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Open AccessArticle

Elevated PDK1 Expression Drives PI3K/AKT/MTOR Signaling Promotes Radiation-Resistant and Dedifferentiated Phenotype of Hepatocellular Carcinoma

1
Department of Hematology and Oncology, Cancer Center, Taipei Medical University-Shuang Ho Hospital, New Taipei City 235, Taiwan
2
Department of Medical Research and Education, Taipei Medical University-Shuang Ho Hospital, New Taipei City 235, Taiwan
3
Department of General Surgery, En Chu Kong Hospital, New Taipei City 237, Taiwan
4
Department of Health Care Management, Yuanpei University of Medical Technology, Hsinchu 300, Taiwan
5
Department of Emergency Medicine, School of Medicine, Taipei Medical University, Taipei City 110, Taiwan
6
Department of Emergency Medicine, Taipei Medical University-Shuang Ho Hospital, New Taipei City 235, Taiwan
7
Department of Pathology, Taipei Medical University-Shuang Ho Hospital, New Taipei City 235, Taiwan
8
Department of Medical Laboratory Science and Biotechnology, Yuanpei University of Medical Technology, Hsinchu City 300, Taiwan
9
Department of Radiology, School of Medicine, College of Medicine, Taipei Medical University, Taipei City 110, Taiwan
10
Department of Radiology, Taipei Medical University-Shuang Ho Hospital, New Taipei City 235, Taiwan
11
Graduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University, Taipei City 110, Taiwan
*
Author to whom correspondence should be addressed.
These authors have contributed equally to this work.
Cells 2020, 9(3), 746; https://doi.org/10.3390/cells9030746
Received: 21 January 2020 / Revised: 6 March 2020 / Accepted: 13 March 2020 / Published: 18 March 2020
(This article belongs to the Special Issue PI3K/AKT/mTOR Signaling Network in Human Health and Diseases)
Resistance to radiotherapy (IR), with consequent disease recurrence, continues to limit the efficacy of contemporary anticancer treatment for patients with hepatocellular carcinoma (HCC), especially in late stage. Despite accruing evidence implicating the PI3K/AKT signaling pathway in cancer-promoting hypoxia, cancerous cell proliferation and radiotherapy-resistance, it remains unclear which molecular constituent of the pathway facilitates adaptation of aggressive HCC cells to tumoral stress signals and drives their evasion of repeated IR-toxicity. This present study investigated the role of PDK1 signaling in IR-resistance, enhanced DNA damage repair and post-IR relapse, characteristic of aggressive HCC cells, while exploring potential PDK1-targetability to improve radiosensitivity. The study employed bioinformatics analyses of gene expression profile and functional protein–protein interaction, generation of IR-resistant clones, flow cytometry-based ALDH activity and side-population (SP) characterization, siRNA-mediated loss-of-PDK1function, western-blotting, immunohistochemistry and functional assays including cell viability, migration, invasion, clonogenicity and tumorsphere formation assays. We showed that the aberrantly expressed PDK1 characterizes poorly differentiated HCC CVCL_7955, Mahlavu, SK-HEP1 and Hep3B cells, compared to the well-differentiated Huh7 or normal adult liver epithelial THLE-2 cells, and independently activates the PI3K/AKT/mTOR signaling. Molecular ablation of PDK1 function enhanced susceptibility of HCC cells to IR and was associated with deactivated PI3K/AKT/mTOR signaling. Additionally, PDK1-driven IR-resistance positively correlated with activated PI3K signaling, enhanced HCC cell motility and invasiveness, augmented EMT, upregulated stemness markers ALDH1A1, PROM1, SOX2, KLF4 and POU5F1, increased tumorsphere-formation efficiency and suppressed biomarkers of DNA damage—RAD50, MSH3, MLH3 and ERCC2. Furthermore, the acquired IR-resistant phenotype of Huh7 cells was strongly associated with significantly increased ALDH activity, SP-enrichment, and direct ALDH1-PDK1 interaction. Moreover, BX795-mediated pharmacological inhibition of PDK1 synergistically enhances the radiosensitivity of erstwhile resistant cells, increased Bax/Bcl-2 apoptotic ratio, while suppressing oncogenicity and clonogenicity. We provide preclinical evidence implicating PDK1 as an active driver of IR-resistance by activation of the PI3K/AKT/mTOR signaling, up-modulation of cancer stemness signaling and suppression of DNA damage, thus, projecting PDK1-targeting as a putative enhancer of radiosensitivity and a potential new therapeutic approach for patients with IR-resistant HCC. View Full-Text
Keywords: hepatocellular cancer; HCC; LIHC; PDK1; PI3K/AKT/mTOR pathway; BX795; selective inhibitor; radiotherapy; radioresistance; combination therapy; stemness; DNA damage hepatocellular cancer; HCC; LIHC; PDK1; PI3K/AKT/mTOR pathway; BX795; selective inhibitor; radiotherapy; radioresistance; combination therapy; stemness; DNA damage
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Bamodu, O.A.; Chang, H.-L.; Ong, J.-R.; Lee, W.-H.; Yeh, C.-T.; Tsai, J.-T. Elevated PDK1 Expression Drives PI3K/AKT/MTOR Signaling Promotes Radiation-Resistant and Dedifferentiated Phenotype of Hepatocellular Carcinoma. Cells 2020, 9, 746.

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