Abstract 2639: Discovery of novel small molecule inhibitors of Hsp90 for oral treatment
Authors: Sirrenberg Christian , Eggenweiler Hans-Michael , Buchstaller Hans-Peter , Wolf Michael , Grädler Ulrich , Musil Djordje , Hoppe Edmund , Zimmermann Astrid , Amendt Christiane , Schwartz Harry , März Joachim , Bomke Jörg , Ansgar Wegener , Gleitz Johannes 
Published: February 2011

Abstract
Hsp90 is a molecular chaperone that plays an important role in the folding and function of client protein substrates in the cell. Many of these client proteins are well known oncogenes that are often over-expressed and mutated in tumor cells and association of Hsp90 with these client proteins maintains their ability to function in the deregulated state. Therefore Hsp90 inhibitors target tumor growth by multiple parallel mechanisms involving several oncogenes, which are critically involved in cell cycle, tumor growth, angiogenesis and apoptosis.
Here we will present HTS and structure based design efforts to identify novel chemotypes that show potent binding to HSP90. In particular a 5-methyl-1H-pyrazol-3-yl-benzene-1,3-diol derivative (1) was identified during an HTS as a potent starting point for further optimization. This initial hit already showed an IC50 of 240nM in a HSP90 binding assay. The HSP90 co-crystal structure of this initial HTS hit revealed its binding mode to the N-terminal ATPase pocket and thus guided subsequent optimization of this structural series towards potency and oral bioavailability. Especially the identification of a new binding pocket for the ethyl group in (1) opened the route for the introduction of a broad variety of substituents. In addition to these variations the pyrazole core in (1) was replaced by a triazolone system. Although the SAR with respect to in vitro potency is comparable for both series, this fine tuning led to compounds with improved bioavailability and in vivo potency.
The most promising candidates from both series were screened for selectivity against other ATPases, kinases and ion channels where they turned out to be completely HSP90 selective. In vivo tumor xenografts using prostate cancer cell line PC3, colon carcinoma HCT116 (KRAS mutation: mechanism of resistance against EGFR inhibitor in mCRC) and NSCLC tumor H1975 (EGFR-T790M mutation: mechanism of resistance against EGFR TKI) demonstrated excellent anti-tumor activity under different schedules after oral and iv administration. Biomarker analysis demonstrated efficient dose and time-dependent down-regulation of EGFR, phospho-Erk, cyclin D1 and up-regulation of Hsp70 as well as induction of apoptosis as determined by the increase of cleaved PARP and cleaved caspase 3. Favorably PK/PD studies revealed extraordinary long lasting compound levels in tumor tissue and long lasting pharmacodynamic effects (for more than 5 days in the tumors), indicating prolonged Hsp90 inhibition in tumor tissue in contrast to healthy tissue, from which the triazolones were rapidly cleared. The overall profile of the triazolones including synthesis, structure activity relationships, X-ray structures as well as a comprehensive pharmacological in vitro and in vivo characterization, together with potential pharmacodynamic and predictive biomarkers will be presented.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2639.