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Trevor M. Penning - One of the best experts on this subject based on the ideXlab platform.

  • potent and highly selective aldo keto reductase 1c3 akr1c3 inhibitors act as chemotherapeutic potentiators in acute myeloid leukemia and t cell acute lymphoblastic leukemia
    Journal of Medicinal Chemistry, 2019
    Co-Authors: Kshitij Verma, Tianzhu Zang, Trevor M. Penning, Paul C Trippier
    Abstract:

    Aldo–keto reductase 1C3 (AKR1C3) catalyzes the synthesis of 9α,11β-prostaglandin (PG) F2α and PGF2α prostanoids that sustain the growth of myeloid precursors in the bone marrow. The enzyme is overexpressed in acute myeloid leukemia (AML) and T-cell acute lymphoblastic leukemia (T-ALL). Moreover, AKR1C3 confers chemotherapeutic resistance to the anthracyclines: first-line agents for the treatment of leukemias. The highly homologous isoforms AKR1C1 and AKR1C2 inactivate 5α-dihydrotestosterone, and their inhibition would be undesirable. We report herein the identification of AKR1C3 inhibitors that demonstrate exquisite isoform selectivity for AKR1C3 over the other closely related isoforms to the order of >2800-fold. Biological evaluation of our isoform-selective inhibitors revealed a high degree of synergistic drug action in combination with the clinical leukemia therapeutics daunorubicin and cytarabine in in vitro cellular models of AML and primary patient-derived T-ALL cells. Our developed compounds exhibi...

  • Role of Human Aldo-Keto Reductases in the Metabolic Activation of the Carcinogenic Air Pollutant 3-Nitrobenzanthrone.
    Chemical research in toxicology, 2018
    Co-Authors: Jessica R. Murray, Ian A. Blair, Volker M. Arlt, Clementina Mesaros, Albrecht Seidel, Trevor M. Penning
    Abstract:

    3-Nitrobenzanthrone (3-NBA) is a potent mutagen and suspected human carcinogen detected in diesel exhaust particulate and ambient air pollution. It requires metabolic activation via nitroreduction to promote DNA adduct formation and tumorigenesis. NAD(P)H:quinone oxidoreductase 1 (NQO1) has been previously implicated as the major nitroreductase responsible for 3-NBA activation, but it has recently been reported that human aldo-keto reductase 1C3 (AKR1C3) displays nitroreductase activity toward the chemotherapeutic agent PR-104A. We sought to determine whether AKR1C isoforms could display nitroreductase activity toward other nitrated compounds and bioactivate 3-NBA. Using discontinuous enzymatic assays monitored by UV-HPLC, we determined that AKR1C1-1C3 catalyze three successive two-electron nitroreductions toward 3-NBA to form the reduced product 3-aminobenzanthrone (3-ABA). Evidence of the nitroso- and hydroxylamino- intermediates were obtained by UPLC-HRMS. Km, kcat, and kcat/ Km values were determined for recombinant AKR1C and NQO1 and compared. We found that AKR1C1, AKR1C3, and NQO1 have very similar apparent catalytic efficiencies (8 vs 7 min-1 mM-1) despite the higher kcat of NQO1 (0.058 vs 0.012 min-1). AKR1C1-1C3 possess a Km much lower than that of NQO1, which suggests that they may be more important than NQO1 at the low concentrations of 3-NBA to which humans are exposed. Given that inhalation represents the primary source of 3-NBA exposure, we chose to evaluate the relative importance of AKR1C1-1C3 and NQO1 in human lung epithelial cell lines. Our data suggest that the combined activities of AKR1C1-1C3 and NQO1 contribute equally to the reduction of 3-NBA in A549 and HBEC3-KT cell lines and together represent approximately 50% of the intracellular nitroreductase activity toward 3-NBA. These findings have significant implications for the metabolism of nitrated polycyclic aromatic hydrocarbons and suggest that the hitherto unrecognized nitroreductase activity of AKR1C enzymes should be further investigated.

  • Abstract 4580A: Metabolic activation of 3-nitrobenzanthrone by human aldo-keto reductases (AKR1C1-AKR1C4)
    Cancer Chemistry, 2015
    Co-Authors: Jessica R. Murray, Tianzhu Zang, Meng Huang, Volker M. Arlt, Heinz H. Schmeiser, Trevor M. Penning
    Abstract:

    In 2012, the International Agency for Research on Cancer (IARC) classified diesel exhaust as a Group 1 carcinogen due to sufficient evidence that exposure is associated with increased risk for lung cancer in humans. However, only a subset of individuals exposed to diesel exhaust develops cancer, indicating the need to identify the genes involved in metabolic activation of these compounds and their genetic variants. Nitro-polycyclic aromatic hydrocarbons (NO2-PAH) are a major component of diesel exhaust and require metabolic activation to exert their carcinogenic activity. A representative NO2-PAH, 3-nitrobenzanthrone (3-NBA), is metabolically activated to 3-aminobenzanthrone (3-ABA) via a 6-electron nitroreduction catalyzed by NQO1 and POR. The reaction leads to the formation of 3-aminobenzanthrone (3-ABA) derived DNA adducts which promote G to T transversions. Building upon previous data that shows human aldo-keto reductase 1C3 (AKR1C3) contains nitroreductase activity towards chemotherapeutic agents (Guise, C.P., Abbattista, M.R., et al., Cancer Res, 70(4), 2010), we chose to examine the nitroreductase activity of AKR1C1-AKR1C4 towards NO2-PAH. We have demonstrated here for the first time that AKR1C enzymes catalyze the nitroreduction of 3-NBA to 3-ABA. We monitored reactions with reverse phase HPLC coupled to in-line photo-diode-array detection (PDA) and fluorescence detection (FLD) to quantify 3-NBA and 3-ABA levels. Fluorescence and UV spectroscopy were used to validate the identity of the compounds. This method was adapted for discontinuous enzymatic assays to measure steady state kinetic parameters for the nitoreductase activity of AKR1C1-AKR1C4 and NQO1. Results indicate that the NQO1 catalyzed reduction of 3-NBA has a higher specific activity, but the combined specific activities of AKR1C catalyzed reduction may play a more significant role in the overall production of 3-ABA. These results suggest that the relative expression of NQO1 and AKR1C enzymes will determine their respective contribution to 3-NBA reduction, especially since all the aforementioned enzymes are inducible by the Nrf2-Keap1 system. This work is supported by P30E513508 and RO1 CA39504 to TMP. Citation Format: Jessica R. Murray, Meng Huang, Tianzhu Zang, Volker M. Arlt, Heinz H. Schmeiser, Trevor M. Penning. Metabolic activation of 3-nitrobenzanthrone by human aldo-keto reductases (AKR1C1-AKR1C4). [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4580A. doi:10.1158/1538-7445.AM2015-4580A

  • screening baccharin analogs as selective inhibitors against type 5 17β hydroxysteroid dehydrogenase akr1c3
    Chemico-Biological Interactions, 2015
    Co-Authors: Tianzhu Zang, Kshitij Verma, Paul C Trippier, Mo Chen, Trevor M. Penning
    Abstract:

    Abstract Aldo–keto reductase 1C3 (AKR1C3), also known as type 5 17β-hydroxysteroid dehydrogenase, is a downstream steroidogenic enzyme and converts androgen precursors to the potent androgen receptor ligands: testosterone and 5α-dihydrotestosterone. Studies have shown that AKR1C3 is involved in the development of castration resistant prostate cancer (CRPC) and that it is a rational drug target for the treatment of CRPC. Baccharin, a component of Brazilian propolis, has been observed to exhibit a high inhibitory potency and selectivity for AKR1C3 over other AKR1C isoforms and is a promising lead compound for developing more potent and selective inhibitors. Here, we report the screening of fifteen baccharin analogs as selective inhibitors against AKR1C3 versus AKR1C2 (type 3 3α-hydroxysteroid dehydrogenase). Among these analogs, the inhibitory activity and selectivity of thirteen compounds were evaluated for the first time. The substitution of the 4-dihydrocinnamoyloxy group of baccharin by an acetate group displayed nanomolar inhibitory potency (IC50: 440 nM) and a 102-fold selectivity over AKR1C2. By contrast, when the cinnamic acid group of baccharin was esterified, there was a dramatic decrease in potency and selectivity for AKR1C3 in comparison to baccharin. Low or sub-micromolar inhibition was observed when the 3-prenyl group of baccharin was removed, and the selectivity over AKR1C2 was low. Although unsubstituted baccharin was still the most potent (IC50: 100 nM) and selective inhibitor for AKR1C3, these data provide structure–activity relationships required for the optimization of new baccharin analogs. They suggest that the carboxylate group on cinnamic acid, the prenyl group, and either retention of 4-dihydrocinnamoyloxy group or acetate substituent on cinnamic acid are important to maintain the high potency and selectivity for AKR1C3.

  • akr1c3 as a target in castrate resistant prostate cancer
    The Journal of Steroid Biochemistry and Molecular Biology, 2013
    Co-Authors: Adegoke O Adeniji, Mo Chen, Trevor M. Penning
    Abstract:

    Abstract Aberrant androgen receptor (AR) activation is the major driver of castrate resistant prostate cancer (CRPC). CRPC is ultimately fatal and more therapeutic agents are needed to treat this disease. Compounds that target the androgen axis by inhibiting androgen biosynthesis and or AR signaling are potential candidates for use in CRPC treatment and are currently being pursued aggressively. Aldo-keto reductase 1C3 (AKR1C3) plays a pivotal role in androgen biosynthesis within the prostate. It catalyzes the 17-ketoreduction of weak androgen precursors to give testosterone and 5α-dihydrotestosterone. AKR1C3 expression and activity has been implicated in the development of CRPC, making it a rational target. Selective inhibition of AKR1C3 will be important, however, due to the presence of closely related isoforms, AKR1C1 and AKR1C2 that are also involved in androgen inactivation. We examine the evidence that supports the vital role of AKR1C3 in CRPC and recent developments in the discovery of potent and selective AKR1C3 inhibitors. This article is part of a Special Issue entitled ‘CSR 2013’.

M E Burczynski - One of the best experts on this subject based on the ideXlab platform.

  • the reactive oxygen species and michael acceptor inducible human aldo keto reductase AKR1C1 reduces the α β unsaturated aldehyde 4 hydroxy 2 nonenal to 1 4 dihydroxy 2 nonene
    Journal of Biological Chemistry, 2001
    Co-Authors: M E Burczynski, N Palackal, Gopishetty R. Sridhar, T M Penning
    Abstract:

    Abstract The human aldo-keto reductase AKR1C1 (20α(3α)-hydroxysteroid dehydrogenase) is induced by electrophilic Michael acceptors and reactive oxygen species (ROS) via a presumptive antioxidant response element (Burczynski, M. E., Lin, H. K., and Penning, T. M. (1999) Cancer Res. 59, 607–614). Physiologically, AKR1C1 regulates progesterone action by converting the hormone into its inactive metabolite 20α-hydroxyprogesterone, and toxicologically this enzyme activates polycyclic aromatic hydrocarbon trans-dihydrodiols to redox-cycling o-quinones. However, the significance of its potent induction by Michael acceptors and oxidative stress is unknown. 4-Hydroxy-2-nonenal (HNE) and other α,β-unsaturated aldehydes produced during lipid peroxidation were reduced by AKR1C1 with high catalytic efficiency. Kinetic studies revealed that AKR1C1 reduced HNE (K m = 34 μm,k cat = 8.8 min− 1) with a k cat/K m similar to that for 20α-hydroxysteroids. Six other homogeneous recombinant AKRs were examined for their ability to reduce HNE. Of these, AKR1C1 possessed one of the highest specific activities and was the only isoform induced by oxidative stress and by agents that deplete glutathione (ethacrynic acid). Several hydroxysteroid dehydrogenases of the AKR1C subfamily catalyzed the reduction of HNE with higher activity than aldehyde reductase (AKR1A1). NMR spectroscopy identified the product of the NADPH-dependent reduction of HNE as 1,4-dihydroxy-2-nonene. The K m of recombinant AKR1C1 for nicotinamide cofactors (K m NADPH ∼6 μm,K m(app) NADH >6 mm) suggested that it is primed for reductive metabolism of HNE. Isoform-specific reverse transcription-polymerase chain reaction showed that exposure of HepG2 cells to HNE resulted in elevated levels of AKR1C1 mRNA. Thus, HNE induces its own metabolism via AKR1C1, and this enzyme may play a hitherto unrecognized role in a response mounted to counter oxidative stress. AKRs represent alternative GSH-independent/NADPH-dependent routes for the reductive elimination of HNE. Of these, AKR1C1 provides an inducible cytosolic barrier to HNE following ROS exposure.

  • structure function aspects and inhibitor design of type 5 17β hydroxysteroid dehydrogenase akr1c3
    Molecular and Cellular Endocrinology, 2001
    Co-Authors: Trevor M. Penning, M E Burczynski, Margaret Moore, Kavitha Ratnam, N Palackal
    Abstract:

    Abstract 17β-Hydroxysteroid dehydrogenase (17β-HSD) type 5 has been cloned from human prostate and is identical to type 2 3α-HSD and is a member of the aldo-keto reductase (AKR) superfamily; it is formally AKR1C3. In vitro the homogeneous recombinant enzyme expressed in Escherichia coli functions as a 3-keto-, 17-keto- and 20-ketosteroid reductase and as a 3α-, 17β- and 20α-hydroxysteroid oxidase. The enzyme will reduce 5α-DHT, Δ4-androstene-3,17-dione, estrone and progesterone to produce 3α-androstanediol, testosterone, 17β-estradiol and 20α-hydroxprogesterone, respectively. It will also oxidize 3α-androstanediol, testosterone, 17β-estradiol and 20α-hydroxyprogesterone to produce 5α-androstane-3,17-dione, Δ4-androstene-3,17-dione, and progesterone, respectively. Many of these properties are shared by the related AKR1C1, AKR1C2 and AKR1C4 isoforms. RT-PCR shows that AKR1C3 is dominantly expressed in the human prostate and mammary gland. Examination of kcat/Km for these reactions indicates that as a reductase it prefers 5α-dihydrotestosterone and 5α-androstane-3,17-dione as substrates to Δ4-androstene-3,17-dione, suggesting that in the prostate it favors the formation of inactive androgens. Its concerted reductase activity may, however, lead to a pro-estrogenic state in the breast since it will convert estrone to 17β-estradiol; convert Δ4-androstene-3,17-dione to testosterone (which can be aromatized to 17β-estradiol); and it will reduce progesterone to its inactive metabolite 20α-hydroxyprogesterone. Drawing on detailed structure-function analysis of the related rat 3α-HSD (AKR1C9), which shares 69% sequence identity with AKR1C3, it is predicted that AKR1C3 catalyzes an ordered bi bi mechanism, that the rate determining step is kchem, and that an oxyanion prevails in the transition state. Based on these relationships steroidal-based inhibitors that compete with the steroid product would be desirable since they would act as uncompetitive inhibitors. With regards to transition state analogs steroid carboxylates and pyrazoles may be preferred while 3α, 17β or 20α-spiro-oxiranes may act as mechanism-based inactivators.

