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1.
Sci Rep ; 14(1): 18283, 2024 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-39112499

RESUMO

Renal fibrosis (RF) represents the most widespread pathological condition in chronic kidney disease (CKD). Recently, protein prenylation has been implicated in the fibrosis's progression. The research examined the renoprotective effect of zoledronic acid (ZA) (50 µg/kg/week) in a rat model of carbon tetrachloride (CCl4)-induced RF through targeting protein prenylation. Forty Wistar male rats were split up into the control group, vehicle-treated group, model-RF group, and RF-ZA group. Mean arterial blood pressure (MBP), BUN, serum creatinine, and urine albumin-creatinine ratio (uACR), protein levels of farnesyl pyrophosphate (FPP), tumour necrosis factor-alpha (TNF-α), transforming growth factor-ß (TGF-ß), and malondialdehyde (MDA), and catalase and gene expression of farnesyl pyrophosphate synthase (FPPS) and nuclear factor-kB (NF-κB) were measured. Immunohistochemical staining for renal interleukin-6 (IL-6), α-smooth muscle actin (α-SMA), and caspase-3, as well as histopathological alterations, were assessed. ZA considerably ceased the reduction in MBP, markedly reduced uACR, serum creatinine, BUN, and expression of FPPS, FPP, NF-κB, TGF-ß, TNF-α, and MDA, and significantly increased catalase levels compared to the model-RF rats. ZA ameliorated the CCl4-induced histopathological alterations and suppressed the expression of caspase-3, α-SMA, and IL-6. In conclusion, ZA preserved renal function and prevented renal fibrosis in a rat model. These were achieved through targeting protein prenylation mainly by inhibiting FPPS.


Assuntos
Fibrose , Geraniltranstransferase , Rim , Prenilação de Proteína , Ratos Wistar , Ácido Zoledrônico , Animais , Ácido Zoledrônico/farmacologia , Masculino , Ratos , Prenilação de Proteína/efeitos dos fármacos , Geraniltranstransferase/metabolismo , Rim/efeitos dos fármacos , Rim/patologia , Rim/metabolismo , Tetracloreto de Carbono , Fosfatos de Poli-Isoprenil/metabolismo , Fosfatos de Poli-Isoprenil/farmacologia , Modelos Animais de Doenças , Fator de Crescimento Transformador beta/metabolismo , Sesquiterpenos/farmacologia , Sesquiterpenos/uso terapêutico , Fator de Necrose Tumoral alfa/metabolismo
2.
J Biol Chem ; 299(11): 105269, 2023 11.
Artigo em Inglês | MEDLINE | ID: mdl-37739036

RESUMO

Prenylation is an irreversible post-translational modification that supports membrane interactions of proteins involved in various cellular processes, including migration, proliferation, and survival. Dysregulation of prenylation contributes to multiple disorders, including cancers and vascular and neurodegenerative diseases. Prenyltransferases tether isoprenoid lipids to proteins via a thioether linkage during prenylation. Pharmacological inhibition of the lipid synthesis pathway by statins is a therapeutic approach to control hyperlipidemia. Building on our previous finding that statins inhibit membrane association of G protein γ (Gγ) in a subtype-dependent manner, we investigated the molecular reasoning for this differential inhibition. We examined the prenylation of carboxy-terminus (Ct) mutated Gγ in cells exposed to Fluvastatin and prenyl transferase inhibitors and monitored the subcellular localization of fluorescently tagged Gγ subunits and their mutants using live-cell confocal imaging. Reversible optogenetic unmasking-masking of Ct residues was used to probe their contribution to prenylation and membrane interactions of the prenylated proteins. Our findings suggest that specific Ct residues regulate membrane interactions of the Gγ polypeptide, statin sensitivity, and extent of prenylation. Our results also show a few hydrophobic and charged residues at the Ct are crucial determinants of a protein's prenylation ability, especially under suboptimal conditions. Given the cell and tissue-specific expression of different Gγ subtypes, our findings indicate a plausible mechanism allowing for statins to differentially perturb heterotrimeric G protein signaling in cells depending on their Gγ-subtype composition. Our results may also provide molecular reasoning for repurposing statins as Ras oncogene inhibitors and the failure of using prenyltransferase inhibitors in cancer treatment.


Assuntos
Proteínas Heterotriméricas de Ligação ao GTP , Prenilação de Proteína , Humanos , Motivos de Aminoácidos , Resistência a Medicamentos/genética , Células HeLa , Proteínas Heterotriméricas de Ligação ao GTP/química , Proteínas Heterotriméricas de Ligação ao GTP/genética , Proteínas Heterotriméricas de Ligação ao GTP/metabolismo , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Modelos Moleculares , Mutação , Prenilação de Proteína/efeitos dos fármacos , Estrutura Terciária de Proteína , Transporte Proteico/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
3.
Sci Rep ; 11(1): 17990, 2021 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-34504210

