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1.
Nat Commun ; 12(1): 4566, 2021 07 27.
Artigo em Inglês | MEDLINE | ID: mdl-34315881

RESUMO

The airway epithelium serves as the interface between the host and external environment. In many chronic lung diseases, the airway is the site of substantial remodeling after injury. While, idiopathic pulmonary fibrosis (IPF) has traditionally been considered a disease of the alveolus and lung matrix, the dominant environmental (cigarette smoking) and genetic (gain of function MUC5B promoter variant) risk factor primarily affect the distal airway epithelium. Moreover, airway-specific pathogenic features of IPF include bronchiolization of the distal airspace with abnormal airway cell-types and honeycomb cystic terminal airway-like structures with concurrent loss of terminal bronchioles in regions of minimal fibrosis. However, the pathogenic role of the airway epithelium in IPF is unknown. Combining biophysical, genetic, and signaling analyses of primary airway epithelial cells, we demonstrate that healthy and IPF airway epithelia are biophysically distinct, identifying pathologic activation of the ERBB-YAP axis as a specific and modifiable driver of prolongation of the unjammed-to-jammed transition in IPF epithelia. Furthermore, we demonstrate that this biophysical state and signaling axis correlates with epithelial-driven activation of the underlying mesenchyme. Our data illustrate the active mechanisms regulating airway epithelial-driven fibrosis and identify targets to modulate disease progression.


Assuntos
Epitélio/fisiopatologia , Fibrose Pulmonar Idiopática/fisiopatologia , Pulmão/fisiopatologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Anfirregulina/genética , Anfirregulina/metabolismo , Fenômenos Biofísicos/efeitos dos fármacos , Epitélio/efeitos dos fármacos , Receptores ErbB/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Predisposição Genética para Doença , Humanos , Fibrose Pulmonar Idiopática/genética , Queratina-5/genética , Queratina-5/metabolismo , Pulmão/efeitos dos fármacos , Mucina-5B/genética , Mucina-5B/metabolismo , Quinazolinas/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fatores de Risco , Transdução de Sinais/efeitos dos fármacos , Fatores de Transcrição/metabolismo , Tirfostinas/farmacologia , Verteporfina/farmacologia
2.
Int J Mol Sci ; 22(12)2021 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-34204001

RESUMO

Radiodynamic therapy (RDT) is a recent extension of conventional photodynamic therapy, in which visible/near infrared light irradiation is replaced by a well-tolerated dose of high-energy X-rays. This enables greater tissue penetration to allow non-invasive treatment of large, deep-seated tumors. We report here the design and testing of a drug delivery system for RDT that is intended to enhance intra- or peri-nuclear localization of the photosensitizer, leading to DNA damage and resulting clonogenic cell kill. This comprises a photosensitizer (Verteporfin, VP) incorporated into poly (lactic-co-glycolic acid) nanoparticles (PLGA NPs) that are surface-functionalized with a cell-penetrating HIV trans-activator of transcription (TAT) peptide. In addition to a series of physical and photophysical characterization studies, cytotoxicity tests in pancreatic (PANC-1) cancer cells in vitro under 4 Gy X-ray exposure from a clinical 6 MV linear accelerator (LINAC) showed that TAT targeting of the nanoparticles markedly enhances the effectiveness of RDT treatment, particularly when assessed by a clonogenic, i.e., DNA damage-mediated, cell kill.


Assuntos
Composição de Medicamentos , Produtos do Gene tat/química , Nanopartículas/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Verteporfina/farmacologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Células Clonais , DNA/metabolismo , Endocitose/efeitos dos fármacos , Humanos , Lipídeos de Membrana/metabolismo , Nanopartículas/ultraestrutura , Oxigênio Singlete/metabolismo
3.
J Photochem Photobiol B ; 222: 112261, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34330081

RESUMO

It is crucial to develop nanocarrier systems to detect and treat drug-resistant micro tumors to prevent recurrence and/or metastasis of cancer. Due to their exceptional features such as biocompatibility, easy surface modification, serving as imaging and therapeutic agent, gold nanoparticles (AuNPs) draw attention as theranostic agents. It is beneficial to combine AuNPs with a second imaging and/or treatment modality such as photodynamic therapy (PDT). PDT is a non-mutagenic treatment approach in which photosensitizer is activated with light, generating reactive oxygen species and/or free radicals to destroy tumor cells. With the aim of developing "off-on" theranostic system, citrate stabilized spherical 13 nm AuNPs were densely coated with polyethylene glycol (PEG). To advance the theranostic feature of PEGylated AuNPs, they were further functionalized with FDA-Approved photosensitizer, Verteporfin (BPD-MA). Due to static quenching between BPD-MA and AuNPs as well as in between nearby BPD-MA molecules, the fluorescence of the ground state complex is quenched and the system is in "off" state. When BPD-MA molecules are cleaved from the AuNPs surface and diffuse away, fluorescence is recovered. Consequently, the system switches to the "on" state. Among the various mole ratios of BPD-MA carrying conjugates prepared, the most promising candidate was selected based on stability, quenching factor, and fluorescence recovery rate. The conjugate was further decorated with D-α-Tocopherol succinate (VitES) to increase the therapeutic efficacy of the theranostic agent via enhancing cellular uptake. Our results showed that it was possible to achieve as high as 80 times fluorescence quenching when the system was "off". As the system switched from "off" to "on" state, 51% of the fluorescence was recovered. When BPD-MA was immobilized on the PEGylated AuNPs, the phototoxic effect of BPD-MA increased twice against the MCF-7 cell line. Moreover, the developed system showed four times more phototoxicity than BPD-MA alone after it was decorated with VitES. Since the developed system is capable of dual imaging (computed tomography and fluorescence) and dual treatment (PDT and hyperthermia), it potentially offers superior imaging and therapy options for various types of in vitro/in vivo applications.


