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1.
Oncol Rep ; 42(1): 231-242, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31059070

RESUMO

Epidermal growth factor receptor (EGFR) is a receptor tyrosine kinase and mutations in this gene are major drivers of lung cancer development. EGFR tyrosine kinase inhibitors (TKIs) are standard first­line therapies for patients with advanced non­small cell lung cancer (NSCLC) with activating EGFR mutations, but are not effective in patients with wild­type EGFR. In the present study, the cytotoxic effects of various TKIs against EGFR were investigated in wild­type NSCLC cells as single treatments or in combination with Fingolimod (FTY720), which has been approved for treating multiple sclerosis and has cytotoxic effects against several tumor cell lines. It was found that the combined treatment with TKIs lapatinib (Lap) or sorafenib (Sor) and FTY720 synergistically suppressed the viability of the NSCLC cell lines A549 and H596. Additionally, FTY720 inhibited lysosomal acidification and suppressed autophagy flux. Immunoblotting and reverse transcription­quantitative polymerase chain reaction showed that FTY720 combined with Lap or Sor, enhanced endoplasmic reticulum (ER) stress loading and cell cycle arrest in A549 cells. The enhancement of ER stress loading and cell cycle arrest induced by combined treatment with Lap or Sor and FTY720, which was associated with the cytotoxicity induced by the combination of these drugs. These findings suggested that FTY720 improved TKI therapy in NSCLC patients with wild­type EGFR, by sensitizing NSCLC cells to TKIs.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Ciclo Celular/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Cloridrato de Fingolimode/farmacologia , Neoplasias Pulmonares/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Células A549 , Proteínas Relacionadas à Autofagia/metabolismo , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Reposicionamento de Medicamentos , Sinergismo Farmacológico , Receptores ErbB/genética , Humanos , Lapatinib/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Sorafenibe/farmacologia
2.
J Neuroinflammation ; 16(1): 54, 2019 Mar 02.
Artigo em Inglês | MEDLINE | ID: mdl-30825874

RESUMO

BACKGROUND: Chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) is an autoimmune-mediated inflammatory disease of the peripheral nervous system characterized by a response directed against certain myelin proteins and for which therapies are limited. Previous studies have suggested a beneficial role of FTY720, a sphingosine 1-phosphate (S1P) receptor agonist, known to deplete lymphocytes from the peripheral blood by sequestering them into lymph nodes, in the treatment of experimental autoimmune neuritis (EAN). Therefore, we investigated whether FTY720 is also beneficial in chronic experimental autoimmune neuritis (c-EAN), a recently developed rat model mimicking human CIDP. METHODS: c-EAN was induced in Lewis rats by immunization with S-palm P0(180-199) peptide. Rats were treated with FTY720 (1 mg/kg) or vehicle intraperitoneally once daily from the onset of clinical signs for 18 days; clinical signs were assessed daily until 60 days post-immunization (dpi). Electrophysiological and histological features were examined at different time points. We also evaluated the serum levels of different pro- and anti-inflammatory cytokines by ELISA or flow cytometry at 18, 40, and 60 dpi. RESULTS: Our data demonstrate that FTY720 decreased the severity and abolished the chronicity of the disease in c-EAN rats. Therapeutic FTY720 treatment reversed electrophysiological and histological anomalies, suggesting that myelinated fibers were subsequently preserved, it inhibited macrophage and IL-17+ cell infiltration in PNS, and it significantly reduced circulating pro-inflammatory cytokines. CONCLUSIONS: FTY720 treatment has beneficial effects on c-EAN, a new animal model mimicking human CIDP. We have shown that FTY720 is an effective immunomodulatory agent, improving the disease course of c-EAN, preserving the myelinated fibers, attenuating the axonal degeneration, and decreasing the number of infiltrated inflammatory cells in peripheral nerves. These data confirm the interest of testing FTY720 or molecules targeting S1P in human peripheral neuropathies.


