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1.
J Med Chem ; 64(14): 10371-10392, 2021 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-34255518

RESUMO

Constitutive activation of the canonical Wnt signaling pathway, in most cases driven by inactivation of the tumor suppressor APC, is a hallmark of colorectal cancer. Tankyrases are druggable key regulators in these malignancies and are considered as attractive targets for therapeutic interventions, although no inhibitor has been progressed to clinical development yet. We continued our efforts to develop tankyrase inhibitors targeting the nicotinamide pocket with suitable drug-like properties for investigating effects of Wnt pathway inhibition on tumor growth. Herein, the identification of a screening hit series and its optimization through scaffold hopping and SAR exploration is described. The systematic assessment delivered M2912, a compound with an optimal balance between excellent TNKS potency, exquisite PARP selectivity, and a predicted human PK compatible with once daily oral dosing. Modulation of cellular Wnt pathway activity and significant tumor growth inhibition was demonstrated with this compound in colorectal xenograft models in vivo.


Assuntos
Antineoplásicos/farmacologia , Neoplasias Colorretais/tratamento farmacológico , Inibidores Enzimáticos/farmacologia , Tanquirases/antagonistas & inibidores , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Feminino , Humanos , Camundongos , Camundongos SCID , Estrutura Molecular , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Relação Estrutura-Atividade , Tanquirases/metabolismo
2.
Int J Mol Sci ; 22(14)2021 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-34298950

RESUMO

More than 80% of colorectal cancer patients have adenomatous polyposis coli (APC) mutations, which induce abnormal WNT/ß-catenin activation. Tankyrase (TNKS) mediates the release of active ß-catenin, which occurs regardless of the ligand that translocates into the nucleus by AXIN degradation via the ubiquitin-proteasome pathway. Therefore, TNKS inhibition has emerged as an attractive strategy for cancer therapy. In this study, we identified pyridine derivatives by evaluating in vitro TNKS enzyme activity and investigated N-([1,2,4]triazolo[4,3-a]pyridin-3-yl)-1-(2-cyanophenyl)piperidine-4-carboxamide (TI-12403) as a novel TNKS inhibitor. TI-12403 stabilized AXIN2, reduced active ß-catenin, and downregulated ß-catenin target genes in COLO320DM and DLD-1 cells. The antitumor activities of TI-12403 were confirmed by the viability of the colorectal cancer cells and its lack of visible toxicity in DLD-1 xenograft mouse model. In addition, combined 5-FU and TI-12403 treatment synergistically inhibited proliferation to a greater extent than that in a single drug treatment. Our observations suggest that TI-12403, a novel selective TNKS1 inhibitor, may be a suitable compound for anticancer drug development.


Assuntos
Antineoplásicos , Neoplasias Colorretais , Descoberta de Drogas , Inibidores Enzimáticos , Proteínas de Neoplasias/antagonistas & inibidores , Piridinas , Tanquirases/antagonistas & inibidores , Tiazóis , Antineoplásicos/química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/enzimologia , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Humanos , Proteínas de Neoplasias/metabolismo , Piridinas/química , Piridinas/farmacologia , Tanquirases/metabolismo , Tiazóis/química , Tiazóis/farmacologia
3.
J Med Chem ; 64(8): 4257-4288, 2021 04 22.
Artigo em Inglês | MEDLINE | ID: mdl-33822624

RESUMO

Canonical WNT signaling is an important developmental pathway that has attracted increased attention for anticancer drug discovery. From the production and secretion of WNT ligands, their binding to membrane receptors, and the ß-catenin destruction complex to the expansive ß-catenin transcriptional complex, multiple components have been investigated as drug targets to modulate WNT signaling. Significant progress in developing WNT inhibitors such as porcupine inhibitors, tankyrase inhibitors, ß-catenin/coactivators, protein-protein interaction inhibitors, casein kinase modulators, DVL inhibitors, and dCTPP1 inhibitors has been made, with several candidates (e.g., LGK-974, PRI-724, and ETC-159) in human clinical trials. Herein we summarize recent progress in the drug discovery and development of small-molecule inhibitors targeting the canonical WNT pathway, focusing on their specific target proteins, in vitro and in vivo activities, physicochemical properties, and therapeutic potential. The relevant opportunities and challenges toward maintaining the balance between efficacy and toxicity in effectively targeting this pathway are also highlighted.