Assuntos
Ouro/química , Nanopartículas Metálicas/química , Nanomedicina Teranóstica , alfa-Tocoferol/química , Sobrevivência Celular/efeitos dos fármacos , Humanos , Células MCF-7 , Nanopartículas Metálicas/toxicidade , Microscopia Confocal , Fármacos Fotossensibilizantes/química , Fármacos Fotossensibilizantes/farmacologia , Polietilenoglicóis/química , Teoria Quântica , Oxigênio Singlete/metabolismo , Espectrometria de Fluorescência , Verteporfina/química , Verteporfina/farmacologia
4.
Mol Carcinog ; 60(7): 440-454, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34003522

RESUMO

Aberrant expression of kinesin family member 4A (KIF4A), which is associated with tumor progression, has been reported in several types of cancer. However, its expression and the underlying molecular mechanisms regulating the transcription of KIF4A in esophageal squamous cell carcinoma (ESCC) remain largely unclear. Here, we found that high KIF4A expression was positively correlated with tumor stage and poor prognosis in ESCC patients. KIF4A silencing significantly inhibited the growth and migration of ESCC cells, arrested cell cycle, and induced apoptosis. Interestingly, KIF4A expression was positively related to the expression of YAP in human ESCC tissues. YAP knockdown or disrupting YAP/TEAD4 interaction by verteporfin repressed KIF4A expression. Also, KIF4A knockdown significantly inhibited the cell growth induced by YAP overexpression. Mechanistically, YAP activated KIF4A transcriptional expression by TEAD4-mediated direct binding to KIF4A promoter. Finally, KIF4A knockdown and verteporfin treatment synergistically inhibited tumor growth in xenograft models. Together, these results indicated that KIF4A, a novel target gene of YAP/TEAD4, may be a progression and prognostic biomarker of ESCC. Targeting drugs for KIF4A combined with YAP inhibitor may be a novel therapeutic strategy for ESCC.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas de Ligação a DNA/metabolismo , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago/patologia , Cinesina/genética , Proteínas Musculares/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Idoso , Animais , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Proteínas de Ligação a DNA/genética , Neoplasias Esofágicas/tratamento farmacológico , Neoplasias Esofágicas/metabolismo , Neoplasias Esofágicas/mortalidade , Carcinoma de Células Escamosas do Esôfago/tratamento farmacológico , Carcinoma de Células Escamosas do Esôfago/metabolismo , Carcinoma de Células Escamosas do Esôfago/mortalidade , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Cinesina/metabolismo , Masculino , Camundongos Nus , Pessoa de Meia-Idade , Proteínas Musculares/genética , Prognóstico , Verteporfina/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
5.
Science ; 372(6540)2021 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-33888614

RESUMO

Skin scarring, the end result of adult wound healing, is detrimental to tissue form and function. Engrailed-1 lineage-positive fibroblasts (EPFs) are known to function in scarring, but Engrailed-1 lineage-negative fibroblasts (ENFs) remain poorly characterized. Using cell transplantation and transgenic mouse models, we identified a dermal ENF subpopulation that gives rise to postnatally derived EPFs by activating Engrailed-1 expression during adult wound healing. By studying ENF responses to substrate mechanics, we found that mechanical tension drives Engrailed-1 activation via canonical mechanotransduction signaling. Finally, we showed that blocking mechanotransduction signaling with either verteporfin, an inhibitor of Yes-associated protein (YAP), or fibroblast-specific transgenic YAP knockout prevents Engrailed-1 activation and promotes wound regeneration by ENFs, with recovery of skin appendages, ultrastructure, and mechanical strength. This finding suggests that there are two possible outcomes to postnatal wound healing: a fibrotic response (EPF-mediated) and a regenerative response (ENF-mediated).


