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1.
Subcell Biochem ; 101: 41-80, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36520303

RESUMO

The Hsp90 chaperone is known to interact with a diverse array of client proteins. However, in every case examined, Hsp90 is also accompanied by a single or several co-chaperone proteins. One class of co-chaperone contains a tetratricopeptide repeat (TPR) domain that targets the co-chaperone to the C-terminal region of Hsp90. Within this class are Hsp90-binding peptidylprolyl isomerases, most of which belong to the FK506-binding protein (FKBP) family. Despite the common association of FKBP co-chaperones with Hsp90, it is abundantly clear that the client protein influences, and is often influenced by, the particular FKBP bound to Hsp90. Examples include Xap2 in aryl hydrocarbon receptor complexes and FKBP52 in steroid receptor complexes. In this chapter, we discuss the known functional roles played by FKBP co-chaperones and, where possible, relate distinctive functions to structural differences between FKBP members.


Assuntos
Proteínas de Choque Térmico HSP90 , Proteínas de Ligação a Tacrolimo , Humanos , Proteínas de Choque Térmico HSP90/química , Proteínas de Choque Térmico HSP90/genética , Proteínas de Choque Térmico HSP90/metabolismo , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Peptidilprolil Isomerase/metabolismo , Ligação Proteica , Proteínas de Ligação a Tacrolimo/genética , Proteínas de Ligação a Tacrolimo/química , Proteínas de Ligação a Tacrolimo/metabolismo , Imunofilinas/genética , Imunofilinas/metabolismo
2.
Pharmaceuticals (Basel) ; 13(11)2020 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-33202977

RESUMO

BACKGROUND: GMC1 (2-(1H-benzimidazol-2-ylsulfanyl)-N-[(Z)-(4-methoxyphenyl) methylideneamino] acetamide) effectively inhibits androgen receptor function by binding directly to FKBP52. This is a novel mechanism for the treatment of castration resistant prostate cancer (CRPC). METHODS: an LC-MS/MS method was developed and validated to quantify GMC1 in plasma and urine from pharmacokinetics studies in rats. An ultra-high-performance liquid chromatography (UHPLC) system equipped with a Waters XTerra MS C18 column was used for chromatographic separation by gradient elution with 0.1% (v/v) formic acid in water and methanol. A Sciex 4000 QTRAP® mass spectrometer was used for analysis by multiple reaction monitoring (MRM) in positive mode; the specific ions [M+H]+m/z 340.995 → m/z 191.000 and [M+H]+ m/z 266.013 → m/z 234.000 were monitored for GMC1 and internal standard (albendazole), respectively. RESULTS: GMC1 and albendazole had retention times of 1.68 and 1.66 min, respectively. The calibration curves for the determination of GMC1 in rat plasma and urine were linear from 1-1000 ng/mL. The LC-MS/MS method was validated with intra- and inter-day accuracy and precision within the 15% acceptance limit. The extraction recovery values of GMC1 from rat plasma and urine were greater than 95.0 ± 2.1% and 97.6 ± 4.6%, respectively, with no significant interfering matrix effect. GMC1 is stable under expected sample handling, storage, preparation and LC-MS/MS analysis conditions. CONCLUSIONS: Pharmacokinetic evaluation of GMC1 revealed that the molecule has a biexponential disposition in rats, is distributed rapidly and extensively, has a long elimination half-life, and appears to be eliminated primarily by first order kinetics.

3.
Cell Chem Biol ; 27(3): 292-305.e6, 2020 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-32017918

RESUMO

Hsp90 plays an important role in health and is a therapeutic target for managing misfolding disease. Compounds that disrupt co-chaperone delivery of clients to Hsp90 target a subset of Hsp90 activities, thereby minimizing the toxicity of pan-Hsp90 inhibitors. Here, we have identified SEW04784 as a first-in-class inhibitor of the Aha1-stimulated Hsp90 ATPase activity without inhibiting basal Hsp90 ATPase. Nuclear magnetic resonance analysis reveals that SEW84 binds to the C-terminal domain of Aha1 to weaken its asymmetric binding to Hsp90. Consistent with this observation, SEW84 blocks Aha1-dependent Hsp90 chaperoning activities, including the in vitro and in vivo refolding of firefly luciferase, and the transcriptional activity of the androgen receptor in cell-based models of prostate cancer and promotes the clearance of phosphorylated tau in cellular and tissue models of neurodegenerative tauopathy. We propose that SEW84 provides a novel lead scaffold for developing therapeutic approaches to treat proteostatic disease.