  • Structure-function aspects and inhibitor design of type 5 17beta-hydroxysteroid dehydrogenase (AKR1C3).
    Molecular and cellular endocrinology, 2001
    Co-Authors: Trevor M. Penning, M E Burczynski, Kavitha Ratnam, M. Moore, H. Ma, N Palackal
    Abstract:

    17beta-Hydroxysteroid dehydrogenase (17beta-HSD) type 5 has been cloned from human prostate and is identical to type 2 3alpha-HSD and is a member of the aldo-keto reductase (AKR) superfamily; it is formally AKR1C3. In vitro the homogeneous recombinant enzyme expressed in Escherichia coli functions as a 3-keto-, 17-keto- and 20-ketosteroid reductase and as a 3alpha-, 17beta- and 20alpha-hydroxysteroid oxidase. The enzyme will reduce 5alpha-DHT, Delta(4)-androstene-3,17-dione, estrone and progesterone to produce 3alpha-androstanediol, testosterone, 17beta-estradiol and 20alpha-hydroxprogesterone, respectively. It will also oxidize 3alpha-androstanediol, testosterone, 17beta-estradiol and 20alpha-hydroxyprogesterone to produce 5alpha-androstane-3,17-dione, Delta(4)-androstene-3,17-dione, and progesterone, respectively. Many of these properties are shared by the related AKR1C1, AKR1C2 and AKR1C4 isoforms. RT-PCR shows that AKR1C3 is dominantly expressed in the human prostate and mammary gland. Examination of k(cat)/K(m) for these reactions indicates that as a reductase it prefers 5alpha-dihydrotestosterone and 5alpha-androstane-3,17-dione as substrates to Delta(4)-androstene-3,17-dione, suggesting that in the prostate it favors the formation of inactive androgens. Its concerted reductase activity may, however, lead to a pro-estrogenic state in the breast since it will convert estrone to 17beta-estradiol; convert Delta(4)-androstene-3,17-dione to testosterone (which can be aromatized to 17beta-estradiol); and it will reduce progesterone to its inactive metabolite 20alpha-hydroxyprogesterone. Drawing on detailed structure-function analysis of the related rat 3alpha-HSD (AKR1C9), which shares 69% sequence identity with AKR1C3, it is predicted that AKR1C3 catalyzes an ordered bi bi mechanism, that the rate determining step is k(chem), and that an oxyanion prevails in the transition state. Based on these relationships steroidal-based inhibitors that compete with the steroid product would be desirable since they would act as uncompetitive inhibitors. With regards to transition state analogs steroid carboxylates and pyrazoles may be preferred while 3alpha, 17beta or 20alpha-spiro-oxiranes may act as mechanism-based inactivators.

  • Structure-function aspects and inhibitor design of type 5 17β-hydroxysteroid dehydrogenase (AKR1C3)☆
    Molecular and Cellular Endocrinology, 2001
    Co-Authors: Trevor M. Penning, M E Burczynski, Margaret Moore, Kavitha Ratnam, Haiching Ma, N Palackal
    Abstract:

    Abstract 17β-Hydroxysteroid dehydrogenase (17β-HSD) type 5 has been cloned from human prostate and is identical to type 2 3α-HSD and is a member of the aldo-keto reductase (AKR) superfamily; it is formally AKR1C3. In vitro the homogeneous recombinant enzyme expressed in Escherichia coli functions as a 3-keto-, 17-keto- and 20-ketosteroid reductase and as a 3α-, 17β- and 20α-hydroxysteroid oxidase. The enzyme will reduce 5α-DHT, Δ4-androstene-3,17-dione, estrone and progesterone to produce 3α-androstanediol, testosterone, 17β-estradiol and 20α-hydroxprogesterone, respectively. It will also oxidize 3α-androstanediol, testosterone, 17β-estradiol and 20α-hydroxyprogesterone to produce 5α-androstane-3,17-dione, Δ4-androstene-3,17-dione, and progesterone, respectively. Many of these properties are shared by the related AKR1C1, AKR1C2 and AKR1C4 isoforms. RT-PCR shows that AKR1C3 is dominantly expressed in the human prostate and mammary gland. Examination of kcat/Km for these reactions indicates that as a reductase it prefers 5α-dihydrotestosterone and 5α-androstane-3,17-dione as substrates to Δ4-androstene-3,17-dione, suggesting that in the prostate it favors the formation of inactive androgens. Its concerted reductase activity may, however, lead to a pro-estrogenic state in the breast since it will convert estrone to 17β-estradiol; convert Δ4-androstene-3,17-dione to testosterone (which can be aromatized to 17β-estradiol); and it will reduce progesterone to its inactive metabolite 20α-hydroxyprogesterone. Drawing on detailed structure-function analysis of the related rat 3α-HSD (AKR1C9), which shares 69% sequence identity with AKR1C3, it is predicted that AKR1C3 catalyzes an ordered bi bi mechanism, that the rate determining step is kchem, and that an oxyanion prevails in the transition state. Based on these relationships steroidal-based inhibitors that compete with the steroid product would be desirable since they would act as uncompetitive inhibitors. With regards to transition state analogs steroid carboxylates and pyrazoles may be preferred while 3α, 17β or 20α-spiro-oxiranes may act as mechanism-based inactivators.

  • The ROS and Michael-acceptor inducible human aldo-Keto reductase AKR1C1 reduces the alpha,beta-unsaturated aldehyde 4-Hydroxy-2-nonenal to 1,4-dihydroxy-2-nonene
    The Journal of biological chemistry, 2000
    Co-Authors: M E Burczynski, N Palackal, Gopishetty R. Sridhar, Trevor M. Penning
    Abstract:

