Abstract
Toll-like receptors (TLRs) are expressed on both immune cells and tumor cells, triggering both anti-tumor and pro-tumor responses. Therefore, TLRs have potential as prognostic biomarkers and immunotherapeutic targets. The aim of this study was to investigate TLR1, TLR2, TLR4, TLR5, and TLR6 expression and association with clinicopathological variables and survival in gastric cancer. Immunohistochemical study on cancer specimens from 564 resected gastric cancer patients was performed using tissue microarrays. The association between patient survival and TLR expression was calculated with Cox regression adjusted for confounding factors. Patients with high cytoplasmic TLR2 expression had significantly poorer 5-year survival than the low cytoplasmic TLR2 expression group in multivariate analysis (adjusted HR 1.38, 95% CI 1.11–1.71), and this estimate was similar in intestinal type (adjusted HR 1.33, 95% CI 0.98–1.80) and diffuse type (adjusted HR 1.48, 95% CI 1.06–2.05) histology subgroups. Patients with high cytoplasmic TLR6 expression group had significantly better 5-year survival compared with low cytoplasmic TLR6 expression group in multivariate analysis (adjusted HR 0.74, 95% CI 0.60–0.91). In the subgroup analysis of diffuse type of histology, the 5-year survival was better in high cytoplasmic TLR6 expression group in multivariable analysis (HR 0.62, 95% CI 0.46–0.83). In the intestinal type of histology subgroup, no significant differences between the groups were present. TLR1, TLR4, and TLR5 expression were not associated with 5-year survival. In conclusion, cytoplasmic TLR2 and TLR6 expression seem to have independent prognostic impact in gastric cancer, while TLR1, TLR4, and TLR5 do not.
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Data are available upon reasonable request from the corresponding author.
References
Sung H, Ferlay J, Siegel RL, Laversanne M, Soerjomataram I, Jemal A, Bray F (2021) Global cancer statistics 2020: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin 71(3):209–249
Asplund J, Kauppila JH, Mattsson F, Lagergren J (2018) Survival trends in gastric adenocarcinoma: a population-based study in Sweden. Ann Surg Oncol 25:2693–2702. https://doi.org/10.1245/s10434-018-6627-y
Sexton RE, Al Hallak MN, Diab M, Azmi AS (2020) Gastric cancer: a comprehensive review of current and future treatment strategies. Cancer Metastasis Rev 39:1179–1203. https://doi.org/10.1007/s10555-020-09925-3
Castaño-Rodríguez N, Kaakoush NO, Mitchell HM (2014) Pattern-recognition receptors and gastric cancer. Front Immunol 22(5):336. https://doi.org/10.3389/fimmu.2014.00336
Pimentel-Nunes P, Afonso L, Lopes P, Roncon-Albuquerque R, Gonçalves N, Henrique R, Moreira-Dias L, Leite-Moreira AF, Dinis-Ribeiro M (2011) Increased expression of Toll-like receptors (TLR) 2, 4 and 5 in gastric dysplasia. Pathol Oncol Res 17:677–683. https://doi.org/10.1007/s12253-011-9368-9
Braunstein MJ, Kucharczyk J, Adams S (2018) Targeting Toll-like receptors for cancer therapy. Target Oncol 13:583–598. https://doi.org/10.1007/s11523-018-0589-7
Cui L, Wang X, Zhang D (2020) TLRs as a promise target along with immune checkpoint against gastric cancer. Front Cell Dev Biol 8:611444. https://doi.org/10.3389/fcell.2020.611444
Diakowska D, Nienartowicz M, Grabowski K, Rosińczuk J, Krzystek-Korpacka M (2019) Toll-like receptors TLR-2, TLR-4, TLR-7, and TLR-9 in tumor tissue and serum of the patients with esophageal squamous cell carcinoma and gastro-esophageal junction cancer. Adv Clin Exp Med 28:515–522. https://doi.org/10.17219/acem/87012
Shcheblyakov DV, Logunov DY, Tukhvatulin AI, Shmarov MM, Naroditsky BS, Gintsburg AL (2010) Toll-like receptors (TLRs): the role in tumor progression. Actanaturae 2:21–29. https://doi.org/10.32607/20758251-2010-2-3-21-29
Sitarz R, Skierucha M, Mielko J, Offerhaus GJA, Maciejewski R, Polkowski WP (2018) Gastric cancer: epidemiology, prevention, classification, and treatment. Cancer Manag Res 10:239–248. https://doi.org/10.2147/CMAR.S149619
Javaid N, Choi S (2020) Toll-like receptors from the perspective of cancer treatment. Cancers 12(2):297. https://doi.org/10.3390/cancers12020297