Assuntos
Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Chaperonas Moleculares/antagonistas & inibidores , Bibliotecas de Moléculas Pequenas/farmacologia , Células HEK293 , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Chaperonas Moleculares/metabolismo , Estrutura Molecular , Dobramento de Proteína/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/química
4.
PLoS One ; 10(7): e0134015, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26207810

RESUMO

FKBP52 and ß-catenin have emerged in recent years as attractive targets for prostate cancer treatment. ß-catenin interacts directly with the androgen receptor (AR) and has been characterized as a co-activator of AR-mediated transcription. FKBP52 is a positive regulator of AR in cellular and whole animal models and is required for the development of androgen-dependent tissues. We previously characterized an AR inhibitor termed MJC13 that putatively targets the AR BF3 surface to specifically inhibit FKBP52-regulated AR signaling. Predictive modeling suggests that ß-catenin interacts with the AR hormone binding domain on a surface that overlaps with BF3. Here we demonstrate that FKBP52 and ß-catenin interact directly in vitro and act in concert to promote a synergistic up-regulation of both hormone-independent and -dependent AR signaling. Our data demonstrate that FKBP52 promotes ß-catenin interaction with AR and is required for ß-catenin co-activation of AR activity in prostate cancer cells. MJC13 effectively blocks ß-catenin interaction with the AR LBD and the synergistic up-regulation of AR by FKBP52 and ß-catenin. Our data suggest that co-regulation of AR by FKBP52 and ß-catenin does not require FKBP52 PPIase catalytic activity, nor FKBP52 binding to Hsp90. However, the FKBP52 proline-rich loop that overhangs the PPIase pocket is critical for synergy.


Assuntos
Receptores Androgênicos/metabolismo , Sistemas do Segundo Mensageiro , Proteínas de Ligação a Tacrolimo/metabolismo , beta Catenina/metabolismo , Sequência de Aminoácidos , Sítios de Ligação , Células HEK293 , Células HeLa , Humanos , Dados de Sequência Molecular , Ligação Proteica , Proteínas de Ligação a Tacrolimo/química , beta Catenina/química
5.
Curr Mol Pharmacol ; 9(2): 109-25, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25986565

RESUMO

Steroid hormone receptors are ligand-dependent transcription factors that require the dynamic, ordered assembly of multimeric chaperone complexes to reach a functional conformation. Heat shock protein (Hsp) 70 and Hsp90 serve as the central chaperones that mediate this process in conjunction with a variety of co-chaperones. Many of these cochaperones represent potential therapeutic targets for the disruption of Hsp90 client protein function. FKBP52 is an Hsp90-associated co-chaperone that has emerged as a promising therapeutic candidate due to its functional specificity for a small subset of Hsp90 client proteins including androgen (AR), glucocorticoid (GR), and progesterone (PR) receptors. Given its Hsp90-client protein specificity, the targeting of FKBP52 should be more specific and less toxic than the Hsp90- targeting drugs. Additionally, the fkbp52-deficient mice display specific phenotypes related to androgen, progesterone, and glucocorticoid insensitivity suggesting minimal off-target effects. Finally, the fact that FKBP52 is already a validated target of the clinically approved immunosuppressive drug, FK506 (Tacrolimus), indicates that FKBP52 is a "druggable" protein. Thus, the development of FKBP52-specific small molecule inhibitors is predicted to be a highly targeted strategy with potential for the treatment of any disease that is dependent on a functional AR, GR, and/or PR signaling pathway. Much progress has been made in understanding the residues and domains critical for FKBP52 function. The proline-rich loop overhanging the FKBP52 FK1 catalytic domain is functionally important and likely represents an interaction surface within the receptor-chaperone complex. Thus, the targeting of FKBP52 proline-rich loop interactions is the most attractive therapeutic approach to disrupt FKBP52 regulation of receptor activity in steroid hormone receptor-dependent physiology and disease.


