PF04929113 (SNX-5422) is a synthetic, novel, small molecule Hsp90 Inhibitor. As an oral formulation that demonstrates strong efficacy and tolerability, PF04929113 is positioned as a breakthrough therapy with broad applicability across a wide range of cancers.

HER2:37 nM, HSP90:41 nM

PF-04929113 showed potent effects on Her-2 stability and led to the expected upregulation of Hsp70.PF-04929113 showed potent antiproliferative activity against a wide range of cancer cell types, e.g., MCF-7 (IC50 = 16 nM), SW620 (IC50 = 19 nM), K562 (IC50 = 23 nM), SK- MEL-5 (IC50 = 25 nM) and A375 (IC50 = 51 nM).

PF-04929113 showed potent effects on Her-2 stability and led to the expected upregulation of Hsp70.PF-04929113 showed potent antiproliferative activity against a wide range of cancer cell types, e.g., MCF-7 (IC50 = 16 nM), SW620 (IC50 = 19 nM), K562 (IC50 = 23 nM), SK- MEL-5 (IC50 = 25 nM) and A375 (IC50 = 51 nM).

The Hsp90 loaded media is challenged with PF-04929113 at a given concentration, ranging from 0.8 to 500 μM, and the amount of Hsp90 liberated at each concentration is determined by Bradford protein assay. The resulting IC50 values are corrected for the ATP ligand concentration and presented as apparent Kd values.

Cell lines: MCF-7,SW620,K562,SK-MEL-5 and A375 cancer cell lines. Concentrations: 0-300 nM. Method: Proliferation rates are measured by seeding cells into 96-well plates,followed by compound addition 24 h later.After addition of PF-04929113,cells are allowed to grow for either an additional 72 or 144 h depending on the rate of growth.At harvest,media is removed and DNA content for individual wells is determined using CyQuant DNA dye.Levels of Hsp90 client proteins and phosphor-regulated proteins in A375 are measured by high content analysis (HCA) using an ArrayScan 4.5 instrument after 24 hours of treatment with PF-04929113,followed by methanol fixation.After fixation in 4% PBS-buffered formalin and permeablization with 0.1% TX-100,cells are probed with anti-Her2,antiphospho-S6 (pS6),antipERK,and anti-Hsp70 primary antibodies,followed by TRITC or FITC conjugated secondary antibodies.Nuclei are also stained with Hoechst DNA binding dye.For each well,250-500 individual nuclei are identified along with the average staining intensity for the client and phospho-proteins for each cell.Average client staining intensities are then calculated for each well.

Animal Models: 5 ×106 MM.1S cells are inoculated subcutaneously in the Fox Chase SCID mice (6-7 weeks old). Formulation: PF-04929113 is dissolved in 1% carboxy methylcellulose/0.5% Tween 80 at 10 mg/mL and stored at 4 °C. Dosages: 20 or 40 mg/kg. Administration: orally 3 times per week,3 weeks in total.