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Background: There is no established standard 3rd line treatment for patients with advanced non-small cell lung cancer (NSCLC). Although cytotoxic chemotherapeutic agents that are not used as 1st or 2nd line treatment are administrated as 3rd line treatment, their anti-tumor efficacy is insufficient. Anti-programmed death ligand-1 (PD-L1)/programmed death-1 (PD1) treatment is more effective and less toxic than chemotherapy in anti-PD-L1/PD-1 treatment-naïve patients with NSCLC. Therefore, anti-PD-L1/PD-1 therapy is considered an appropriate 3rd line treatment. However, the anti-tumor efficacy is limited in patients previously treated with anti-PD-L1/PD-1 antibody. Today, new drugs are needed to increase the efficacy of anti-PD-L1/PD-1 antibodies. Methods: This open-label, single-arm, investigator-initiated phase II study is designed to evaluate combination treatment of nivolumab and TM5614, a plasminogen activator inhibitor (PAI-1) inhibitor as 3rd or more line treatment in NSCLC patients who underwent standard treatment. The primary endpoint is the objective response rate and the secondary endpoints are progression-free survival (PFS), overall survival (OS), duration of response (DOR) and safety. Recruitment began in September 2023 and is expected to continue for approximately three years. Discussion: Currently, there is no standard 3rd line treatment for advanced NSCLC, and we hope that the findings of this study will facilitate more effective treatments in this setting. Ethics and dissemination: the study protocol conformed to the ethical principles outlined in the Declaration of Helsinki. All patients will provide written informed consent prior to enrollment. Results will be published in a peer-reviewed publication. Trial Registration: This study is registered to Japan Registry of Clinical Trials with number: jRCT2061230039 (19/July/2023).
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BACKGROUND: Anti-programmed cell death 1 antibodies (anti-PD-1 Abs) are widely used for advanced melanoma, but the efficacy of an anti-PD-1 Abs is limited in the Asian population. There remains an unmet need to improve the therapeutic effects of anti-PD-1 Abs treatment, particularly in melanoma patients who are refractory to anti-PD-1 Abs. The aim was to evaluate anti-PD-1 Abs treatment in combination with TM5614 (plasminogen activator inhibitor-1: PAI-1 inhibitor) in patients with unresectable melanoma. METHODS: The TM5614-MM study was a multicentre, open-label, single-arm, phase 2 clinical trial to evaluate the efficacy and safety of nivolumab in combination with TM5614 in patients with advanced, unresectable malignant melanoma recruited at 7 Japanese institutes between 13 September 2021 and 31 March 2023. Patients with metastatic or unresectable melanoma previously treated with anti-PD-1 Abs were enrolled. Nivolumab 480 mg was administered intravenously every 4 weeks for 8 weeks, while TM5614 was administered orally at a dose of 120 mg (0-4 weeks) and 180 mg once daily (5-8 weeks). The primary endpoint was the overall response rate after 8 weeks of concomitant use of TM5614. RESULTS: Thirty nine patients were enrolled, and 34 patients in the anti-PD-1 Abs-refractory cohort. The overall response rate at 8 weeks was 25.9% (95% CI: 12.9-44.9%; P = .027) in 27 anti-PD-1-Abs refractory patients by investigator assessment in the protocol per set cohort. Seven patients discontinued treatment due to progressive disease or adverse events. Treatment-related grade 3 or higher adverse events occurred in 3 of 39 patients (7.7%) in the intention-to-treat cohort. CONCLUSIONS: TM5614 in combination with nivolumab is well-tolerated and effective in anti-PD-1 Abs-refractory, unresectable melanoma. TRIAL REGISTRATION: This trial was registered with Clinical Trial gov, jRCT2021210029.
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Background: Increased levels of plasminogen activator inhibitor-1 (PAI-1) in tumors have been found to correlate with poor clinical outcomes in patients with cancer. Although abundant data support the involvement of PAI-1 in cancer progression, whether PAI-1 contributes to tumor immune surveillance remains unclear. The purposes of this study are to determine whether PAI-1 regulates the expression of immune checkpoint molecules to suppresses the immune response to cancer and demonstrate the potential of PAI-1 inhibition for cancer therapy. Methods: The effects of PAI-1 on the expression of the immune checkpoint molecule programmed cell death ligand 1 (PD-L1) were investigated in several human and murine tumor cell lines. In addition, we generated tumor-bearing mice and evaluated the effects of a PAI-1 inhibitor on tumor progression or on the tumor infiltration of cells involved in tumor immunity either alone or in combination with immune checkpoint inhibitors. Results: PAI-1 induces PD-L1 expression through the JAK/STAT signaling pathway in several types of tumor cells and surrounding cells. Blockade of PAI-1 impedes PD-L1 induction in tumor cells, significantly reducing the abundance of immunosuppressive cells at the tumor site and increasing cytotoxic T-cell infiltration, ultimately leading to tumor regression. The anti-tumor effect elicited by the PAI-1 inhibitor is abolished in immunodeficient mice, suggesting that PAI-1 blockade induces tumor regression by stimulating the immune system. Moreover, combining a PAI-1 inhibitor with an immune checkpoint inhibitor significantly increases tumor regression. Conclusions: PAI-1 protects tumors from immune surveillance by increasing PD-L1 expression; hence, therapeutic PAI-1 blockade may prove valuable in treating malignant tumors.