Assuntos
Bibliotecas de Moléculas Pequenas/farmacologia , Proteínas Wnt/metabolismo , Via de Sinalização Wnt/efeitos dos fármacos , Animais , Sítios de Ligação , Descoberta de Drogas , Inibidores Enzimáticos/química , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Humanos , Simulação de Dinâmica Molecular , Neoplasias/tratamento farmacológico , Peptídeos/química , Peptídeos/metabolismo , Mapas de Interação de Proteínas/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/metabolismo , Bibliotecas de Moléculas Pequenas/uso terapêutico , Fatores de Transcrição TCF/química , Fatores de Transcrição TCF/metabolismo , Tanquirases/antagonistas & inibidores , Tanquirases/metabolismo , Proteínas Wnt/química , beta Catenina/química , beta Catenina/metabolismo
4.
Int J Mol Sci ; 22(8)2021 Apr 16.
Artigo em Inglês | MEDLINE | ID: mdl-33923443

RESUMO

During cell migration, protrusion of the leading edge is driven by the polymerization of Arp2/3-dependent branched actin networks. Migration persistence is negatively regulated by the Arp2/3 inhibitory protein Arpin. To better understand Arpin regulation in the cell, we looked for its interacting partners and identified both Tankyrase 1 and 2 (TNKS) using a yeast two-hybrid screening and coimmunoprecipitation with full-length Arpin as bait. Arpin interacts with ankyrin repeats of TNKS through a C-terminal-binding site on its acidic tail, which overlaps with the Arp2/3-binding site. Arpin was found to dissolve the liquid-liquid phase separation of TNKS upon overexpression. To uncouple the interactions of Arpin with TNKS and Arp2/3, we introduced point mutations in the Arpin tail and attempted to rescue the increased migration persistence of the Arpin knockout cells using random plasmid integration or compensating knock-ins at the ARPIN locus. Arpin mutations impairing interactions with either Arp2/3 or TNKS were insufficient to fully abolish Arpin activity. Only the mutation that affected both interactions rendered Arpin completely inactive, suggesting the existence of two independent pathways, whereby Arpin controls the migration persistence.


Assuntos
Proteínas de Transporte/metabolismo , Movimento Celular , Tanquirases/metabolismo , Sítios de Ligação , Proteínas de Transporte/química , Células HEK293 , Células HeLa , Humanos , Ligação Proteica , Tanquirases/química , Técnicas do Sistema de Duplo-Híbrido
5.
Biochem Biophys Res Commun ; 552: 66-72, 2021 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-33743349

RESUMO

Transplantation of retinal pigment epithelium (RPE) cells derived from human embryonic stem cells (hESCs) or induced pluripotent stem cells (hiPSCs) hold great promise as a new therapeutic modality for age-related macular degeneration and Stargardt disease. The development of hESC/hiPSC-derived RPE cells as cell-based therapeutic products requires a robust, scalable production for every hiPSC line congruent for patients. However, individual hESC/hiPSC lines show bias in differentiation. Here we report an efficient, robust method that induces RPE cells regardless of the differentiation propensity of the hiPSC lines. Application of the tankyrase inhibitor IWR-1-endo, which potentially inhibits Wnt signaling, promoted retinal differentiation in dissociated hiPSCs under feeder-free, two-dimensional culture conditions. The other tankyrase inhibitor, XAV939, also promoted retinal differentiation. However, Wnt signaling inhibitors, IWP-2 and iCRT3, that target porcupine and ß-catenin/TCF, respectively, did not. Further treatment with the GSK3ß inhibitor CHIR99021 and FGF receptor inhibitor SU5402 induced hexagonal pigmented cells with phagocytotic ability. Notably, the IWR-1-endo-based differentiation method induced RPE cells even in an hiPSC line that expresses a lower level of the differentiation propensity marker SALL3, which is indicative of resistance to ectoderm differentiation. The present study demonstrated that tankyrase inhibitors cause efficient and robust RPE differentiation, irrespective of the SALL3 expression levels in hiPSC lines. This differentiation method will resolve line-to-line variations of hiPSCs in RPE production and facilitate clinical application and industrialization of RPE cell products for regenerative medicine.


Assuntos
Diferenciação Celular , Células-Tronco Embrionárias Humanas/citologia , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes/citologia , Epitélio Pigmentado da Retina/citologia , Tanquirases/metabolismo , Transplante de Células/métodos , Células Cultivadas , Compostos Heterocíclicos com 3 Anéis/farmacologia , Células-Tronco Embrionárias Humanas/efeitos dos fármacos , Células-Tronco Embrionárias Humanas/metabolismo , Humanos , Imidas/farmacologia , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/metabolismo , Degeneração Macular/terapia , Células-Tronco Pluripotentes/efeitos dos fármacos , Células-Tronco Pluripotentes/metabolismo , Quinolinas/farmacologia , Epitélio Pigmentado da Retina/metabolismo , Tanquirases/antagonistas & inibidores
6.
Expert Opin Ther Pat ; 31(7): 645-661, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33567917

