FH535 Suppresses Osteosarcoma Growth In Vitro and Inhibits Wnt Signaling through Tankyrases.

Carl T Gustafson,Tewodros Mamo,Avudaiappan Maran,Michael J Yaszemski,Kristen L Shogren

doi:10.3389/fphar.2017.00285

Carl T Gustafson, Tewodros Mamo + Show 3 more

Open Access

https://doi.org/10.3389/fphar.2017.00285

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Abstract

Osteosarcoma (OS) is an aggressive primary bone tumor which exhibits aberrantly activated Wnt signaling. The canonical Wnt signaling cascade has been shown to drive cancer progression and metastasis through the activation of β-catenin. Hence, small molecule inhibitors of Wnt targets are being explored as primary or adjuvant chemotherapy. In this study, we have investigated the ability of FH535, an antagonist of Wnt signaling, to inhibit the growth of OS cells. We found that FH535 was cytotoxic in all OS cell lines which were tested (143b, U2OS, SaOS-2, HOS, K7M2) but well tolerated by normal human osteoblast cells. Additionally, we have developed an in vitro model of doxorubicin-resistant OS and found that these cells were highly responsive to FH535 treatment. Our analysis provided evidence that FH535 strongly inhibited markers of canonical Wnt signaling. In addition, our findings demonstrate a reduction in PAR-modification of Axin2 indicating inhibition of the tankyrase 1/2 enzymes. Moreover, we observed inhibition of auto-modification of PARP1 in the presence of FH535, indicating inhibition of PARP1 enzymatic activity. These data provide evidence that FH535 acts through the tankyrase 1/2 enzymes to suppress Wnt signaling and could be explored as a potent chemotherapeutic agent for the control of OS.

Highlights

Osteosarcoma (OS) is the most common primary bone tumor, with approximately 1000 new cases in the United States every year (Fletcher et al, 2002)
We found that FH535 is a potent inhibitor of Wnt signaling in OS, and is cytotoxic to the OS cell lines which were tested
Further experiments revealed that treatment with FH535 decreased PARylation of Axin2, as well as PARP1 autoPARylation, indicating that FH535 acts on Wnt signaling through inhibition of the TNKS1/2 enzymes. These findings provide a conclusive mechanism for FH535 inhibition of canonical Wnt signaling, and suggest that non-specific blockade of PARP1 may confound Wnt-specific claims based on the effects of FH535

Summary

Introduction

Osteosarcoma (OS) is the most common primary bone tumor, with approximately 1000 new cases in the United States every year (Fletcher et al, 2002). Lack of response to the standard chemotherapy regimen is the major cause for disease progression in OS (Bacci et al, 1993; Fletcher et al, 2002). Canonical Wnt signaling has been frequently tied to chemotherapy resistance and poor prognosis in OS; the current literature supports the concept that Wnt signaling is involved in tumor metastasis and proliferation in OS (Flores et al, 2012; Chen et al, 2015). Overexpression of Wnt promoting factors, and under-expression of endogenous Wnt inhibitors, are correlated with disease intensity and poor prognosis (Mandal et al, 2007; Lu et al, 2012). The chemotherapy regimen in OS has remained constant in the past several decades, and newly developed Wnt-targeting chemotherapeutics have not yet reached approval for clinical use in cancers

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