Assuntos
Cicatriz/patologia , Fibroblastos/fisiologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Regeneração , Pele/lesões , Cicatrização , Animais , Cicatriz/prevenção & controle , Fibroblastos/transplante , Regulação da Expressão Gênica , Técnicas de Inativação de Genes , Mecanotransdução Celular , Camundongos , Camundongos Transgênicos , Proteínas Proto-Oncogênicas c-yes/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-yes/genética , Proteínas Proto-Oncogênicas c-yes/metabolismo , Transdução de Sinais , Estresse Mecânico , Ativação Transcricional , Transcriptoma , Verteporfina/farmacologia
6.
Cell Death Dis ; 12(1): 8, 2021 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-33414428

RESUMO

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer, accounting for the majority of breast cancer-related death. Due to the lack of specific therapeutic targets, chemotherapeutic agents (e.g., paclitaxel) remain the mainstay of systemic treatment, but enrich a subpopulation of cells with tumor-initiating capacity and stem-like characteristics called cancer stem cells (CSCs); thus development of a new and effective strategy for TNBC treatment is an unmet medical need. Cancer nanomedicine has transformed the landscape of cancer drug development, allowing for a high therapeutic index. In this study, we developed a new therapy by co-encapsulating clinically approved drugs, such as paclitaxel, verteporfin, and combretastatin (CA4) in polymer-lipid hybrid nanoparticles (NPs) made of FDA-approved biomaterials. Verteporfin is a drug used in the treatment of macular degeneration and has recently been found to inhibit the Hippo/YAP (Yes-associated protein) pathway, which is known to promote the progression of breast cancer and the development of CSCs. CA4 is a vascular disrupting agent and has been tested in phase II/III of clinical trials. We found that our new three drug-NP not only effectively inhibited TNBC cell viability and cell migration, but also significantly diminished paclitaxel-induced and/or CA4-induced CSC enrichment in TNBC cells, partially through inhibiting the upregulated Hippo/YAP signaling. Combination of verteporfin and CA4 was also more effective in suppressing angiogenesis in an in vivo zebrafish model than single drug alone. The efficacy and application potential of our triple drug-NPs were further assessed by using clinically relevant patient-derived xenograft (PDX) models. Triple drug-NP effectively inhibited the viability of PDX organotypic slide cultures ex vivo and stopped the growth of PDX tumors in vivo. This study developed an approach capable of simultaneously inhibiting bulk cancer cells, CSCs, and angiogenesis.


Assuntos
Bibenzilas/farmacologia , Nanopartículas/uso terapêutico , Paclitaxel/farmacologia , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Verteporfina/farmacologia , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Combinação de Medicamentos , Feminino , Humanos , Camundongos Nus , Células-Tronco Neoplásicas , Ratos , Peixe-Zebra
7.
Oncol Rep ; 45(1): 83-94, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33416116

RESUMO

Platelet­derived growth factor (PDGF) is a potent mitogen and chemoattractant that serves a role in the development of several types of solid cancer, and abnormal PDGF activity has been reported in numerous human tumors. Tumor­derived PDGF ligands are considered to act in either a paracrine or autocrine manner, serving roles in the phosphorylation of receptors on tumor and stromal cells in the tumor microenvironment. Despite the well­established association between PDGF and tumor progression, the precise mechanisms of autocrine PDGF signaling in pancreatic tumor cells remain elusive. Therefore, the present study aimed to analyze the influence of PDGF­BB in pancreatic cancer. Pancreatic adenocarcinoma BxPC­3 cells were cultured and treated with recombinant human PDGF­BB in vitro. Cell proliferation was tested using an MTT assay. Cell apoptosis was measured using flow cytometry. Tumor cell migration and invasion were examined via wound­healing and Transwell assays, respectively. The expression and subcellular localization of Yes­associated protein (YAP) was determined using western blotting and immunofluorescence. The transcriptional activity of target genes was tested using a luciferase assay and reverse transcription­quantitative PCR. The present study revealed that PDGF­BB significantly promoted cell proliferation in pancreatic adenocarcinoma BxPC­3 cells and enhanced the aggressiveness of this cell line, as demonstrated by Transwell and wound­healing assays. Anoikis resistance is an important mechanism by which metastatic cells avoid apoptosis when detaching from adjacent cells or the extracellular matrix. PDGF­BB treatment inhibited anoikis under anchorage­independent conditions. Mechanistic experiments revealed that PDGF­BB promoted the upregulation and activation of the transcriptional coactivator YAP, an effector of the Hippo signaling pathway. RhoA or protein phosphatase­1 (PP­1) inhibition partially abolished the accumulation and activation of YAP, suggesting PDGF­BB­mediated YAP dephosphorylation and transactivation via the RhoA/PP­1 cascade. Pharmacologic inhibition of the PDGF receptor directly downregulated YAP activity and the expression levels of downstream genes. Furthermore, verteporfin, a small molecular inhibitor of the Hippo/YAP signaling pathway, partially reversed the effects of PDGF­BB on cell proliferation, anoikis resistance and cell migration. In conclusion, the present study revealed that the Hippo/YAP signaling pathway may be involved in the tumor­promoting activity of PDGF­BB in pancreatic cancer.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Adenocarcinoma/genética , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogênicas c-sis/metabolismo , Fatores de Transcrição/genética , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/patologia , Anoikis/efeitos dos fármacos , Anoikis/genética , Apoptose/efeitos dos fármacos , Apoptose/genética , Becaplermina/metabolismo , Carcinogênese/genética , Carcinogênese/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Meios de Cultura/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/patologia , Isoformas de Proteínas , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/metabolismo , Ativação Transcricional , Regulação para Cima , Verteporfina/farmacologia , Verteporfina/uso terapêutico
8.
Eur J Pharmacol ; 893: 173829, 2021 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-33347823

