pevonedistat and Osteosarcoma

Neddylation inhibitor Pevonedistat (MLN4924) is a novel anticancer drug and has demonstrated broad-spectrum anticancer activity. Nevertheless, we found that Pevonedistat had only a modest apoptotic effect in osteosarcoma (OS) cells. Moreover, we noted that inhibition of neddylation by Pevonedistat led to accumulation of Mcl-1 protein in OS cells. Because Mcl-1 is an important anti-apoptotic protein and also because apoptosis has been shown to be a major cell death pathway, we hypothesized that Mcl-1 accumulation negatively impacted Pevonedistat-mediated anticancer activity in OS cells. In this regard, we employed genetic or pharmacological approaches to inhibit Mcl-1 expression and to examine the effect on Pevonedistat-induced apoptosis in OS cells. We found that inhibition of Mcl-1 expression by siRNA considerably enhanced Pevonedistat-triggered the activation of caspase-3, PARP cleavage and apoptosis, and also dramatically promoted the ability of Pevonedistat to inhibit colony formation of OS cells. Moreover, we observed that flavopiridol, a FDA approved drug, inhibited Mcl-1 expression and substantially enhanced Pevonedistat-mediated activation of apoptosis signaling and significantly augmented cell killing effect in OS cells. Altogether, our study shows that Mcl-1 is a critical resistance factor to Pevonedistat monotherapy, and suggests that Pevonedistat in combinations with flavopiridol may achieve better anticancer therapy.

Topics: Antineoplastic Agents; Apoptosis; Caspase 3; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cyclopentanes; Humans; Myeloid Cell Leukemia Sequence 1 Protein; Osteosarcoma; Proto-Oncogene Proteins c-bcl-2; Pyrimidines

Neddylation is a post-translational protein modification process associated with carcinogenesis and cancer development. MLN4924, a pharmaceutical neddylation inhibitor, induces potent anti-cancer effects in multiple types of cancers. In this study, we investigated the effects of MLN4924 on human osteosarcoma (OS). Levels of both NEDD8 activating enzyme E1 (NAE1) and ubiquitin-conjugating enzyme E2M (Ube2M), two critical components of the neddylation pathway, were much higher in OS tissues and cells than in normal osseous tissues and cells. MLN4924 treatment led to DNA damage, reduced cell viability, senescence and apoptosis in OS cells. Moreover, MLN4924 inhibited OS xenograft tumor growth in mice. Mechanistically, MLN4924 blocked the neddylation of cullins and induced accumulation of several tumor-suppressive substrates of Cullin-RING E3 ubiquitin ligases (CRLs), including CDT1, Wee1, p21, p27, Noxa, and p16. These results suggest clinical studies investigating the utility of MLN4924 for the treatment of OS are warranted.

Topics: Animals; Apoptosis; Bone Neoplasms; Cellular Senescence; Cullin Proteins; Cyclopentanes; DNA Damage; G2 Phase Cell Cycle Checkpoints; Humans; Mice; Osteosarcoma; Protein Processing, Post-Translational; Pyrimidines; Ubiquitin-Activating Enzymes; Xenograft Model Antitumor Assays

The anticancer small molecule MLN4924, a Nedd8-activating enzyme (NAE) inhibitor, triggers cell-cycle arrest, apoptosis, and senescence in cancer cells. In this study, we demonstrate that MLN4924 suppresses osteosarcoma cell proliferation by inducing G2/M cell cycle arrest and apoptosis. Our results indicate that MLN4924 stabilizes the retinoid orphan nuclear receptor alpha (RORα) by decreasing its ubiquitination. RNA interference of RORα attenuates the anti-proliferative effect of MLN4924 in U2OS osteosarcoma cells. MLN4924 up-regulates the expression of p21 and Bmal1, two transcriptional targets of RORα. However, p21 plays a minimal role in the anti-proliferative effect of MLN4924 in U2OS osteosarcoma cells. In contrast, Bmal1 suppression by siRNA attenuates the anti-proliferative effect of MLN4924 in U2OS osteosarcoma cells, indicating that the MLN4924-mediated cell growth inhibition is mediated by Bmal1. These results show MLN4924 to be a promising therapeutic agent for the treatment of osteosarcoma and suggest that MLN4924-induced tumor growth inhibition is mediated by the circadian clock components RORα and Bmal1.

Topics: Animals; Apoptosis; ARNTL Transcription Factors; Biomarkers, Tumor; Bone Neoplasms; Cell Proliferation; Circadian Clocks; Cyclopentanes; Enzyme Inhibitors; Humans; Mice; Mice, Nude; Nuclear Receptor Subfamily 1, Group F, Member 1; Osteosarcoma; Pyrimidines; Tumor Cells, Cultured; Xenograft Model Antitumor Assays