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Antígeno B7-H1 , Inibidor 1 de Ativador de Plasminogênio , Evasão Tumoral , Animais , Feminino , Humanos , Camundongos , Antígeno B7-H1/metabolismo , Antígeno B7-H1/genética , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Evasão da Resposta Imune , Camundongos Endogâmicos C57BL , Neoplasias/imunologia , Neoplasias/metabolismo , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Inibidor 1 de Ativador de Plasminogênio/genética , Transdução de Sinais , Microambiente Tumoral/imunologiaRESUMO
Cutaneous angiosarcoma (CAS) is an endothelial cell-derived, highly aggressive type of vascular tumour. Although chemoradiotherapy with paclitaxel (PTX) is recognized as a first-line therapy for CAS, second-line therapy for CAS remains controversial, and there is no standard therapy for taxane-resistant CAS. Plasminogen activator inhibitor-1 (PAI-1) is associated with poor clinical outcomes, and elevated levels of PAI-1 in both tissue and serum are correlated with poor response to therapy in various cancers, including skin cancers. Since PAI-1 protects endothelial cells from Fas ligand-mediated apoptosis, PAI-1 inhibition might induce apoptosis of endothelial cell-derived tumours such as CAS. This is a single-arm, open-label, multi-institutional, Phase 2 clinical trial to assess the efficacy and safety of PTX in combination with TM5614 (PAI-1 inhibitor) in patients with PTX-resistant CAS. PTX will be administered for 28 weeks, with oral administration of TM5614. The primary endpoint of this study will be the overall response rate (ORR) at 28 weeks after starting treatment (central image evaluation). The secondary endpoint will include the ORR at 28 weeks after starting treatment (investigator evaluation), ORR at 8 weeks and 16 weeks after initiation of treatment (central and investigator evaluation), progression-free survival, overall survival, disease control rate and safety profiles. Assuming the null hypothesis of a response rate of 13.6% and an alternative hypothesis of 45%, a minimum of 15 patients are required to achieve a two-sided, Type I error of 5% and power of 70% based on the exact binomial distribution. Data quality control will be conducted by a combination of centralized (remote) and on-site monitoring. This study will contribute to the development of novel combination therapy for PTX-resistant CAS patients, which remains an unmet clinical need.
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Protocolos de Quimioterapia Combinada Antineoplásica , Hemangiossarcoma , Neoplasias Cutâneas , Humanos , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Ensaios Clínicos Fase II como Assunto , Células Endoteliais , Hemangiossarcoma/tratamento farmacológico , Paclitaxel/farmacologia , Paclitaxel/uso terapêutico , Inibidor 1 de Ativador de Plasminogênio , Neoplasias Cutâneas/tratamento farmacológico , Estudos Multicêntricos como AssuntoRESUMO
Congenital fibroblast growth factor 23 (FGF23)-related hypophosphatemic rickets/osteomalacia is a rare bone metabolism disorder characterized by hypophosphatemia and caused by genetic abnormalities that result in excessive secretion of FGF23. Hyp mice are a model of X-linked hypophosphatemia (XLH) caused by deletion of the PHEX gene and excessive production of FGF23. The purpose of this study was to investigate the potential of TM5614 as a therapeutic agent for the treatment of congenital FGF23-related hypophosphatemic rickets and osteomalacia in humans by administering TM5614 to Hyp mice and examining its curative effect on hypophosphatemia. After a single oral administration of TM5614 10 mg·kg-1 to female Hyp mice starting at 17 weeks of age, the serum phosphate concentration increased with a peak at 6 h after administration. ELISA confirmed that TM5614 administration decreased the intact FGF23 concentration in the blood. Expression of 25-hydroxyvitamin D-1α-hydroxylase protein encoded by Cyp27b1 mRNA in the kidney was suppressed in Hyp mice, and treatment with 10 mg·kg-1 of TM5614 normalized the expression of 25-hydroxyvitamin D-1α-hydroxylase protein and Cyp27b1 mRNA in the kidneys of these mice. Our data indicate that oral administration of TM5614 ameliorates hypophosphatemia in Hyp mice, suggesting that TM5614 may be an effective treatment for congenital FGF23-related hypophosphatemic rickets and osteomalacia.