Zheng R, Ma J (2022) Immunotherapeutic implications of Toll-like receptors activation in tumor microenvironment. Pharmaceutics 14(11):2285. https://doi.org/10.3390/pharmaceutics14112285
Farooq M, Batool M, Kim MS, Choi S (2021) Toll-like receptors as a therapeutic target in the era of immunotherapies. Front Cell Dev Biol 9:756315. https://doi.org/10.3389/fcell.2021.756315
Urban-Wojciuk Z, Khan MM, Oyler BL, Fåhraeus R, Marek-Trzonkowska N, Nita-Lazar A, Hupp TR, Goodlett DR (2019) The role of TLRs in anti-cancer immunity and tumor rejection. Front Immunol 22(10):2388. https://doi.org/10.3389/fimmu.2019.02388
Chen X, Zhang Y, Fu Y (2022) The critical role of Toll-like receptor-mediated signaling in cancer immunotherapy. Med Drug Discov 14. https://doi.org/10.1016/j.medidd.2022.100122
Vacchelli E, Eggermont A, Sautès-Fridman C, Galon J, Zitvogel L, Kroemer G, Galluzzi L (2013) Trial watch Toll-like receptor agonists for cancer therapy. Oncoimmunology 2(8):e25238. https://doi.org/10.4161/onci.25238
Kasurinen A, Hagström J, Laitinen A, Kokkola A, Böckelman C, Haglund C (2019) Evaluation of Toll-like receptors as prognostic biomarkers in gastric cancer: high tissue TLR5 predicts a better outcome. Sci Rep 9(1):12553. https://doi.org/10.1038/s41598-019-49111-2
Schmaußer B, Andrulis M, Endrich S, Müller-Hermelink HK, Eck M (2005) Toll-like receptors TLR4, TLR5 and TLR9 on gastric carcinoma cells: an implication for interaction with Helicobacter pylori. Int J Med Microbiol 295:179–185. https://doi.org/10.1016/j.ijmm.2005.02.009
Yang H, Wang B, Yan J, Wang T, Zhou X-N, Wen H-Y, Zhu X-M (2014) Toll-like receptor 2 promotes invasion by SGC-7901 human gastric carcinoma cells and is associated with gastric carcinoma metastasis. Ann Clin Lab Sci 44(2):158–66
Santini D, Angeletti S, Ruzzo A et al (2008) Toll-like receptor 4 Asp299Gly and Thr399Ile polymorphisms in gastric cancer of intestinal and diffuse histotypes. Clin Exp Immunol 154:360–364. https://doi.org/10.1111/j.1365-2249.2008.03776.x
Song EJ, Kang MJ, Kim YS, Kim SM, Lee SE, Kim CH, Kim DJ, Park JH (2011) Flagellin promotes the proliferation of gastric cancer cells via the Toll-like receptor 5. Int J Mol Med 28:115–119. https://doi.org/10.3892/ijmm.2011.656
Huhta H, Helminen O, Kauppila JH, Salo T, Porvari K, Saarnio J, Lehenkari PP, Karttunen TJ (2016) The expression of Toll-like receptors in normal human and murine gastrointestinal organs and the effect of microbiome and cancer. Histochem Cytochem 64(8):470–82. https://doi.org/10.1369/0022155416656154
Fernandez-Garcia B, Eiró N, González-Reyes S, González L, Aguirre A, González LO, Del Casar JM, García-Muñiz JL (1997) Vizoso FJ (2014) Clinical significance of Toll-like receptor 3, 4, and 9 in gastric cancer. J Immunother 37:77–83. https://doi.org/10.1097/CJI.0000000000000016
Yuan X, Zhou Y, Wang W, Li J, Xie G, Zhao Y, Xu D, Shen L (2013) Activation of TLR4 signaling promotes gastric cancer progression by inducing mitochondrial ROS production. Cell Death Dis 4:e794–e794. https://doi.org/10.1038/cddis.2013.334
Bankhead P, Loughrey MB, Fernández JA, Dombrowski Y, McArt DG, Dunne PD, McQuaid S, Gray RT, Murray LJ, Coleman HG, James JA, Salto-Tellez M, Hamilton PW (2017) QuPath: open source software for digital pathology image analysis. Sci Rep 7:16877–16878. https://doi.org/10.1038/s41598-017-17204-5
Kim SW, Roh J, Park CS (2016) Immunohistochemistry for pathologists: protocols, pitfalls, and tips. J Pathol Transl Med 50:411–418. https://doi.org/10.4132/jptm.2016.08.08
Eskuri M, Kemi N, Helminen O, Huhta H, Kauppila JH (2023) Toll-like receptors 3, 7, 8, and 9 in gastric cancer. APMIS 131:92–99. https://doi.org/10.1111/apm.13281
Uno K, Kato K, Shimosegawa T (2014) Novel role of toll-like receptors in Helicobacter pylori-induced gastric malignancy. World J Gastroenterol 20:5244–5251. https://doi.org/10.3748/wjg.v20.i18.5244
Zhao D, Wu YH, Zhao TC, Jia ZF, Cao DH, Yang NA, Wang YQ, Cao XY, Jiang J (2019) Single-nucleotide polymorphisms in Toll-like receptor genes are associated with the prognosis of gastric cancer and are not associated with Helicobacter pylori infection. Infect Genet Evol 73:384–389. https://doi.org/10.1016/j.meegid.2019.06.005