RESUMO

INTRODUCTION: Tankyrase inhibitors gained significant attention as therapeutic targets in oncology because of their potency. Their primary role in inhibiting the Wnt signaling pathway makes them an important class of compounds with the potential to be used as a combination therapy in future treatments of colorectal cancer. AREAS COVERED: This review describes pertinent work in the development of tankyrase inhibitors with a great emphasis on the recently patented TNKS inhibitors published from 2013 to 2020. This article also highlights a couple of promising candidates having tankyrase inhibitory effects and are currently undergoing clinical trials. EXPERT OPINION: Following the successful clinical applications of PARP inhibitors, tankyrase inhibition has gained significant attention in the research community as a target with high therapeutic potential. The ubiquitous role of tankyrase in cellular homeostasis and Wnt-dependent tumor proliferation brought difficulties for researchers to strike the right balance between potency and on-target toxicity. The need for novel tankyrase inhibitors with a better ADMET profile can introduce an additional regimen in treating various malignancies in monotherapy or adjuvant therapy. The development of combination therapies, including tankyrase inhibitors with or without PARP inhibitory properties, can potentially benefit the larger population of patients with unmet medical needs.


Assuntos
Antineoplásicos/farmacologia , Neoplasias Colorretais/tratamento farmacológico , Tanquirases/antagonistas & inibidores , Animais , Antineoplásicos/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Desenvolvimento de Medicamentos , Inibidores Enzimáticos/administração & dosagem , Inibidores Enzimáticos/farmacologia , Humanos , Patentes como Assunto , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Tanquirases/metabolismo , Via de Sinalização Wnt/efeitos dos fármacos
7.
Biochem Biophys Res Commun ; 537: 85-92, 2021 01 22.
Artigo em Inglês | MEDLINE | ID: mdl-33387887

RESUMO

Transactive response DNA-binding protein of 43 kDa (TDP-43) abnormally forms aggregates in certain subtypes of frontotemporal lobar degeneration (FTLD) and in amyotrophic lateral sclerosis (ALS). The pathological forms of TDP-43 have reported to be associated with poly(ADP-ribose) (PAR), which regulates the properties of these aggregates. A recent study has indicated that tankyrase, a member of the PAR polymerase (PARP) family, regulates pathological TDP-43 formation under conditions of stress, and tankyrase inhibitors suppress TDP-43 aggregate formation and cytotoxicity. Since we reported the development of tankyrase inhibitors that are more specific than conventional inhibitors, in this study, we examined their effects on the formation of TDP-43 aggregates in cultured cells. Time-lapse imaging showed that TDP-43 aggregates appeared in the nucleus within 30 min of treatment with sodium arsenite. Several tankyrase inhibitors suppressed the formation of aggregates and decreased the levels of the tankyrase protein. Immunohistochemical studies demonstrated that tankyrase was localized to neuronal cytoplasmic inclusions in the spinal cords of patients with ALS. Moreover, the tankyrase protein levels were significantly higher in the brains of patients with FTLD than in the brains of control subjects. These findings suggest that the inhibition of tankyrase activity protects against TDP-43 toxicity. Tankyrase inhibitors may be a potential treatment to suppress the progression of TDP-43 proteinopathies.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Inibidores Enzimáticos/farmacologia , Agregados Proteicos , Tanquirases/antagonistas & inibidores , Arsenitos/toxicidade , Linhagem Celular Tumoral , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Células HEK293 , Humanos , Poli Adenosina Difosfato Ribose/toxicidade , Agregados Proteicos/efeitos dos fármacos , Proteinopatias TDP-43/patologia , Tanquirases/metabolismo
8.
Cell Mol Life Sci ; 78(5): 2299-2314, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32970185

RESUMO

Glioblastoma (GBM) is the most aggressive cancer of central nervous system with worst patient outcome. Telomere maintenance is a crucial mechanism governing GBM initiation and progression making it an attractive target. microRNAs (miRNAs) have shown therapeutic potential in GBM. Earlier, we showed miR-490 is downregulated in GBM patients and plays a tumor suppressive role. Here, we show that miR-490 regulates telomere maintenance program in GBM by directly targeting Telomeric Repeat-binding Factor 2 (TERF2) of the shelterin complex, Tankyrase 2 (TNKS2) and Serine/Threonine-protein kinase, SMG1. Overexpression of miR-490 resulted in effects characteristic to hampered telomere maintenance via TERF2 inhibition. These include induction of telomere dysfunction-induced foci and global DNA damage (53BP1 foci), along with an increase in p-γH2AX levels. Further, it led to inhibition of telomere maintenance hallmarks via reduced stemness (SOX2 and SOX4 downregulation) and induction of senescence (H3K9me3 marks gain and SIRT1 downregulation). It also initiated downstream DNA damage response (DDR) leading to p53 pathway activation. Moreover, microarray data analysis highlighted an overlap between miR-490 expression and REST-inhibition responses in GBM. Thus, miR-490-mediated targeting of telomere maintenance could be therapeutically important in GBM.