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Raquitismo Hipofosfatêmico Familiar , Hipofosfatemia , Osteomalacia , Camundongos , Feminino , Humanos , Animais , Raquitismo Hipofosfatêmico Familiar/tratamento farmacológico , Raquitismo Hipofosfatêmico Familiar/metabolismo , Inibidor 1 de Ativador de Plasminogênio , Osteomalacia/tratamento farmacológico , Osteomalacia/metabolismo , 25-Hidroxivitamina D3 1-alfa-Hidroxilase/uso terapêutico , Hipofosfatemia/tratamento farmacológico , Hipofosfatemia/metabolismo , RNA Mensageiro/metabolismoRESUMO
BACKGROUND: We recently showed that pharmacological inhibition of plasminogen activator inhibitor-1 (PAI-1) activity, based on TM5614, increases cell motility and induces the detachment of hematopoietic stem cells from their niches. In this TM5614 phase II clinical trial, we investigated whether the combination of a PAI-1 inhibitor and tyrosine kinase inhibitors (TKIs) would induce a deep molecular response (DMR) in patients affected by chronic myeloid leukemia (CML) by quantifying BCR-ABL1 transcripts. METHODS: Patients with chronic phase CML treated with a stable daily dose of TKIs for at least 1 year and yielding a major molecular response (MMR) but not achieving MR4.5 were eligible for this study. After inclusion, patients began to receive TM5614 as well as a TKI. The primary objective was an evaluation of the cumulative incidence of patient progression from an MMR/MR4 to MR4.5 by 12 months. RESULTS: Thirty-three patients were enrolled in the study. The median age was 59.0 years and 58% were male. No Sokal high-risk patients were enrolled in this trial. The median TKI treatment duration was 4.8 years. At the start of this study, seven patients and 26 patients received imatinib and second-generation TKIs, respectively. The cumulative MR4.5 incidence by 12 months was 33.3% (95% confidence interval, 18.0%-51.8%). The cumulative MR4.5 spontaneous conversion over 12 months was estimated as 8% with TKIs alone based on historical controls. The halving time of BCR-ABL1 at 2 months was significantly shorter for patients who achieved an MR4.5 , by 12 months than for the other patients (cutoff value: 48 days; sensitivity: 0.80; specificity: 0.91; ROC-AUC: 0.83). During this study, bleeding events and abnormal coagulation related to the drug were not reported, and TM5614 was found to be highly safe. CONCLUSION: TM5614 combined with TKI was well tolerated and induced MR4.5 in more patients than stand-alone TKI treatment.
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Leucemia Mielogênica Crônica BCR-ABL Positiva , Leucemia Mieloide de Fase Crônica , Humanos , Masculino , Pessoa de Meia-Idade , Feminino , Proteínas de Fusão bcr-abl/genética , Inibidor 1 de Ativador de Plasminogênio/genética , Inibidores de Proteínas Quinases/efeitos adversos , Leucemia Mieloide de Fase Crônica/tratamento farmacológico , Leucemia Mieloide de Fase Crônica/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genéticaRESUMO
Background: A diagnosis with histological classification by pathologists is very important for appropriate treatments to improve the prognosis of patients with breast cancer. However, the number of pathologists is limited, and assisting the pathological diagnosis by artificial intelligence becomes very important. Here, we presented an automatic breast lesions detection model using microscopic histopathological images based on a Single Shot Multibox Detector (SSD) for the first time and evaluated its significance in assisting the diagnosis. Methods: We built the data set and trained the SSD model with 1361 microscopic images and evaluated using 315 images. Pathologists and medical students diagnosed the images with or without the assistance of the model to investigate the significance of our model in assisting the diagnosis. Results: The model achieved 88.3% and 90.5% diagnostic accuracies in 3-class (benign, non-invasive carcinoma, or invasive carcinoma) or 2-class (benign or malignant) classification tasks, respectively, and the mean intersection over union was 0.59. Medical students achieved a remarkably higher diagnostic accuracy score (average 84.7%) with the assistance of the model compared to those without assistance (average 67.4%). Some people diagnosed images in a short time using the assistance of the model (shorten by average 6.4â¯min) while others required a longer time (extended by 7.2â¯min). Conclusion: We presented the automatic breast lesions detection method at high speed using histopathological micrographs. The present system may conveniently support the histological diagnosis by pathologists in laboratories.
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Chronic myeloid leukemia (CML) is triggered by t(9;22)(q34;q11.2) translocation, leading to the formation of the BCR-ABL1 fusion gene. Although the development of BCR-ABL1 tyrosine kinase inhibitors (TKIs) has dramatically improved the prognosis of CML, the disease could often relapse, presumably because leukemic stem cell fraction of CML (CML-LSC) may reside in specific niches, and also acquire an ability to resist the cytotoxic agents. Recently a study indicated that pharmacological inhibition of plasminogen activator inhibitor-1 (PAI-1, also known as SERPINE1) would cause detachment of CML-LSCs from their niche by inducing maturation of membrane-type matrix metalloprotease-1 (MT1-MMP), leading to increased susceptibility of CML-LSCs against TKIs. However, the direct antitumor effect of PAI-1 inhibition in CML remains unclear. Because PAI-1 mRNA expression was lower in CML cell line (K562) than bone marrow mononuclear cells derived from CML patients, we established K562 cell clones stably expressing exogenous PAI-1 (K562/PAI-1). We found that TM5614 treatment significantly suppressed cell proliferation and induced apoptosis in K562/PAI-1 cells, accompanied by increased activity of Furin protease, which is a known target of PAI-1. Besides processing mature MT1-MMP, Furin is in charge of cleaving the NOTCH receptor to form a heterodimer before exporting it to the cell surface membrane. In K562/PAI-1 cells, TM5614 treatment increased NOTCH1 intracellular domain (NICD) protein expression as well as NOTCH1 target of HEY1 mRNA levels. Finally, forced expression of either Furin or NICD in K562/PAI-1 cells significantly inhibited cell proliferation and induced apoptosis. Collectively, PAI-1 inhibition may have an antitumor effect by modulating the Furin/NICD pathway.
