TAS4464, A Highly Potent and Selective Inhibitor of NEDD8-Activating Enzyme, Suppresses Neddylation and Shows Antitumor Activity in Diverse Cancer Models

Abstract

NEDD8-activating enzyme (NAE) is an essential E1 enzyme of the NEDD8 conjugation (neddylation) pathway, which controls cancer cell growth and survival through activation of cullin-RING ubiquitin ligase complexes (CRL). In this study, we describe the preclinical profile of a novel, highly potent, and selective NAE inhibitor, TAS4464. TAS4464 selectively inhibited NAE relative to the other E1s UAE and SAE. TAS4464 treatment inhibited cullin neddylation and subsequently induced the accumulation of CRL substrates such as CDT1, p27, and phosphorylated IkBa in human cancer cell lines. TAS4464 showed greater inhibitory effects than those of the known NAE inhibitor MLN4924 both in enzyme assay and in cells. Cytotoxicity profiling revealed that TAS4464 is highly potent with wide-spread antiproliferative activity not only for cancer cell lines, but also patient-derived tumor cells. TAS4464 showed prolonged target inhibition in human tumor xenograft mouse models; weekly or twice a week TAS4464 administration led to prominent antitumor activity in multiple human tumor xenograft mouse models including both hematologic and solid tumors without marked weight loss. As a conclusion, TAS4464 is the most potent and highly selective NAE inhibitor reported to date, showing superior antitumor activity with prolonged target inhibition. It is, therefore, a promising agent for the treatment of a variety of tumors including both hematologic and solid tumors. These results support the clinical evaluation of TAS4464 in hematologic and solid tumors.

Activity of TAS4464, a novel NEDD8 activating enzyme E1 inhibitor, against multiple myeloma via inactivation of nuclear factor κB pathways

“…We have reported that TAS4464 inhibits the neddylation pathway via inhibition of NAE, which leads to accumulation of CRL substrates in several solid and hematologic tumor cell lines . In the present study, TAS4464 also reduced the levels of neddylated UBC12 (an E2 for NEDD8) and neddylated cullin1 (a component of CRL) and selectively induced the accumulation of the CRL substrates p‐IκBα, CDT1, NRF2 and p21, but not substrates of non–CRL E3s (GADD34 and PTTG), in MM.1S cells in a dose‐dependent and time‐dependent manner (Figure ).…”

“…TAS4464 is one of the most selective and potent NAE inhibitors reported to date both in vitro and in vivo . TAS4464 exhibits widespread antiproliferative activity in various tumor cell lines and is especially active against most hematologic malignancy cell lines, including myeloma‐derived cell lines …”

Dynamics of replication origin over-activation

“…The interaction of CDT1 with geminin also serves to prevent re-replication during normal mitotic growth 12,15,17,45,47 . Interfering with the degradation and inactivation of CDT1, either by geminin depletion 47 or by inhibiting the activity of cullinanchored ubiquitin ligases 28,31,33,46 , can be exploited to trigger re-initiation and subsequent selective killing of cancer cells. We observed that cancer cells that undergo partial genome duplication due to the persistence of pre-RCs on chromatin exhibit similar DNA synthesis characteristics regardless of the mechanisms that prevent pre-RC dissociation.…”

“…However, excess initiation of DNA replication can have deleterious consequences, including oncogenic transformation of normal cells and increased genomic instability in cancer 28,44,45,47 , consistent with our observations suggesting that massive overreplication can lead to senescence. Those consequences can be exploited therapeutically to induce selective killing of cancer cells 28,[31][32][33]47 . The results reported here imply that circumventing the strong inhibitory interactions that normally prevent excess DNA synthesis can occur via at least two pathways, each activating a distinct set of replication origins.…”

“…Partial genome re-replication can also be caused by inhibition of DOT1L, a methyltransferase which catalyzes the demethylation of histone H3 on lysine 79 (H3K79Me2), thereby removing a histone modification typically associated with a group of replication origins 27 . Because massive re-replication can drive cell death specifically in checkpointcompromised cancer cells, both CDT1 stabilization by inactivation of ubiquitin-mediated degradation and inhibition of DOT1L are currently being explored as novel anti-cancer therapeutic strategies [28][29][30][31][32][33] .Given that genome re-replication is a common avenue to genomic instability and considering its potential as a strategy for chemotherapy, it is important to understand in detail its mechanics and downstream consequences. Here, we report that re-replication exhibits aberrant replication fork dynamics and occurs more slowly than the routine genome duplication that takes place during normal growth.…”