    Abstract The human aldo-keto reductase AKR1C1 (20α(3α)-hydroxysteroid dehydrogenase) is induced by electrophilic Michael acceptors and reactive oxygen species (ROS) via a presumptive antioxidant response element (Burczynski, M. E., Lin, H. K., and Penning, T. M. (1999) Cancer Res. 59, 607–614). Physiologically, AKR1C1 regulates progesterone action by converting the hormone into its inactive metabolite 20α-hydroxyprogesterone, and toxicologically this enzyme activates polycyclic aromatic hydrocarbon trans-dihydrodiols to redox-cycling o-quinones. However, the significance of its potent induction by Michael acceptors and oxidative stress is unknown. 4-Hydroxy-2-nonenal (HNE) and other α,β-unsaturated aldehydes produced during lipid peroxidation were reduced by AKR1C1 with high catalytic efficiency. Kinetic studies revealed that AKR1C1 reduced HNE (K m = 34 μm,k cat = 8.8 min− 1) with a k cat/K m similar to that for 20α-hydroxysteroids. Six other homogeneous recombinant AKRs were examined for their ability to reduce HNE. Of these, AKR1C1 possessed one of the highest specific activities and was the only isoform induced by oxidative stress and by agents that deplete glutathione (ethacrynic acid). Several hydroxysteroid dehydrogenases of the AKR1C subfamily catalyzed the reduction of HNE with higher activity than aldehyde reductase (AKR1A1). NMR spectroscopy identified the product of the NADPH-dependent reduction of HNE as 1,4-dihydroxy-2-nonene. The K m of recombinant AKR1C1 for nicotinamide cofactors (K m NADPH ∼6 μm,K m(app) NADH >6 mm) suggested that it is primed for reductive metabolism of HNE. Isoform-specific reverse transcription-polymerase chain reaction showed that exposure of HepG2 cells to HNE resulted in elevated levels of AKR1C1 mRNA. Thus, HNE induces its own metabolism via AKR1C1, and this enzyme may play a hitherto unrecognized role in a response mounted to counter oxidative stress. AKRs represent alternative GSH-independent/NADPH-dependent routes for the reductive elimination of HNE. Of these, AKR1C1 provides an inducible cytosolic barrier to HNE following ROS exposure.

N Palackal - One of the best experts on this subject based on the ideXlab platform.

  • the reactive oxygen species and michael acceptor inducible human aldo keto reductase AKR1C1 reduces the α β unsaturated aldehyde 4 hydroxy 2 nonenal to 1 4 dihydroxy 2 nonene
    Journal of Biological Chemistry, 2001
    Co-Authors: M E Burczynski, N Palackal, Gopishetty R. Sridhar, T M Penning
    Abstract:

    Abstract The human aldo-keto reductase AKR1C1 (20α(3α)-hydroxysteroid dehydrogenase) is induced by electrophilic Michael acceptors and reactive oxygen species (ROS) via a presumptive antioxidant response element (Burczynski, M. E., Lin, H. K., and Penning, T. M. (1999) Cancer Res. 59, 607–614). Physiologically, AKR1C1 regulates progesterone action by converting the hormone into its inactive metabolite 20α-hydroxyprogesterone, and toxicologically this enzyme activates polycyclic aromatic hydrocarbon trans-dihydrodiols to redox-cycling o-quinones. However, the significance of its potent induction by Michael acceptors and oxidative stress is unknown. 4-Hydroxy-2-nonenal (HNE) and other α,β-unsaturated aldehydes produced during lipid peroxidation were reduced by AKR1C1 with high catalytic efficiency. Kinetic studies revealed that AKR1C1 reduced HNE (K m = 34 μm,k cat = 8.8 min− 1) with a k cat/K m similar to that for 20α-hydroxysteroids. Six other homogeneous recombinant AKRs were examined for their ability to reduce HNE. Of these, AKR1C1 possessed one of the highest specific activities and was the only isoform induced by oxidative stress and by agents that deplete glutathione (ethacrynic acid). Several hydroxysteroid dehydrogenases of the AKR1C subfamily catalyzed the reduction of HNE with higher activity than aldehyde reductase (AKR1A1). NMR spectroscopy identified the product of the NADPH-dependent reduction of HNE as 1,4-dihydroxy-2-nonene. The K m of recombinant AKR1C1 for nicotinamide cofactors (K m NADPH ∼6 μm,K m(app) NADH >6 mm) suggested that it is primed for reductive metabolism of HNE. Isoform-specific reverse transcription-polymerase chain reaction showed that exposure of HepG2 cells to HNE resulted in elevated levels of AKR1C1 mRNA. Thus, HNE induces its own metabolism via AKR1C1, and this enzyme may play a hitherto unrecognized role in a response mounted to counter oxidative stress. AKRs represent alternative GSH-independent/NADPH-dependent routes for the reductive elimination of HNE. Of these, AKR1C1 provides an inducible cytosolic barrier to HNE following ROS exposure.

  • structure function aspects and inhibitor design of type 5 17β hydroxysteroid dehydrogenase akr1c3
    Molecular and Cellular Endocrinology, 2001
    Co-Authors: Trevor M. Penning, M E Burczynski, Margaret Moore, Kavitha Ratnam, N Palackal
    Abstract:

    Abstract 17β-Hydroxysteroid dehydrogenase (17β-HSD) type 5 has been cloned from human prostate and is identical to type 2 3α-HSD and is a member of the aldo-keto reductase (AKR) superfamily; it is formally AKR1C3. In vitro the homogeneous recombinant enzyme expressed in Escherichia coli functions as a 3-keto-, 17-keto- and 20-ketosteroid reductase and as a 3α-, 17β- and 20α-hydroxysteroid oxidase. The enzyme will reduce 5α-DHT, Δ4-androstene-3,17-dione, estrone and progesterone to produce 3α-androstanediol, testosterone, 17β-estradiol and 20α-hydroxprogesterone, respectively. It will also oxidize 3α-androstanediol, testosterone, 17β-estradiol and 20α-hydroxyprogesterone to produce 5α-androstane-3,17-dione, Δ4-androstene-3,17-dione, and progesterone, respectively. Many of these properties are shared by the related AKR1C1, AKR1C2 and AKR1C4 isoforms. RT-PCR shows that AKR1C3 is dominantly expressed in the human prostate and mammary gland. Examination of kcat/Km for these reactions indicates that as a reductase it prefers 5α-dihydrotestosterone and 5α-androstane-3,17-dione as substrates to Δ4-androstene-3,17-dione, suggesting that in the prostate it favors the formation of inactive androgens. Its concerted reductase activity may, however, lead to a pro-estrogenic state in the breast since it will convert estrone to 17β-estradiol; convert Δ4-androstene-3,17-dione to testosterone (which can be aromatized to 17β-estradiol); and it will reduce progesterone to its inactive metabolite 20α-hydroxyprogesterone. Drawing on detailed structure-function analysis of the related rat 3α-HSD (AKR1C9), which shares 69% sequence identity with AKR1C3, it is predicted that AKR1C3 catalyzes an ordered bi bi mechanism, that the rate determining step is kchem, and that an oxyanion prevails in the transition state. Based on these relationships steroidal-based inhibitors that compete with the steroid product would be desirable since they would act as uncompetitive inhibitors. With regards to transition state analogs steroid carboxylates and pyrazoles may be preferred while 3α, 17β or 20α-spiro-oxiranes may act as mechanism-based inactivators.