Assuntos
Proteínas de Choque Térmico HSP90/metabolismo , Neoplasias da Próstata/metabolismo , Receptores Androgênicos/metabolismo , Receptores de Glucocorticoides/metabolismo , Receptores de Progesterona/metabolismo , Proteínas de Ligação a Tacrolimo/metabolismo , Animais , Descoberta de Drogas , Humanos , Masculino , Modelos Moleculares , Terapia de Alvo Molecular , Próstata/efeitos dos fármacos , Próstata/metabolismo , Neoplasias da Próstata/tratamento farmacológico , Proteínas de Ligação a Tacrolimo/química
6.
Int J Biol Sci ; 11(4): 434-47, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25798063

RESUMO

Tetratricopeptide repeat domain 9A (TTC9A) is a target gene of estrogen and progesterone. It is over-expressed in breast cancer. However, little is known about the physiological function of TTC9A. The objectives of this study were to establish a Ttc9a knockout mouse model and to study the consequence of Ttc9a gene inactivation. The Ttc9a targeting vector was generated by replacing the Ttc9a exon 1 with a neomycin cassette. The mice homozygous for Ttc9a exon 1 deletion appear to grow normally and are fertile. However, further characterization of the female mice revealed that Ttc9a deficiency is associated with greater body weight, bigger thymus and better mammary development in post-pubertal mice. Furthermore, Ttc9a deficient mammary gland was more responsive to estrogen treatment with greater mammary ductal lengthening, ductal branching and estrogen target gene induction. Since Ttc9a is induced by estrogen in estrogen target tissues, these results suggest that Ttc9a is a negative regulator of estrogen function through a negative feedback mechanism. This is supported by in vitro evidence that TTC9A over-expression attenuated ERα activity in MCF-7 cells. Although TTC9A does not bind to ERα or its chaperone protein Hsp90 directly, TTC9A strongly interacts with FKBP38 and FKBP51, both of which interact with ERα and Hsp90 and modulate ERα activity. It is plausible therefore that TTC9A negatively regulates ERα activity through interacting with co-chaperone proteins such as FKBP38 and FKBP51.


Assuntos
Receptor alfa de Estrogênio/metabolismo , Proteínas dos Microfilamentos/metabolismo , Animais , Receptor alfa de Estrogênio/genética , Feminino , Humanos , Células MCF-7 , Glândulas Mamárias Humanas/metabolismo , Camundongos , Camundongos Knockout , Proteínas dos Microfilamentos/genética , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Ligação Proteica , Transdução de Sinais , Proteínas de Ligação a Tacrolimo/metabolismo
7.
Subcell Biochem ; 78: 35-68, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25487015

RESUMO

Hsp90 functionally interacts with a broad array of client proteins, but in every case examined Hsp90 is accompanied by one or more co-chaperones. One class of co-chaperone contains a tetratricopeptide repeat domain that targets the co-chaperone to the C-terminal region of Hsp90. Within this class are Hsp90-binding peptidylprolyl isomerases, most of which belong to the FK506-binding protein (FKBP) family. Despite the common association of FKBP co-chaperones with Hsp90, it is now clear that the client protein influences, and is influenced by, the particular FKBP bound to Hsp90. Examples include Xap2 in aryl hydrocarbon receptor complexes and FKBP52 in steroid receptor complexes. In this chapter, we discuss the known functional roles played by FKBP co-chaperones and, where possible, relate distinctive functions to structural differences between FKBP members.


