CLEFMA Activates the Extrinsic and Intrinsic Apoptotic Processes through JNK1/2 and p38 Pathways in Human Osteosarcoma Cells.

Jia-Sin Yang,Renn-Chia Lin,Shun-Fa Yang,Ya-Chiu Lin,Ko-Hsiu Lu,Yi-Hsien Hsieh,Geng-Chung Li,Heng-Hsiung Wu

doi:10.3390/molecules24183280

Abstract

Due to the poor prognosis of metastatic osteosarcoma, chemotherapy is usually employed in the adjuvant situation to improve the prognosis and the chances of long-term survival. 4-[3,5-Bis(2-chlorobenzylidene)-4-oxo-piperidine-1-yl]-4-oxo-2-butenoic acid (CLEFMA) is a synthetic analog of curcumin and possesses anti-inflammatory and anticancer properties. To further obtain information regarding the apoptotic pathway induced by CLEFMA in osteosarcoma cells, microculture tetrazolium assay, annexin V-FITC/PI apoptosis staining assay, human apoptosis array, and Western blotting were employed. CLEFMA dose-dependently decreased the cell viabilities of human osteosarcoma U2OS and HOS cells and significantly induced apoptosis in human osteosarcoma cells. In addition to the effector caspase 3, CLEFMA significantly activated both extrinsic caspase 8 and intrinsic caspase 9 initiators. Moreover, CLEFMA increased the phosphorylation of extracellular signal-regulated protein kinases (ERK)1/2, c-Jun N-terminal kinases (JNK)1/2 and p38. Using inhibitors of JNK (JNK-in-8) and p38 (SB203580), CLEFMA’s increases of cleaved caspases 3, 8, and 9 could be expectedly suppressed, but they could not be affected by co-treatment with the ERK inhibitor (U0126). Conclusively, CLEFMA activates both extrinsic and intrinsic apoptotic pathways in human osteosarcoma cells through JNK and p38 signaling. These findings contribute to a better understanding of the mechanisms responsible for CLEFMA’s apoptotic effects on human osteosarcoma cells.

Highlights

Osteosarcoma, the most common histological form of primary bone cancer, is most prevalent in teenagers and young adults [1,2]
A 24 h treatment with 20 μM of CLEFMA showed about a 50% reduction, while a 24 h treatment with 80 μM of showed a 50% reduction, 24 habout treatment decreased theabout cell viability of U2OSwhile cellsa by
Antitumor effects in retinoblastoma cells by regulating the Jun N-terminal kinase (JNK) and p38 pathways [23], while this occurs curcumin exerts antitumor effects in retinoblastoma cells by regulating the JNK and p38 pathways through ERK1/2 and p38 signaling in malignant mesothelioma cells [24]

Summary

Introduction

Osteosarcoma, the most common histological form of primary bone cancer, is most prevalent in teenagers and young adults [1,2]. Surgical en bloc resection of the cancer to achieve a complete radical excision has been the treatment of choice for osteosarcoma [2], but its prognosis is poor because of its highly metastatic potential. Novel agents that target particular intracellular signaling pathways related to the distinctive properties of osteosarcoma cells need to be developed. Multiple stress-inducible molecules, such as mitogen-activated protein kinase (MAPK)/extracellular signal-regulated protein kinase (ERK), c-Jun N-terminal kinase (JNK), and nuclear factor kappa B (NF-κB), have been implied in transmitting the apoptotic pathway [5,6]. Failure to undergo apoptosis may result in treatment resistance. Thereby, understanding the molecular events that regulate apoptosis in response to chemotherapy provides novel opportunities to develop molecular-targeted therapy through the intrinsic and/or extrinsic pathways for osteosarcoma, which is very difficult to cure

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