  • Structure-function aspects and inhibitor design of type 5 17beta-hydroxysteroid dehydrogenase (AKR1C3).
    Molecular and cellular endocrinology, 2001
    Co-Authors: Trevor M. Penning, M E Burczynski, Kavitha Ratnam, M. Moore, H. Ma, N Palackal
    Abstract:

    17beta-Hydroxysteroid dehydrogenase (17beta-HSD) type 5 has been cloned from human prostate and is identical to type 2 3alpha-HSD and is a member of the aldo-keto reductase (AKR) superfamily; it is formally AKR1C3. In vitro the homogeneous recombinant enzyme expressed in Escherichia coli functions as a 3-keto-, 17-keto- and 20-ketosteroid reductase and as a 3alpha-, 17beta- and 20alpha-hydroxysteroid oxidase. The enzyme will reduce 5alpha-DHT, Delta(4)-androstene-3,17-dione, estrone and progesterone to produce 3alpha-androstanediol, testosterone, 17beta-estradiol and 20alpha-hydroxprogesterone, respectively. It will also oxidize 3alpha-androstanediol, testosterone, 17beta-estradiol and 20alpha-hydroxyprogesterone to produce 5alpha-androstane-3,17-dione, Delta(4)-androstene-3,17-dione, and progesterone, respectively. Many of these properties are shared by the related AKR1C1, AKR1C2 and AKR1C4 isoforms. RT-PCR shows that AKR1C3 is dominantly expressed in the human prostate and mammary gland. Examination of k(cat)/K(m) for these reactions indicates that as a reductase it prefers 5alpha-dihydrotestosterone and 5alpha-androstane-3,17-dione as substrates to Delta(4)-androstene-3,17-dione, suggesting that in the prostate it favors the formation of inactive androgens. Its concerted reductase activity may, however, lead to a pro-estrogenic state in the breast since it will convert estrone to 17beta-estradiol; convert Delta(4)-androstene-3,17-dione to testosterone (which can be aromatized to 17beta-estradiol); and it will reduce progesterone to its inactive metabolite 20alpha-hydroxyprogesterone. Drawing on detailed structure-function analysis of the related rat 3alpha-HSD (AKR1C9), which shares 69% sequence identity with AKR1C3, it is predicted that AKR1C3 catalyzes an ordered bi bi mechanism, that the rate determining step is k(chem), and that an oxyanion prevails in the transition state. Based on these relationships steroidal-based inhibitors that compete with the steroid product would be desirable since they would act as uncompetitive inhibitors. With regards to transition state analogs steroid carboxylates and pyrazoles may be preferred while 3alpha, 17beta or 20alpha-spiro-oxiranes may act as mechanism-based inactivators.

  • Structure-function aspects and inhibitor design of type 5 17β-hydroxysteroid dehydrogenase (AKR1C3)☆
    Molecular and Cellular Endocrinology, 2001
    Co-Authors: Trevor M. Penning, M E Burczynski, Margaret Moore, Kavitha Ratnam, Haiching Ma, N Palackal
    Abstract:

    Abstract 17β-Hydroxysteroid dehydrogenase (17β-HSD) type 5 has been cloned from human prostate and is identical to type 2 3α-HSD and is a member of the aldo-keto reductase (AKR) superfamily; it is formally AKR1C3. In vitro the homogeneous recombinant enzyme expressed in Escherichia coli functions as a 3-keto-, 17-keto- and 20-ketosteroid reductase and as a 3α-, 17β- and 20α-hydroxysteroid oxidase. The enzyme will reduce 5α-DHT, Δ4-androstene-3,17-dione, estrone and progesterone to produce 3α-androstanediol, testosterone, 17β-estradiol and 20α-hydroxprogesterone, respectively. It will also oxidize 3α-androstanediol, testosterone, 17β-estradiol and 20α-hydroxyprogesterone to produce 5α-androstane-3,17-dione, Δ4-androstene-3,17-dione, and progesterone, respectively. Many of these properties are shared by the related AKR1C1, AKR1C2 and AKR1C4 isoforms. RT-PCR shows that AKR1C3 is dominantly expressed in the human prostate and mammary gland. Examination of kcat/Km for these reactions indicates that as a reductase it prefers 5α-dihydrotestosterone and 5α-androstane-3,17-dione as substrates to Δ4-androstene-3,17-dione, suggesting that in the prostate it favors the formation of inactive androgens. Its concerted reductase activity may, however, lead to a pro-estrogenic state in the breast since it will convert estrone to 17β-estradiol; convert Δ4-androstene-3,17-dione to testosterone (which can be aromatized to 17β-estradiol); and it will reduce progesterone to its inactive metabolite 20α-hydroxyprogesterone. Drawing on detailed structure-function analysis of the related rat 3α-HSD (AKR1C9), which shares 69% sequence identity with AKR1C3, it is predicted that AKR1C3 catalyzes an ordered bi bi mechanism, that the rate determining step is kchem, and that an oxyanion prevails in the transition state. Based on these relationships steroidal-based inhibitors that compete with the steroid product would be desirable since they would act as uncompetitive inhibitors. With regards to transition state analogs steroid carboxylates and pyrazoles may be preferred while 3α, 17β or 20α-spiro-oxiranes may act as mechanism-based inactivators.

  • The ROS and Michael-acceptor inducible human aldo-Keto reductase AKR1C1 reduces the alpha,beta-unsaturated aldehyde 4-Hydroxy-2-nonenal to 1,4-dihydroxy-2-nonene
    The Journal of biological chemistry, 2000
    Co-Authors: M E Burczynski, N Palackal, Gopishetty R. Sridhar, Trevor M. Penning
    Abstract:

    Abstract The human aldo-keto reductase AKR1C1 (20α(3α)-hydroxysteroid dehydrogenase) is induced by electrophilic Michael acceptors and reactive oxygen species (ROS) via a presumptive antioxidant response element (Burczynski, M. E., Lin, H. K., and Penning, T. M. (1999) Cancer Res. 59, 607–614). Physiologically, AKR1C1 regulates progesterone action by converting the hormone into its inactive metabolite 20α-hydroxyprogesterone, and toxicologically this enzyme activates polycyclic aromatic hydrocarbon trans-dihydrodiols to redox-cycling o-quinones. However, the significance of its potent induction by Michael acceptors and oxidative stress is unknown. 4-Hydroxy-2-nonenal (HNE) and other α,β-unsaturated aldehydes produced during lipid peroxidation were reduced by AKR1C1 with high catalytic efficiency. Kinetic studies revealed that AKR1C1 reduced HNE (K m = 34 μm,k cat = 8.8 min− 1) with a k cat/K m similar to that for 20α-hydroxysteroids. Six other homogeneous recombinant AKRs were examined for their ability to reduce HNE. Of these, AKR1C1 possessed one of the highest specific activities and was the only isoform induced by oxidative stress and by agents that deplete glutathione (ethacrynic acid). Several hydroxysteroid dehydrogenases of the AKR1C subfamily catalyzed the reduction of HNE with higher activity than aldehyde reductase (AKR1A1). NMR spectroscopy identified the product of the NADPH-dependent reduction of HNE as 1,4-dihydroxy-2-nonene. The K m of recombinant AKR1C1 for nicotinamide cofactors (K m NADPH ∼6 μm,K m(app) NADH >6 mm) suggested that it is primed for reductive metabolism of HNE. Isoform-specific reverse transcription-polymerase chain reaction showed that exposure of HepG2 cells to HNE resulted in elevated levels of AKR1C1 mRNA. Thus, HNE induces its own metabolism via AKR1C1, and this enzyme may play a hitherto unrecognized role in a response mounted to counter oxidative stress. AKRs represent alternative GSH-independent/NADPH-dependent routes for the reductive elimination of HNE. Of these, AKR1C1 provides an inducible cytosolic barrier to HNE following ROS exposure.

Michael C Byrns - One of the best experts on this subject based on the ideXlab platform.