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Antineoplásicos , Furina , Leucemia Mielogênica Crônica BCR-ABL Positiva , Antineoplásicos/farmacologia , Apoptose , Resistencia a Medicamentos Antineoplásicos , Humanos , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Metaloproteinase 14 da Matriz/metabolismo , Inibidor 1 de Ativador de Plasminogênio/genética , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , RNA MensageiroRESUMO
Bone marrow adipocytes (BMAs) have recently been recognized as a niche component with a suppressive function. Obese individuals with abundant BMAs exhibit impaired hematopoietic regeneration after hematopoietic stem cell transplantation (HSCT). We hypothesized that plasminogen activator inhibitor type-1 (PAI-1), an adipokine that regulates the fibrinolytic system, contributes to impaired hematopoiesis in bone marrow (BM) microenvironment with abundant BMAs. We demonstrated that BMAs differentiated in vitro could secrete PAI-1 and were positive for PAI-1 in vivo. In addition, the abundance of BMAs was associated with high levels of PAI-1 expression. The BMA-rich microenvironment exhibited impaired hematopoietic regeneration after HSCT when compared with a BMA-less microenvironment. The impaired hematopoietic regeneration in BMA-rich microenvironment was significantly alleviated by PAI-1 knockout or PAI-1 inhibitor treatment. Obese mice with abundant BMAs, compared with normal-weight mice, exhibited higher bone marrow PAI-1 concentrations, increased fibrinolytic system suppression, and lower stem cell factor (SCF) concentrations after HSCT. PAI-1 inhibitor administration significantly activated the fibrinolytic system in obese mice, contributing to the higher SCF concentration. Moreover, PAI-1 inhibitor treatment significantly alleviated the impaired hematopoietic regeneration in obese mice both after 5-fluorouracil injection and HSCT. These results indicate that PAI-1 hinders hematopoietic regeneration in BMA-rich microenvironments. The blockade of PAI-1 activity could be a novel therapeutic means of facilitating hematopoietic reconstitution in BMA-rich patients.
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Adipócitos/metabolismo , Medula Óssea/efeitos dos fármacos , Hematopoese/efeitos dos fármacos , Células-Tronco Hematopoéticas/efeitos dos fármacos , Obesidade/metabolismo , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Inibidor 1 de Ativador de Plasminogênio/farmacologia , Animais , Antimetabólitos/farmacologia , Medula Óssea/metabolismo , Fluoruracila/farmacologia , Técnicas de Inativação de Genes , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Obesidade/genética , Obesidade/terapia , Inibidor 1 de Ativador de Plasminogênio/genética , Regeneração/efeitos dos fármacos , Fator de Células-Tronco/metabolismo , Nicho de Células-Tronco/efeitos dos fármacosRESUMO
Plasminogen activator inhibitor-1 (PAI-1), a key regulator of the fibrinolytic system, is the main physiological inhibitor of plasminogen activators. By interacting with matrix components, including vitronectin (Vn), PAI-1 plays a regulatory role in tissue remodeling, cell migration, and intracellular signaling. Emerging evidence points to a role for PAI-1 in various pathological conditions, including cardiovascular diseases, cancer, and fibrosis. Targeting PAI-1 is therefore a promising therapeutic strategy in PAI-1-related pathologies. A class of small molecule inhibitors including TM5441 and TM5484, designed to bind the cleft in the central ß-sheet A of PAI-1, showed to be potent PAI-1 inhibitors in vivo. However, their binding site has not yet been confirmed. Here, we report two X-ray crystallographic structures of PAI-1 in complex with TM5484. The structures revealed a binding site at the flexible joint region, which is distinct from the presumed binding site. Based on the structural analysis and biochemical data we propose a mechanism for the observed dose-dependent two-step mechanism of PAI-1 inhibition. By binding to the flexible joint region in PAI-1, TM5484 might restrict the structural flexibility of this region, thereby inducing a substrate form of PAI-1 followed by a conversion to an inert form.
