PDF(Pubmed)
Abstract：
Osteosarcoma is a frequent and extremely aggressive type of pediatric cancer. New therapeutic approaches are needed to improve the overall survival of osteosarcoma patients. Our previous results suggest that NMNAT1, a key enzyme in nuclear NAD+ synthesis, facilitates the survival of cisplatin-treated osteosarcoma cells. A high-throughput cytotoxicity screening was performed to identify novel pathways or compounds linked to the cancer-promoting role of NMNAT1. Nine compounds caused higher toxicity in the NMNAT1 KO U2OS cells compared to their wild type counterparts, and actinomycin D (ActD) was the most potent. ActD-treatment of NMNAT1 KO cells increased caspase activity and secondary necrosis. The reduced NAD+ content in NMNAT1 KO cells was further decreased by ActD, which partially inhibited NAD+-dependent enzymes, including the DNA nick sensor enzyme PARP1 and the NAD+-dependent deacetylase SIRT1. Impaired PARP1 activity increased DNA damage in ActD-treated NMNAT1 knockout cells, while SIRT1 impairment increased acetylation of the p53 protein, causing the upregulation of pro-apoptotic proteins (NOXA, BAX). Proliferation was decreased through both PARP- and SIRT-dependent pathways. On the one hand, PARP inhibitors sensitized wild type but not NMNAT1 KO cells to ActD-induced anti-clonogenic effects; on the other hand, over-acetylated p53 induced the expression of the anti-proliferative p21 protein leading to cell cycle arrest. Based on our results, NMNAT1 acts as a survival factor in ActD-treated osteosarcoma cells. By inhibiting both PARP1- and SIRT1-dependent cellular pathways, NMNAT1 inhibition can be a promising new tool in osteosarcoma chemotherapy.
摘要：
骨肉瘤是一种常见且极具侵袭性的儿科癌症。需要新的治疗方法来提高骨肉瘤患者的总体生存率。我们之前的结果表明，NMNAT1是核NAD+合成的关键酶，促进顺铂治疗的骨肉瘤细胞的存活。进行高通量细胞毒性筛选以鉴定与NMNAT1的癌症促进作用相关的新的途径或化合物。与野生型对应物相比，九种化合物在NMNAT1KOU2OS细胞中引起更高的毒性，放线菌素D（ActD）是最有效的。NMNAT1KO细胞的ActD处理增加了caspase活性和继发性坏死。NMNAT1KO细胞中NAD+含量的降低被ActD进一步降低，部分抑制NAD+依赖性酶，包括DNA缺口传感器酶PARP1和NAD+依赖性脱乙酰酶SIRT1。PARP1活性受损增加ActD处理的NMNAT1敲除细胞的DNA损伤，而SIRT1损伤增加了p53蛋白的乙酰化，引起促凋亡蛋白的上调（NOXA，巴克斯）。增殖通过PARP和SIRT依赖性途径降低。一方面,PARP抑制剂使野生型而非NMNAT1KO细胞对ActD诱导的抗克隆作用敏感；另一方面，过乙酰化p53诱导抗增殖p21蛋白的表达，导致细胞周期停滞。根据我们的结果,NMNAT1在ActD处理的骨肉瘤细胞中充当存活因子。通过抑制PARP1-和SIRT1依赖性细胞通路，NMNAT1抑制可能是骨肉瘤化疗中一个有前途的新工具。