Assuntos
Cloridrato de Fingolimode/farmacologia , Imunossupressores/farmacologia , Neurite Autoimune Experimental/patologia , Nervo Isquiático/efeitos dos fármacos , Nervo Isquiático/patologia , Animais , Masculino , Neuritos/efeitos dos fármacos , Polirradiculoneuropatia Desmielinizante Inflamatória Crônica , Ratos , Ratos Endogâmicos Lew , Receptores de Lisoesfingolipídeo/agonistas , Índice de Gravidade de Doença
3.
Mol Neurobiol ; 56(9): 5971-5986, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30701416

RESUMO

Astroglia, the primary homeostatic cells of the central nervous system, play an important role in neuroinflammation. They act as facultative immunocompetent antigen-presenting cells (APCs), expressing major histocompatibility complex (MHC) class II antigens upon activation with interferon (IFN)-γ and possibly other proinflammatory cytokines that are upregulated in disease states, including multiple sclerosis (MS). We characterized the anti-inflammatory effects of fingolimod (FTY720), an established drug for MS, and its phosphorylated metabolite (FTY720-P) in IFN-γ-activated cultured rat astrocytes. The expression of MHC class II compartments, ß2 adrenergic receptor (ADR-ß2), and nuclear factor kappa-light-chain enhancer of activated B cells subunit p65 (NF-κB p65) was quantified in immunofluorescence images acquired by laser scanning confocal microscopy. In addition, MHC class II-enriched endocytotic vesicles were labeled by fluorescent dextran and their mobility analyzed in astrocytes subjected to different treatments. FTY720 and FTY720-P treatment significantly reduced the number of IFN-γ-induced MHC class II compartments and substantially increased ADR-ß2 expression, which is otherwise small or absent in astrocytes in MS. These effects could be partially attributed to the observed decrease in NF-κB p65 expression, because the NF-κB signaling cascade is activated in inflammatory processes. We also found attenuated trafficking and secretion from dextran-labeled endo-/lysosomes that may hinder efficient delivery of MHC class II molecules to the plasma membrane. Our data suggest that FTY720 and FTY720-P at submicromolar concentrations mediate anti-inflammatory effects on astrocytes by suppressing their action as APCs, which may further downregulate the inflammatory process in the brain, constituting the therapeutic effect of fingolimod in MS.


Assuntos
Astrócitos/patologia , Cloridrato de Fingolimode/uso terapêutico , Inflamação/tratamento farmacológico , Inflamação/patologia , Interferon gama/farmacologia , Trifosfato de Adenosina/farmacologia , Animais , Astrócitos/efeitos dos fármacos , Contagem de Células , Células Cultivadas , Vesículas Citoplasmáticas/efeitos dos fármacos , Vesículas Citoplasmáticas/metabolismo , Dextranos/metabolismo , Feminino , Cloridrato de Fingolimode/farmacologia , Antígenos de Histocompatibilidade Classe II/metabolismo , Ratos Wistar , Receptores Adrenérgicos beta 2/metabolismo , Fator de Transcrição RelA/metabolismo
4.
J Neurol ; 266(3): 726-734, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30661133

RESUMO

BACKGROUND: It has been described that treating relapsing-remitting multiple sclerosis (RRMS) patients with alemtuzumab following fingolimod could be less effective due to the different dynamics of lymphocyte repopulation. Effectiveness and safety of alemtuzumab compared to rituximab after fingolimod withdrawal were analyzed. PATIENTS AND METHODS: A follow-up of a cohort of RRMS patients treated with alemtuzumab or rituximab after fingolimod withdrawal was accomplished. Effectiveness, measured by the percentage of patients with no evidence of disease activity (NEDA), and the presence of side effects (SE) were registered. RESULTS: Fifty-five patients, 28 with alemtuzumab and 27 with rituximab, were analyzed. No differences in the washout period or in the baseline lymphocytes counts were observed. After a mean follow-up period of 28.8 months, the annualized relapsing rate was significantly reduced in the alemtuzumab group from 1.29 to 0.004 (p < 0.001) and in the rituximab group from 1.24 to 0.02 (p < 0.001), without differences. A significant reduction of the median EDSS from 2.8 to 2.0 in the alemtuzumab group and from 3.5 to 2.5 (p < 0.01) in the rituximab group was observed, without differences. Eighty-two per cent (n = 28) of patients in alemtuzumab group and 69.2% (n = 26) in rituximab group achieved NEDA criteria, without differences (p = 0.3). Symptoms related to the infusion were the most frequent SE in both groups. No serious SE were registered. CONCLUSION: Treating RRMS patients with alemtuzumab or rituximab after fingolimod withdrawal is effective and safe, without significant differences between both groups in our series.


