Initial testing of the investigational NEDD8-activating enzyme inhibitor MLN4924 by the pediatric preclinical testing program

Abstract

Background: MLN4924 is an investigational first-in-class small molecule inhibitor of NEDD8-activating enzyme (NAE). NAE is an essential component of the NEDD8 conjugation pathway, controlling the activity of a subset of ubiquitin-proteasome system (UPS) E3 ligases, multiprotein complexes that transfer ubiquitin molecules to substrate proteins.

Procedures: MLN4924 was tested against the PPTP in vitro panel using 96-hour exposure time at concentrations ranging from 1.0 nM to 10 µM. It was tested in vivo at a dose of 100 mg/kg [66 mg/kg for the acute lymphoblastic leukemia (ALL) xenografts] administered orally twice daily × 5 days. Treatment duration was 3 weeks.

Results: The median relative IC(50) for MLN4924 against the PPTP cell lines was 143 nM, (range: 15-678 nM) with that for the Ewing panel being significantly lower (31 nM). MLN4924 induced significant differences in EFS distribution compared to control in 20 of 34 (59%) evaluable solid tumor xenografts. MLN4924 induced intermediate activity (EFS T/C values >2) in 9 of the 33 evaluable xenografts (27%), including 4 of 4 glioblastoma xenografts, 2 of 3 Wilm's tumor xenografts, 2 of 5 rhabdomyosarcoma xenografts, and 1 of 4 neuroblastoma xenografts. For the ALL panel, 5 of 8 evaluable xenografts showed intermediate activity for the EFS T/C measure. MLN4924 did not induce objective responses in the PPTP solid tumor or ALL panels.

Conclusions: MLN4924 showed potent activity in vitro and in vivo showed tumor growth inhibitory activity against a subset of the PPTP solid tumor and ALL xenografts.

Figure 1

MLN4924 in vivo objective response activity, left: The colored heat map depicts group response scores. A high level of activity is indicated by a score of 6 or more, intermediate activity by a score of > 2 but < 6, and low activity by a score of < 2. Right: representation of tumor sensitivity based on the difference of individual tumor lines from the midpoint response (stable disease). Bars to the right of the median represent lines that are more sensitive, and to the left are tumor models that are less sensitive. Red bars indicate lines with a significant difference in EFS distribution between treatment and control groups, while blue bars indicate lines for which the EFS distributions were not significantly different.

Figure 2

MLN4924 activity against individual solid tumor xenografts, Kaplan-Meier curves for EFS, median relative tumor volume graphs, and individual tumor volume graphs (controls, gray and treated, black lines) are shown for selected lines: Wilms tumor KT-10, glioblastomas BT-39 and GBM2, and neuroblastoma NB-1771.

Figure 3

MLN4924 activity against individual ALL xenografts, Kaplan-Meier curves for EFS and graphs of median and individual percentages of hCD45 cells (controls, gray and treated, black lines), are shown for selected lines: (A) ALL-2, and (B) ALL-3.

Figure 4

MLN4924 decreases total neddylated cullin proteins in vivo. Mice were treated with a single administration of MLN4924 (100 mg/kg S.C.), or compound vehicle (Veh). Tumors were excised from 0.5 to 24 hr after MLN4924 administration. A. Total neddylated cullin proteins; B. Relative levels of neddylated cullin proteins normalized to α-tubulin.