RESUMO

Neuroblastoma is an embryonal malignancy of early childhood arising from the embryonic sympatho-adrenal lineage of the neural crest. About half of all cases are currently classified as high-risk of disease recurrence, with an overall survival rate of less than 40% at 5 years despite intensive therapy. Recent studies on matched primary tumours and at the relapse revealed downregulation of genes transcriptionally silenced by YAP as significant association with neuroblastoma relapse. Here, we evaluated the pharmacological targeting of YAP/TAZ with the YAP/TAZ-TEAD inhibitor Verteporfin (VP) in Tumour Initiating Cells (TICs) derived from High-Risk Neuroblastoma patients. VP treatment suppresses YAP/TAZ expression, induces apoptosis and causes the re-organization of the cytoskeleton reducing cells migration and clonogenic ability. Moreover, VP reduces the percentage of side population cells and ABC transporters involved in drug resistance, and the percentage of stem cell subpopulations CD133+ and CD44+ of TICs. Finally, we demonstrated that VP sensitizes TICs to the standard drugs used for neuroblastoma therapy etoposide and cis-platin opening the way to use VP as drug repositioning candidate for recurrent neuroblastoma.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Apoptose/efeitos dos fármacos , Células-Tronco Neoplásicas/efeitos dos fármacos , Neuroblastoma/tratamento farmacológico , Células da Side Population/efeitos dos fármacos , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Verteporfina/farmacologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Cisplatino/farmacologia , Reposicionamento de Medicamentos , Etoposídeo/farmacologia , Humanos , Invasividade Neoplásica , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Neuroblastoma/metabolismo , Neuroblastoma/patologia , Células da Side Population/metabolismo , Células da Side Population/patologia , Transdução de Sinais
9.
Int J Mol Sci ; 21(21)2020 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-33142952

RESUMO

Yes-associated protein (YAP) activation after acute ischemic kidney injury might be related to interstitial fibrosis and impaired renal tubular regeneration. Verteporfin (VP) is a photosensitizer used in photodynamic therapy to treat age-related macular degeneration. In cancer cells, VP inhibits TEA domain family member (TEAD)-YAP interactions without light stimulation. The protective role of VP in unilateral ureteral obstruction (UUO)-induced renal fibrosis and related mechanisms remains unclear. In this study, we investigate the protective effects of VP on UUO-induced renal tubulointerstitial inflammation and fibrosis and its regulation of the transforming growth factor-ß1 (TGF-ß1)/Smad signaling pathway. We find that VP decreased the UUO-induced increase in tubular injury, inflammation, and extracellular matrix deposition in mice. VP also decreased myofibroblast activation and proliferation in UUO kidneys and NRK-49F cells by modulating Smad2 and Smad3 phosphorylation. Therefore, YAP inhibition might have beneficial effects on UUO-induced tubulointerstitial inflammation and fibrosis by regulating the TGF-ß1/Smad signaling pathway.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas de Ciclo Celular/antagonistas & inibidores , Fibrose/prevenção & controle , Inflamação/prevenção & controle , Nefropatias/prevenção & controle , Obstrução Ureteral/complicações , Verteporfina/farmacologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteínas de Ciclo Celular/metabolismo , Células Cultivadas , Modelos Animais de Doenças , Fibrose/etiologia , Fibrose/metabolismo , Fibrose/patologia , Inflamação/etiologia , Inflamação/metabolismo , Inflamação/patologia , Nefropatias/etiologia , Nefropatias/metabolismo , Nefropatias/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fármacos Fotossensibilizantes/farmacologia , Ratos
10.
Int J Mol Sci ; 21(22)2020 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-33218077

RESUMO

Liver fibrosis represents the wound healing response to sustained hepatic injury with activation of hepatic stellate cells (HSCs). The I148M variant of the PNPLA3 gene represents a risk factor for development of severe liver fibrosis. Activated HSCs carrying the I148M variant display exacerbated pro-inflammatory and pro-fibrogenic features. We aimed to examine whether the I148M variant may impair Hedgehog and Yap signaling, as key pathways implicated in the control of energy expenditure and maintenance of myofibroblastic traits. First, we show that TGF-ß rapidly up-regulated the PNPLA3 transcript and protein and Yap/Hedgehog target gene expression. In addition, HSCs overexpressing PNPLA3 I148M boosted anaerobic glycolysis, as supported by higher lactate release and decreased phosphorylation of the energy sensor AMPK. These cells displayed higher Yap and Hedgehog signaling, due to accumulation of total Yap protein, Yap promoter activity and increased downstream targets expression, compared to WT cells. HSCs exposed to TGF-ß and leptin rapidly increased total Yap, together with a reduction in its inhibited form, phosphorylated Yap. In line, Yap-specific inhibitor Verteporfin strongly abolished Yap-mediated genes expression, at baseline as well as after TGF-ß and leptin treatments in HSCs with I148M PNPLA3. Finally, Yap transcriptional activity was strongly reduced by a combination of Verteporfin and Rosiglitazone, a PPARγ synthetic agonist. In conclusion, HSCs carrying the PNPLA3 variant show activated Yap/Hedgehog pathways, resulting in altered anaerobic glycolysis and enhanced synthesis of Hedgehog markers and sustained Yap signaling. TGF-ß and leptin exacerbate Yap/Hedgehog-related fibrogenic genes expression, while Yap inhibitors and PPARγ agonists abrogate these effects in PNPLA3 I148M carrying HSCs.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Hedgehog/genética , Células Estreladas do Fígado/metabolismo , Lipase/genética , Proteínas de Membrana/genética , Polimorfismo de Nucleotídeo Único , Transdução de Sinais/genética , Fatores de Transcrição/genética , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Linhagem Celular , Células Cultivadas , Expressão Gênica/efeitos dos fármacos , Glicólise/efeitos dos fármacos , Glicólise/genética , Proteínas Hedgehog/metabolismo , Células Estreladas do Fígado/citologia , Células Estreladas do Fígado/efeitos dos fármacos , Humanos , Leptina/farmacologia , Lipase/metabolismo , Proteínas de Membrana/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/farmacologia , Regulação para Cima/efeitos dos fármacos , Verteporfina/farmacologia
11.
Molecules ; 25(22)2020 Nov 12.
Artigo em Inglês | MEDLINE | ID: mdl-33198255