Assuntos
Alemtuzumab/farmacologia , Cloridrato de Fingolimode/farmacologia , Fatores Imunológicos/farmacologia , Esclerose Múltipla Recidivante-Remitente/tratamento farmacológico , Rituximab/farmacologia , Adulto , Alemtuzumab/efeitos adversos , Estudos de Coortes , Feminino , Cloridrato de Fingolimode/efeitos adversos , Humanos , Fatores Imunológicos/efeitos adversos , Masculino , Pessoa de Meia-Idade , Rituximab/efeitos adversos
5.
Glia ; 67(3): 498-511, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30484906

RESUMO

Accumulating evidence indicates that neuroinflammation contributes to the pathogenesis and exacerbation of neurodegenerative disorders, such as Alzheimer's disease (AD). Sphingosine-1-phosphate (S1P) is a pleiotropic bioactive lipid that regulates many pathophysiological processes including inflammation. We present evidence here that the spinster homolog 2 (Spns2), a S1P transporter, promotes microglia pro-inflammatory activation in vitro and in vivo. Spns2 knockout (Spns2KO) in primary cultured microglia resulted in significantly reduced levels of pro-inflammatory cytokines induced by lipopolysaccharide (LPS) and amyloid-beta peptide 1-42 oligomers (Aß42) when compared with littermate controls. Fingolimod (FTY720), a S1P receptor 1 (S1PR1) functional antagonist and FDA approved drug for relapsing-remitting multiple sclerosis, partially blunted Aß42-induced pro-inflammatory cytokine generation, suggesting that Spns2 promotes microglia pro-inflammatory activation through S1P-signaling. Spns2KO significantly reduced Aß42-induced nuclear factor kappa B (NFκB) activity. S1P increased, while FTY720 dampened, Aß42-induced NFκB activity, suggesting that Spns2 activates microglia inflammation through, at least partially, NFκB pathway. Spns2KO mouse brains showed significantly reduced Aß42-induced microglia activation/accumulation and reduced levels of pro-inflammatory cytokines when compared with age-matched controls. More interestingly, Spns2KO ameliorated Aß42-induced working memory deficit detected by Y-Maze. In summary, these results suggest that Spns2 promotes pro-inflammatory polarization of microglia and may play a crucial role in AD pathogenesis.


Assuntos
Peptídeos beta-Amiloides/farmacologia , Proteínas de Transporte de Ânions/metabolismo , Inflamação/metabolismo , Microglia/metabolismo , Animais , Proteínas de Transporte de Ânions/genética , Citocinas/metabolismo , Cloridrato de Fingolimode/farmacologia , Lipopolissacarídeos/farmacologia , Lisofosfolipídeos/metabolismo , Aprendizagem em Labirinto/fisiologia , Memória de Curto Prazo/fisiologia , Camundongos , Camundongos Knockout , Microglia/efeitos dos fármacos , NF-kappa B/metabolismo , Receptores de Lisoesfingolipídeo/metabolismo , Transdução de Sinais/efeitos dos fármacos , Esfingosina/análogos & derivados , Esfingosina/metabolismo
6.
Mol Neurobiol ; 56(1): 174-185, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-29687345