RESUMO

Macrothrombocytopenia is a common pathology of missense mutations in genes regulating actin dynamics. Takenouchi-Kosaki syndrome (TKS) harboring the c.191A > G, Tyr64Cys (Y64C) variant in Cdc42 exhibits a variety of clinical manifestations, including immunological and hematological anomalies. In the present study, we investigated the functional abnormalities of the Y64C mutant in HEK293 cells and elucidated the mechanism of macrothrombocytopenia, one of the symptoms of TKS patients, by monitoring the production of platelet-like particles (PLP) using MEG-01 cells. We found that the Y64C mutant was concentrated at the membrane compartment due to impaired binding to Rho-GDI and more active than the wild-type. The Y64C mutant also had lower association with its effectors Pak1/2 and N-WASP. Y64C mutant-expressing MEG-01 cells demonstrated short cytoplasmic protrusions with aberrant F-actin and microtubules, and reduced PLP production. This suggested that the Y64C mutant facilitates its activity and membrane localization, resulting in impaired F-actin dynamics for proplatelet extension, which is necessary for platelet production. Furthermore, such dysfunction was ameliorated by either suppression of Cdc42 activity or prenylation using chemical inhibitors. Our study may lead to pharmacological treatments for TKS patients.


Assuntos
Megacariócitos/efeitos dos fármacos , Megacariócitos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Trombocitopenia/metabolismo , Proteína cdc42 de Ligação ao GTP/antagonistas & inibidores , Proteína cdc42 de Ligação ao GTP/metabolismo , Actinas/metabolismo , Alquil e Aril Transferases/antagonistas & inibidores , Benzamidas/farmacologia , Plaquetas/metabolismo , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Células HEK293 , Humanos , Mutação , Prenilação de Proteína/efeitos dos fármacos , Pirazóis/farmacologia , Transdução de Sinais/genética , Sulfonamidas/farmacologia , Síndrome , Trombocitopenia/genética , Trombopoese/efeitos dos fármacos , Trombopoese/genética , Transfecção , Proteína Neuronal da Síndrome de Wiskott-Aldrich/metabolismo , Proteína cdc42 de Ligação ao GTP/genética , Quinases Ativadas por p21/metabolismo , Inibidor alfa de Dissociação do Nucleotídeo Guanina rho/metabolismo
4.
Arch Immunol Ther Exp (Warsz) ; 69(1): 10, 2021 Apr 03.
Artigo em Inglês | MEDLINE | ID: mdl-33811524

RESUMO

The review discusses a new approach to the prevention and treatment of viral infections based on the use of pine needles polyprenyl phosphate (PPP) and associated with the infringement of prenylation process-the attachment of farnesol or geranyl geraniol to the viral protein. Currently, prenylation has been detected in type 1 adenovirus, hepatitis C virus, several herpes viruses, influenza virus, HIV. However, this list is far from complete, given that prenylated proteins play an extremely important role in the activity of the virus. We assume that the interferon produced in response to PPP may suppress expression of the SREBP2 transcription factor. As a result, the mevalonic acid pathway is violated and, as a result, the formation of early polyprenols precursors (geraniol, geranyl geraniol, farnesol), which are necessary for the prenylation of viral proteins, is blocked and the formation of mature, virulent virus particles is broken. As a consequence, the maturation of viral particles is inhibited, and defective particles are formed. Polyprenol was extracted from greenery (pine, fir and spruce needles, mulberry leaves, etc.), purified by chromatography, phosphorylated and identified by HPLC and NMR. Obtained PPP was used as antiviral in some experimental models in vitro and in vivo. During numerous studies, it was found that PPP manifested versatile antiviral effects, both in vitro and in vivo. The maximum effect was observed with viruses in which the presence of prenylated proteins was established, namely influenza A virus, HIV-1, tick-borne encephalitis virus, hepatitis A and C viruses, herpes simplex viruses type 1 and 2, some coronavirus. The available data obtained both in the experimental conditions and during clinical trials allow us to regard PPPs as safe and effective medicine for prevention and treatment of viral diseases.


Assuntos
Antivirais/farmacologia , Pinus/química , Fosfatos de Poli-Isoprenil/farmacologia , Prenilação de Proteína/efeitos dos fármacos , Viroses/tratamento farmacológico , Animais , Antivirais/uso terapêutico , Ensaios Clínicos como Assunto , Modelos Animais de Doenças , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/imunologia , Humanos , Interferons/metabolismo , Microscopia Eletrônica , Fosfatos de Poli-Isoprenil/uso terapêutico , Proteína de Ligação a Elemento Regulador de Esterol 2/metabolismo , Resultado do Tratamento , Proteínas Virais/metabolismo , Vírion/efeitos dos fármacos , Vírion/ultraestrutura , Viroses/imunologia , Viroses/prevenção & controle , Replicação Viral/efeitos dos fármacos , Replicação Viral/imunologia
5.
Pharmacol Res ; 167: 105528, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33667685

