Nutlin-3a is the active enantiomer of Nutlin-3, an MDM2 antagonist that inhibits MDM2-p53 interaction (Ki=90 nM) and activates p53. Nutlin-3a binds preferentially to the p53-binding pocket of MDM2, leading to stabilization of p53 and activation of the p53 pathway. Nutlin-3a has antitumor activity.

MDM2-p53 interaction:90 nM

METHODS: N1E-115 cells were treated with Ionomycin calcium (0.2-10 µM) for 3-24 h. Cell viability was determined by trypan blue dye exclusion assay.
RESULTS: Ionomycin calcium induced cell death in a concentration and time dependent manner. [1]
METHODS: Fura-2-loaded mouse cerebellar astrocytes were treated with Ionomycin calcium (0.1-10 µM) for 35 min, and cell membrane Ca(2+) concentration was monitored by changes in the Fura-2 340/380 ratio.
RESULTS: At concentrations ≤1 µM [Ca2+]c slowly declined after the initial peak, reaching significantly lower levels after 35 min. However, at 2 µM of ionomycin the peak level of [Ca2+]c was sustained in the plateau phase, and at concentrations >2 µM ionomycin [Ca2+]c was significantly higher after 35 min than at the initial peak. [2]

METHODS: To investigate the mechanism of demyelination induced in vivo, Ionomycin calcium (2-50 µM, 0.5 µL) was dorsal column injected into SD rats.
RESULTS: Rats injected with Ionomycin calcium showed varying degrees of lesions observed from days 1-21, including areas of localized edema, a small number of scattered demyelinated axons, and larger lesions containing up to 200 demyelinated or degenerated axons. [3]

Biacore studies: Competition assays are performed on a Biacore S51. A Series S Sensor chip CM5 is derivatized for immobilization of a PentaHis antibody for capture of the His-tagged p53. The level of capture is ~ 200 response units (1 response unit corresponds to 1 pg of protein per mm 2). The concentration of MDM2 protein is kept constant at 300 nM. Test compounds are dissolved in DMSO at 10 mM and further diluted to make a concentration series of inhibitor in each MDM2 test sample. The assays are run at 25 °C in running buffer (10 mM Hepes, 0.15 M NaCl, 2% DMSO). MDM2-p53 binding in the presence of inhibitor is calculated as a percentage of binding in the absence of inhibitor and IC50 is calculated using Microsoft Excel

SRB(Only for Reference)