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Inibidor 1 de Ativador de Plasminogênio/efeitos dos fármacos , Sítios de Ligação , Domínio Catalítico , Cristalização , Cristalografia por Raios X , Relação Dose-Resposta a Droga , Humanos , Modelos Moleculares , Inibidor 1 de Ativador de Plasminogênio/química , Conformação Proteica , Relação Estrutura-AtividadeRESUMO
Plasminogen activator inhibitor 1 (PAI-1) is a functional biomarker of the metabolic syndrome. Previous studies have demonstrated that PAI-1 is a mechanistic contributor to several elements of the syndrome, including obesity, hypertension and insulin resistance. Here we show that PAI-1 is also a critical regulator of hepatic lipid metabolism. RNA sequencing revealed that PAI-1 directly regulates the transcriptional expression of numerous genes involved in mammalian lipid homeostasis, including PCSK9 and FGF21. Pharmacologic or genetic reductions in plasma PAI-1 activity ameliorates hyperlipidemia in vivo. These experimental findings are complemented with the observation that genetic deficiency of PAI-1 is associated with reduced plasma PCSK9 levels in humans. Taken together, our findings identify PAI-1 as a novel contributor to mammalian lipid metabolism and provides a fundamental mechanistic insight into the pathogenesis of one of the most pervasive medical problems worldwide.
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Dislipidemias/genética , Fígado Gorduroso/genética , Inibidor 1 de Ativador de Plasminogênio/fisiologia , Animais , Células Cultivadas , Estudos de Coortes , Dislipidemias/metabolismo , Fígado Gorduroso/metabolismo , Feminino , Fatores de Crescimento de Fibroblastos/genética , Células Hep G2 , Humanos , Metabolismo dos Lipídeos/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Pró-Proteína Convertase 9/genéticaRESUMO
Therapeutic strategies that target leukemic stem cells (LSCs) provide potential advantages in the treatment of chronic myeloid leukemia (CML). Here, we show that selective blockade of plasminogen activator inhibitor-1 (PAI-1) enhances the susceptibility of CML-LSCs to tyrosine kinase inhibitor (TKI), which facilitates the eradication of CML-LSCs and leads to sustained remission of the disease. We demonstrated for the first time that TGF-ß-PAI-1 axis was selectively augmented in CML-LSCs in the bone marrow (BM), whereby protecting CML-LSCs from TKI treatment. Furthermore, the combined administration of TKI plus a PAI-1 inhibitor, in a mouse model of CML, significantly enhanced the eradication of CML cells in the BM and prolonged the survival of CML mice. The combined therapy of imatinib and a PAI-1 inhibitor prevented the recurrence of CML-like disease in serially transplanted recipients, indicating the elimination of CML-LSCs. Interestingly, PAI-1 inhibitor treatment augmented membrane-type matrix metalloprotease-1 (MT1-MMP)-dependent motility of CML-LSCs, and the anti-CML effect of PAI-1 inhibitor was extinguished by the neutralizing antibody for MT1-MMP, underlining the mechanistic importance of MT1-MMP. Our findings provide evidence of, and a rationale for, a novel therapeutic tactic, based on the blockade of PAI-1 activity, for CML patients.
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Proteínas de Fusão bcr-abl , Leucemia Mielogênica Crônica BCR-ABL Positiva , Animais , Humanos , Mesilato de Imatinib , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Camundongos , Células-Tronco Neoplásicas , Inibidor 1 de Ativador de Plasminogênio , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Serpina E2RESUMO
An orally bioavailable small molecule inhibitor of plasminogen activator inhibitor1 (PAI1) is currently being clinically assessed as a novel antithrombotic agent. Although PAI1 is known to serve a key role in the pathogenesis of metabolic syndrome (MetS) including nonalcoholic steatohepatitis (NASH), the pharmacological action of an oral PAI1 inhibitor against the development of MetSrelated liver fibrosis remains unclear. The current study was designed to explicate the effect of TM5275, an oral PAI1 inhibitor, on MetSrelated hepatic fibrogenesis. The in vivo antifibrotic effect of orally administered TM5275 was investigated in two different rat MetS models. Fischer 344 rats received a cholinedeficient Laminoaciddefined diet for 12 weeks to induce steatohepatitis with development of severe hepatic fibrosis. Otsuka LongEvans Tokushima Fatty rats, used to model congenital diabetes, underwent intraperitoneal injection of porcine serum for 6 weeks to induce hepatic fibrosis under diabetic conditions. In each experimental model, TM5275 markedly ameliorated the development of hepatic fibrosis and suppressed the proliferation of activated hepatic stellate cells (HSCs). Additionally, the hepatic production of tumor growth factor (TGF)ß1 and total collagen was suppressed. In vitro assays revealed that TGFß1 stimulated the upregulation of Serpine1 mRNA expression, which was inhibited by TM5275 treatment in cultured HSCT6 cells, a rat HSC cell line. Furthermore, TM5275 substantially attenuated the TGFß1stimulated proliferative and fibrogenic activity of HSCs by inhibiting AKT phosphorylation. Collectively, TM5275 demonstrated an antifibrotic effect on liver fibrosis in different rat MetS models, suppressing TGFß1induced HSC proliferation and collagen synthesis. Thus, PAI1 inhibitors may serve as effective future therapeutic agents against NASHbased hepatic fibrosis.