RESUMO

Geranylgeranyl diphosphate synthase (GGDPS), an enzyme in the isoprenoid biosynthetic pathway (IBP), produces the isoprenoid (geranylgeranyl pyrophosphate, GGPP) used in protein geranylgeranylation reactions. Our prior studies utilizing triazole bisphosphonate-based GGDPS inhibitors (GGSIs) have revealed that these agents represent a novel strategy by which to induce cancer cell death, including multiple myeloma and pancreatic cancer. Statins inhibit the rate-limiting enzyme in the IBP and potentiate the effects of GGSIs in vitro. The in vivo effects of combination therapy with statins and GGSIs have not been determined. Here we evaluated the effects of combining VSW1198, a novel GGSI, with a statin (lovastatin or pravastatin) in CD-1 mice. Twice-weekly dosing with VSW1198 at the previously established maximally tolerated dose in combination with a statin led to hepatotoxicity, while once-weekly VSW1198-based combinations were feasible. No abnormalities in kidney, spleen, brain or skeletal muscle were observed with combination therapy. Combination therapy disrupted protein geranylgeranylation in vivo. Evaluation of hepatic isoprenoid levels revealed decreased GGPP levels in the single drug groups and undetectable GGPP levels in the combination groups. Additional studies with combinations using 50% dose-reductions of either VSW1198 or lovastatin revealed minimal hepatotoxicity with expected on-target effects of diminished GGPP levels and disruption of protein geranylgeranylation. Combination statin/GGSI therapy significantly slowed tumor growth in a myeloma xenograft model. Collectively, these studies are the first to demonstrate that combination IBP inhibitor therapy alters isoprenoid levels and disrupts protein geranylgeranylation in vivo as well as slows tumor growth in a myeloma xenograft model, thus providing the framework for future clinical exploration.


Assuntos
Vias Biossintéticas/efeitos dos fármacos , Diterpenos/administração & dosagem , Sistemas de Liberação de Medicamentos/métodos , Inibidores de Hidroximetilglutaril-CoA Redutases/administração & dosagem , Prenilação de Proteína/efeitos dos fármacos , Terpenos/metabolismo , Triazóis/administração & dosagem , Animais , Vias Biossintéticas/fisiologia , Linhagem Celular Tumoral , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/patologia , Diterpenos/toxicidade , Avaliação Pré-Clínica de Medicamentos/métodos , Quimioterapia Combinada , Inibidores Enzimáticos/administração & dosagem , Inibidores Enzimáticos/toxicidade , Farnesiltranstransferase/antagonistas & inibidores , Farnesiltranstransferase/metabolismo , Feminino , Inibidores de Hidroximetilglutaril-CoA Redutases/toxicidade , Lovastatina/administração & dosagem , Lovastatina/toxicidade , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Pravastatina/administração & dosagem , Pravastatina/toxicidade , Prenilação de Proteína/fisiologia , Terpenos/antagonistas & inibidores , Triazóis/toxicidade , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
6.
ACS Chem Biol ; 15(7): 1930-1941, 2020 07 17.
Artigo em Inglês | MEDLINE | ID: mdl-32573189

RESUMO

Fungal pathogens pose an increasing threat to global food security through devastating effects on staple crops and contamination of food supplies with carcinogenic toxins. Widespread deployment of agricultural fungicides has increased crop yields but is driving increasingly frequent resistance to available agents and creating environmental reservoirs of drug-resistant fungi that can also infect susceptible human populations. To uncover non-cross-resistant modes of antifungal action, we leveraged the unique chemical properties of boron chemistry to synthesize novel 6-thiocarbamate benzoxaboroles with broad spectrum activity against diverse fungal plant pathogens. Through whole genome sequencing of Saccharomyces cerevisiae isolates selected for stable resistance to these compounds, we identified mutations in the protein prenylation-related genes, CDC43 and ERG20. Allele-swapping experiments confirmed that point mutations in CDC43, which encodes an essential catalytic subunit within geranylgeranyl transferase I (GGTase I) complex, were sufficient to confer resistance to the benzoxaboroles. Mutations in ERG20, which encodes an upstream farnesyl pyrophosphate synthase in the geranylgeranylation pathway, also conferred resistance. Consistent with impairment of protein prenylation, the compounds disrupted membrane localization of the classical geranylgeranylation substrate Cdc42. Guided by molecular docking predictions, which favored Cdc43 as the most likely direct target, we overexpressed and purified functional GGTase I complex to demonstrate direct binding of benzoxaboroles to it and concentration-dependent inhibition of its transferase activity. Further development of the boron-containing scaffold described here offers a promising path to the development of GGTase I inhibitors as a mechanistically distinct broad spectrum fungicide class with reduced potential for cross-resistance to antifungals in current use.


Assuntos
Antifúngicos/farmacologia , Compostos de Boro/farmacologia , Prenilação de Proteína/efeitos dos fármacos , Proteínas de Saccharomyces cerevisiae/metabolismo , Tiocarbamatos/farmacologia , Alquil e Aril Transferases/antagonistas & inibidores , Alquil e Aril Transferases/genética , Alquil e Aril Transferases/metabolismo , Antifúngicos/síntese química , Antifúngicos/metabolismo , Compostos de Boro/síntese química , Compostos de Boro/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Membrana Celular/efeitos dos fármacos , Dimetilaliltranstransferase/genética , Dimetilaliltranstransferase/metabolismo , Farmacorresistência Fúngica/genética , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacologia , Fungos/efeitos dos fármacos , Fungos/genética , Simulação de Acoplamento Molecular , Mutação Puntual , Ligação Proteica , Proteínas de Saccharomyces cerevisiae/genética , Tiocarbamatos/síntese química , Tiocarbamatos/metabolismo , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/metabolismo
7.
J Intern Med ; 288(2): 183-191, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32176377