RESUMO

Verteporfin, a free base benzoporphyrin derivative monoacid ring A, is a photosensitizing drug for photodynamic therapy (PDT) used in the treatment of the wet form of macular degeneration and activated by red light of 689 nm. Here, we present the first direct study of its photofragmentation channels in the gas phase, conducted using a laser interfaced mass spectrometer across a broad photoexcitation range from 250 to 790 nm. The photofragmentation channels are compared with the collision-induced dissociation (CID) products revealing similar dissociation pathways characterized by the loss of the carboxyl and ester groups. Complementary solution-phase photolysis experiments indicate that photobleaching occurs in verteporfin in acetonitrile; a notable conclusion, as photoinduced activity in Verteporfin was not thought to occur in homogenous solvent conditions. These results provide unique new information on the thermal break-down products and photoproducts of this light-triggered drug.


Assuntos
Lasers , Fármacos Fotossensibilizantes/farmacologia , Verteporfina/farmacologia , Ésteres , Gases , Luz , Espectrometria de Massas , Fotoquimioterapia , Fotólise , Porfirinas/farmacologia , Espectrometria de Massas por Ionização por Electrospray , Espectrofotometria
12.
J Photochem Photobiol B ; 212: 112039, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-33002779

RESUMO

In this study we report a novel theranostic lipid-polymer liposome, obtained from DPPC and the triblock copolymer F127 covalently modified with 5(6)-carboxyfluorescein (CF) for photodynamic applications. Due to the presence of F127, small unilamellar vesicle (SUV) liposomes were synthesized by a simple and fast thin-film hydration method without the need for an extrusion process. The vesicles have around 100 nm, low polydispersity and superb solution stability. The clinically used photosensitizer verteporfin (VP) was entrapped into the vesicles, mostly in monomeric form, with 90% loading efficiency. Stern-Volmer and fluorescence lifetime assays showed heterogeneous distribution of the VP and CF into the vesicles, ensuring the integrity of their individual photophysical properties. The theranostic properties were entirely photoactivatable and can be trigged by a unique wavelength (470 nm). The feasibility of the system was tested against the Glioblastoma multiforme cell line T98G. Cellular uptake by time-resolved fluorescence microscopy showed monomerized VP (monoexponential decay, 6.0 ns) at nucleus level, while CF was detected at the membrane by fluorescence microscopy. The strategy's success was supported by the reduction of 98% in the viability of T98G cells by the photoactivated lipid-polymer liposome with [VP] = 1.0 µmol L-1. Therefore, the novel theranostic liposome is a potential system for use in cancer and ocular disease therapies.


Assuntos
Fotoquimioterapia/métodos , Verteporfina/administração & dosagem , Verteporfina/farmacologia , Linhagem Celular Tumoral , Estabilidade de Medicamentos , Humanos , Cinética , Lipossomos , Verteporfina/uso terapêutico
13.
BMC Cancer ; 20(1): 1042, 2020 Oct 29.
Artigo em Inglês | MEDLINE | ID: mdl-33121449