RESUMO

Sphingolipid signaling disturbances correlate with Alzheimer's disease (AD) progression. We examined the influence of FTY720/fingolimod, a sphingosine analog and sphingosine-1-phosphate (S1P) receptor modulator, on the expression of sphingolipid metabolism and signaling genes in a mouse transgenic AD model. Our results demonstrated that AßPP (V717I) transgene led with age to reduced mRNA expression of S1P receptors (S1PRs), sphingosine kinase SPHK2, ceramide kinase CERK, and the anti-apoptotic Bcl2 in the cerebral cortex and hippocampus, suggesting a pro-apoptotic shift in 12-month old mice. These changes largely emulated alterations we observed in the human sporadic AD hippocampus: reduced SPHK1, SPHK2, CERK, S1PR1, and BCL2. We observed that the responses to FTY720 treatment were modified by age and notably differed between control (APP-) and AD transgenic (APP+) animals. AßPP (V717I)-expressing 12-month-old animals reacted to fingolimod with wide changes in the gene expression program in cortex and hippocampus, including increased pro-survival SPHKs and CERK. Moreover, BCL2 was elevated by FTY720 in the cortex at all ages (3, 6, 12 months) while in hippocampus this increase was observed at 12 months only. In APP- mice, fingolimod did not induce any significant mRNA changes at 12 months. Our results indicate significant effect of FTY720 on the age-dependent transcription of genes involved in sphingolipid metabolism and pro-survival signaling, suggesting its neuroprotective role in AD animal model.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/genética , Cloridrato de Fingolimode/uso terapêutico , Regulação da Expressão Gênica/efeitos dos fármacos , Esfingolipídeos/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Região CA1 Hipocampal/metabolismo , Região CA1 Hipocampal/patologia , Modelos Animais de Doenças , Feminino , Cloridrato de Fingolimode/administração & dosagem , Cloridrato de Fingolimode/farmacologia , Humanos , Camundongos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
7.
Neurosci Lett ; 690: 178-180, 2019 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-30359694

RESUMO

In searching for Parkinson's disease (PD) pharmacotherapies we began studying FTY720, a food and drug administration (FDA) approved drug. We also created derivatives, FTY720-C2 and FTY720-Mitoxy, and began assessing them. Here we treated dopaminergic MN9D cells with FTY720s then measured microRNA (miRNA) levels by PCR arrays. We discovered that all three FTY720s increased miR376b-3p, while FTY720-C2 also increased miR-128-3p, miR-146b-5p, miR-7a-5p, and miR-9-5p, and FTY720-Mitoxy also increased miR-30d-5p. Investigations revealed that some miRNAs downregulate alpha-synuclein, while others reduce apoptosis, suggesting that FTY720s may act to reduce synucleinopathy and dopaminergic neuron loss in PD and related disorders.


Assuntos
Ceramidas/farmacologia , Neurônios Dopaminérgicos/metabolismo , Cloridrato de Fingolimode/análogos & derivados , Cloridrato de Fingolimode/farmacologia , MicroRNAs/metabolismo , Fármacos Neuroprotetores/metabolismo , Regulação para Cima/efeitos dos fármacos , Animais , Células Cultivadas , Camundongos
8.
Life Sci ; 217: 243-250, 2019 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-30550889

RESUMO

AIMS: Pancreatic stellate cells (PSCs) play a critical role in the development of pancreatic fibrosis. Any agents that can affect PSC activation could become potential candidates for treating pancreatic fibrosis. FTY720 can attenuate chronic pancreatic fibrosis by suppressing T-cell infiltration, but its effect on PSCs remains unknown. This study was conducted to investigate the effects of FTY720 on PSC activation in cultured rat PSCs. MAIN METHODS: The viability of PSCs after FTY720 treatment was detected by MTT. Cell proliferation and migration analysis was performed using the iCELLigence System and a Transwell assay. Cell apoptosis was assessed by flow cytometry, western blot and an activity assay. The mitochondrial membrane potential (MMP) was assessed by JC-1 staining. The expression of α-SMA, collagen I, fibronectin, Beclin-1, Atg5, P62 and LC3B were analysed by immunofluorescence, quantitative real-time PCR and western blot. Rapamycin and phenformin hydrochloride were used to determine whether FTY720 inhibits PSC autophagy by the AMPK/mTOR pathway. KEY FINDINGS: FTY720 supressed PSC viability, proliferation and migration. FTY720 inhibited PSC activation, induced PSC apoptosis and supressed PSC autophagy. We also confirmed that FTY720 inhibited PSC autophagy via the AMPK/mTOR pathway. SIGNIFICANCE: Our results indicated that FTY720 inhibited PSC activation by promoting cell apoptosis and inhibiting PSC autophagy by suppressing AMPK and activating the mTOR pathway. These findings may explain the therapeutic mechanisms of FTY720 in treating pancreatic fibrosis and further suggest that targeting autophagy and the related signalling pathways may provide new strategies for the treatment of pancreatic fibrosis.