Assuntos
Neoplasias Encefálicas/genética , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Glioblastoma/genética , MicroRNAs/genética , Homeostase do Telômero/genética , Regiões 3' não Traduzidas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Fatores de Transcrição SOXC/genética , Fatores de Transcrição SOXC/metabolismo , Tanquirases/genética , Tanquirases/metabolismo , Proteína 2 de Ligação a Repetições Teloméricas/genética , Proteína 2 de Ligação a Repetições Teloméricas/metabolismo
9.
Eur J Med Chem ; 207: 112712, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-32877803

RESUMO

Tankyrases are the group of enzymes belonging to a class of Poly (ADP-ribose) polymerase (PARP) recently named ADP-ribosyltransferase (ARTD). The two isoforms of tankyrase i.e. tankyrase1 (TNKS1) and tankyrase2 (TNKS2) were abundantly expressed in various biological functions in telomere regulation, Wnt/ß-catenin signaling pathway, viral replication, endogenous hormone regulation, glucose transport, cherubism disease, erectile dysfunction, and apoptosis. The structural analysis, mechanistic information, in vitro and in vivo studies led identification and development of several classes of tankyrase inhibitors under clinical phases. In the nutshell, this review will drive future research on tankyrase as it enlighten the structural and functional features of TNKS 1 and TNKS 2, different classes of inhibitors with their structure-activity relationship studies, molecular modeling studies, as well as past, current and future perspective of the different class of tankyrase inhibitors.


Assuntos
Descoberta de Drogas , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Tanquirases/antagonistas & inibidores , Tanquirases/metabolismo , Animais , Domínio Catalítico/efeitos dos fármacos , Desenvolvimento de Medicamentos , Humanos , Isoenzimas/análise , Isoenzimas/antagonistas & inibidores , Isoenzimas/metabolismo , Modelos Moleculares , Conformação Proteica/efeitos dos fármacos , Tanquirases/análise
10.
Biochem Biophys Res Commun ; 529(4): 970-976, 2020 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-32819607

RESUMO

In this study, the role of ubiquitin conjugating enzyme E2 M (UBE2M) and molecular mechanisms associated with osteoarthritis (OA) were explored. Cartilage tissues and corresponding healthy tissues from OA patients were isolated. Our data suggested that the expression level of UBE2M in OA patients was significantly higher compared to that in healthy individuals (P < 0.01). The apoptosis of human OA chondrocytes was inhibited when silencing UBE2M and increased when overexpressing UBE2M. XAV939, as a tankyrase 1 inhibitor, could block the signaling pathway of Wnt/ß-catenin, which significantly reversed the change introduced by UBE2M. The expression level of cytoplasmic ß-catenin in siUBE2M cells dramatically increased, and the expression levels of nuclear ß-catenin, cleaved caspase-3 (C-caspase-3), and MMP13 remarkably downregulated. Moreover, the ubiquitination of Axin was enhanced by the overexpression of UBE2M. The expression level of Axin significantly decreased in OA chondrocytes with UBE2M overexpression and increased after MG132 treatment. Moreover, UBE2M enhanced the apoptosis of OA chondrocytes by activating the Axin-dependent Wnt/ß-catenin pathway. In this process, UBE2M downregulated Axin in an ubiquitination-dependent degradation pathway and subsequently activated Wnt/ß-catenin signaling.


Assuntos
Condrócitos/metabolismo , Osteoartrite/genética , Tanquirases/genética , Enzimas de Conjugação de Ubiquitina/genética , Proteínas Wnt/genética , beta Catenina/genética , Apoptose/efeitos dos fármacos , Apoptose/genética , Proteína Axina/genética , Proteína Axina/metabolismo , Cartilagem Articular/metabolismo , Cartilagem Articular/patologia , Estudos de Casos e Controles , Caspase 3/genética , Caspase 3/metabolismo , Condrócitos/efeitos dos fármacos , Condrócitos/patologia , Fêmur/metabolismo , Fêmur/patologia , Regulação da Expressão Gênica , Compostos Heterocíclicos com 3 Anéis/farmacologia , Humanos , Leupeptinas/farmacologia , Metaloproteinase 13 da Matriz/genética , Metaloproteinase 13 da Matriz/metabolismo , Osteoartrite/metabolismo , Osteoartrite/patologia , Cultura Primária de Células , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Tanquirases/antagonistas & inibidores , Tanquirases/metabolismo , Enzimas de Conjugação de Ubiquitina/antagonistas & inibidores , Enzimas de Conjugação de Ubiquitina/metabolismo , Proteínas Wnt/metabolismo , beta Catenina/antagonistas & inibidores , beta Catenina/metabolismo
11.
Cell Rep ; 32(3): 107922, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32698014