Assuntos
Proteínas de Choque Térmico HSP90/metabolismo , Proteínas de Ligação a Tacrolimo/metabolismo , Animais , Sítios de Ligação , Proteínas de Choque Térmico HSP90/química , Humanos , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Transdução de Sinais , Relação Estrutura-Atividade , Proteínas de Ligação a Tacrolimo/química
8.
Nutr Cancer ; 59(2): 217-27, 2007.
Artigo em Inglês | MEDLINE | ID: mdl-18001217

RESUMO

Barrett's esophagus (BE) is a premalignant lesion in which columnar epithelium (containing goblet cells) replaces esophageal squamous cells. Previous evidence suggested that hydrophobic bile acids and zinc deficiency each play a role in BE development. We fed wild-type C57BL/6 mice a zinc-deficient diet containing the hydrophobic bile acid, deoxycholic acid for various times up to 152 days. All mice fed this diet developed esophagitis by 69 days on the diet and 63% of the mice on this diet for 88 to 152 days also developed a BE-like lesion. Esophageal tissues showed thickened mucosa, increased proliferation, and increased expression of markers associated with oxidative and nitrosative stress. The newly formed BE-like lesions expressed Mucin-2, a marker of columnar differentiation. They also showed translocation of the p65 subunit of nuclear factor-kappaB and beta -catenin to the nucleus and typical histological changes associated with BE lesions. This mouse model of esophagitis and BE is expected to contribute to a deeper understanding of BE pathogenesis and to strategies for prevention of BE progression to cancer.


Assuntos
Esôfago de Barrett/patologia , Ácido Desoxicólico/farmacologia , Dieta , Esofagite/patologia , Zinco/deficiência , Animais , Esôfago de Barrett/etiologia , Divisão Celular/efeitos dos fármacos , Modelos Animais de Doenças , Progressão da Doença , Esofagite/etiologia , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Estresse Oxidativo , Zinco/administração & dosagem
9.
Gut ; 56(6): 763-71, 2007 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-17145738

RESUMO

BACKGROUND: Barrett's oesophagus is a premalignant condition associated with an increased risk for the development of oesophageal adenocarcinoma (ADCA). Previous studies indicated that oxidative damage contributes to the development of ADCA. OBJECTIVE: To test the hypothesis that bile acids and gastric acid, two components of refluxate, can induce oxidative stress and oxidative DNA damage. METHODS: Oxidative stress was evaluated by staining Barrett's oesophagus tissues with different degrees of dysplasia with 8-hydroxy-deoxyguanosine (8-OH-dG) antibody. The levels of 8-OH-dG were also evaluated ex vivo in Barrett's oesophagus tissues incubated for 10 min with control medium and medium acidified to pH 4 and supplemented with 0.5 mM bile acid cocktail. Furthermore, three oesophageal cell lines (Seg-1 cells, Barrett's oesophagus cells and HET-1A cells) were exposed to control media, media containing 0.1 mM bile acid cocktail, media acidified to pH 4, and media at pH 4 supplemented with 0.1 mM bile acid cocktail, and evaluated for induction of reactive oxygen species (ROS). RESULTS: Immunohistochemical analysis showed that 8-OH-dG is formed mainly in the epithelial cells in dysplastic Barrett's oesophagus. Importantly, incubation of Barrett's oesophagus tissues with the combination of bile acid cocktail and acid leads to increased formation of 8-OH-dG. An increase in ROS in oesophageal cells was detected after exposure to pH 4 and bile acid cocktail. CONCLUSIONS: Oxidative stress and oxidative DNA damage can be induced in oesophageal tissues and cells by short exposures to bile acids and low pH. These alterations may underlie the development of Barrett's oesophagus and tumour progression.


Assuntos
Esôfago de Barrett/metabolismo , Ácidos e Sais Biliares/fisiologia , Dano ao DNA , Estresse Oxidativo , 8-Hidroxi-2'-Desoxiguanosina , Adulto , Idoso , Idoso de 80 Anos ou mais , Apoptose/efeitos dos fármacos , Esôfago de Barrett/genética , Esôfago de Barrett/patologia , Ácidos e Sais Biliares/farmacologia , Biópsia , Meios de Cultura , Desoxiguanosina/análogos & derivados , Desoxiguanosina/metabolismo , Progressão da Doença , Esôfago/efeitos dos fármacos , Esôfago/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Microscopia de Fluorescência , Pessoa de Meia-Idade , Estresse Oxidativo/efeitos dos fármacos , Células Tumorais Cultivadas
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