Fukata M, Abreu MT (2008) Role of Toll-like receptors in gastrointestinal malignancies. Oncogene 27:234–243. https://doi.org/10.1038/sj.onc.1210908
Van Bergenhenegouwen J, Plantinga TS, Joosten LAB, Netea MG, Folkerts G, Kraneveld AD, Garssen J, Vos AP (2013) TLR2 & Co: a critical analysis of the complex interactions between TLR2 and coreceptors. J Leukoc Biol 94:885–902. https://doi.org/10.1189/jlb.0113003
Oliveira-Nascimento L, Massari P, Wetzler LM (2012) The role of TLR2 infection and immunity. Front Immunol 18(3):79. https://doi.org/10.3389/fimmu.2012.00079
Huhta H, Helminen O, Lehenkari PP, Saarnio J, Karttunen TJ, Kauppila JH (2016) Toll-like receptors 1, 2, 4 and 6 in esophageal epithelium, Barrett’s esophagus, dysplasia and adenocarcinoma. Oncotarget 7(17):23658–67. https://doi.org/10.18632/oncotarget.8151
Semlali A, Almutairi M, Pathan AAK, Azzi A, Parine NR, AlAmri A, Arafah M, Aljebreen AM, Alharbi O, Almadi MA, Azzam NA, Alanazi M, Rouabhia M (2019) Toll-like receptor 6 expression, sequence variants, and their association with colorectal cancer risk. J Cancer 10:2969–2981. https://doi.org/10.7150/jca.31011
Sato Y, Wakita A, Maeda E, Nagaki Y, Sasamori R, Kemuriyama K, Nozaki S, Satoru I, Terata K, Imai K, Nanjo H, Nomura K, Minamiya Y (2023) High TLR6 expression status predicts a more favorable prognosis after esophagectomy for locally advanced thoracic esophageal squamous cell carcinoma. Curr Oncol 30:4724–4735. https://doi.org/10.3390/curroncol30050356
Acknowledgements
We thank Erja Tomperi and Riitta Vuento for important technical assistance. The study benefited from samples/data from Northern Finland Biobank Borealis, Oulu, Finland.
Funding
This study was supported by grants from The Finnish Medical Foundation (M.E.), The Maud Kuistila Memorial Foundation (M.E.), Orion Research Foundation (J.H.K.), Thelma Mäkikyrö Foundation (J.H.K.), and Mary and Georg C. Ehrnroot Foundation (J.H.K.), Päivikki and Sakari Sohlberg Foundation (J.H.K.), The Finnish Cancer Foundation (J.H.K.), and Sigrid Juselius Foundation (J.H.K.).
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All the authors conceived and designed the study. N.K., O.H., H.H., and J.H.K. acquired the data. M.E. and N.K. performed the experiments. M.E. analyzed the data, drafted the manuscript, and prepared the original figures and tables. All the authors critically reviewed, edited, and approved the manuscript. J.H.K provided funding, supervised the study, and is the guarantor of the study.
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The use of patient samples and the data inquiry were approved by the Oulu University Hospital Ethics Committee (15.2.2016 §51). The need to obtain a written or oral consent from the patients was waived by the Finnish National Authority for Medicolegal Affairs (VALVIRA). This study was performed in accordance with the Declaration of Helsinki.
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Supplementary Figure 1.
Representative images of TLR1, TLR4 and TLR5 expression immunostaining in gastric adenocarcinoma. Low cytoplasmic TLR1 expression (A), high cytoplasmic TLR1 expression (B), low cytoplasmic TLR4 expression (C), high cytoplasmic TLR4 expression (D), low membranous TLR4 expression (E), high membranous TLR4 expression (F), low cytoplasmic TLR5 expression (G), high cytoplasmic TLR5 expression (H), low nuclear TLR5 expression (I), and high nuclear TLR5 expression (J). (PNG 2.57 Mb).
Supplementary Figure 2.
The Kaplan-Meier figures presenting 5-year survival stratified by Toll-like receptor expressions in gastric adenocarcinoma, with p values for significance based on the log-rank test. Cytoplasmic TLR1 expression (A), cytoplasmic TLR4 expression (B), membranous TLR4 expression (C), cytoplasmic TLR5 expression (D), and nuclear TLR5 expression (E). (PNG 9.21 mb).
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Eskuri, M., Kemi, N., Helminen, O. et al. Toll-like receptors 1, 2, 4, 5, and 6 in gastric cancer. Virchows Arch (2023). https://doi.org/10.1007/s00428-023-03635-1
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DOI: https://doi.org/10.1007/s00428-023-03635-1