RESUMO

Spatiotemporal control of Wnt/ß-catenin signaling is critical for organism development and homeostasis. The poly-(ADP)-ribose polymerase Tankyrase (TNKS1) promotes Wnt/ß-catenin signaling through PARylation-mediated degradation of AXIN1, a component of the ß-catenin destruction complex. Although Wnt/ß-catenin is a niche-restricted signaling program, tissue-specific factors that regulate TNKS1 are not known. Here, we report prostate-associated gene 4 (PAGE4) as a tissue-specific TNKS1 inhibitor that robustly represses canonical Wnt/ß-catenin signaling in human cells, zebrafish, and mice. Structural and biochemical studies reveal that PAGE4 acts as an optimal substrate decoy that potently hijacks substrate binding sites on TNKS1 to prevent AXIN1 PARylation and degradation. Consistently, transgenic expression of PAGE4 in mice phenocopies TNKS1 knockout. Physiologically, PAGE4 is selectively expressed in stromal prostate fibroblasts and functions to establish a proper Wnt/ß-catenin signaling niche through suppression of autocrine signaling. Our findings reveal a non-canonical mechanism for TNKS1 inhibition that functions to establish tissue-specific control of the Wnt/ß-catenin pathway.


Assuntos
Antígenos de Neoplasias/metabolismo , Especificidade de Órgãos , Tanquirases/antagonistas & inibidores , Via de Sinalização Wnt , Motivos de Aminoácidos , Sequência de Aminoácidos , Animais , Antígenos de Neoplasias/química , Proteína Axina , Fibroblastos/metabolismo , Células HEK293 , Humanos , Masculino , Camundongos Knockout , Modelos Biológicos , Poli ADP Ribosilação , Próstata/metabolismo , Domínios Proteicos , Proteólise , Células Estromais/metabolismo , Especificidade por Substrato , Tanquirases/química , Tanquirases/metabolismo , Ubiquitinação , Peixe-Zebra
12.
Nat Commun ; 11(1): 3321, 2020 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-32620872

RESUMO

Human telomeres are bound by the telomere repeat binding proteins TRF1 and TRF2. Telomere shortening in human cells leads to a DNA damage response that signals replicative senescence. While insufficient loading of TRF2 at shortened telomeres contributes to the DNA damage response in senescence, the contribution of TRF1 to senescence induction has not been determined. Here we show that counter to TRF2 deficiency-mediated induction of DNA damage, TRF1 deficiency serves a protective role to limit induction of DNA damage induced by subtelomere recombination. Shortened telomeres recruit insufficient TRF1 and as a consequence inadequate tankyrase 1 to resolve sister telomere cohesion. Our findings suggest that the persistent cohesion protects short telomeres from inappropriate recombination. Ultimately, in the final division, telomeres are no longer able to maintain cohesion and subtelomere copying ensues. Thus, the gradual loss of TRF1 and concomitant persistent cohesion that occurs with telomere shortening ensures a measured approach to replicative senescence.


Assuntos
Encurtamento do Telômero/genética , Telômero/genética , Proteína 1 de Ligação a Repetições Teloméricas/genética , Proteína 2 de Ligação a Repetições Teloméricas/genética , Sequência de Bases , Linhagem Celular , Linhagem Celular Tumoral , Senescência Celular/genética , Dano ao DNA , Células HEK293 , Heterocromatina/genética , Heterocromatina/metabolismo , Humanos , Hibridização in Situ Fluorescente , Mutação , Interferência de RNA , Tanquirases/metabolismo , Telômero/metabolismo , Proteína 1 de Ligação a Repetições Teloméricas/deficiência , Proteína 1 de Ligação a Repetições Teloméricas/metabolismo , Proteína 2 de Ligação a Repetições Teloméricas/deficiência , Proteína 2 de Ligação a Repetições Teloméricas/metabolismo
13.
Mol Cell Biochem ; 471(1-2): 15-27, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32472322

