Hsp90 is a molecular chaperone with important functions in regulating the function of several proteins with potential pathogenic activity. to inhibit Hsp90. Even before Hsp90 was confirmed as its target it was known to possess potent tumoricidal activity and [17]. However it suffers from a number of drawbacks which have prevented its clinical development including limited aqueous solubility and dose Nepafenac limiting hepatotoxicity. The latter is thought to stem from GM’s benzoquinone moiety which has significant Michael acceptor activity. Analogs with reduced electrophilicity have been developed including 17-AAG (2) and 17-DMAG (3) and these have demonstrated proof of concept for Hsp90 inhibition. 17-AAG (2) was the first Hsp90 inhibitor to enter clinical studies and has shown promising results in HER2-overexpressing tumors [18]. A number of drawbacks including difficulty to formulate cost of manufacture and the difficulty to administer pharmacologically relevant doses without toxicity has limited its development in other cancers. 17-DMAG (3) has similar an activity to 17-AAG (2) but is usually water soluble. This agent as well as a reduced form of 17-AAG (2) IPI-504 (4) [19] have also entered clinical trials. Because of the limitations of GM-based inhibitors novel inhibitors of Hsp90 with more drug-like properties were actively sought. Structure-based design high throughput screening fragment-based design and virtual testing have all been utilized to identify small molecules that bind to the N-terminal ATP pocket of Hsp90. These efforts have identified a number of unique chemotypes including purine (i.e. 16 and 26) isoxazole (i.e. 5) and 6 7 (i.e. 6) as potent Nepafenac and selective Hsp90 inhibitors which have already or will soon enter into clinical trials [20-23]. The remainder of this evaluate will focus on the purine class of inhibitors with special emphasis on their discovery and development into clinical brokers for the treatment of cancer but will also IKK2 touch upon their potential usefulness in neuro degenerative diseases. PURINE-SCAFFOLD HSP90 INHIBITORS 1 Discovery of PU3 The first identified synthetic Hsp90 inhibitor was based on the purine (PU)-scaffold [24]. The unique structural features of the N-terminal nucleotide pocket as well as the shape adopted by ATP when Hsp90-bound were used to rationally design a molecule to fit into Nepafenac this pocket. The initial lead molecule PU3 (7 Fig. 2) bound to purified Hsp90 with an EC50 = 15-20 μM (1 μM for 17-AAG) and exhibited phenotypic effects in breast malignancy cells much like those observed for GM (1). In MCF-7 and SKBr3 breast malignancy cells 7 caused the degradation of HER2 HER3 Raf-1 and estrogen receptor (ER) onco-proteins at a concentration as low as 10 μM to 50 μM. In a typical feed-back heat shock response due to Hsp90 inhibition it induced the synthesis of Hsp90 and Hsp70 in these cells. 7 also exhibited anti- proliferative effects against genetically unique breast malignancy cells (i.e. MCF-7 ER+; SKBr3 HER2+; MDA-MB-468 ER- and HER2-) at low micromolar concentrations (≤ 50 Nepafenac μM) and caused G1 cell cycle arrest. G1-block was followed by morphological and functional differentiation. Fig. (2) Structure of PU3 (7) and initial SAR of methylene linker series leading to PU24FCl (8). The co-crystal structure of 7 bound to human N-terminal Hsp90u (observe 1UY6.pdb) suggested that this purine ring binds in the same position as that of ADP with the C6-NH2 making a key conversation with Asp93 [25]. There is also a network of hydrogen bonds between N1 N7 and C6-NH2 of 7 with Asn51 Nepafenac Ser52 Thr284 and Gly97 through three water molecules. The phenyl ring of 7 is usually stacked between the side chains of Phe138 and Leu107 and makes additional hydrophobic interactions with Met98 and Leu103. The methoxy groups make hydrophobic contacts with the aromatic rings of Trp162 and Tyr139 as well as with the aliphatic carbons of Ala111 and Val150. The first and second methylene groups of the Nepafenac N9-butyl chain provide additional hydrophobic contacts with Leu107 and Met98. The discovery of PU3 (7) as an Hsp90 inhibitor served to initiate medicinal chemistry efforts round the PU-scaffold in an effort to improve both potency and physical/ chemical properties. Major efforts have focused on probing the structure-activity relationship (SAR) of the aromatic moiety to the purine at C8-position the nature of the linker between the PU-scaffold and the substituted aromatic ring and the alkyl chain at N9 position. Substitutions to the 2-position of the adenine ring have also been.