RESUMO

BACKGROUND: Breast cancer (BC) can be divided into five subtypes: Lumina1A, Lumina1B, HER-2 overexpression, Basal-like and Normal breast-like subtype, based on the differently expressed genes in breast cancer tissue. The Hippo signaling pathway plays an indispensable role in BC. The YAP gene is a terminal effector of Hippo pathway, and hyperactivation of YAP mediates tumorigenesis. As an inhibitor of YAP, non-photoactivated verteporfin (VP) can inhibit YAP-mediated tumor proliferation and angiogenesis by eliminating its interaction with TEAD. This study aimed to determine the effect and molecular mechanisms of VP-mediated inhibition of YAP in different subtypes of BC. METHODS: Luminal A, Luminal B and Basal-like BC cells were cultivated in vitro to study effects of VP on proliferation and apoptosis of these three molecular BC subtypes. RESULTS: Our experimental results showed that VP inhibited cell proliferation, YAP-TEAD interaction and expression of its downstream targets. VP also induced tumor cell apoptosis, and promoted the cleavage of Caspase-9 and PARP in the cells of various molecular subtypes of BC. CONCLUSION: These findings provide a basis for the use of VP as a potential anti-tumor therapeutic for BC by targeting the Hippo pathway effector YAP.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Apoptose , Neoplasias da Mama/patologia , Proliferação de Células , Fármacos Fotossensibilizantes/farmacologia , Fatores de Transcrição/metabolismo , Verteporfina/farmacologia , Neoplasias da Mama/classificação , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Feminino , Humanos , Transdução de Sinais , Células Tumorais Cultivadas
14.
Sci Rep ; 10(1): 17939, 2020 10 21.
Artigo em Inglês | MEDLINE | ID: mdl-33087792

RESUMO

Infections caused by Staphylococcus aureus pose a serious and sometimes fatal health issue. With the aim of exploring a novel therapeutic approach, we chose GraXRS, a Two-Component System (TCS) that determines bacterial resilience against host innate immune barriers, as an alternative target to disarm S. aureus. Following a drug repurposing methodology, and taking advantage of a singular staphylococcal strain that lacks the whole TCS machinery but the target one, we screened 1.280 off-patent FDA-approved drug for GraXRS inhibition. Reinforcing the connection between this signaling pathway and redox sensing, we found that antioxidant and redox-active molecules were capable of reducing the expression of the GraXRS regulon. Among all the compounds, verteporfin (VER) was really efficient in enhancing PMN-mediated bacterial killing, while topical administration of such drug in a murine model of surgical wound infection significantly reduced the bacterial load. Experiments relying on the chemical mimicry existing between VER and heme group suggest that redox active residue C227 of GraS participates in the inhibition exerted by this FDA-approved drug. Based on these results, we propose VER as a promising candidate for sensitizing S. aureus that could be helpful to combat persistent or antibiotic-resistant infections.


Assuntos
Proteínas de Bactérias/antagonistas & inibidores , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/uso terapêutico , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/genética , Infecção da Ferida Cirúrgica/tratamento farmacológico , Infecção da Ferida Cirúrgica/microbiologia , Verteporfina/farmacologia , Verteporfina/uso terapêutico , Animais , Modelos Animais de Doenças , Reposicionamento de Medicamentos , Humanos , Camundongos , Terapia de Alvo Molecular , Neutrófilos/imunologia , Fagocitose , Staphylococcus aureus/imunologia
15.
Exp Mol Pathol ; 117: 104546, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32976820

RESUMO

RNA binding motif protein 3 (RBM3) has been shown to be upregulated in several types of human tumors. Using tissue microarrays and immunohistochemistry, we showed here that both nuclear and cytoplasmic RBM3 expression levels were higher in hepatocellular carcinoma (HCC) tissues than in adjacent non-tumorous tissues. High nuclear RBM3 was found to be correlated with larger tumor size (P = 0.030), high serum AFP levels (P = 0.011), and advanced Edmonson grading (P = 0.006). Cytoplasmic RBM3 was associated with advanced Edmonson grading (P = 0.003). Kaplan-Meier survival analysis revealed that, although not statistically significant, there was a trend toward shortened overall survival in the subset of HCC patients with high RBM3 expression (both nuclear and cytoplasmic). In addition, we found that RBM3 could promote YAP1 expression in HCC cells. Moreover, we found that YAP1 played an essential part in RBM3-induced proliferation of HCC cells. Furthermore, we demonstrated that Verteporfin, a YAP1 inhibitor, could repress RBM3-induced proliferation of HCC cells. Our findings provide a new experimental basis for further understanding of the possible role of RBM3-YAP1 in the regulation of HCC proliferation.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , Proteínas de Ligação a RNA/genética , Fatores de Transcrição/genética , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Idoso , Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Intervalo Livre de Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Neoplasias Hepáticas/patologia , Masculino , Pessoa de Meia-Idade , Transdução de Sinais , Fatores de Transcrição/antagonistas & inibidores , Verteporfina/farmacologia , alfa-Fetoproteínas/genética
16.
Mol Med Rep ; 22(5): 3955-3961, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32901856