RESUMO

The biological activity of vascular endothelial growth factor (VEGF), the major cytokine regulating the process of angiogenesis is tightly controlled at multiple levels including processes involving post-translational modification such as ADP-ribosylation and glycosylation. ADP-ribosylation is a reversible NAD+-dependent modification, catalyzed by poly ADP-ribose polymerase (PARP) or ADP-ribosyl transferase (ADPRTs) and has been reported by us and others as a modification that reduces the biological activity of VEGF. The factors responsible for any such modification should occur in the secretory pathway, i.e., in the endoplasmic reticulum and Golgi. Our investigation carried out in this direction revealed that ADP-ribosylation of VEGF requires the interplay between members of poly ADP-ribose polymerase (PARP) family in the secretory pathway, viz., ER associated PARP-16 and Golgi associated Tankyrase-2 (TNKS-2). The data presented in this manuscript suggest that PARP-16 catalysis the priming mono ADP-ribosylation of VEGF which is a prerequisite for poly ADP-ribosylation of VEGF by TNKS-2.


Assuntos
Poli ADP Ribosilação , Poli(ADP-Ribose) Polimerases/metabolismo , Processamento de Proteína Pós-Traducional , Tanquirases/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Células Cultivadas , Embrião de Galinha , Humanos , Poli(ADP-Ribose) Polimerases/genética , Tanquirases/genética
14.
Dev Cell ; 53(5): 545-560.e7, 2020 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-32442396

RESUMO

Embryonic genome activation (EGA) is orchestrated by an intrinsic developmental program initiated during oocyte maturation with translation of stored maternal mRNAs. Here, we show that tankyrase, a poly(ADP-ribosyl) polymerase that regulates ß-catenin levels, undergoes programmed translation during oocyte maturation and serves an essential role in mouse EGA. Newly translated TNKS triggers proteasomal degradation of axin, reducing targeted destruction of ß-catenin and promoting ß-catenin-mediated transcription of target genes, including Myc. MYC mediates ribosomal RNA transcription in 2-cell embryos, supporting global protein synthesis. Suppression of tankyrase activity using knockdown or chemical inhibition causes loss of nuclear ß-catenin and global reductions in transcription and histone H3 acetylation. Chromatin and transcriptional profiling indicate that development arrests prior to the mid-2-cell stage, mediated in part by reductions in ß-catenin and MYC. These findings indicate that post-transcriptional regulation of tankyrase serves as a ligand-independent developmental mechanism for post-translational ß-catenin activation and is required to complete EGA.


Assuntos
Blastocisto/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Tanquirases/metabolismo , beta Catenina/genética , Animais , Blastocisto/citologia , Histonas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Embrionárias Murinas/metabolismo , Oócitos/citologia , Oócitos/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , RNA Ribossômico/genética , RNA Ribossômico/metabolismo , Tanquirases/genética , Regulação para Cima , beta Catenina/metabolismo
15.
Cell Death Dis ; 11(5): 356, 2020 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-32393760

RESUMO

Circular RNAs (circRNAs), non-coding RNAs generated by precursor mRNA back-splicing of exons, have been reported to fulfill multiple roles in cancer. However, the role of quite a lot circRNAs in colorectal cancer (CRC) remains mostly unknown. Herein, we explored the expression profiles of circRNAs in 5 paired samples of CRC patients by microarray and noted a circRNA, hsa_circ_0005615 (circ5615), was significantly upregulated in CRC tissues. Circ5615 was derived from exon 2 of NFATC3 and its upregulation was tightly correlated with higher T stage and poor prognosis in CRC patients. Studies in vitro and in vivo demonstrated that knockdown of circ5615 in cancer cells inhibited proliferation and cell cycle acceleration, while overexpression promoted malignant phenotypes. Mechanistically, RNA immunoprecipitation, biotin-coupled probe pull-down and luciferase reporter assays revealed circ5615 effectively bound to miR-149-5p and might play a role like miR-149-5p sponge. Additionally, tankyrase (TNKS), regulator of ß-catenin stabilization, was identified as circ5615 downstream and the potential miR-149-5p targets by RNA-seq and bioinformatics analysis. We further verified the upregulation of ß-catenin and cyclin D1 induced by circ5615. Our results indicated that circ5615 exerted oncogenic function as competing endogenous RNA (ceRNA) of miR-149-5p to release TNKS and activated Wnt/ß-catenin pathway.


