Abstract
Osteosarcoma (OS) is the most common primary bone tumor in children and young adults. Several studies have confirmed the involvement of the insulin-like growth factor (IGF) system in the regulation of OS cell proliferation and differentiation as well as in the protection of cells from chemotherapy. Insulin receptor substrate (IRS)-1 is a critical mediator of IGF-1R signaling, and we recently reported that its overexpression in OS cells increases proliferation, migration, and metastasis both in vitro and in vivo. In this study, we evaluated the efficacy of NT157, a selective inhibitor of IRS-1/2, in a panel of OS cells. A strong dose-dependent inhibition of growth was observed in the MG-63, OS-19, and U-2OS OS cell lines, displaying IC50 values at sub-micromolar doses after 72 h of treatment. Exposure to NT157 elicited dose- and time-dependent decreases in IRS-1 levels. Moreover, a protein analysis showed that the degradation of IRS-1 inhibited the activation of principal downstream mediators of the IGF pathway. NT157 significantly affected the cells’ migratory ability, as confirmed by a wound-healing assay. The inhibitor induced cytostatic effects, as evidenced by G2/M cell cycle arrest, and did not affect apoptosis. Consequently, NT157 was combined with drugs used to treat OS in order to capitalize on its therapeutic potential. Simultaneous treatments were made in association with chemotherapeutic agents in a fixed ratio for 72 h and cell proliferation was determined by MTT assay. Synergistic or addictive effects with respect to single agents are expressed as the combination index. Significant synergistic effects were obtained with several targeted drugs, such as Everolimus, a mammalian target of rapamycin (mTOR) inhibitor, and NVP-BEZ235, a dual inhibitor of PI-3K/mTOR. Overall, these findings provide evidence for the effectiveness of a selected inhibitor of IRS-1/2 NT157 in OS cells, displaying a promising approach based on the targeting of IRS-1 combined with other therapies for the treatment of this pediatric solid tumor.
Highlights
The activation of the insulin-like growth factor (IGF) system regulates several aspects of the malignant phenotype, including the development and progression of cancer and metastasis [1, 2]
In vitro Activity of NT157 in OS Cells We previously demonstrated that the IGF system, including its critical mediator Insulin receptor substrate (IRS)-1, is involved in the regulation of OS cell proliferation [26]
The efficacy of the selective inhibitor of IRS-1/2 NT157 was investigated in three representative OS cell lines
Summary
The activation of the insulin-like growth factor (IGF) system regulates several aspects of the malignant phenotype, including the development and progression of cancer and metastasis [1, 2]. A number of different physiologic pathways involved in both mitogenic and metabolic responses, such as steroids, hormones, cytokines, and integrins, can regulate IRS protein expression [5,6,7,8] Many of these effectorsignaling pathways have been implicated in tumorigenesis and cancer progression. IRS1 is constitutively activated in a variety of solid tumors, including breast cancers, leiomyomas, Wilms’ tumors, rhabdomyosarcomas, liposarcomas, leiomyosarcomas, and adrenal cortical carcinomas [9]. In addition to their canonical function as cytosolic signal transduction molecules, IRS proteins can be shuttled to the nucleus and may contribute to the process of malignant transformation. IRS-1 can interact with transcription factors, such as β-catenin, ER-α, and the androgen receptor (AR), to modulate the promoter activity of several genes involved in malignant transformation [14,15,16,17]
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