RESUMO

The RAS genes, which include H, N, and KRAS, comprise the most frequently mutated family of oncogenes in cancer. Mutations in KRAS - such as the G12C mutation - are found in most pancreatic, half of colorectal and a third of lung cancer cases and is thus responsible for a substantial proportion of cancer deaths. Consequently, KRAS has been the subject of exhaustive drug-targeting efforts over the past 3-4 decades. These efforts have included targeting the KRAS protein itself but also its posttranslational modifications, membrane localization, protein-protein interactions and downstream signalling pathways. Most of these strategies have failed and no KRAS-specific drugs have yet been approved. However, for one specific mutation, KRASG12C , there is light on the horizon. MRTX849 was recently identified as a potent, selective and covalent KRASG12C inhibitor that possesses favourable drug-like properties. MRTX849 selectively modifies the mutant cysteine residue in GDP-bound KRASG12C and inhibits GTP-loading and downstream KRAS-dependent signalling. The drug inhibits the in vivo growth of multiple KRASG12C -mutant cell line xenografts, causes tumour regression in patient-derived xenograft models and shows striking responses in combination with other agents. It has also produced objective responses in patients with mutant-specific lung and colorectal cancer. In this review, we discuss the history of RAS drug-targeting efforts, the discovery of MRTX849, and how this drug provides an exciting and long-awaited opportunity to selectively target mutant KRAS in patients.


Assuntos
Antineoplásicos/farmacologia , Neoplasias/tratamento farmacológico , Neoplasias/genética , Proteínas Proto-Oncogênicas p21(ras)/antagonistas & inibidores , Proteínas Proto-Oncogênicas p21(ras)/genética , Ensaios Clínicos como Assunto , Inibidores Enzimáticos/farmacologia , Humanos , Mutação , Prenilação de Proteína/efeitos dos fármacos , Proteínas Proto-Oncogênicas p21(ras)/metabolismo
8.
Fundam Clin Pharmacol ; 34(1): 32-40, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31058344

RESUMO

Aberrant activation of GTPases is common in cervical cancer, and their proper biological functions largely depend on a post-translational modification termed prenylation. Simvastatin is a cholesterol-lowering drug via inhibiting HMG-CoA reductase, thereby inhibiting protein prenylation. In this study, we show that simvastatin selectively inhibits proliferation and induces apoptosis in cervical cancer cells while sparing normal cervical epithelial cells. This is achieved by depleting geranylgeranyl pyrophosphate, inhibiting prenylation, decreasing GTPases activities and suppressing the activation of downstream Ras and RhoA signaling. The combination of simvastatin and paclitaxel remarkably augments in vitro as well as in vivo efficacy of either drug alone in cellular system and xenograft mouse model. Importantly, we show that cervical cancer cells have higher level of HMG-CoA reductase and elevated activities of GTPases, suggesting that cervical cancer cells may be more dependent on prenylation than normal cervical epithelial cells. This might explain the selective inhibitory effects of simvastatin in cervical cancer. Since simvastatin is already available for clinic use, these results suggest that simvastatin is a promising drug candidate in combination with chemotherapy for the treatment of cervical cancer. Our findings also emphasize the therapeutic value of prenylation inhibition and provide preclinical evidence to evaluate prenylation-targeted drugs in cervical cancer.


Assuntos
GTP Fosfo-Hidrolases/metabolismo , Paclitaxel/farmacologia , Sinvastatina/farmacologia , Neoplasias do Colo do Útero/tratamento farmacológico , Animais , Antineoplásicos Fitogênicos/administração & dosagem , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Hidroximetilglutaril-CoA Redutases/efeitos dos fármacos , Hidroximetilglutaril-CoA Redutases/metabolismo , Inibidores de Hidroximetilglutaril-CoA Redutases/administração & dosagem , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Camundongos SCID , Paclitaxel/administração & dosagem , Fosfatos de Poli-Isoprenil/metabolismo , Prenilação de Proteína/efeitos dos fármacos , Sinvastatina/administração & dosagem , Neoplasias do Colo do Útero/enzimologia , Neoplasias do Colo do Útero/patologia , Ensaios Antitumorais Modelo de Xenoenxerto
9.
Bioorg Med Chem Lett ; 29(24): 126757, 2019 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-31699606

RESUMO

Benzimidazole carboxyphosphonates and bisphosphonates have been prepared and evaluated for their activity as inhibitors of protein prenylation or isoprenoid biosynthesis. The nature of the phosphonate head group was found to dictate enzyme specificity. The lead carboxyphosphonate inhibits geranylgeranyl transferase II while its corresponding bisphosphonate analogue potently inhibits farnesyl diphosphate synthase. The most active inhibitors effectively disrupted protein prenylation in human multiple myeloma cells.


Assuntos
Benzimidazóis/antagonistas & inibidores , Benzimidazóis/uso terapêutico , Organofosfonatos/antagonistas & inibidores , Organofosfonatos/uso terapêutico , Prenilação de Proteína/efeitos dos fármacos , Benzimidazóis/farmacologia , Humanos , Organofosfonatos/farmacologia
10.
Biochem Biophys Res Commun ; 517(3): 439-444, 2019 09 24.
Artigo em Inglês | MEDLINE | ID: mdl-31376932

RESUMO

The clinical management of pediatric acute myeloid leukemia (AML) is still challenging and identification of drugs that can enhance the efficacy of standard of care is a potential therapeutic strategy. We show that pamidronate, a FDA-approved drug used for bone disorders, is an attractive candidate for AML treatment. Pamidronate inhibits proliferation and induces apoptosis of AML cells regardless of cellular and genetic heterogeneity. Pamidronate displays selective anti-AML activity by preferentially inhibiting survival and colony formation of AML CD34+ cells while normal bone marrow CD34+ cells are largely unaffected. Importantly, pamidronate remarkably enhances the inhibitory effects of all tested AML standard of care at subtoxic concentration. Mechanism studies show that pamidronate inhibits protein prenylation via dual action on geranylgeranylation and farnesylation, and subsequently decreases Ras activity. The rescue studies using overexpression of constitutively active Ras further confirm that pamidronate augments the efficacy of AML standard of care through inhibiting Ras. Since pamidronate is already used in clinic, our preclinical findings suggest that it may be an effective addition to treatment armamentarium for AML.