Assuntos
Neoplasias Colorretais/enzimologia , RNA Circular/metabolismo , Tanquirases/metabolismo , Animais , Ciclo Celular , Proliferação de Células , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Ciclina D1/genética , Ciclina D1/metabolismo , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Células HCT116 , Células HT29 , Humanos , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Circular/genética , Tanquirases/genética , Carga Tumoral , Regulação para Cima , Via de Sinalização Wnt , beta Catenina/genética , beta Catenina/metabolismo
16.
Commun Biol ; 3(1): 196, 2020 04 24.
Artigo em Inglês | MEDLINE | ID: mdl-32332858

RESUMO

The development of immune checkpoint inhibitors represents a major breakthrough in cancer therapy. Nevertheless, a substantial number of patients fail to respond to checkpoint pathway blockade. Evidence for WNT/ß-catenin signaling-mediated immune evasion is found in a subset of cancers including melanoma. Currently, there are no therapeutic strategies available for targeting WNT/ß-catenin signaling. Here we show that a specific small-molecule tankyrase inhibitor, G007-LK, decreases WNT/ß-catenin and YAP signaling in the syngeneic murine B16-F10 and Clone M-3 melanoma models and sensitizes the tumors to anti-PD-1 immune checkpoint therapy. Mechanistically, we demonstrate that the synergistic effect of tankyrase and checkpoint inhibitor treatment is dependent on loss of ß-catenin in the tumor cells, anti-PD-1-stimulated infiltration of T cells into the tumor and induction of an IFNγ- and CD8+ T cell-mediated anti-tumor immune response. Our study uncovers a combinatorial therapeutical strategy using tankyrase inhibition to overcome ß-catenin-mediated resistance to immune checkpoint blockade in melanoma.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Inibidores Enzimáticos/farmacologia , Inibidores de Checkpoint Imunológico/farmacologia , Melanoma Experimental/tratamento farmacológico , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Neoplasias Cutâneas/tratamento farmacológico , Sulfonas/farmacologia , Tanquirases/antagonistas & inibidores , Triazóis/farmacologia , Via de Sinalização Wnt/efeitos dos fármacos , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Citotoxicidade Imunológica/efeitos dos fármacos , Sinergismo Farmacológico , Feminino , Células HEK293 , Humanos , Interferon gama/metabolismo , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Melanoma Experimental/enzimologia , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Camundongos Transgênicos , Receptor de Morte Celular Programada 1/metabolismo , Neoplasias Cutâneas/enzimologia , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/patologia , Tanquirases/metabolismo , Carga Tumoral/efeitos dos fármacos , beta Catenina/genética , beta Catenina/metabolismo
17.
Nat Commun ; 11(1): 1195, 2020 03 05.
Artigo em Inglês | MEDLINE | ID: mdl-32139672

RESUMO

Here, we report that the functionality of vascular progenitors (VP) generated from normal and disease-primed conventional human induced pluripotent stem cells (hiPSC) can be significantly improved by reversion to a tankyrase inhibitor-regulated human naïve epiblast-like pluripotent state. Naïve diabetic vascular progenitors (N-DVP) differentiated from patient-specific naïve diabetic hiPSC (N-DhiPSC) possessed higher vascular functionality, maintained greater genomic stability, harbored decreased lineage-primed gene expression, and were more efficient in migrating to and re-vascularizing the deep neural layers of the ischemic retina than isogenic diabetic vascular progenitors (DVP). These findings suggest that reprogramming to a stable naïve human pluripotent stem cell state may effectively erase dysfunctional epigenetic donor cell memory or disease-associated aberrations in patient-specific hiPSC. More broadly, tankyrase inhibitor-regulated naïve hiPSC (N-hiPSC) represent a class of human stem cells with high epigenetic plasticity, improved multi-lineage functionality, and potentially high impact for regenerative medicine.


Assuntos
Vasos Sanguíneos/patologia , Diabetes Mellitus/patologia , Células-Tronco Pluripotentes Induzidas/patologia , Isquemia/terapia , Retina/patologia , Células-Tronco/patologia , Tanquirases/antagonistas & inibidores , Adulto , Animais , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Linhagem da Célula/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Senescência Celular/efeitos dos fármacos , Dano ao DNA , Inibidores Enzimáticos/farmacologia , Epigênese Genética/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Fibroblastos/patologia , Código das Histonas , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Isquemia/patologia , Camundongos , Organoides/efeitos dos fármacos , Organoides/patologia , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Regiões Promotoras Genéticas/genética , Células-Tronco/efeitos dos fármacos , Células-Tronco/ultraestrutura , Tanquirases/metabolismo , Teratoma/patologia , Transcrição Genética/efeitos dos fármacos
18.
Exp Cell Res ; 390(1): 111935, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32151493

