Direct conversion of osteosarcoma to adipocytes by targeting TNIK
Toru Hirozane, … , Eisuke Kobayashi, Tesshi Yamada
Toru Hirozane, … , Eisuke Kobayashi, Tesshi Yamada
Published January 5, 2021
Citation Information: JCI Insight. 2021;6(3):e137245. https://doi.org/10.1172/jci.insight.137245.
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Research Article Cell biology Therapeutics

Direct conversion of osteosarcoma to adipocytes by targeting TNIK

Abstract

Osteosarcoma (OS) is an aggressive mesenchymal tumor for which no molecularly targeted therapies are available. We have previously identified TRAF2- and NCK-interacting protein kinase (TNIK) as an essential factor for the transactivation of Wnt signal target genes and shown that its inhibition leads to eradication of colorectal cancer stem cells. The involvement of Wnt signaling in the pathogenesis of OS has been implicated. The aim of the present study was to examine the potential of TNIK as a therapeutic target in OS. RNA interference or pharmacological inhibition of TNIK suppressed the proliferation of OS cells. Transcriptome analysis suggested that a small-molecule inhibitor of TNIK upregulated the expression of genes involved in OS cell metabolism and downregulated transcription factors essential for maintaining the stem cell phenotype. Metabolome analysis revealed that this TNIK inhibitor redirected the metabolic network from carbon flux toward lipid accumulation in OS cells. Using in vitro and in vivo OS models, we confirmed that TNIK inhibition abrogated the OS stem cell phenotype, simultaneously driving conversion of OS cells to adipocyte-like cells through induction of PPARγ. In relation to potential therapeutic targeting in clinical practice, TNIK was confirmed to be in an active state in OS cell lines and clinical specimens. From these findings, we conclude that TNIK is applicable as a potential target for treatment of OS, affecting cell fate determination.

Authors

Toru Hirozane, Mari Masuda, Teppei Sugano, Tetsuya Sekita, Naoko Goto, Toru Aoyama, Takato Sakagami, Yuko Uno, Hideki Moriyama, Masaaki Sawa, Naofumi Asano, Masaya Nakamura, Morio Matsumoto, Robert Nakayama, Tadashi Kondo, Akira Kawai, Eisuke Kobayashi, Tesshi Yamada

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Figure 1

TNIK inhibition suppresses OS cell growth.

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TNIK inhibition suppresses OS cell growth. (A) U2OS and NOS-1 OS cells w...
(A) U2OS and NOS-1 OS cells were transfected with control siRNA (Ctrl) and siRNA against TNIK (siTNIK) in triplicate, and their expression of the TNIK gene (normalized to ACTB) was quantified by real-time RT-PCR. Data represent the mean ± SD of 3 replicates. One-way ANOVA, ****P < 0.0001. (B) Real-time growth monitoring of U2OS and NOS-1 OS cells transfected with control siRNA and siTNIK. Values at the time of transfection (0 hour) were set to 1. Data represent the mean ± SD of 4 replicates. One-way ANOVA, ****P < 0.0001. (C) The expression of TNIK (normalized to ACTB) in U2OS and NOS-10 cells treated with DMSO (Ctrl), NCB-0846 (846), or NCB-0970 (970) for 24 hours was quantified by RT-PCR. Data represent the mean ± SD of 3 replicates. One-way ANOVA, **P < 0.01, ****P < 0.0001. (D) Nine OS (U2OS, NOS-10, MNNG/HOS, NOS-1, HsOS1, G292, HuO9N2, HuO3N1, and NY) and (E) two osteoblast (NHOst and HOB-c) cell lines were cultured in the presence of 0.003–10 μM NCB-0846 or NCB-0970 for 72 hours, and their relative viability (IC50) was assessed in terms of ATP production. Data represent the mean ± SD of 3 replicates. (F) NOS-10 cells were inoculated into the subcutaneous tissues of 6-week-old female NOD/SCID mice. When the average volume of the xenografts reached 100 mm3, twice daily administration of water (vehicle, n = 10) or 80 mg/kg NCB-0846 hydrochloride (NCB-0846, n = 10) was initiated with a day off every 3 days and continued for a total of 15 days. Tumor growth and body weight in the vehicle group were compared with those in the NCB-0846 group at the same time points on a daily basis. Multiple t test corrected using the Holm-Sidak method, **P < 0.01, ***P < 0.001. Data represent the mean ± SEM.