Assuntos
Antineoplásicos/farmacologia , Células-Tronco Neoplásicas/efeitos dos fármacos , Pamidronato/farmacologia , Prenilação de Proteína/efeitos dos fármacos , Processamento de Proteína Pós-Traducional , Proteínas ras/antagonistas & inibidores , Anilidas/farmacologia , Anexina A5/genética , Anexina A5/metabolismo , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/genética , Benzotiazóis/farmacologia , Conservadores da Densidade Óssea/farmacologia , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Criança , Citarabina/farmacologia , Daunorrubicina/farmacologia , Reposicionamento de Medicamentos , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/patologia , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Compostos de Fenilureia/farmacologia , Cultura Primária de Células , Piridinas/farmacologia , Proteínas ras/genética , Proteínas ras/metabolismo
11.
Nat Chem ; 11(6): 552-561, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30936521

RESUMO

Post-translational farnesylation or geranylgeranylation at a C-terminal cysteine residue regulates the localization and function of over 100 proteins, including the Ras isoforms, and is a therapeutic target in diseases including cancer and infection. Here, we report global and selective profiling of prenylated proteins in living cells enabled by the development of isoprenoid analogues YnF and YnGG in combination with quantitative chemical proteomics. Eighty prenylated proteins were identified in a single human cell line, 64 for the first time at endogenous abundance without metabolic perturbation. We further demonstrate that YnF and YnGG enable direct identification of post-translationally processed prenylated peptides, proteome-wide quantitative analysis of prenylation dynamics and alternative prenylation in response to four different prenyltransferase inhibitors, and quantification of defective Rab prenylation in a model of the retinal degenerative disease choroideremia.


Assuntos
Alcinos/química , Sondas Moleculares/química , Prenilação de Proteína , Proteínas/análise , Proteoma/análise , Proteômica/métodos , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Linhagem Celular , Técnicas de Inativação de Genes , Humanos , Espectrometria de Massas , Camundongos Knockout , Prenilação de Proteína/efeitos dos fármacos , Proteínas/química , Proteoma/química
12.
Expert Opin Ther Pat ; 29(5): 315-325, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31023104

RESUMO

INTRODUCTION: Bisphosphonates (BPs) are widely used to manage a variety of bone disorders, including osteoporosis, metastatic bone disease and myeloma bone disease. The nitrogen-containing BPs (NBPs) target osteoclast activity by disrupting protein prenylation via inhibition of farnesyl diphosphate synthase (FDPS). AREAS COVERED: This review summarizes the recent advances in BPs with a focus on the latest patents (2015-2018). Patents involving novel BPs, new modes of BP delivery, as well as use of BPs to deliver other drugs to bone are discussed. A review of phosphonate-based drugs targeting geranylgeranyl diphosphate synthase (GGDPS) or geranylgeranyl transferase II (GGTase II) as alternative strategies to disrupt protein geranylgeranylation is provided. EXPERT OPINION: While the NBPs remain the mainstay of treatment for most bone disorders, further understanding of their pharmacological properties could lead to further refinement of their chemical structures and optimization of efficacy and safety profiles. In addition, the development of NBP analogs or drug delivery mechanisms that allow for nonbone tissue exposure could allow for the use of these drugs as direct anticancer agents. The development of GGDPS and GGTase II inhibitors represents alternative heterocycle phosphonate-based strategies to disrupt protein geranylgeranylation and may have potential as anticancer agents and/or as bone-targeted therapies.


Assuntos
Doenças Ósseas/tratamento farmacológico , Difosfonatos/uso terapêutico , Prenilação de Proteína/efeitos dos fármacos , Animais , Doenças Ósseas/fisiopatologia , Difosfonatos/farmacologia , Sistemas de Liberação de Medicamentos , Desenvolvimento de Medicamentos , Farnesiltranstransferase/antagonistas & inibidores , Geraniltranstransferase/antagonistas & inibidores , Humanos , Osteoclastos/efeitos dos fármacos , Patentes como Assunto , Transferases/antagonistas & inibidores
13.
Mol Pharmacol ; 95(4): 361-375, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30765461