  • development of potent and selective indomethacin analogues for the inhibition of akr1c3 type 5 17β hydroxysteroid dehydrogenase prostaglandin f synthase in castrate resistant prostate cancer
    Journal of Medicinal Chemistry, 2013
    Co-Authors: Andy J Liedtke, Adegoke O Adeniji, David W. Christianson, Michael C Byrns, Lawrence J Marnett, Mo Chen, Trevor M. Penning
    Abstract:

    Castrate-resistant prostate cancer (CRPC) is a fatal, metastatic form of prostate cancer. CRPC is characterized by reactivation of the androgen axis due to changes in androgen receptor signaling and/or adaptive intratumoral androgen biosynthesis. AKR1C3 is upregulated in CRPC where it catalyzes the formation of potent androgens. This makes AKR1C3 a target for the treatment of CRPC. AKR1C3 inhibitors should not inhibit AKR1C1/AKR1C2, which inactivate 5α-dihydrotestosterone. Indomethacin, used to inhibit cyclooxygenase, also inhibits AKR1C3 and displays selectivity over AKR1C1/AKR1C2. Parallel synthetic strategies were used to generate libraries of indomethacin analogues, which exhibit reduced cyclooxygenase inhibitory activity but retain AKR1C3 inhibitory potency and selectivity. The lead compounds inhibited AKR1C3 with nanomolar potency, displayed >100-fold selectivity over AKR1C1/AKR1C2, and blocked testosterone formation in LNCaP-AKR1C3 cells. The AKR1C3·NADP+·2′-des-methyl-indomethacin crystal structur...

  • Development of Potent and Selective Indomethacin Analogues for the Inhibition of AKR1C3 (Type 5 17β-Hydroxysteroid Dehydrogenase/Prostaglandin F Synthase) in Castrate-Resistant Prostate Cancer
    Journal of Medicinal Chemistry, 2013
    Co-Authors: Andy J Liedtke, Adegoke O Adeniji, David W. Christianson, Michael C Byrns, Lawrence J Marnett, Mo Chen, Trevor M. Penning
    Abstract:

    Castrate-resistant prostate cancer (CRPC) is a fatal, metastatic form of prostate cancer. CRPC is characterized by reactivation of the androgen axis due to changes in androgen receptor signaling and/or adaptive intratumoral androgen biosynthesis. AKR1C3 is upregulated in CRPC where it catalyzes the formation of potent androgens. This makes AKR1C3 a target for the treatment of CRPC. AKR1C3 inhibitors should not inhibit AKR1C1/AKR1C2, which inactivate 5α-dihydrotestosterone. Indomethacin, used to inhibit cyclooxygenase, also inhibits AKR1C3 and displays selectivity over AKR1C1/AKR1C2. Parallel synthetic strategies were used to generate libraries of indomethacin analogues, which exhibit reduced cyclooxygenase inhibitory activity but retain AKR1C3 inhibitory potency and selectivity. The lead compounds inhibited AKR1C3 with nanomolar potency, displayed >100-fold selectivity over AKR1C1/AKR1C2, and blocked testosterone formation in LNCaP-AKR1C3 cells. The AKR1C3·NADP+·2′-des-methyl-indomethacin crystal structur...

  • abstract b16 development of potent and selective indomethacin analogs for the inhibition of akr1c3 type 5 17β hydroxysteroid dehydrogenase in crpc
    Cancer Research, 2012
    Co-Authors: Adegoke O Adeniji, Andrew Liedkte, David W. Christianson, Michael C Byrns, Lawrence J Marnett, Mo Chen, Trevor M. Penning
    Abstract:

    Abstract Castrate-resistant prostate cancer (CRPC) is a fatal metastatic form of prostate cancer that accounts for 30,000 deaths in the U.S. annually. Patients with advanced prostate cancer that initially respond to hormonal ablative therapy (orchiectomy or chemical castration) invariably develop CRPC. CRPC is characterized by reactivation of the androgen axis due to changes in androgen receptor (AR) signaling and/or adaptive intratumoral androgen biosynthesis. The latter is targeted with the new agent abiraterone acetate (a P450 17α-hydroxylase/17,20-lyase inhibitor) which prevents the production of the adrenal androgen precursor, dehydroepiandrosterone but has the unintended consequence of building up the potent mineralocortcoid, desoxycorticosterone. To circumvent this side effect, abiraterone acetate is co-administered with prednisone. AKR1C3 is among the most upregulated genes in CRPC and in soft tissue metastasis where its 17β-hydroxysteroid dehydrogenase activity converts the weak androgens, 4-androstene-3,17-dione and 5α-androstane-3,17-dione to the potent androgens, testosterone and 5α-dihydrotestosterone, respectively. The role of AKR1C3 in the pre-receptor regulation of ligands for the AR makes it a more selective target than abiraterone for the treatment of CRPC. Inhibitors of AKR1C3 should not inhibit the related isoforms, AKR1C1 and AKR1C2 since these enzymes inactivate 5α-dihydrotestosterone. Indomethacin, a therapeutic agent used to inhibit cyclooxygenase (COX) also inhibits AKR1C3 potently and displays selectivity over AKR1C1 and AKR1C2. Parallel synthetic strategies were used to generate libraries of indomethacin analogs which based on known structure-activity relationships should no longer inhibit COX enzymes but may retain AKR1C3 inhibitory potency and selectivity. Three classes of AKR1C3 inhibitors were discovered, that had the desired properties: indomethacin analogs (in which the 3′-side chain was modified), 2′-desmethyl-indomethacin analogs (which had either an acid or trifluoromethylsulfonamide substituent at the 3′-position), and 3′-alkyl-indomethacin analogs (which had either an acid or trifluoromethylsulfonamide substituent at the 2′-position). The lead compounds inhibited AKR1C3 with nanomolar potency, displayed over 100-fold selectivity for AKR1C3 over AKR1C1/AKR1C2 and produced robust inhibition of testosterone formation in an LNCaP-AKR1C3 prostate cancer cell line. Several had minimal inhibitory affects on COX-1 and COX-2. In a separate study, indomethacin blocked PSA and ERG expression, and cell proliferation in a VCaP xenograft model of CRPC providing in vivo efficacy data (Cai at al., Cancer Res. 71: 6503, 2011). 2′-Desmethyl-indomethacin 1 was crystallized in complex with AKR1C3 and NADP+, and diffraction data were obtained at 1.8 A resolution. The structure showed that in the absence of the 2′-methyl group the compound assumes a different binding mode to indomethacin. Compound 1 binds perpendicular to indomethacin such that the carboxylic acid on its indole ring is tethered to the oxyanion site and protrudes into a SP1 pocket as seen with the other NSAIDs like flufenamic acid. This model provides a binding pose for the 2′-desmethyl indomethacin analogs and also predicts a possible binding pose for the 3′-alkyl indomethacin series. The compounds reported are promising agents for the preclinical development of therapeutics for CRPC, which by targeting AKR1C3 will be more selective than P450 17α-hydroxylase/17,20-lyase inhibitors, and will not have to be co-administered with prednisone. The agents disclosed in this abstract are protected by US Provisional Patent Application No. 61/548,004. [Supported by R01-CA90744 and a Prostate Cancer Foundation Challenge Award to TMP and R01-CA889450 to LJM] Citation Format: Adegoke Adeniji, Andrew Liedkte, Michael C. Byrns, M Chen, Yi Jin, David Christianson, Lawrence J. Marnett, Trevor M. Penning. Development of potent and selective indomethacin analogs for the inhibition of AKR1C3 (type 5 17β-hydroxysteroid dehydrogenase) in CRPC [abstract]. In: Proceedings of the AACR Special Conference on Advances in Prostate Cancer Research; 2012 Feb 6-9; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(4 Suppl):Abstract nr B16.

