Novel agent DMAMCL suppresses osteosarcoma growth and decreases the stemness of osteosarcoma stem cell

Osteosarcoma (OS) is the most common primary malignancy of bone that mostly affects children, adolescents, and young people. Despite advances have been made in multimodal therapy of OS, the long-term survival rate has reached a plateau, and the main obstacles are bad response to chemotherapy and gained chemoresistance. In this study, we tested the therapeutic effect of a newly reported drug, DMAMCL, on OS. Five human OS cell lines (143B, MNNG, MG63, Saos-2, U-2OS), and the mouse fibroblast cell line (NIH3T3) and human retinal epithelial cell (ARPE19) were used. The anti-tumor effect of DMAMCL was studied by MTS assay or IncuCyte-Zoom (<i>in vitro</i>), and Xenograft-mice-model (<i>in vivo</i>). Changes of cell cycle, apoptotic cells, caspase3/7 activities, and stemness after DMAMCL treatment were investigated. BAX siRNAs were used to knockdown the expression of BAX. Expressions of CyclinB1, CDC2, BCL-2 family, PARP, CD133, and Nanog were measured by Western Blotting. DMAMCL-induced dose-dependent OS cell death <i>in vitro</i>, and suppressed tumor growth and extended the survival of xenograft-bearing mice. DMAMCL-induced G2/M phase arrest <i>in vitro</i>, and apoptosis both <i>in vitro</i> and <i>in vivo</i>. Down-regulation of BAX expression attenuated the DMAMCL-induced OS cell death <i>in vitro</i>. We also found that DMAMCL inhibited the stemness in OS cells. These results indicated that DMAMCL possess therapeutic value in OS and may be a promising candidate for the new drug discovery for OS therapy.骨肉瘤（Osteosarcoma, OS）是最常见的原发性骨恶性肿瘤，主要发病人群为儿童、青少年及青年群体。尽管骨肉瘤的多模态治疗已取得诸多进展，但其长期生存率已进入平台期，主要治疗瓶颈在于化疗应答不佳以及获得性化疗耐药。本研究针对新近报道的药物DMAMCL对骨肉瘤的治疗效果展开了评估。实验采用了5株人骨肉瘤细胞系（143B、MNNG、MG63、Saos-2、U-2OS）、小鼠成纤维细胞系（NIH3T3）以及人视网膜上皮细胞（ARPE19）。通过MTS检测、IncuCyte-Zoom活细胞成像系统（体外实验）以及异种移植小鼠模型（体内实验），探究DMAMCL的抗肿瘤活性；同时检测了DMAMCL处理后细胞周期、凋亡细胞比例、caspase3/7活性及细胞干性的变化。使用BAX小干扰RNA（BAX siRNA）敲低BAX的表达，并通过蛋白质印迹法（Western Blotting）检测CyclinB1、CDC2、BCL-2家族蛋白、PARP、CD133及Nanog的表达水平。实验结果显示，DMAMCL可在体外诱导剂量依赖性骨肉瘤细胞死亡，同时抑制异种移植肿瘤生长并延长荷瘤小鼠的生存期；DMAMCL可在体外诱导细胞G2/M期阻滞，并在体内外均引发细胞凋亡；敲低BAX的表达可减弱DMAMCL诱导的骨肉瘤细胞死亡；此外本研究还发现，DMAMCL可抑制骨肉瘤细胞的干性。上述结果表明，DMAMCL在骨肉瘤治疗中具备潜在治疗价值，或可成为骨肉瘤新药研发的极具前景的候选药物。