tumor growth of colorectal cancer in vivo. Int J Clin Exp Pathol 8:13267–13272
Castegna A, Menga A (2018) Glutamine synthetase: localization dictates outcome. Genes 9:108–116. https://doi.org/10.3390/genes9020108
Chen J, Chen JK, Conway EM, Harris RC (2013) Survivin mediates renal proximal tubule recovery from AKI. J Am Soc Nephrol 24:2023–2033. https://doi.org/10.1681/ASN.2013010076
Claesson-Welsh L (2016) VEGF receptor signal transduction—a brief update. Vascul Pharmacol 86:14–17. https://doi.org/10.1016/j.vph.2016.05.011
Costa FGB, Stefano JT, Faria DP, Levy CS, Chammas MC, Carneiro CG, Pereira IA, Cogliati B, Carrilho FJ, Oliveira CP (2018) [18F]FDG PET imaging evaluation on non-alcoholic fatty liver disease and hepatocellular carcinoma model treated with sorafenib. Hepatoma Res 4:35–43. https://doi.org/10.20517/2394-5079.2018.06
De Lima VM, Oliveira CP, Alves VA, Chammas MC, Oliveira EP, Stefano JT, de Mello ES, Cerri GG, Carrilho FJ, Caldwell SH (2008) A rodent model of NASH with cirrhosis, oval cell proliferation and hepatocellular carcinoma. J Hepatol 49:1055–1061. https://doi.org/10.1016/j.jhep.2008.07.024
De Lorenzo S, Tovoli F, Barbera MA, Garuti F, Palloni A, Frega G, Garajovà I, Rizzo A, Trevisani F, Brandi G (2018) Metronomic capecitabine vs. best supportive care in Child-Pugh B hepatocellular carcinoma: a proof of concept. Sci Rep 8:4–10. https://doi.org/10.1038/s41598-018-28337-6
Di Tommaso L, Roncalli M (2017) Tissue biomarkers in hepatocellular tumors: which, when and how. Front Med 4:10–18. https://doi.org/10.3389/fmed.2017.00010
Divella R, Mazzocca A, Daniele A, Sabbà C, Paradiso A (2019) Obesity, nonalcoholic fatty liver disease and adipocytokines network in promotion of cancer. Int J Biol Sci 15:610–616. https://doi.org/10.7150/ijbs.29599
Dolganiuc A (2015) Alcohol and viral hepatitis: role of lipid rafts. Alcohol Res 37:299–309. https://doi.org/10.3949/ccjm.67.s1.42
Edmondson HA, Steiner PE (1954) Primary carcinoma of the liver: a study of 100 cases among 48,900 necropsies. Cancer 7:462–503. https://doi.org/10.1002/1097-0142(195405)7:3%3c462::aid-cncr2820070308%3e3.0.co;2-e
Farzaneh Behelgardi M, Zahri S, Mashayekhi F, Mansouri K, Asghari SM (2018) A peptide mimicking the binding sites of VEGF-A and VEGF-B inhibits VEGFR-1/-2 driven angiogenesis, tumor growth and metastasis. Sci Rep 8:17924. https://doi.org/10.1038/s41598-018-36394-0
Febbraio MA, Reibe S, Shalapour S, Ooi GJ, Watt MJ, Karin M (2019) Preclinical models for studying NASH-driven HCC: how useful are they? Cell Metab 29:18–26. https://doi.org/10.1016/j.cmet.2018.10.012
Garg H, Suri P, Gupta JC, Talwar GP, Dubey S (2016) Survivin: a unique target for tumor therapy. Cancer Cell Int 16:49–58. https://doi.org/10.1186/s12935-016-0326-1
Geh D, Anstee QM, Reeves HL (2021) NAFLD-associated HCC: progress and opportunities. J Hepatocell Carcinoma 8:223–239. https://doi.org/10.2147/JHC.S272213
Gokmen Karasu AF, Sonmez FC, Aydin S, Adanir I, Marasli M, Ilhan GK (2018) Survivin expression in simple endometrial polyps and tamoxifen-associated endometrial polyps. Int J Gynecol Pathol 37:27–31. https://doi.org/10.1097/PGP.0000000000000376
Hanahan D, Weinberg RA (2011) Hallmarks of cancer: the next generation. Cell 144:646–674. https://doi.org/10.1016/j.cell.2011.02.013
Hernández-Morales J, Hernández-Coronado CG, Guzmán A, Zamora-Gutiérrez D, Fierro F, Gutiérrez CG, Rosales-Torres AM (2021) Hypoxia up-regulates VEGF ligand and downregulates VEGF soluble receptor mRNA expression in bovine granulosa cells in vitro. Theriogenology 165:76–83. https://doi.org/10.1016/j.theriogenology.2021.02.006