  • inhibitors of type 5 17β hydroxysteroid dehydrogenase akr1c3 overview and structural insights
    The Journal of Steroid Biochemistry and Molecular Biology, 2011
    Co-Authors: Michael C Byrns, Trevor M. Penning
    Abstract:

    Abstract There is considerable interest in the development of an inhibitor of aldo–keto reductase (AKR) 1C3 (type 5 17β-hydroxysteroid dehydrogenase and prostaglandin F synthase) as a potential therapeutic for both hormone-dependent and hormone-independent cancers. AKR1C3 catalyzes the reduction of 4-androstene-3,17-dione to testosterone and estrone to 17β-estradiol in target tissues, which will promote the proliferation of hormone dependent prostate and breast cancers, respectively. AKR1C3 also catalyzes the reduction of prostaglandin (PG) H2 to PGF2α and PGD2 to 9α,11β-PGF2, which will limit the formation of anti-proliferative prostaglandins, including 15-deoxy-Δ12,14-PGJ2, and contribute to proliferative signaling. AKR1C3 is overexpressed in a wide variety of cancers, including breast and prostate cancer. An inhibitor of AKR1C3 should not inhibit the closely related isoforms AKR1C1 and AKR1C2, as they are involved in other key steroid hormone biotransformations in target tissues. Several structural leads have been explored as inhibitors of AKR1C3, including non-steroidal anti-inflammatory drugs, steroid hormone analogues, flavonoids, cyclopentanes, and benzodiazepines. Inspection of the available crystal structures of AKR1C3 with multiple ligands bound, along with the crystal structures of the other AKR1C isoforms, provides a structural basis for the rational design of isoform specific inhibitors of AKR1C3. We find that there are subpockets involved in ligand binding that are considerably different in AKR1C3 relative to the closely related AKR1C1 or AKR1C2 isoforms. These pockets can be used to further improve the binding affinity and selectivity of the currently available AKR1C3 inhibitors. Article from the special issue on Targeted Inhibitors.

  • discovery of substituted 3 phenylamino benzoic acids as potent and selective inhibitors of type 5 17β hydroxysteroid dehydrogenase akr1c3
    Bioorganic & Medicinal Chemistry Letters, 2011
    Co-Authors: Adegoke O Adeniji, Barry M Twenter, Jeffrey D. Winkler, Michael C Byrns, Trevor M. Penning
    Abstract:

    Abstract Aldo-keto reductase 1C3 (AKR1C3) also known as type 5 17β-hydroxysteroid dehydrogenase has been implicated as one of the key enzymes driving the elevated intratumoral androgen levels observed in castrate resistant prostate cancer (CRPC). AKR1C3 inhibition therefore presents a rational approach to managing CRPC. Inhibitors should be selective for AKR1C3 over other AKR1C enzymes involved in androgen metabolism. We have synthesized 2-, 3-, and 4-(phenylamino)benzoic acids and identified 3-(phenylamino)benzoic acids that have nanomolar affinity and exhibit over 200-fold selectivity for AKR1C3 versus other AKR1C isoforms. The AKR1C3 inhibitory potency of the 4′-substituted 3-(phenylamino)benzoic acids shows a linear correlation with both electronic effects of substituents and the pKa of the carboxylic acid and secondary amine groups, which are interdependent. These compounds may be useful in treatment and/or prevention of CRPC as well as understanding the role of AKR1C3 in endocrinology.

T M Penning - One of the best experts on this subject based on the ideXlab platform.

  • role of aldo keto reductase family 1 akr1 enzymes in human steroid metabolism
    Steroids, 2014
    Co-Authors: Tea Lanišnik Rižner, T M Penning
    Abstract:

    Abstract Human aldo–keto reductases AKR1C1–AKR1C4 and AKR1D1 play essential roles in the metabolism of all steroid hormones, the biosynthesis of neurosteroids and bile acids, the metabolism of conjugated steroids, and synthetic therapeutic steroids. These enzymes catalyze NADPH dependent reductions at the C3, C5, C17 and C20 positions on the steroid nucleus and side-chain. AKR1C1–AKR1C4 act as 3-keto, 17-keto and 20-ketosteroid reductases to varying extents, while AKR1D1 acts as the sole Δ 4 -3-ketosteroid-5β-reductase (steroid 5β-reductase) in humans. AKR1 enzymes control the concentrations of active ligands for nuclear receptors and control their ligand occupancy and trans -activation, they also regulate the amount of neurosteroids that can modulate the activity of GABA A and NMDA receptors. As such they are involved in the pre-receptor regulation of nuclear and membrane bound receptors. Altered expression of individual AKR1C genes is related to development of prostate, breast, and endometrial cancer. Mutations in AKR1C1 and AKR1C4 are responsible for sexual development dysgenesis and mutations in AKR1D1 are causative in bile-acid deficiency.

  • human cytosolic hydroxysteroid dehydrogenases of the aldo ketoreductase superfamily catalyze reduction of conjugated steroids implications for phase i and phase ii steroid hormone metabolism
    Journal of Biological Chemistry, 2009
    Co-Authors: Yi Jin, Ian A. Blair, Seon Hwa Lee, Ling Duan, H J Kloosterboer, T M Penning
    Abstract:

    Abstract Aldo-ketoreductase 1C (AKR1C) enzymes catalyze the NADPH-dependent reduction of ketosteroids to hydroxysteroids. They are Phase I metabolizing enzymes for natural and synthetic steroid hormones. They convert 5α-dihydrotestosterone (Dht, potent androgen) to 3α/β-androstanediols (inactive androgens) and the prodrug tibolone (Tib) to estrogenic 3α/β-hydroxytibolones. Herein we demonstrate for the first time that human AKR1C enzymes (AKR1C1-4) are able to reduce conjugated steroids such as Dht-17β-glucuronide (DhtG), Dht-17β-sulfate (DhtS), and Tib-17β-sulfate (TibS). Product identities were characterized by liquid chromatography-mass spectrometry, and kinetic parameters of the reactions were determined. The product profile of the reduction of each steroid conjugate by the individual AKR1C isoform was similar to that of the corresponding free steroid except for the reduction of DhtG catalyzed by AKR1C2, where a complete inversion in stereochemical preference to 3β-reduction (with DhtG) from 3α-reduction (with Dht and DhtS) was observed. The catalytic efficiency of 3-keto reduction was modestly affected by the presence of a 17-sulfate group but severely impaired by the presence of a 17-glucuronide group for AKR1C1-3 isoforms. AKR1C4, however, showed superior catalytic efficiencies versus the other isoforms, and those were unaffected by steroid conjugation. Our findings provide evidence for alternative pathways of steroid metabolism where the phase I reaction (reduction) occurs after the phase II reaction (conjugation). Specifically, it is indicated that Dht is metabolized to its metabolite 3α-androstanediol-17-glucuronide via the previously unrecognized “conjugation pathway” involving the sequential reactions of UGT2B17 and AKR1C4 in liver but via the conventional “reduction pathway” involving the sequential reactions of AKR1C2 and UGT2B15/17 in prostate.

