Abstract
Nucleus accumbens-associated protein-1 (NAC1) is a transcriptional repressor encoded by the NACC1 gene, which is amplified and overexpressed in various human cancers and plays critical roles in tumor development, progression, and drug resistance. NAC1 has therefore been explored as a potential therapeutic target for managing malignant tumors. However, effective approaches for effective targeting of this nuclear protein remain elusive. In this study, we identified a core unit consisting of Met7 and Leu90 in NAC1's N-terminal domain (amino acids 1-130), which is critical for its homodimerization and stability. Furthermore, using a combination of computational analysis of the NAC1 dimerization interface and high-throughput screening (HTS) for small molecules that inhibit NAC1 homodimerization, we identified a compound (NIC3) that selectively binds to the conserved Leu-90 of NAC1 and prevents its homodimerization, leading to proteasomal NAC1 degradation. Moreover, we demonstrate that NIC3-mediated down-regulation of NAC1 protein sensitizes drug-resistant tumor cells to conventional chemotherapy and enhances the antimetastatic effect of the antiangiogenic agent bevacizumab both in vitro and in vivo These results suggest that small-molecule inhibitors of NAC1 homodimerization may effectively sensitize cancer cells to some anticancer agents and that NAC1 homodimerization could be further explored as a potential therapeutic target in the development of antineoplastic agents.
Highlights
Nucleus accumbens–associated protein-1 (NAC1) is a transcriptional repressor encoded by the NACC1 gene, which is amplified and overexpressed in various human cancers and plays critical roles in tumor development, progression, and drug resistance
We found that the biotin-tagged positive probe possessed the ability to effectively inhibit NAC1 expression, whereas the biotin-tagged negative probe did not have this ability at the same concentration (Fig. 3E), and reduction of NAC1 protein caused by PP could be rescued by the proteasome inhibitor MG132 (Fig. 3F)
We show that this type of homodimerization is important for the stability of NAC1 protein (Figs. 1 and S2C)
Summary
The conserved BTB/POZ domain is essential for NAC1 dimerization, which plays important roles in tumor development [11]; no evidence has been provided about the mechanisms and biologic consequences of NAC1 dimerization. We observed that combination treatment of NIC3 with Adriamycin or cisplatin significantly augmented the cytotoxic effect in the resistant cancer cell lines (Fig. 6, A–D). As compared with either vehicle control, NIC3, or Adriamycin alone, the combination treatment of NIC3 with Adriamycin caused a significant increase in the terminal deoxynucleotidyltransferase dUTP nick end labeling (TUNEL) staining–positive cells and a decrease in Ki67, a proliferation marker (Fig. 6H), and in NAC1 protein expression No significant changes in body weight were observed in the mice treated with the combination therapy (Fig. 6G) These results suggest that NIC3 is synergistic with cisplatin or Adriamycin in suppressing cancer cell growth, which is associated with induction of apoptotic cell death in drug-resistant tumor cells both in vitro and in vivo. These observations suggest that NIC3 could be used as a sensitizer in antimetastatic therapy
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