RESUMO

Breast cancer is one of the most aggressive malignant tumors in women. According to the expression differences of estrogen receptor, progesterone receptor, human epidermal growth factor receptor­2 (HER­2) and cell proliferation antigen Ki­67, breast cancer can be divided into four molecular subtypes: Luminal A, Luminal B, HER­2 overexpression and Basal­like. Yes­associated protein (YAP), a downstream effector of the Hippo pathway, is overexpressed in human cancers and is associated with proliferation, apoptosis, migration, invasion and resistance to chemotherapy drugs in breast cancer cells. Verteporfin (VP) is used as a photosensitizer in the treatment of neovascular macular degeneration. VP is also identified as an inhibitor of YAP/TEA domain transcription factor (TEAD) interaction in the absence of light activation. However, detailed structural information about VP and YAP interactions is relatively scarce and VP research targeting YAP in different molecular subtypes of breast cancer cells is also rare. The aims of the present study were to structurally describe the VP binding site in the YAP crystal structure and to verify the non­photoreactive VP effect targeting YAP on the migration of different molecular subtypes of breast cancer cells. The crystal structure of VP and YAP was calculated by AutoDock 4.2 and the result was illustrated using PyMOL. The non­photoactivated VP effect on the migration of Luminal A MCF­7, Luminal B BT­474 and triple­negative breast cancer BT­549 breast cancer cells was evaluated by wound healing and Transwell migration experiments. Results from molecular docking experiments demonstrated that VP could interact through hydrogen bonds and hydrophobic interactions with important YAP residues involved in TEADs binding (Gln82, Val84, Met86 and Arg89). Migration experiments revealed that the non­photoinduced VP could inhibit the migration of different molecular subtypes of breast cancer cells. The results of the present study indicated that VP may be a novel repositioned drug for breast cancer treatment in the future.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/química , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Neoplasias da Mama/metabolismo , Fármacos Fotossensibilizantes/farmacologia , Fatores de Transcrição/química , Fatores de Transcrição/metabolismo , Verteporfina/farmacologia , Sítios de Ligação , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Cristalografia por Raios X , Feminino , Humanos , Células MCF-7 , Modelos Moleculares , Simulação de Acoplamento Molecular , Domínios Proteicos/efeitos dos fármacos
17.
Oncogene ; 39(43): 6647-6663, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32934314

RESUMO

Fibroblast growth factor receptor type 2 (FGFR2) has emerged as a key oncogenic factor that regulates gastric cancer (GC) progression, but the underlying mechanism of FGF-FGFR2 signaling pathway remains largely unknown. To identify the potential molecular mechanisms of the oncogenic FGFR2 in gastric carcinogenesis and convey a novel therapeutic strategy, we profiled the FGFR alterations and analyzed their clinical associations in TCGA and Hong Kong GC cohorts. We found that FGFR2 overexpression in GC cell lines and primary tumors predicted poor survival and was associated with advanced stages of GC. Functionally, growth abilities and cell cycle progression of GC were inhibited by inactivation of ERK-MAPK signal transduction after FGFR2 knockdown, while apoptosis was promoted. Meanwhile, the first-line anti-cancer drug sensitivity was enhanced. RNA-seq analysis further revealed that YAP1 signaling serves as a significant downstream modulator and mediates the oncogenic signaling of FGFR2. When stimulating FGFR2 by rhFGF18, we observed intensified F-actin, nuclear accumulation of YAP1, and overexpression of YAP1 targets, but these effects were attenuated by either FGFR2 depletion or AZD4547 administration. Additionally, the FGF18-FGFR2 signaling upregulated YAP1 expression through activating c-Jun, an effector of MAPK signaling. In our cohort, 28.94% of GC cases were characterized as FGFR2, c-Jun, and YAP1 co-positive and demonstrated worse clinical outcomes. Remarkably, we also found that co-targeting FGFR2 and YAP1 by AZD4547 and Verteporfin synergistically enhanced the antitumor effects in vitro and in vivo. In conclusion, we have identified the oncogenic FGF-FGFR2 regulates YAP1 signaling in GC. The findings also highlight the translational potential of FGFR2-c-Jun-YAP1 axis, which may serve as a prognostic biomarker and therapeutic target for GC.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Fatores de Crescimento de Fibroblastos/metabolismo , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/metabolismo , Neoplasias Gástricas/patologia , Fatores de Transcrição/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Benzamidas/farmacologia , Benzamidas/uso terapêutico , Carcinogênese/efeitos dos fármacos , Carcinogênese/patologia , Linhagem Celular Tumoral , Estudos de Coortes , Variações do Número de Cópias de DNA , Conjuntos de Dados como Assunto , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Humanos , Estimativa de Kaplan-Meier , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Camundongos , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Piperazinas/farmacologia , Piperazinas/uso terapêutico , Prognóstico , Intervalo Livre de Progressão , Proteínas Proto-Oncogênicas c-jun/metabolismo , Pirazóis/farmacologia , Pirazóis/uso terapêutico , RNA-Seq , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/antagonistas & inibidores , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/genética , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/genética , Neoplasias Gástricas/mortalidade , Fatores de Transcrição/antagonistas & inibidores , Regulação para Cima , Verteporfina/farmacologia , Verteporfina/uso terapêutico , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Biosci Rep ; 40(8)2020 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-32779719