  • increased expression of genes converting adrenal androgens to testosterone in androgen independent prostate cancer
    Cancer Research, 2006
    Co-Authors: Michael Stanbrough, T M Penning, Glenn J Bubley, Kenneth N Ross, Todd R Golub, Mark A Rubin, Phillip G Febbo, Steven P Balk
    Abstract:

    Androgen receptor (AR) plays a central role in prostate cancer, and most patients respond to androgen deprivation therapies, but they invariably relapse with a more aggressive prostate cancer that has been termed hormone refractory or androgen independent. To identify proteins that mediate this tumor progression, gene expression in 33 androgen-independent prostate cancer bone marrow metastases versus 22 laser capture-microdissected primary prostate cancers was compared using Affymetrix oligonucleotide microarrays. Multiple genes associated with aggressive behavior were increased in the androgen-independent metastatic tumors (MMP9, CKS2, LRRC15, WNT5A, EZH2, E2F3, SDC1, SKP2, and BIRC5), whereas a candidate tumor suppressor gene (KLF6) was decreased. Consistent with castrate androgen levels, androgen-regulated genes were reduced 2- to 3-fold in the androgen-independent tumors. Nonetheless, they were still major transcripts in these tumors, indicating that there was partial reactivation of AR transcriptional activity. This was associated with increased expression of AR (5.8-fold) and multiple genes mediating androgen metabolism (HSD3B2, AKR1C3, SRD5A1, AKR1C2, AKR1C1, and UGT2B15). The increase in aldo-keto reductase family 1, member C3 (AKR1C3), the prostatic enzyme that reduces adrenal androstenedione to testosterone, was confirmed by real-time reverse transcription-PCR and immunohistochemistry. These results indicate that enhanced intracellular conversion of adrenal androgens to testosterone and dihydrotestosterone is a mechanism by which prostate cancer cells adapt to androgen deprivation and suggest new therapeutic targets.

  • competing roles of cytochrome p450 1a1 1b1 and aldo keto reductase 1a1 in the metabolic activation of 7 8 dihydroxy 7 8 dihydro benzo a pyrene in human bronchoalveolar cell extracts
    Chemical Research in Toxicology, 2005
    Co-Authors: Hao Jiang, Yumin Shen, Amy M Quinn, T M Penning
    Abstract:

    : (+/-)-7,8-Dihydroxy-7,8-dihydrobenzo[a]pyrene (BP-7,8-diol), a proximate carcinogen derived from benzo[a]pyrene (BP) requires further metabolic activation to exert its carcinogenic effects. Two principal pathways have been implicated, and these involve either the formation of (+/-)-trans-7,8-dihydroxy-9alpha,10alpha-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (anti-BPDE) catalyzed by P450 1A1/P450 1B1 (NADPH-dependent monoxygenases) or the formation of benzo[a]pyrene-7,8-dione (BP-7,8-dione) catalyzed by human aldo-keto reductases AKR1A1 and AKR1C1-AKR1C4 [NAD(P)(H)-dependent oxidoreductases]. The relative contributions of the two pathways to PAH activation are unknown. In this study, BP-7,8-diol metabolism was studied in human bronchoalveolar H358 cell extracts. Parental H358 cells do not constitutively express P450 1A1/P450 1B1 or AKRs but were manipulated by induction with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to express P450 1A1/P450 1B1 or by stable transfection to express AKR1A1 (aldehyde reductase). TCDD induction of AKR1A1 transfectants provided a cell line that expressed both pathways. Extracts derived from parental H358 cells plus TCDD (P450 induction) produced electrophilic anti-BPDE, which hydrolyzed to benzo[a]pyrene tetrahydrotetrols (BP-tetrols), extracts derived from AKR1A1-transfected cells (AKR1A1 expression) produced reactive and redox-active BP-7,8-dione, which was trapped in situ as its mono(thioether) conjugate, and extracts derived from AKR1A1 transfectants plus TCDD (coexpression of P450 1A1/P450 1B1 and AKR1A1) produced both anti-BPDE and BP-7,8-dione. The competing activation of BP-7,8-diol by P450 1A1/P450 1B1 and AKR1A1 was studied with varied NADPH:NAD+ ratios. The system with a relatively higher concentration of NADPH favored formation of anti-BPDE via P450 1A1/P450 1B1, while the system with the higher concentration of NAD+ favored formation of BP-7,8-dione via AKR1A1. Under conditions that mimic the cellular redox state, 10 microM NADPH and 1 mM NAD+, equal amounts of BP-tetrols and BP-7,8-dione were formed. This suggests that P450 1A1/P450 1B1 and AKR1A1 play competing roles in the metabolic activation of BP-7,8-diol and that the dominant pathway of BP-7,8-diol activation depends on the redox state of the cells. These model systems provide a cellular context in which the dominant DNA adducts/lesions formed by either pathway may be compared.

  • development of nonsteroidal anti inflammatory drug analogs and steroid carboxylates selective for human aldo keto reductase isoforms potential antineoplastic agents that work independently of cyclooxygenase isozymes
    Molecular Pharmacology, 2005
    Co-Authors: David R. Bauman, Sridhar Gopishetty, Stephen I Rudnick, Lawrence M Szewczuk, T M Penning
    Abstract:

    Human aldo-keto reductases (AKRs) regulate nuclear receptors by controlling ligand availability. Enzymes implicated in regulating ligand occupancy and trans -activation of the nuclear receptors belong to the AKR1C family (AKR1C1-AKR1C3). Nuclear receptors regulated by AKR1C members include the steroid hormone receptors (androgen, estrogen, and progesterone receptors) and the orphan peroxisome proliferator-activated receptor (PPARγ). In human myeloid leukemia (HL-60) cells, ligand access to PPARγ is regulated by AKR1C3, which diverts PGD2 metabolism away from J-series prostanoids ([Desmond et al., 2003][1]). Inhibition of AKR1C3 by indomethacin, a nonsteroidal anti-inflammatory drug (NSAID), caused PPARγ-mediated terminal differentiation of the HL-60 cells. To discriminate between antineoplastic effects of NSAIDs that are mediated by either AKR1C or cyclooxygenase (COX) isozymes, selective inhibitors are required. We report a structural series of N -phenylanthranilic acid derivatives and steroid carboxylates that selectively inhibit recombinant AKR1C isoforms but do not inhibit recombinant COX-1 or COX-2. The inhibition constants, IC50, K I values, and inhibition patterns were determined for the NSAID analogs and steroid carboxylates against AKR1C and COX isozymes. Lead compounds, 4-chloro- N -phenylanthranilic acid and 4-benzoyl-benzoic acid for the N -phenylanthranilic acid analogs and most steroid carboxylates, exhibited IC50 values that had greater than 500-fold selectivity for AKR1C isozymes compared with COX-1 and COX-2. Crystallographic and molecular modeling studies showed that the carboxylic acid of the inhibitor ligand was tethered by the catalytic Tyr55-OH2+ and explained why A-ring substituted N -phenylanthranilates inhibited only AKR1C enzymes. These compounds can be used to dissect the role of the AKR1C isozymes in neoplastic diseases and may have cancer chemopreventive roles independent of COX inhibition. [1]: #ref-9