Assuntos
Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Cloridrato de Fingolimode/farmacologia , Imunossupressores/farmacologia , Células Estreladas do Pâncreas/efeitos dos fármacos , Células Estreladas do Pâncreas/patologia , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Células Cultivadas , Fibrose , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Pâncreas/citologia , Pâncreas/efeitos dos fármacos , Pâncreas/metabolismo , Pâncreas/patologia , Células Estreladas do Pâncreas/citologia , Células Estreladas do Pâncreas/metabolismo , Ratos , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismo
9.
Mult Scler ; 25(1): 72-80, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-28933245

RESUMO

OBJECTIVE: Study aims were to determine the frequency of highly active disease in pediatric multiple sclerosis (MS), the response to natalizumab (NTZ) and fingolimod (FTY) treatment, and the impact of current treatment modalities on the clinical course. METHODS: Retrospective single-center study in the German Center for MS in Childhood and Adolescence. RESULTS: Of 144 patients with first MS manifestation between 2011 and 2015, 41.6% fulfilled the criteria for highly active MS. In total, 55 patients treated with NTZ and 23 with FTY demonstrated a significant reduction in relapse rate (NTZ: 95.2%, FTY: 75%), new T2 lesions (NTZ: 97%, FTY: 81%), and contrast-enhancing lesions (NTZ: 97%, FTY: 93%). However, seven patients switched from NTZ to FTY experienced an increase in disease activity. Comparing pediatric MS patients treated in 2005 with those treated in 2015 showed a 46% reduction in relapse rate and a 44% reduction in mean Expanded Disability Status Scale (EDSS). CONCLUSION: The rate of highly active disease among pediatric MS patients is high; more than 40% in our cohort. Response to NTZ and FTY treatment is similar if not better than observed in adults. Current treatment modalities including earlier treatment initiation and the introduction of NTZ and FTY have significantly improved the clinical course of pediatric MS.


Assuntos
Cloridrato de Fingolimode/farmacologia , Fatores Imunológicos/farmacologia , Esclerose Múltipla/tratamento farmacológico , Natalizumab/farmacologia , Adolescente , Criança , Progressão da Doença , Feminino , Cloridrato de Fingolimode/administração & dosagem , Seguimentos , Humanos , Fatores Imunológicos/administração & dosagem , Imagem por Ressonância Magnética , Masculino , Esclerose Múltipla/patologia , Esclerose Múltipla/fisiopatologia , Natalizumab/administração & dosagem , Recidiva , Estudos Retrospectivos , Índice de Gravidade de Doença
10.
Wound Repair Regen ; 27(1): 59-68, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30368971

RESUMO

In this study, rat models of wound closure by first and second intention were developed to evaluate the influence that two immunosuppressants for treating multiple sclerosis (fingolimod, azathioprine) have on wound healing. Sixty-three Sprague-Dawley rats were daily treated with fingolimod (0.6 mg/kg), azathioprine (2.5 mg/kg), or placebo (saline). Following 6 weeks of treatment, a linear incision (1.5 cm) or a circular excisional defect (diameter 1.5 cm) was made on the dorsal skin. The treatments were uninterrupted and after 7 days (incisional) or 21 days (incisional, excisional), animals were euthanized (n = 7 per group and time-point). Morphometric (wound closure), histological (stainings), and immunofluorescent studies (macrophages) were performed to evaluate the healing process. For both the incisional and excisional defects, animals treated with fingolimod exhibited a healing process equivalent to that of placebo in terms of collagenization, wound closure, and macrophage response. By comparison, groups treated with azathioprine displayed a delay in healing times which was especially evident in the excisional defect, where inflammatory reaction and collagen deposition in the repair tissue remained active by day 21. These results show that immunosuppressants with a selective mechanism of action (fingolimod) can have less impact on wound healing than their classical nonselective counterparts (azathioprine).


