Dennis S. Yamashita

Identification of 4-(2-(4-amino-1,2,5-oxadiazol-3-yl)-1-ethyl-7-{[(3S)-3-piperidinylmethyl]oxy}-1H-imidazo[4,5-c]pyridin-4-yl)-2-methyl-3-butyn-2-ol (GSK690693), a novel inhibitor of AKT kinase.

Overexpression of AKT has an antiapoptotic effect in many cell types, and expression of dominant negative AKT blocks the ability of a variety of growth factors to promote survival. Therefore, inhibitors of AKT kinase activity might be useful as monotherapy for the treatment of tumors with activated AKT. Herein, we describe our lead optimization studies culminating in the discovery of compound 3g (GSK690693). Compound 3g is a novel ATP competitive, pan-AKT kinase inhibitor with IC 50 values of 2, 13, and 9 nM against AKT1, 2, and 3, respectively. An X-ray cocrystal structure was solved with 3g and the kinase domain of AKT2, confirming that 3g bound in the ATP binding pocket. Compound 3g potently inhibits intracellular AKT activity as measured by the inhibition of the phosphorylation levels of GSK3beta. Intraperitoneal administration of 3g in immunocompromised mice results in the inhibition of GSK3beta phosphorylation and tumor growth in human breast carcinoma (BT474) xenografts.

Structure-activity studies of rapamycin analogs: evidence that the C-7 methoxy group is part of the effector domain and positioned at the FKBP12-FRAP interface

BACKGROUNDnRapamycin is an immunosuppressant natural product, which blocks T-cell mitogenesis and yeast proliferation. In the cytoplasm, rapamycin binds to the immunophilin FKBP12 and the complex of these two molecules binds to a recently discovered protein, FRAP. The rapamycin molecule has two functional domains, defined by their interaction with FKBP12 (binding domain) or with FRAP (effector domain). We previously showed that the allylic methoxy group at C-7 of rapamycin could be replaced by a variety of different substituents. We set out to examine the effects of such substitutions on FKBP12 binding and on biological activity.nnnRESULTSnRapamycin C-7-modified analogs of both R and S configurations were shown to have high affinities for FKBP12, yet these congeners displayed a wide range of potencies in splenocyte and yeast proliferation assays. The X-ray crystal structures of four rapamycin analogs in complexes with FKBP12 were determined and revealed that protein and ligand backbone conformations were essentially the same as those observed for the parent rapamycin-FKBP12 complex and that the C-7 group remained exposed to solvent. We then prepared a rapamycin analog with a photoreactive functionality as part of the C-7 substituent. This compound specifically labeled, in an FKBP12-dependent manner, a protein of approximately 250 kDa, which comigrates with recombinant FRAP.nnnCONCLUSIONSnWe conclude that the C-7 methoxy group of rapamycin is part of the effector domain. In the ternary complex, this group is situated in close proximity to FRAP, at the interface between FRAP and FKBP12.

Design, synthesis and evaluation of dual domain FKBP ligands

Abstract A number of dual domain acyclic and macrocyclic pipecolyl α-ketoamide derivatives were prepared which possess the elements of previously described high-affinity FKBP binding domains as well as simplified mimics of the FK506 effector domain, a critical feature for immunosuppressive activity of the FKBP12-FK506 complex. Compounds of this study exhibited a range of FKBP cis-trans peptidyl-prolyl isomerase inhibitory activities but no activity in a splenocyte mitogenesis assay for immunosuppression.

Synthesis of I‐125 labeled photoaffinity rapamycin analogs

Two no-carrier-added 125 I-labeled photoaffinity rapamycin analogs were prepared : 7-demethoxy-7-(4-azido-3- 125 I-benzyloxy)rapamycin (2) and its C 28 -C 29 seco analog 3. The key reactions of the synthesis were substitution of the C 7 methoxyl of rapamycin (1) with 4-azido-3-tributylstannylbenzyloxy group, exchange of tributyltin with 125 I using Na 125 I and Chloramine-T, and a ZnCl 2 mediated retro-Aldol cleavage of the C 28 -C 29 bond of rapamycin.

Discovery of 5-pyrrolopyridinyl-2-thiophenecarboxamides as potent AKT kinase inhibitors.

A pyrrolopyridinyl thiophene carboxamide 7 was discovered as a tractable starting point for a lead optimization effort in an AKT kinase inhibition program. SAR studies aided by a co-crystal structure of 7 in AKT2 led to the identification of AKT inhibitors with subnanomolar potency. Representative compounds showed antiproliferative activity as well as inhibition of phosphorylation of the downstream target GSK3beta.

Preparative chiral HPLC separation of all possible stereoisomers of LY191704 and LY266111 and their in vitro inhibition of human types 1 and 2 steroid 5α-reductases

Abstract A preparative chiral HPLC separation of each of the four stereoisomers of LY191704 [(±)-1, and (±)-3] and LY266111 [(±)-2, and (±)-4] is reported. All eight compounds have been evaluated in vitro as inhibitors of recombinant human type 1 and type 2 steroid 5α-reductase. The trans enantiomers of LY266111, (+)-2 and (−)-2, show equal and potent inhibition of the type 1 isozyme. The cis enantiomers of LY266111, (+)-4 and (−)-4, and the unsaturated analogue 6 show significantly reduced type 1 inhibitory activity. The cis and trans enantiomeric pairs of LY191704 [(+)-1, (−)-1, (+)-3, and (−)-3] and the unsaturated analog 5 display similar and potent activity against the type 1 isozyme. All compounds display relatively poor activity against the human type 2 isozyme.

3-Carboxy-20-keto steroids are dual uncompetitive inhibitors of human steroid 5α-reductase types 1 and 2

Steroidal 3-carboxy-20-ketones have been prepared within two structural series, the androsta-3,5-dienes and the estra-1,3,5-trienes, as potential inhibitors of types 1 and 2 steroid 5 alpha-reductase, the enzyme activity responsible for the final step in biosynthesis of dihydrotestosterone. These compounds are shown to be potent uncompetitive inhibitors of both human recombinant enzyme activities, defining a novel class of dual steroid 5 alpha-reductase inhibitors.

2,3,5-Trisubstituted pyridines as selective AKT inhibitors. Part II: Improved drug-like properties and kinase selectivity from azaindazoles

A novel series of AKT inhibitors containing 2,3,5-trisubstituted pyridines with novel azaindazoles as hinge binding elements are described. Among these, the 4,7-diazaindazole compound 2c has improved drug-like properties and kinase selectivity than those of indazole 1, and displays greater than 80% inhibition of GSK3beta phosphorylation in a BT474 tumor xenograft model in mice.

Azepanone-based inhibitors of human cathepsin S: optimization of selectivity via the P2 substituent.

A series of azepanone inhibitors of cathepsin S is described. Selectivity over both cathepsin K and cathepsin L was achieved by varying the P2 substituent. Ultimately, a balanced potency and selectivity profile was achieved in compound 39 possessing a 1-methylcyclohexyl alanine at P2 and nicotinamide as the P substituent. The cellular potency of selected analogs is also described.

2,3,5-Trisubstituted pyridines as selective AKT inhibitors-Part I: Substitution at 2-position of the core pyridine for ROCK1 selectivity.

2,3,5-Trisubstituted pyridines have been designed as potent AKT inhibitors that are selective against ROCK1 based on the comparison between AKT and ROCK1 structures. Substitution at the 2-position of the core pyridine is the key element to provide selectivity against ROCK1. An X-ray co-crystal structure of 9p in PKA supports the proposed rationale of ROCK1 selectivity.