RESUMO

Platelet transfusion is required for life-threatening thrombocytopenic bleeding, and single donor platelet concentrate is the ideal transfusion product. However, due to the inadequate number of donors that can donate a large volume of platelets, in vitro platelets production could be an alternative. We developed an in vitro production system designed to increase the platelet production yield from cultured cells. Previously, we reported that depletion of a Hippo pathway core kinase (LATS1/2) inhibited platelet production from cultured megakaryocytes. In the present study, we further investigated the role of the Hippo pathway in megakaryocyte proliferation and platelet production by focusing on the role of its effector proteins (YAP and TAZ), which are down-stream targets of LATS1/2 kinase. We found that YAP plays an essential role in megakaryoblastic cell proliferation, maturation, and platelet production, while TAZ showed minor effect. Knockdown of YAP, either by genetic manipulation or pharmaceutical molecule, significantly increased caspase-3-mediated apoptosis in cultured megakaryocytes, and increased platelet production as opposed to overexpressing YAP. We, therefore, demonstrate a paradigm for the regulation of megakaryocyte development and platelet production via the Hippo signaling pathway, and suggest the potential use of an FDA-approved drug to induce higher platelet production in cultured cells.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Plaquetas/metabolismo , Megacariócitos/metabolismo , Trombopoese , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/genética , Apoptose , Plaquetas/efeitos dos fármacos , Linhagem Celular Tumoral , Dobutamina/farmacologia , Regulação da Expressão Gênica , Humanos , Megacariócitos/efeitos dos fármacos , Transdução de Sinais , Trombopoese/efeitos dos fármacos , Transativadores/genética , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genética , Verteporfina/farmacologia
19.
Exp Neurol ; 333: 113431, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32750359

RESUMO

Astrocytic Yes-associated protein (YAP) has been implicated in astrocytic proliferation and differentiation in the developing neocortex. However, the role of astrocytic YAP in diseases of the nervous system remains poorly understood. Here, we hypothesized that astrocytic YAP exerted a neuroprotective effect against cerebral ischemic injury in rats by regulating signal transducer and activator of transcription 3 (STAT3) signaling. In this study, we investigated whether the expression of nuclear YAP in the astrocytes of rats increased significantly after middle cerebral artery occlusion (MCAO) and its effect on cerebral ischemic injury. We used XMU-MP-1 to trigger localization of YAP into the nucleus and found that XMU-MP-1 treatment decreased ischemia/stroke-induced brain injury including reduced neuronal death and reactive astrogliosis, and extenuated release of interleukin-1ß (IL-1ß), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α). Mechanically, XMU-MP-1 treatment suppressed the expression of phospho-STAT3 (P-STAT3). We established an in-vitro oxygen-glucose deprivation/reperfusion (OGD/R) model to simulate an ischemic condition and further explore the function of astrocytic YAP. We found that nuclear translocation of astrocytic YAP in rats could improve cell vitality, decrease the release of inflammatory cytokines and reduce the expression of P-STAT3 in vitro. In contrast, we also found that inhibition of YAP by verteporfin further aggravated the injury induced by OGD/R via STAT3 signaling. In summary, our results showed that nuclear localization of astrocytic YAP exerted a neuroprotective effect after cerebral ischemic injury in rats via inhibition of the STAT3 signaling.


Assuntos
Proteínas Reguladoras de Apoptose/farmacologia , AVC Isquêmico/prevenção & controle , Fármacos Neuroprotetores , Fator de Transcrição STAT3/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Astrócitos/efeitos dos fármacos , Citocinas/metabolismo , Glucose/deficiência , Hipóxia Encefálica/prevenção & controle , Infarto da Artéria Cerebral Média/prevenção & controle , AVC Isquêmico/patologia , Masculino , Ratos , Ratos Sprague-Dawley , Traumatismo por Reperfusão/prevenção & controle , Verteporfina/farmacologia
20.
Mol Cell Biochem ; 475(1-2): 79-91, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32761300

RESUMO

Hepatocellular carcinoma (HCC) is one of the most lethal malignancies worldwide. However, the immune tolerance limits the effect of chemotherapeutic drugs. Therefore, the mechanism of cisplatin in promoting PD-L1 expression by YAP1 was investigated in the present study, and we found that cisplatin increased the expression level of YAP1 in the mouse liver with H22 cells. Meanwhile, cisplatin improved the expression level of PD-L1, IL-1ß and CCL2 in the tumor microenvironment. Further, cisplatin also enhanced the expression level of YAP1 in shYAP1 HepG2215 cells. The expression of PD-L1 was decreased by Verteporfin, YAP1 inhibitor, during the treatment of DEN/TCPOBOP-induced liver cancer in C57BL/6 mice. These results suggested that cisplatin could deteriorate the immunosuppressive microenvironment through increasing PD-L1, CCL2, IL-1ß by upregulated YAP1 expression. Therefore, the study suggested that YAP1 blockade destroyed the immunosuppressive microenvironment of cancer to improve the effect of chemotherapy in HCC.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Antígeno B7-H1/biossíntese , Carcinoma Hepatocelular/metabolismo , Cisplatino/farmacologia , Neoplasias Hepáticas/metabolismo , Fatores de Transcrição/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Antineoplásicos/farmacologia , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Células Hep G2 , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas Experimentais/tratamento farmacológico , Neoplasias Hepáticas Experimentais/metabolismo , Neoplasias Hepáticas Experimentais/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Fármacos Fotossensibilizantes/farmacologia , Transdução de Sinais , Fatores de Transcrição/genética , Microambiente Tumoral , Verteporfina/farmacologia
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