RESUMO

Guanine nucleotide-binding proteins (G proteins) facilitate the transduction of external signals to the cell interior, regulate most eukaryotic signaling, and thus have become crucial disease drivers. G proteins largely function at the inner leaflet of the plasma membrane (PM) using covalently attached lipid anchors. Both small monomeric and heterotrimeric G proteins are primarily prenylated, either with a 15-carbon farnesyl or a 20-carbon geranylgeranyl polyunsaturated lipid. The mevalonate [3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase] pathway synthesizes lipids for G-protein prenylation. It is also the source of the precursor lipids for many biomolecules, including cholesterol. Consequently, the rate-limiting enzymes of the mevalonate pathway are major targets for cholesterol-lowering medications and anticancer drug development. Although prenylated G protein γ (Gγ) is essential for G protein-coupled receptor (GPCR)-mediated signaling, how mevalonate pathway inhibitors, statins, influence subcellular distribution of Gßγ dimer and Gαßγ heterotrimer, as well as their signaling upon GPCR activation, is poorly understood. The present study shows that clinically used statins not only significantly disrupt PM localization of Gßγ but also perturb GPCR-G protein signaling and associated cell behaviors. The results also demonstrate that the efficiency of prenylation inhibition by statins is Gγ subtype-dependent and is more effective toward farnesylated Gγ types. Since Gγ is required for Gßγ signaling and shows a cell- and tissue-specific subtype distribution, the present study can help understand the mechanisms underlying clinical outcomes of statin use in patients. This work also reveals the potential of statins as clinically usable drugs to control selected GPCR-G protein signaling.


Assuntos
Subunidades beta da Proteína de Ligação ao GTP/metabolismo , Subunidades gama da Proteína de Ligação ao GTP/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Receptores Acoplados a Proteínas G/metabolismo , Animais , Linhagem Celular , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Colesterol/metabolismo , Células HeLa , Humanos , Ácido Mevalônico/farmacologia , Camundongos , Prenilação de Proteína/efeitos dos fármacos , Células RAW 264.7 , Transdução de Sinais/efeitos dos fármacos
14.
Crit Rev Biochem Mol Biol ; 54(1): 41-60, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30773935

RESUMO

Dysregulation of isoprenoid biosynthesis is implicated in numerous biochemical disorders that play a role in the onset and/or progression of age-related diseases, such as hypercholesterolemia, osteoporosis, various cancers, and neurodegeneration. The mevalonate metabolic pathway is responsible for the biosynthesis of the two key isoprenoid metabolites, farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP). Post-translational prenylation of various proteins, including the small GTP-binding proteins (GTPases), with either FPP or GGPP is vital for proper localization and activation of these proteins. Prenylated GTPases play a critical role in cell signaling, proliferation, cellular plasticity, oncogenesis, and cancer metastasis. Pre-clinical and clinical studies strongly suggest that inhibition of protein prenylation can be an effective treatment for non-skeletal cancers. In this review, we summarize the most recent drug discovery efforts focusing on blocking protein farnesylation and/or geranylgeranylation and the biochemical and structural data available in guiding the current on-going studies in drug discovery. Furthermore, we provide a summary on the biochemical association between disruption of protein prenylation, endoplasmic reticulum (ER) stress, unfolded protein response (UPR) signaling, and cancer.


Assuntos
Vias Biossintéticas/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Farnesiltranstransferase/antagonistas & inibidores , Geraniltranstransferase/antagonistas & inibidores , Neoplasias/tratamento farmacológico , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Descoberta de Drogas , Inibidores Enzimáticos/uso terapêutico , Farnesiltranstransferase/metabolismo , Geraniltranstransferase/metabolismo , Humanos , Ácido Mevalônico/metabolismo , Modelos Moleculares , Neoplasias/metabolismo , Fosfatos de Poli-Isoprenil/antagonistas & inibidores , Fosfatos de Poli-Isoprenil/metabolismo , Prenilação de Proteína/efeitos dos fármacos , Sesquiterpenos/antagonistas & inibidores , Sesquiterpenos/metabolismo
15.
J Med Chem ; 61(15): 6904-6917, 2018 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-30016091

RESUMO

Post-translational prenylation of the small GTP-binding proteins (GTPases) is vital to a plethora of biological processes, including cellular proliferation. We have identified a new class of thienopyrimidine-based bisphosphonate (ThP-BP) inhibitors of the human geranylgeranyl pyrophosphate synthase (hGGPPS) that block protein prenylation in multiple myeloma (MM) cells leading to cellular apoptosis. These inhibitors are also effective in blocking the proliferation of other types of cancer cells. We confirmed intracellular target engagement, demonstrated the mechanism of action leading to apoptosis, and determined a direct correlation between apoptosis and intracellular inhibition of hGGPPS. Administration of a ThP-BP inhibitor to a MM mouse model confirmed in vivo downregulation of Rap1A geranylgeranylation and reduction of monoclonal immunoglobulins (M-protein, a biomarker of disease burden) in the serum. These results provide the first proof-of-principle that hGGPPS is a valuable therapeutic target in oncology and more specifically for the treatment of multiple myeloma.