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Fibrinolíticos/administração & dosagem , Células Estreladas do Fígado/efeitos dos fármacos , Cirrose Hepática Experimental/tratamento farmacológico , Cirrose Hepática Experimental/etiologia , Síndrome Metabólica/complicações , Hepatopatia Gordurosa não Alcoólica/complicações , Piperazinas/administração & dosagem , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , para-Aminobenzoatos/administração & dosagem , Administração Oral , Animais , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Modelos Animais de Doenças , Células Estreladas do Fígado/metabolismo , Masculino , Síndrome Metabólica/induzido quimicamente , Hepatopatia Gordurosa não Alcoólica/induzido quimicamente , Ratos , Ratos Endogâmicos F344 , Ratos Long-Evans , Fator de Crescimento Transformador beta1/metabolismoRESUMO
Senescence of alveolar type II (ATII) cells, progenitors of the alveolar epithelium, is a pathological feature and contributes importantly to the pathogenesis of idiopathic pulmonary fibrosis. Despite recognition of the importance of ATII cell senescence in idiopathic pulmonary fibrosis pathogenesis, how ATII cell senescence is regulated and how senescent ATII cells contribute to lung fibrogenesis remain unclear. In this study, we show that TGF-ß1 (transforming growth factor-ß1), a most ubiquitous and potent profibrotic cytokine, induces plasminogen activator inhibitor-1 (PAI-1), a cell senescence and fibrosis mediator, and p16 as well as senescence, but not apoptosis, in primary mouse ATII cells. We also found that senescent ATII cells secrete various cytokines and chemokines, including IL-4 and IL-13, which stimulate the expression of genes associated with a profibrotic phenotype in alveolar macrophages. Similar responses were also observed in TGF-ß1-treated rat ATII (L2) and rat macrophage NR8383 cells. Deletion of PAI-1 or inhibition of PAI-1 activity with a small molecule PAI-1 inhibitor, however, blocks TGF-ß1-induced senescence as well as a senescence-associated secretory phenotype in ATII and L2 cells and, consequently, the stimulatory effects of the conditioned medium from senescent ATII/L2 cells on macrophages. Moreover, we show that silencing p16 ameliorates PAI-1 protein-induced ATII cell senescence and secretion of profibrotic mediators. Our data suggest that PAI-1 mediates TGF-ß1-induced ATII cell senescence and secretion of profibrotic mediators through inducing p16, and they also suggest that senescent ATII cells contribute to lung fibrogenesis in part by activating alveolar macrophages through secreting profibrotic and proinflammatory mediators.
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Células Epiteliais Alveolares/citologia , Senescência Celular/fisiologia , Ativação de Macrófagos/fisiologia , Macrófagos Alveolares/fisiologia , Serpina E2/fisiologia , Fator de Crescimento Transformador beta1/fisiologia , Células Epiteliais Alveolares/metabolismo , Animais , Células Cultivadas , Quimiocinas/metabolismo , Meios de Cultivo Condicionados/farmacologia , Inibidor p16 de Quinase Dependente de Ciclina/fisiologia , Citocinas/metabolismo , Genes p16 , Camundongos , Camundongos Knockout , Fibrose Pulmonar/patologia , Interferência de RNA , RNA Interferente Pequeno/genética , Ratos , Serpina E2/deficiência , Serpina E2/genéticaRESUMO
Upon hematopoietic stem cell transplantation (HSCT), the availability of recipients' niches in the bone marrow (BM) is one of the factors that influence donor HSC engraftment and hematopoietic reconstitution. Therefore, myeloablative conditioning, such as irradiation and/or chemotherapy, which creates empty niches in the recipients' BM, is required for the success of HSCT. However, the conventional myeloablation causes extensive damages to the patients' BM, which results in the treatment-induced severe complications and even mortality. Thus, alternative and mild conditioning could fulfill the need for safer HSCT-based therapies for hematological and nonhematological disorders. Recently, we have demonstrated that pharmacological inhibition of plasminogen activator inhibitor-1 (PAI-1) activity increases cellular motility and cause detachment of HSCs from the niches. In this study, we performed HSCT using a PAI-1 inhibitor without any myeloablative conditioning. Donor HSCs were transplanted to recipient mice that were pretreated with saline or a PAI-1 inhibitor. Saline pretreated nonmyeloablative recipients showed no engraftment. In contrast, donor cell engraftment was detected in the PAI-1 inhibitor pretreated recipients. Multilineage differentiation, including lymphoid and myeloid cells, was observed in the PAI-1 inhibitor pretreated recipients. Donor-derived cells that exhibited multilineage reconstitution as well as the existence of stem/progenitor cells were detected in the secondary recipients, confirming the maintenance of donor HSCs in the BM of PAI-1 inhibitor pretreated primary recipients. The results indicate that the PAI-1 blockade vacates functional niches in the recipients' BM, which allows the engraftment of long-term multilineage HSCs without myeloablative conditioning.
