CD147 augmented monocarboxylate transporter-1/4 expression through modulation of the Akt-FoxO3-NF-κB pathway promotes cholangiocarcinoma migration and invasion
- 139 Downloads
Cholangiocarcinoma (CCA) is an aggressive type of cancer. The major obstacles for treatment are its late presentation and the occurrence metastases. Targeting the metastatic process may serve as a treatment option. CD147 is a membrane protein that promotes CCA metastasis. High lactate levels in CCA are predicted to result from lactate dehydrogenase A expression and sensitivity to monocarboxylate transporter (MCT) inhibitors. An involvement of CD147 in MCT maturation has been reported, but the exact role of MCT in CCA is not clear. Here, we aimed to assess the mechanism of CD147-promoted CCA progression through MCT regulation.
The expression levels of CD147 and MCT-1/4 in human CCA tissues were determined by immunohistochemistry. Two CD147 knockout (CD147 KO) CCA cell (KKU-213) clones were established using the CRISPR/Cas9 system. Cell migration and invasion were determined using a Boyden chamber assay. Temporal protein levels were modified by siRNA, specific inhibitors and/or activators. The expression of target proteins was determined using Western blot analyses.
CD147 and MCT-1/4 were found to be overexpressed in CCA tissues compared to normal bile duct tissues. In addition, we found that CD147 knockdown significantly alleviated CCA cell migration and invasion, concomitant with decreased pAkt, pFoxO3, pNF-κB (pp65) and MCT-1/4 levels. Conversely, we found that FoxO3 knockdown led to recovered migration/invasion abilities and increased pp65 and MCT-1/4 expression levels. The involvement of Akt in the regulation of MCT-1/4 expression through CD147 was established by inhibition and activation of Akt phosphorylation.
Our data indicate that CD147 promotes the malignant progression of CCA cells by activating the Akt-FoxO3-NF-κB-MCT-1/4 axis. As such, CD147 may serve as a possible target for advanced CCA treatment.
KeywordsCholangiocarcinoma CD147 Akt FoxO3 MCT NF-κB
Protein kinase B (PKB)
Clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9
Dulbecco’s modified Eagle’s medium
- CD147 KO
The authors would like to thank Dr. Brett Stringer for LentiCRISPRv2 puro, Prof. Didier Trono for pCMVR8.74 and pMD2.G, Prof. Kazuo Umezawa, Aichi Medical University, Japan for providing DHMEQ and Prof. James A Will for editing this manuscript via the KKU Publication Clinic, Khon Kaen University, Thailand.
This project was supported by a TRF-MRC (Newton fund) project grant to C. Pairojkul (DBG5980004), TRF Senior Research Scholar Grant to S. Wongkham (RTA5780012) and Khon Kaen University, Thailand (#KKU61003502 to K. Vaeteewoottacharn).
Compliance with ethical standards
The authors declare no potential conflicts of interest.
All procedures performed in studies involving human participants were conducted in accordance with the ethical standards of the Institutional Research Committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards. Human CCA tissues were obtained from the specimen bank of the Cholangiocarcinoma Research Institute, Khon Kaen University, Thailand. The patients had undergone liver resection at Srinagarind Hospital, Thailand. Informed patient consents were obtained prior to initiation of the study. The research protocol (#HE571283 and #HE581369) was approved by the Human Research Ethics Committee at Khon Kaen University.
- 3.S.A. Khan, B.R. Davidson, R.D. Goldin, N. Heaton, J. Karani, S.P. Pereira, W.M. Rosenberg, P. Tait, S.D. Taylor-Robinson, A.V. Thillainayagam, H.C. Thomas, H. Wasan, G. British, Society of, guidelines for the diagnosis and treatment of cholangiocarcinoma: An update. Gut 61(1657-1669) (2012)PubMedCrossRefGoogle Scholar
- 9.W. Schneiderhan, M. Scheler, K.H. Holzmann, M. Marx, J.E. Gschwend, M. Bucholz, T.M. Gress, T. Seufferlein, G. Adler, F. Oswald, CD147 silencing inhibits lactate transport and reduces malignant potential of pancreatic cancer cells in in vivo and in vitro models. Gut 58, 1391–1398 (2009)PubMedCrossRefGoogle Scholar