Assuntos
Inibidores Enzimáticos/farmacologia , Geranil-Geranildifosfato Geranil-Geraniltransferase/antagonistas & inibidores , Mieloma Múltiplo/patologia , Prenilação de Proteína/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Domínio Catalítico , Proliferação de Células/efeitos dos fármacos , Inibidores Enzimáticos/química , Geranil-Geranildifosfato Geranil-Geraniltransferase/química , Geranil-Geranildifosfato Geranil-Geraniltransferase/metabolismo , Humanos , Concentração Inibidora 50 , Modelos Moleculares , Pirimidinas/química , Pirimidinas/farmacologia , Proteínas rap1 de Ligação ao GTP/metabolismo
16.
J Bone Miner Res ; 33(11): 2059-2070, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30001457

RESUMO

Patients with Hutchinson-Gilford progeria syndrome (HGPS) have low bone mass and an atypical skeletal geometry that manifests in a high risk of fractures. Using both in vitro and in vivo models of HGPS, we demonstrate that defects in the canonical WNT/ß-catenin pathway, seemingly at the level of the efficiency of nuclear import of ß-catenin, impair osteoblast differentiation and that restoring ß-catenin activity rescues osteoblast differentiation and significantly improves bone mass. Specifically, we show that HGPS patient-derived iPSCs display defects in osteoblast differentiation, characterized by a decreased alkaline phosphatase activity and mineralizing capacity. We demonstrate that the canonical WNT/ß-catenin pathway, a major signaling cascade involved in skeletal homeostasis, is impaired by progerin, causing a reduction in the active ß-catenin in the nucleus and thus decreased transcriptional activity, and its reciprocal cytoplasmic accumulation. Blocking farnesylation of progerin restores active ß-catenin accumulation in the nucleus, increasing signaling, and ameliorates the defective osteogenesis. Moreover, in vivo analysis of the Zmpste24-/- HGPS mouse model demonstrates that treatment with a sclerostin-neutralizing antibody (SclAb), which targets an antagonist of canonical WNT/ß-catenin signaling pathway, fully rescues the low bone mass phenotype to wild-type levels. Together, this study reveals that the ß-catenin signaling cascade is a therapeutic target for restoring defective skeletal microarchitecture in HGPS. © 2018 American Society for Bone and Mineral Research.


Assuntos
Doenças Ósseas Metabólicas/complicações , Doenças Ósseas Metabólicas/metabolismo , Diferenciação Celular , Osteoblastos/metabolismo , Progéria/complicações , Progéria/metabolismo , Transdução de Sinais , beta Catenina/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Animais , Anticorpos Neutralizantes/farmacologia , Doenças Ósseas Metabólicas/patologia , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Modelos Animais de Doenças , Glicoproteínas/imunologia , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular , Lamina Tipo A/metabolismo , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Camundongos , Modelos Biológicos , Mutação/genética , Osteoblastos/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Fenótipo , Progéria/genética , Progéria/patologia , Prenilação de Proteína/efeitos dos fármacos , Via de Sinalização Wnt/efeitos dos fármacos
17.
Cancer Res ; 78(5): 1347-1357, 2018 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-29229608

RESUMO

The statin family of drugs preferentially triggers tumor cell apoptosis by depleting mevalonate pathway metabolites farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP), which are used for protein prenylation, including the oncoproteins of the RAS superfamily. However, accumulating data indicate that activation of the RAS superfamily are poor biomarkers of statin sensitivity, and the mechanism of statin-induced tumor-specific apoptosis remains unclear. Here we demonstrate that cancer cell death triggered by statins can be uncoupled from prenylation of the RAS superfamily of oncoproteins. Ectopic expression of different members of the RAS superfamily did not uniformly sensitize cells to fluvastatin, indicating that increased cellular demand for protein prenylation cannot explain increased statin sensitivity. Although ectopic expression of HRAS increased statin sensitivity, expression of myristoylated HRAS did not rescue this effect. HRAS-induced epithelial-to-mesenchymal transition (EMT) through activation of zinc finger E-box binding homeobox 1 (ZEB1) sensitized tumor cells to the antiproliferative activity of statins, and induction of EMT by ZEB1 was sufficient to phenocopy the increase in fluvastatin sensitivity; knocking out ZEB1 reversed this effect. Publicly available gene expression and statin sensitivity data indicated that enrichment of EMT features was associated with increased sensitivity to statins in a large panel of cancer cell lines across multiple cancer types. These results indicate that the anticancer effect of statins is independent from prenylation of RAS family proteins and is associated with a cancer cell EMT phenotype.Significance: The use of statins to target cancer cell EMT may be useful as a therapy to block cancer progression. Cancer Res; 78(5); 1347-57. ©2017 AACR.


Assuntos
Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Fluvastatina/farmacologia , Neoplasias/patologia , Prenilação de Proteína/efeitos dos fármacos , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo , Proteínas ras/metabolismo , Apoptose , Biomarcadores Tumorais , Proliferação de Células , Humanos , Ácido Mevalônico/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Fosfatos de Poli-Isoprenil/metabolismo , Sesquiterpenos/metabolismo , Células Tumorais Cultivadas , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética , Proteínas ras/genética
18.
J Biol Chem ; 293(8): 2770-2785, 2018 02 23.
Artigo em Inglês | MEDLINE | ID: mdl-29282289

RESUMO

Protein prenylation is a post-translational modification that has been most commonly associated with enabling protein trafficking to and interaction with cellular membranes. In this process, an isoprenoid group is attached to a cysteine near the C terminus of a substrate protein by protein farnesyltransferase (FTase) or protein geranylgeranyltransferase type I or II (GGTase-I and GGTase-II). FTase and GGTase-I have long been proposed to specifically recognize a four-amino acid CAAX C-terminal sequence within their substrates. Surprisingly, genetic screening reveals that yeast FTase can modify sequences longer than the canonical CAAX sequence, specifically C(x)3X sequences with four amino acids downstream of the cysteine. Biochemical and cell-based studies using both peptide and protein substrates reveal that mammalian FTase orthologs can also prenylate C(x)3X sequences. As the search to identify physiologically relevant C(x)3X proteins begins, this new prenylation motif nearly doubles the number of proteins within the yeast and human proteomes that can be explored as potential FTase substrates. This work expands our understanding of prenylation's impact within the proteome, establishes the biologically relevant reactivity possible with this new motif, and opens new frontiers in determining the impact of non-canonically prenylated proteins on cell function.