Assuntos
Medula Óssea/metabolismo , Transplante de Células-Tronco Hematopoéticas , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Nicho de Células-Tronco , Condicionamento Pré-Transplante , Animais , Linhagem da Célula , Células-Tronco Hematopoéticas/metabolismo , CamundongosRESUMO
Alzheimer's disease (AD) is a major cause of dementia in the elderly with no effective treatment. Accumulation of amyloid-ß peptide (Aß) in the brain is a pathological hallmark of AD and is believed to be a central disease-causing and disease-promoting event. In a previous study, we showed that deletion of plasminogen activator inhibitor 1 (PAI-1), a primary inhibitor of tissue type and urokinase type plasminogen activators (tPA and uPA), significantly reduced brain Aß load in APP/PS1 mice, an animal model of familial AD. In this study, we further show that oral administration of TM5275, a small molecule inhibitor of PAI-1, for a period of 6 weeks, inhibits the activity of PAI-1 and increases the activities of tPA and uPA as well as plasmin, which is associated with a reduction of Aß load in the hippocampus and cortex and improvement of learning/memory function in APP/PS1 mice. Protein abundance of low density lipoprotein related protein-1 (LRP-1), a multi ligand endocytotic receptor involved in transporting Aß out of the brain, as well as plasma Aß42 are increased, whereas the expression and processing of full-length amyloid-ß protein precursor is not affected by TM5275 treatment in APP/PS1 mice. In vitro studies further show that PAI-1 increases, whereas TM5275 reduces, Aß40 level in the culture medium of SHSY5Y-APP neuroblastoma cells. Collectively, our data suggest that TM5275 improves memory function of APP/PS1 mice, probably by reducing brain Aß accumulation through increasing plasmin-mediated degradation and LRP-1-mediated efflux of Aß in the brain.
Assuntos
Doença de Alzheimer/complicações , Peptídeos beta-Amiloides/metabolismo , Encéfalo/efeitos dos fármacos , Transtornos da Memória , Piperazinas/uso terapêutico , para-Aminobenzoatos/uso terapêutico , Doença de Alzheimer/genética , Precursor de Proteína beta-Amiloide/genética , Animais , Encéfalo/metabolismo , Linhagem Celular Tumoral , Modelos Animais de Doenças , Comportamento Exploratório/efeitos dos fármacos , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Masculino , Aprendizagem em Labirinto/efeitos dos fármacos , Transtornos da Memória/tratamento farmacológico , Transtornos da Memória/etiologia , Transtornos da Memória/patologia , Camundongos , Camundongos Transgênicos , Mutação/genética , Neuroblastoma/patologia , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Presenilina-1/genética , Ativador de Plasminogênio Tipo Uroquinase/metabolismoRESUMO
A 56-year-old man with advanced RCC and a past medical history of type 2 diabetes underwent a radical left nephrectomy following a histological diagnosis of papillary RCC, G2, INF b, pT3, V1 in 1999. In 2008, sorafenib was started to treat multiple pulmonary metastases of RCC. In 2011, sorafenib was switched to sunitinib when radiologic progression was observed. In 2014, sunitinib was switched to axitinib when further radiologic progression was observed. In 2015, the patient was referred to Yazawa clinic for homecare urology when hospital visits became difficult due to cancer pain and bilateral lower-extremity muscle weakness. Cancer pain was controlled using acetaminophen and a fentanyl patch. During the administration of axitinib, a CTCAE grade 1 vocal disorder was detected. We reduced the axitinib dose from 10 mg to 6 mg, and valsartan and an antiflatulent were administered due to CTCAE grade 2 hypertension and diarrhea, respectively. Axitinib administration continued until the patient died. He had survived more than 11 years following the detection of lung metastasis. In this patient, a good balance between cancer treatment and palliative care was achieved through the application of homecare urology. In a super-aged society such as Japan, urologists with an awareness of Zaitaku Medicine, a Japanese style of homecare that provides continuing appropriate medical treatment and welfare support to patients with access barriers to hospital treatment to enable them to live out the remainder of their lives with dignity, may play a key role in the development of Zaitaku Medicine.
Assuntos
Carcinoma de Células Renais/tratamento farmacológico , Imidazóis/uso terapêutico , Indazóis/uso terapêutico , Neoplasias Renais/tratamento farmacológico , Inibidores de Proteínas Quinases/uso terapêutico , Axitinibe , Evolução Fatal , Serviços de Assistência Domiciliar , Humanos , Neoplasias Renais/patologia , Masculino , Pessoa de Meia-IdadeRESUMO
Lipoprotein glomerulopathy (LPG) is characterized by the accumulation of lipoprotein thrombi within glomerular capillaries. This rare disorder is associated with various types of mutations in the apolipoprotein E gene (apoE). Herein, we present a case of LPG with a combination of apoE Chicago (Arg147Pro) and apoE (Glu3Lys) mutations. A 51-year-old Japanese woman presented with severe (3+) proteinuria. The initial renal biopsy showed glomerular capillary dilation and occlusion with lipid granules, a specific characteristic of LPG. Phenotype, genotype, and apoE DNA sequence analyses detected 2 mutations as described above within the same allele. Although both mutations had already been reported in 1 case of LPG each, this is the first time that the combination of the 2 mutations was identified in the same case. Familial analysis detected the same mutations in the patient's mother. However, she has not suffered LPG thus far. In addition, a re-analysis of the previous LPG case with apoE (Glu3Lys) also identified the apoE Chicago mutation, as was observed in our case. Treatment with fenofibrate and irbesartan was initiated, and urinary protein excretion ceased within 1 year; recurrence was not observed after an additional 2 years of follow-up. A second biopsy after 2 years showed great improvement, with lipoprotein thrombi identified only in 2 of 18 glomeruli.
