Restoration of p53 activity by inhibition of the p53-MDM2 interaction has been considered an attractive approach for cancer treatment. However, the hydrophobic protein-protein interaction surface represents a significant challenge for the development of small-molecule inhibitors with desirable pharmacological profiles. RG7112 was the first small-molecule p53-MDM2 inhibitor in clinical development. Here, we report the discovery and characterization of a second generation clinical MDM2 inhibitor, RG7388, with superior potency and selectivity.

The cyclin-dependent protein kinases are key regulators of cell cycle progression. Aberrant expression or altered activity of distinct cyclin-dependent kinase (CDK) complexes results in escape of cells from cell cycle control, leading to unrestricted cell proliferation. CDK inhibitors have the potential to induce cell cycle arrest and apoptosis in cancer cells, and identifying small-molecule CDK inhibitors has been a major focus in cancer research. Several CDK inhibitors are entering the clinic, the most recent being selective CDK2 and CDK4 inhibitors. We have identified a diaminopyrimidine compound, R547, which is a potent and selective ATP-competitive CDK inhibitor. In cell-free assays, R547 effectively inhibited CDK1/cyclin B, CDK2/cyclin E, and CDK4/cyclin D1 (K i = 1 -3 nmol/L) and was inactive (K i > 5,000 nmol/L) against a panel of >120 unrelated kinases. In vitro, R547 effectively inhibited the proliferation of tumor cell lines independent of multidrug resistant status, histologic type, retinoblastoma protein, or p53 status, with IC 50 s V 0.60 Mmol/L. The growth-inhibitory activity is characterized by a cell cycle block at G 1 and G 2 phases and induction of apoptosis. R547 reduced phosphorylation of the cellular retinoblastoma protein at specific CDK phosphorylation sites at the same concentrations that induced cell cycle arrest, suggesting a potential pharmacodynamic marker for clinical use. In vivo, R547 showed antitumor activity in all of the models tested to date, including six human tumor xenografts and an orthotopic syngeneic rat model. R547 was efficacious with daily oral dosing as well as with once weekly i.v. dosing in established human tumor models and at the targeted efficacious exposures inhibited phosphorylation of the retinoblastoma protein in the tumors. The selective kinase inhibition profile and the preclinical antitumor activity of R547 suggest that it may be promising for development for use in the treatment of solid tumors. R547 is currently being evaluated in phase I clinical trials.

o-Halosubstituted aromatic triazenes (e.g. I, Scheme 1) react with aryloxides (e.g. II, Scheme 1) in the presence of CuBr´Me 2 S, K 2 CO 3 and pyridine in acetonitrile at reflux to afford biaryl ethers (e.g. V, Scheme 1). This general methodology (Tables 1 and 2) was applied to the construction of the C-O-D and D-O-E vancomycin model systems 37 (Scheme 2) and 50 (Scheme 3), demonstrating its potential in a projected total synthesis of vancomycin (1, Figure 1). For the construction of the vancomycin model AB biaryl ring system, a sequential strategy involving a Suzuki coupling of the C-O-D aryl iodide 74 (Scheme 7) and boronic acid 53 (Scheme 4), followed by macrolactamization was demonstrated, in which the preformed C-O-D ring framework served to preorganize the precursor for cyclization. The latter investigation led to Suzuki-coupling-based asymmetric synthesis of biaryl systems in which 2,2-bis(diphenylphosphino)-1,1'-binaphthyl (BINAP) was found to be the optimum ligand (Tables 3 and 4).

The development of small-molecule MDM2 inhibitors to restore dysfunctional p53 activities represents a novel approach for cancer treatment. In a previous communication, the efforts leading to the identification of a non-imidazoline MDM2 inhibitor, RG7388, was disclosed and revealed the desirable in vitro and in vivo pharmacological properties that this class of pyrrolidinebased inhibitors possesses. Given this richness and the critical need for a wide variety of chemical structures to ensure success in the clinic, research was expanded to evaluate additional derivatives. Here we report two new potent, selective, and orally active p53-MDM2 antagonists, RO5353 and RO2468, as follow-ups with promising potential for clinical development.KEYWORDS: MDM2, p53, wild-type, small molecule, apoptosis, cancer T umor suppressor p53 is a potent transcription factor that is activated in response to cellular stress and regulates downstream genes controlling cell cycle arrest and apoptosis. 1−4 Dysfunction of the p53 pathway is the most frequent alteration observed in human cancers. 5 MDM2 is the primary negative regulator of p53 through binding to its transactivation domain and promoting proteosomal degradation. 6−8 In tumor cells with wild-type p53 (∼50%), reactivation of the p53 pathway by inhibition of MDM2 with small molecules has been considered as potentially an attractive novel therapeutic approach for cancer treatment. 9,10 Currently, several smallmolecule MDM2 inhibitors including RG7112 and RG7388 (Figure 1) are undergoing clinical evaluations. 11−14 To maximize the chance of success in the clinic and derisk any potential idiopathic toxicity associated with specific chemotypes, continued research efforts are required to expand chemodiversity and identify potent and selective MDM2 antagonists with desirable in vitro ADMET and in vivo pharmacokinetic properties. Here we report the discovery of RO5353 and RO2468, two new highly potent and selective MDM2 inhibitors with potential for clinical development.Our exploration initially led to the identification of a potent and selective MDM2 inhibitor RO8994 (Figure 1), which was found to be highly efficacious against established human tumor xenografts in nude mouse models. 15 Two key structural elements of RG7388 were preserved in RO8994. First, it was established that the stereochemical configuration of the pyrrolidine core structure in which the two aryl rings ("A" and "B") adopt a "Trans" orientation was very important for optimal binding to MDM2. 14 The architecture of spiroindolinone-3,3′-pyrrolidine series (as exemplified by MI-219) was first reported by Ding et al. 16−18 Consistent with our findings, this group recently published their latest findings in which the original stereochemistry was found to be unstable and