Abstract
Aldo-keto reductase 1C3 (AKR1C3) catalyses the NADPH dependent reduction of carbonyl groups in a number of important steroid and prostanoid molecules. The enzyme is also over-expressed in prostate and breast cancer and its expression is correlated with the aggressiveness of the disease. The steroid products of AKR1C3 catalysis are important in proliferative signalling of hormone-responsive cells, while the prostanoid products promote prostaglandin-dependent proliferative pathways. In these ways, AKR1C3 contributes to tumour development and maintenance, and suggest that inhibition of AKR1C3 activity is an attractive target for the development of new anti-cancer therapies. Non-steroidal anti-inflammatory drugs (NSAIDs) are one well-known class of compounds that inhibits AKR1C3, yet crystal structures have only been determined for this enzyme with flufenamic acid, indomethacin, and closely related analogues bound. While the flufenamic acid and indomethacin structures have been used to design novel inhibitors, they provide only limited coverage of the NSAIDs that inhibit AKR1C3 and that may be used for the development of new AKR1C3 targeted drugs. To understand how other NSAIDs bind to AKR1C3, we have determined ten crystal structures of AKR1C3 complexes that cover three different classes of NSAID, N-phenylanthranilic acids (meclofenamic acid, mefenamic acid), arylpropionic acids (flurbiprofen, ibuprofen, naproxen), and indomethacin analogues (indomethacin, sulindac, zomepirac). The N-phenylanthranilic and arylpropionic acids bind to common sites including the enzyme catalytic centre and a constitutive active site pocket, with the arylpropionic acids probing the constitutive pocket more effectively. By contrast, indomethacin and the indomethacin analogues sulindac and zomepirac, display three distinctly different binding modes that explain their relative inhibition of the AKR1C family members. This new data from ten crystal structures greatly broadens the base of structures available for future structure-guided drug discovery efforts.
Highlights
Aldo-keto reductase 1C3 (AKR1C3; known as prostaglandin F synthase, type 5 17b-hydroxysteroid dehydrogenase, type 2 3a-hydroxysteroid dehydrogenase, and dihydrodiol dehydrogenase X) is a human enzyme that catalyses the reduction of carbonyl groups on both steroids and prostaglandins (Figure 1)
Overall Structure and Active Site Sub-pockets Human AKR1C3 protein was expressed in E. coli, purified by affinity chromatography, and catalytic activity assayed with 9,10phenanthrenequinone (Figure S1 inset) in the presence of NADPH by following the decrease in absorbance at 380 nm resulting from NADPH consumption
The AKR1C3 protein shows its canonical a8b8 barrel core with an active site formed by the loop structure located at the C-terminal end of the barrel (Figure 2A), while one molecule of NADP+ and one Non-steroidal anti-inflammatory drugs (NSAIDs) molecule were clearly described by electron density present in the active site near the catalytic residues Tyr-55 and His-117
Summary
Aldo-keto reductase 1C3 (AKR1C3; known as prostaglandin F synthase, type 5 17b-hydroxysteroid dehydrogenase, type 2 3a-hydroxysteroid dehydrogenase, and dihydrodiol dehydrogenase X) is a human enzyme that catalyses the reduction of carbonyl groups on both steroids and prostaglandins (Figure 1) It converts 4-androstene-3,17-dione to testosterone, estrone to 17bestradiol, and progesterone to 20a-hydroxyprogesterone, changing receptor affinities; testosterone has increased androgen receptor affinity, 17b-estradiol has increased estrogen receptor affinity, and 20a-hydroxyprogesterone has reduced affinity for progesterone receptors [1,2,3]. The prostaglandin substrates PGH2 and PGD2 are structurally unrelated to the steroid hormones and are reduced to products PGF2a and 9a,11b-PGF2 respectively [4,5,6] These products display increased F prostanoid receptor affinity, and enhanced proliferative activity. AKR1C3 metabolises the anti-cancer prodrug PR-104A; this is the first known example of nitroreductase activity by AKR1C3 [18]
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