Huang DQ, El-Serag HB, Loomba R (2021) Global epidemiology of NAFLD-related HCC: trends, predictions, risk factors and prevention. Nat Rev Gastroenterol Hepatol 18:223–238. https://doi.org/10.1038/s41575-020-00381-6
Ichikawa T, Sano K, Morisaka H (2014) Diagnosis of pathologically early HCC with EOB-MRI: experiences and current consensus. Liver Cancer 3:97–107. https://doi.org/10.1159/000343865
Jha K, Shukla M, Pandey M (2012) Survivin expression and targeting in breast cancer. Surg Oncol 21:125–131. https://doi.org/10.1016/j.suronc.2011.01.001
Kapiris I, Nastos K, Karakatsanis A, Theodosopoulos T, Karandrea D, Kondi Pafiti A, Contis J (2019) Survivin expression in hepatocellular carcinoma. Correlation with clinicopathological characteristics and overall survival. J BUON 24:1934–1942
Kim GW, Lee DH, Jeon YH, Yoo J, Kim SY, Lee SW, Cho HY, Kwon SH (2021) Glutamine synthetase as a therapeutic target for cancer treatment. Int J Mol Sci 22:1701–1708. https://doi.org/10.3390/ijms22041701
Llovet JM, Kelley RK, Villanueva A, Singal AG, Pikarsky E, Roayaie S, Lencioni R, Koike K, Zucman-Rossi J, Finn RS (2021) Hepatocellular carcinoma. Nat Rev Dis Primers 7:6–16. https://doi.org/10.1038/s41572-020-00240-3
Lu JG, Farukhi MA, Mayeda D, French SW (2017) Hepatocellular carcinoma with neuroendocrine differentiation: a case report. Exp Mol Pathol 103:200–203. https://doi.org/10.1016/j.yexmp.2017.09.004
Nguyen T, Phillips D, Jain D, Torbenson M, Wu TT, Yeh MM, Kakar S (2015) Comparison of 5 immunohistochemical markers of hepatocellular differentiation for the diagnosis of hepatocellular carcinoma. Arch Pathol Lab Med 139:1028–1034. https://doi.org/10.5858/arpa.2014-0479-OA
Nguyen TB, Roncalli M, Di Tommaso L, Kakar S (2016) Combined use of heat-shock protein 70 and glutamine synthetase is useful in the distinction of typical hepatocellular adenoma from atypical hepatocellular neoplasms and well-differentiated hepatocellular carcinoma. Mod Pathol 29:283–292. https://doi.org/10.1038/modpathol.2015.162
Pinto Marques H, Gomes da Silva S, De Martin E, Agopian VG, Martins PN (2020) Emerging biomarkers in HCC patients: current status. Int J Surg 82S:70–76. https://doi.org/10.1016/j.ijsu.2020.04.043
Plewka D, Jakubiec-Bartnik B, Morek M, Bogunia E, Bienioszek M, Wolski H, Kotrych D, Dziekan K, Seremak-Mrozikiewicz A, Plewka A (2015) Survivin in ovary tumors. Ginekol Pol 86:525–530. https://doi.org/10.17772/gp/57855
Rizzo A, Nannini M, Novelli M, Ricci AD, Scioscio VD, Pantaleo MA (2020) Dose reduction and discontinuation of standard-dose regorafenib associated with adverse drug events in cancer patients: a systematic review and meta-analysis. Ther Adv Med Oncol 12:1–10. https://doi.org/10.1177/1758835920936932
Rizzo A, Ricci AD, Di Federico A, Frega G, Palloni A, Tavolari S, Brandi G (2021) Predictive biomarkers for checkpoint inhibitor-based immunotherapy in hepatocellular carcinoma: where do we stand? Front Oncol 11:1–7. https://doi.org/10.3389/fonc.2021.803133
Rowe IA, Galsinh SK, Wilson GK, Parker R, Durant S, Lazar C, Branza-Nichita N, Bicknell R, Adams DH, Balfe P, McKeating JA (2014) Paracrine signals from liver sinusoidal endothelium regulate hepatitis C virus replication. Hepatol 59:375–384. https://doi.org/10.1002/hep.26571
Scheer A, Knauer SK, Verhaegh R (2017) Survivin expression pattern in the intestine of normoxic and ischemic rats. BMC Gastroenterol 17:76–85. https://doi.org/10.1186/s12876-017-0625-6