RESUMO

Tankyrase 1 (TNKS1; PARP-5a) and Tankyrase 2 (TNKS2; PARP-5b) are poly-ADP-ribosyl-polymerase (PARP)-domain-containing proteins that regulate the activities of a wide repertoire of target proteins via post-translational addition of poly-ADP-ribose polymers (PARylation). Although tankyrases were first identified as regulators of human telomere elongation, important and expansive roles of tankyrase activity have recently emerged in the development and maintenance of stem cell states. Herein, we summarize the current state of knowledge of the various tankyrase-mediated activities that may promote human naïve and 'extended' pluripotency'. We review the putative role of tankyrase and PARP inhibition in trophectoderm specification, telomere elongation, DNA repair and chromosomal segregation, metabolism, and PTEN-mediated apoptosis. Importantly, tankyrases possess PARP-independent activities that include regulation of MDC1-associated DNA repair by homologous recombination (HR) and autophagy/pexophagy, which is an essential mechanism of protein synthesis in the preimplantation embryo. Additionally, tankyrases auto-regulate themselves via auto-PARylation which augments their cellular protein levels and potentiates their non-PARP tankyrase functions. We propose that these non-PARP-related activities of tankyrase proteins may further independently affect both naïve and extended pluripotency via mechanisms that remain undetermined. We broadly outline a hypothetical framework for how inclusion of a tankyrase/PARP inhibitor in small molecule cocktails may stabilize and potentiate naïve and extended pluripotency via pleiotropic routes and mechanisms.


Assuntos
Células-Tronco Pluripotentes/metabolismo , Tanquirases/metabolismo , Apoptose , Diferenciação Celular , Reparo do DNA , Humanos , Células-Tronco Pluripotentes/citologia , Tanquirases/genética , Homeostase do Telômero
19.
J Med Chem ; 63(8): 4183-4204, 2020 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-32202790

RESUMO

Tankyrases (TNKS/TNKS2) belong to the poly(ADP-ribose) polymerase family. Inhibition of their enzymatic activities attenuates the Wnt/ß-catenin signaling, which plays an important role in cancer pathogenesis. We previously reported the discovery of RK-287107, a spiroindoline-based, highly selective, potent tankyrase inhibitor. Herein we describe the optimization process of RK-287107 leading to RK-582, which exhibits a markedly improved robust tumor growth inhibition in a COLO-320DM mouse xenograft model when orally administered. In addition to the dose-dependent elevation and attenuation of the levels of biomarkers AXIN2 and ß-catenin, respectively, results of the TCF reporter and cell proliferation studies on COLO-320DM are discussed.


Assuntos
Neoplasias do Colo/tratamento farmacológico , Desenho de Fármacos , Descoberta de Drogas/métodos , Inibidores Enzimáticos/administração & dosagem , Tanquirases/antagonistas & inibidores , Administração Oral , Animais , Linhagem Celular Tumoral , Neoplasias do Colo/enzimologia , Inibidores Enzimáticos/química , Feminino , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Estrutura Terciária de Proteína , Ratos , Tanquirases/química , Tanquirases/metabolismo , Resultado do Tratamento , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
20.
Commun Biol ; 3(1): 123, 2020 03 13.
Artigo em Inglês | MEDLINE | ID: mdl-32170109

RESUMO

Sjögren syndrome/scleroderma autoantigen 1 (SSSCA1) was first described as an auto-antigen over-expressed in Sjögren's syndrome and in scleroderma patients. SSSCA1 has been linked to mitosis and centromere association and as a potential marker candidate in diverse solid cancers. Here we characterize SSSCA1 for the first time, to our knowledge, at the molecular, structural and subcellular level. We have determined the crystal structure of a zinc finger fold, a zinc ribbon domain type 2 (ZNRD2), at 2.3 Å resolution. We show that the C-terminal domain serves a dual function as it both behaves as the interaction site to Tankyrase 1 (TNKS1) and as a nuclear export signal. We identify TNKS1 as a direct binding partner of SSSCA1, map the binding site to TNKS1 ankyrin repeat cluster 2 (ARC2) and thus define a new binding sequence. We experimentally verify and map a new nuclear export signal sequence in SSSCA1.


Assuntos
Autoantígenos/química , Autoantígenos/metabolismo , Neoplasias/metabolismo , Ribonucleoproteínas/química , Ribonucleoproteínas/metabolismo , Tanquirases/química , Tanquirases/metabolismo , Transporte Ativo do Núcleo Celular/genética , Autoantígenos/genética , Sítios de Ligação , Cristalografia por Raios X , Células HeLa , Humanos , Neoplasias/patologia , Sinais de Exportação Nuclear , Filogenia , Ligação Proteica/genética , Conformação Proteica em alfa-Hélice , Domínios Proteicos , Mapas de Interação de Proteínas/genética , Ribonucleoproteínas/genética , Transfecção
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