RESUMO
Hematopoietic stem and progenitor cells (HSPCs) reside in the supportive stromal niche in bone marrow (BM); when needed, however, they are rapidly mobilized into the circulation, suggesting that HSPCs are intrinsically highly motile but usually stay in the niche. We questioned what determines the motility of HSPCs. Here, we show that transforming growth factor (TGF)-ß-induced intracellular plasminogen activator inhibitor (PAI)-1 activation is responsible for keeping HSPCs in the BM niche. We found that the expression of PAI-1, a downstream target of TGF-ß signaling, was selectively augmented in niche-residing HSPCs. Functional inhibition of the TGF-ß-PAI-1 signal increased MT1-MMP-dependent cellular motility, causing a detachment of HSPCs from the TGF-ß-expressing niche cells, such as megakaryocytes. Furthermore, consistently high motility in PAI-1-deficient HSPCs was demonstrated by both a transwell migration assay and reciprocal transplantation experiments, indicating that intracellular, not extracellular, PAI-1 suppresses the motility of HSPCs, thereby causing them to stay in the niche. Mechanistically, intracellular PAI-1 inhibited the proteolytic activity of proprotein convertase Furin, diminishing MT1-MMP activity. This reduced expression of MT1-MMP in turn affected the expression levels of several adhesion/deadhesion molecules for determination of HSPC localization, such as CD44, VLA-4, and CXCR4, which then promoted the retention of HSPCs in the niche. Our findings open up a new field for the study of intracellular proteolysis as a regulatory mechanism of stem cell fate, which has the potential to improve clinical HSPC mobilization and transplantation protocols.
Assuntos
Células-Tronco Hematopoéticas/metabolismo , Espaço Intracelular/metabolismo , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Nicho de Células-Tronco , Fator de Crescimento Transformador beta/metabolismo , Animais , Medula Óssea/metabolismo , Movimento Celular , Espaço Extracelular/metabolismo , Furina/metabolismo , Mobilização de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/citologia , Humanos , Metaloproteinase 14 da Matriz/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células-Tronco Multipotentes/metabolismo , Transdução de SinaisRESUMO
Adhesive small bowel obstruction remains a common problem for surgeons. After surgery, platelet aggregation contributes to coagulation cascade and fibrin clot formation. With clotting, fibrin degradation is simultaneously enhanced, driven by tissue plasminogen activator-mediated cleavage of plasminogen to form plasmin. The aim of this study was to investigate the cellular events and proteolytic responses that surround plasminogen activator inhibitor (PAI-1; Serpine1) inhibition of postoperative adhesion. Peritoneal adhesion was induced by gauze deposition in the abdominal cavity in C57BL/6 mice and those that were deficient in fibrinolytic factors, such as Plat-/- and Serpine1-/- In addition, C57BL/6 mice were treated with the novel PAI-1 inhibitor, TM5275. Some animals were treated with clodronate to deplete macrophages. Epidermal growth factor (EGF) experiments were performed to understand the role of macrophages and how EGF contributes to adhesion. In the early phase of adhesive small bowel obstruction, increased PAI-1 activity was observed in the peritoneal cavity. Genetic and pharmacologic PAI-1 inhibition prevented progression of adhesion and increased circulating plasmin. Whereas Serpine1-/- mice showed intra-abdominal bleeding, mice that were treated with TM5275 did not. Mechanistically, PAI-1, in combination with tissue plasminogen activator, served as a chemoattractant for macrophages that, in turn, secreted EGF and up-regulated the receptor, HER1, on peritoneal mesothelial cells, which led to PAI-1 secretion, further fueling the vicious cycle of impaired fibrinolysis at the adhesive site. Controlled inhibition of PAI-1 not only enhanced activation of the fibrinolytic system, but also prevented recruitment of EGF-secreting macrophages. Pharmacologic PAI-1 inhibition ameliorated adhesion formation in a macrophage-dependent manner.-Honjo, K., Munakata, S., Tashiro, Y., Salama, Y., Shimazu, H., Eiamboonsert, S., Dhahri, D., Ichimura, A., Dan, T., Miyata, T., Takeda, K., Sakamoto, K., Hattori, K., Heissig, B. Plasminogen activator inhibitor-1 regulates macrophage-dependent postoperative adhesion by enhancing EGF-HER1 signaling in mice.