Stefano JT, de Oliveira CP, Corrêa-Giannella ML, Soares IC, Kubrusly MS, Bellodi-Privato M, de Mello ES, de Lima VM, Carrilho FJ, Alves VA (2011) Decreased immunoexpression of survivin could be a potential marker in human non-alcoholic fatty liver disease progression? Liver Int 31:377–385. https://doi.org/10.1111/j.1478-3231.2010.02370.x
Su C (2016) Survivin in survival of hepatocellular carcinoma. Cancer Lett 379:184–190. https://doi.org/10.1016/j.canlet.2015.06.016
Thoolen B, Maronpot RR, Harada T, Nyska A, Rousseaux C, Nolte T, Malarkey DE, Kaufmann W, Küttler K, Deschl U, Nakae D, Gregson R, Vinlove MP, Brix AE, Singh B, Belpoggi F, Ward JM (2010) Proliferative and nonproliferative lesions of the rat and mouse hepatobiliary system. Toxicol Pathol 38:5S-81S. https://doi.org/10.1177/0192623310386499
Tremosini S, Forner A, Boix L, Vilana R, Bianchi L, Reig M, Rimola J, Rodríguez-Lope C, Ayuso C, Solé M, Bruix J (2012) Prospective validation of an immunohistochemical panel (glypican 3, heat shock protein 70 and glutamine synthetase) in liver biopsies for diagnosis of very early hepatocellular carcinoma. Gut 61:1481–1487. https://doi.org/10.1136/gutjnl-2011-301862
Wang W, Jia WD, Xu GL, Wang ZH, Li JS, Ma JL, Ge YS, Xie SX, Yu JH (2009) Antitumoral activity of rapamycin mediated through inhibition of HIF-1alpha and VEGF in hepatocellular carcinoma. Dig Dis Sci 54:2128–2136. https://doi.org/10.1007/s10620-008-0605-3
Xue Y, San Luis B, Lane DP (2019) Intratumour heterogeneity of p53 expression: causes and consequences. J Pathol 249:274–285. https://doi.org/10.1002/path.5328
Yang JD, Hainaut P, Gores GJ, Amadou A, Plymoth A, Roberts LR (2019) A global view of hepatocellular carcinoma: trends, risk, prevention and management. Nat Rev Gastroenterol Hepatol 16:589–604. https://doi.org/10.1038/s41575-019-0186-y
Younossi Z, Anstee QM, Marietti M, Hardy T, Henry L, Eslam M, George J, Bugianesi E (2018) Global burden of NAFLD and NASH: trends, predictions, risk factors and prevention. Nat Rev Gastroenterol Hepatol 15:11–20. https://doi.org/10.1038/nrgastro.2017.109
Zhang Q, Bai X, Chen W, Ma T, Hu Q, Liang C, Xie S, Chen C, Hu L, Xu S, Liang T (2013) Wnt/β-catenin signaling enhances hypoxia-induced epithelial-mesenchymal transition in hepatocellular carcinoma via crosstalk with hif-1α signaling. Carcinogenesis 34:962–973. https://doi.org/10.1093/carcin/bgt027
Acknowledgements
The authors thank the Boards of the Institutions involved in the study. The abstract was presented at the European Association for the Study of the Liver’s (EASL) Liver Cancer Summit Feb 3‐Feb 4, 2022, online, and published as abstract PO-170 in LCS 2022 Abstract Book.
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The author João Pedro Nassar Reis, was benefited from the Scientific Initiation Grant: PIBIC-CNPq (nº 2564/2020).
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JPNR, PFU performed immunohistochemical reactions and write the first draft of the manuscript. BC performed the statistical analysis. VAFA and ALF analysed the histological sections and the immunohistochemical evaluations. JTS, FJC AND CPM aided in the study design and discussion of the results. CPM, VAFA, BC and ALF contributed to the discussion of the results, organization and review of the manuscript. All authors read and approved the manuscript.
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Nassar-Reis, J.P., Umeta, P.F., Stefano, J.T. et al. P53 and VEGF are promising biomarkers for sorafenib efficacy in an experimental model of NASH-related HCC. J Mol Histol 54, 473–488 (2023). https://doi.org/10.1007/s10735-023-10142-9
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DOI: https://doi.org/10.1007/s10735-023-10142-9