- 10.J. Hao, H. Chen, M.C. Madigan, P.J. Cozzi, J. Beretov, W. Xiao, W.J. Delprado, P.J. Russell, Y. Li, Co-expression of CD147 (EMMPRIN), CD44v3-10, MDR1 and monocarboxylate transporters is associated with prostate cancer drug resistance and progression. Br J Cancer 103, 1008–1018 (2010)PubMedPubMedCentralCrossRefGoogle Scholar
- 15.P. Zhao, W. Zhang, S.J. Wang, X.L. Yu, J. Tang, W. Huang, Y. Li, H.Y. Cui, Y.S. Guo, J. Tavernier, S.H. Zhang, J.L. Jiang, Z.N. Chen, HAb18G/CD147 promotes cell motility by regulating annexin II-activated RhoA and Rac1 signaling pathways in hepatocellular carcinoma cells. Hepatology 54, 2012–2024 (2011)PubMedCrossRefGoogle Scholar
- 18.P. Huang, S. Chang, X. Jiang, J. Su, C. Dong, X. Liu, Z. Yuan, Z. Zhang, H. Liao, RNA interference targeting CD147 inhibits the proliferation, invasiveness, and metastatic activity of thyroid carcinoma cells by down-regulating glycolysis. Int J Clin Exp Pathol 8, 309–318 (2015)PubMedPubMedCentralGoogle Scholar
- 25.G. Matsumoto, J. Namekawa, M. Muta, T. Nakamura, H. Bando, K. Tohyama, M. Toi, K. Umezawa, Targeting of nuclear factor kappaB pathways by dehydroxymethylepoxyquinomicin, a novel inhibitor of breast carcinomas: Antitumor and antiangiogenic potential in vivo. Clin Cancer Res 11, 1287–1293 (2005)PubMedGoogle Scholar
- 30.W. Seubwai, C. Wongkham, A. Puapairoj, N. Khuntikeo, A. Pugkhem, C. Hahnvajanawong, J. Chaiyagool, K. Umezawa, S. Okada, S. Wongkham, Aberrant expression of NF-kappaB in liver fluke associated cholangiocarcinoma: Implications for targeted therapy. PLoS One 9, e106056 (2014)PubMedPubMedCentralCrossRefGoogle Scholar
- 31.M.G. Thompson, M. Larson, A. Vidrine, K. Barrios, F. Navarro, K. Meyers, P. Simms, K. Prajapati, L. Chitsike, L.M. Hellman, B.M. Baker, S.K. Watkins, FOXO3-NF-kappaB RelA protein complexes reduce Proinflammatory cell signaling and function. J Immunol 195, 5637–5647 (2015)PubMedPubMedCentralCrossRefGoogle Scholar
- 37.V. Miranda-Goncalves, M. Honavar, C. Pinheiro, O. Martinho, M.M. Pires, C. Pinheiro, M. Cordeiro, G. Bebiano, P. Costa, I. Palmeirim, R.M. Reis, F. Baltazar, Monocarboxylate transporters (MCTs) in gliomas: Expression and exploitation as therapeutic targets. Neuro-Oncology 15, 172–188 (2013)PubMedCrossRefGoogle Scholar
- 39.N. Tanaka, M. Zhao, L. Tang, A.A. Patel, Q. Xi, H.T. Van, H. Takahashi, A.A. Osman, J. Zhang, J. Wang, J.N. Myers, G. Zhou, Gain-of-function mutant p53 promotes the oncogenic potential of head and neck squamous cell carcinoma cells by targeting the transcription factors FOXO3a and FOXM1. Oncogene 37, 1279–1292 (2017)PubMedPubMedCentralCrossRefGoogle Scholar
- 41.D. Ni, X. Ma, H.Z. Li, Y. Gao, X.T. Li, Y. Zhang, Q. Ai, P. Zhang, E.L. Song, Q.B. Huang, Y. Fan, X. Zhang, Downregulation of FOXO3a promotes tumor metastasis and is associated with metastasis-free survival of patients with clear cell renal cell carcinoma. Clin Cancer Res 20, 1779–1790 (2014)PubMedCrossRefPubMedCentralGoogle Scholar
- 43.S. Yothaisong, H. Dokduang, A. Techasen, N. Namwat, P. Yongvanit, V. Bhudhisawasdi, A. Puapairoj, G.J. Riggins, W. Loilome, Increased activation of PI3K/AKT signaling pathway is associated with cholangiocarcinoma metastasis and PI3K/mTOR inhibition presents a possible therapeutic strategy. Tumour Biol 34, 3637–3648 (2014)CrossRefGoogle Scholar
- 47.H. Jo, S. Mondal, D. Tan, E. Nagata, S. Takizawa, A.K. Sharma, Q. Hou, K. Shanmugasundaram, A. Prasad, J.K. Tung, A.O. Tejeda, H. Man, A.C. Rigby, H.R. Luo, Small molecule-induced cytosolic activation of protein kinase Akt rescues ischemia-elicited neuronal death. Proc Natl Acad Sci U S A 109, 10581–10586 (2012)PubMedPubMedCentralCrossRefGoogle Scholar
- 50.F. Fei, X. Li, L. Xu, D. Li, Z. Zhang, X. Guo, H. Yang, Z. Chen, J. Xing, CD147-CD98hc complex contributes to poor prognosis of non-small cell lung cancer patients through promoting cell proliferation via the PI3K/Akt signaling pathway. Ann Surg Oncol 21, 4359–4368 (2014)PubMedCrossRefGoogle Scholar