Assuntos
Alquil e Aril Transferases/metabolismo , Modelos Moleculares , Prenilação de Proteína , Alquil e Aril Transferases/antagonistas & inibidores , Alquil e Aril Transferases/química , Alquil e Aril Transferases/genética , Motivos de Aminoácidos , Animais , Bases de Dados de Proteínas , Inibidores Enzimáticos/farmacologia , Genes Reporter , Proteínas de Fluorescência Verde/química , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células HEK293 , Humanos , Microscopia de Fluorescência , Prenilação de Proteína/efeitos dos fármacos , Subunidades Proteicas/antagonistas & inibidores , Subunidades Proteicas/química , Subunidades Proteicas/genética , Subunidades Proteicas/metabolismo , Proteômica/métodos , Ratos , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/metabolismo , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/antagonistas & inibidores , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Especificidade por Substrato
19.
J Cell Physiol ; 233(1): 186-200, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28639275

RESUMO

The MEKK3/MEK5/ERK5 signaling axis is required for cardiovascular development in vivo. We analyzed the physiological role of ERK5 in cardiac endothelial cells and the consequence of activation of this kinase by the statin class of HMG Co-A reductase inhibitor drugs. We utilized human cardiac microvascular endothelial cells (HCMECs) and altered ERK5 expression using siRNA mediated gene silencing or overexpression of constitutively active MEK5 and ERK5 to reveal a role for ERK5 in regulating endothelial tight junction formation and cell permeability. Statin treatment of HCMECs stimulated activation of ERK5 and translocation to the plasma membrane resulting in co-localization with the tight junction protein ZO-1 and a concomitant reduction in endothelial cell permeability. Statin mediated activation of ERK5 was a consequence of reduced isoprenoid synthesis following HMG Co-A reductase inhibition. Statin pretreatment could overcome the effect of doxorubicin in reducing endothelial tight junction formation and prevent increased permeability. Our data provide the first evidence for the role of ERK5 in regulating endothelial tight junction formation and endothelial cell permeability. Statin mediated ERK5 activation and the resulting decrease in cardiac endothelial cell permeability may contribute to the cardioprotective effects of statins in reducing doxorubicin-induced cardiotoxicity.


Assuntos
Permeabilidade Capilar/efeitos dos fármacos , Vasos Coronários/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Cardiopatias/prevenção & controle , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Proteína Quinase 7 Ativada por Mitógeno/metabolismo , Junções Íntimas/efeitos dos fármacos , Antibióticos Antineoplásicos/toxicidade , Cardiotoxicidade , Células Cultivadas , Vasos Coronários/enzimologia , Citoproteção , Relação Dose-Resposta a Droga , Doxorrubicina/toxicidade , Células Endoteliais/enzimologia , Ativação Enzimática , Cardiopatias/induzido quimicamente , Cardiopatias/enzimologia , Cardiopatias/genética , Humanos , Proteína Quinase 7 Ativada por Mitógeno/genética , Prenilação de Proteína/efeitos dos fármacos , Transporte Proteico/efeitos dos fármacos , Quinolinas/farmacologia , Interferência de RNA , Rosuvastatina Cálcica/farmacologia , Transdução de Sinais/efeitos dos fármacos , Sinvastatina/farmacologia , Junções Íntimas/enzimologia , Transfecção , Proteína da Zônula de Oclusão-1/metabolismo
20.
J Med Chem ; 61(8): 3239-3252, 2018 04 26.
Artigo em Inglês | MEDLINE | ID: mdl-28505447

RESUMO

Post-translational modifications (PTMs) allot versatility to the biological functions of highly conserved proteins. Recently, modifications to non-histone proteins such as methylation, acetylation, phosphorylation, glycosylation, ubiquitination, and many more have been linked to the regulation of pivotal pathways related to cellular response and stability. Due to the roles these dynamic modifications assume, their dysregulation has been associated with cancer and many other important diseases such as inflammatory disorders and neurodegenerative diseases. For this reason, we present a review and perspective on important post-translational modifications on non-histone proteins, with emphasis on their roles in diseases and small molecule inhibitors developed to target PTM writers. Certain PTMs' contribution to epigenetics has been extensively expounded; yet more efforts will be needed to systematically dissect their roles on non-histone proteins, especially for their relationships with nononcological diseases. Finally, current research approaches for PTM study will be discussed and compared, including limitations and possible improvements.


Assuntos
Processamento de Proteína Pós-Traducional , Proteínas/metabolismo , Acetilação/efeitos dos fármacos , Animais , Citrulinação/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Glicosilação/efeitos dos fármacos , Humanos , Metilação/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Prenilação de Proteína/efeitos dos fármacos , Ubiquitinação/efeitos dos fármacos
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