However, research and development to provide safe, stable, efficacious, and patient compliant formulation plays a vital role in bringing peptide therapeutics to market. Recent advances in the field of peptide therapeutics have led to the design and synthesis of many promising peptides. In contrast, for RG7112, only modest induction of p21 was observed at all doses and time points. Induction of p21 was observed for tumors in response to ATSP-7041 treatment, with a robust 12-to 19-fold increase in p21 8 h post dose.

Tumors were removed from mice 4, 8, or 24 h after the last dose of either vehicle or ATSP-7041 at 20 or 30 mg/kg, and p21 mRNA levels were determined by quantitative RT-PCR. (C) ATSP-7041 induced greater p21 expression than the MDM2 selective small-molecule inhibitor RG7112.

xenografts received vehicle or 20 or 30 mg/kg ATSP-7041 solution i.v., qod, or 50 or 100 mg/kg of a p.o. Mice (n = 10 per group) bearing established s.c. (B) Treatments resulted in dose-dependent and statistically significant tumor growth inhibition against the MDMX overexpressing MCF-7 human breast cancer xenograft model. Tumor volumes were calipered throughout the study, and data were plotted as mean ± SEM. (A) ATSP-7041 induced statistically significant tumor growth inhibition for both qd and qod dosing schedules against the MDM2 amplified osteosarcoma xenograft model SJSA-1. The side chain is coloredĪTSP-7041 suppresses tumor growth in vivo of multiple human xenograft models and increases expression of p21. ( C ) A high-resolution X-ray structure of ATSP-7041 bound to MDMX delin- eates its intermolecular contacts with the target protein. ( iii ) Other view of the solvent exposed surfaces, after vertical rotation of 90°. ( ii ) Solvent exposed surfaces representation of the three stapled peptides with the same view angle as in ref. Polar residues (Asn, Gln, Ser, and Thr) are colored orange green is for hydrophobic, ole fi n staple, and aromatic residues (Ala, Val, Leu, Ile, Pro, Cba, Phe, Tyr, and Trp), blue for basic residues (Lys and Arg), red for acidic residues (Asp and Glu), and light blue for His. Side chains are represented in stick style whereas the ole fi n staple is in ball and stick style. ( i ) The three stapled peptides are represented as fully α -helical peptides with the viewer facing the long axis of the helix by the N terminus. ( B ) The α − helical wheel projections (3D) of ATSP-7041 versus SAH-p53-8 and the stapled pDI analog ATSP-3900 show the extended hydrophobic surface contributing to the enhance amphipathicity of ATSP-7041. ( A ) Comparative CD spectroscopy of pDI and stapled peptide ATSP-7041in pH 7 buffer illustrates their intrinsic α − helical properties. Overall, ATSP-7041 demonstrates in vitro and in vivo proof-of-concept that stapled peptides can be developed as therapeutically relevant inhibitors of protein-protein interaction and may offer a viable modality for cancer therapy.Ĭonformational and 3D-structural properties of ATSP-7041. Most importantly, ATSP-7041 demonstrates robust p53-dependent tumor growth suppression in MDM2/MDMX-overexpressing xenograft cancer models, with a high correlation to on-target pharmacodynamic activity, and possesses favorable pharmacokinetic and tissue distribution properties. A high resolution (1.7-Å) X-ray crystal structure reveals its molecular interactions with the target protein MDMX, including multiple contacts with key amino acids as well as a role for the hydrocarbon staple itself in target engagement. Specifically, ATSP-7041 binds both MDM2 and MDMX with nanomolar affinities, shows submicromolar cellular activities in cancer cell lines in the presence of serum, and demonstrates highly specific, on-target mechanism of action. Here, we report a potent and selective dual inhibitor of MDM2 and MDMX, ATSP-7041, which effectively activates the p53 pathway in tumors in vitro and in vivo. Stapled α-helical peptides have emerged as a promising new modality for a wide range of therapeutic targets.

Stapled helix