Assuntos
Azatioprina/farmacologia , Cloridrato de Fingolimode/farmacologia , Imunossupressores/farmacologia , Inflamação/tratamento farmacológico , Técnicas de Fechamento de Ferimentos , Cicatrização/efeitos dos fármacos , Animais , Inflamação/patologia , Masculino , Modelos Animais , Ratos , Ratos Sprague-Dawley
11.
FASEB J ; 33(4): 4703-4715, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30592632

RESUMO

Schwann cells promote nerve regeneration by adaptation of a regenerative phenotype referred to as repair mediating Schwann cell. Down-regulation of myelin proteins, myelin clearance, formation of Bungner's bands, and secretion of trophic factors characterize this cell type. We have previously shown that the sphingosine-1-phosphate receptor agonist Fingolimod/FTY720P promotes the generation of this particular Schwann cell phenotype by activation of dedifferentiation markers and concomitant release of trophic factors resulting in enhanced neurite growth of dorsal root ganglion neurons. Despite its biomedical relevance, a detailed characterization of the corresponding Schwann cell secretome is lacking, and the impact of FTY720P on enhancing neurite growth is not defined. Here, we applied a label-free quantitative mass spectrometry approach to characterize the secretomes derived from primary neonatal and adult rat Schwann cells in response to FTY720P. We identified a large proportion of secreted proteins with a high overlap between the neonatal and adult Schwann cells, which can be associated with biologic processes such as development, axon growth, and regeneration. Moreover, FTY720P-treated Schwann cells release proteins downstream of Smad signaling known to support neurite growth. Our results therefore uncover a network of trophic factors involved in glial-mediated repair of the peripheral nervous system.-Schira, J., Heinen, A., Poschmann, G., Ziegler, B., Hartung, H.-P., Stühler, K., Küry, P. Secretome analysis of nerve repair mediating Schwann cells reveals Smad-dependent trophism.


Assuntos
Regeneração Nervosa/fisiologia , Células de Schwann/metabolismo , Proteínas Smad/metabolismo , Animais , Células Cultivadas , Cromatografia Líquida , Biologia Computacional , Cloridrato de Fingolimode/farmacologia , Organofosfatos/farmacologia , Ratos , Células de Schwann/efeitos dos fármacos , Transdução de Sinais/fisiologia , Proteínas Smad/genética , Esfingosina/análogos & derivados , Esfingosina/farmacologia , Espectrometria de Massas em Tandem , Ácido Tricloroacético/química
12.
JCI Insight ; 3(23)2018 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-30518694

RESUMO

PD-1/PD-L1 checkpoint therapy for cancer is commonly considered to act by reactivating T cells in the tumor microenvironment. Here, we present data from 2 mouse tumor models demonstrating an essential involvement of tumor-draining lymph nodes in PD-1 and PD-L1 therapeutic efficacy. Immune activation induced by checkpoint treatment was predominantly observed in the tumor-draining, but not nondraining, lymph nodes and was reflected in local accumulation of CD8+ T cells. Surgical resection of these lymph nodes, but not contralateral lymph nodes, abolished therapy-induced tumor regressions and was associated with decreased immune infiltrate in the tumor microenvironment. Moreover, inhibitor FTY720, which locks lymphocytes in lymph organs, also abrogated checkpoint therapy, suggesting that the tumor-draining lymph nodes function as sites of T cell invigoration required for checkpoint blockade therapy. Now that PD-1/PD-L1 checkpoint treatment is applied in earlier clinical stages of cancer, our preclinical data advocate for enrolling patients with their tumor-draining lymph nodes still in place, to optimally engage the antitumor immune response and thereby enhance clinical benefit.


Assuntos
Antígeno B7-H1/metabolismo , Linfonodos/imunologia , Linfócitos do Interstício Tumoral/imunologia , Receptor de Morte Celular Programada 1/metabolismo , Animais , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Cloridrato de Fingolimode/farmacologia , Humanos , Imunoterapia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Microambiente Tumoral
13.
Nat Commun ; 9(1): 4854, 2018 11 19.
Artigo em Inglês | MEDLINE | ID: mdl-30451860

RESUMO

Natural killer (NK) cells are reported to have immunological memory, with CD49a+ liver-resident NK cells shown to confer hapten-specific memory responses, but how this memory is induced or maintained is unclear. Here we show that memory type I innate lymphoid cells (ILC1s), which express IL-7Rα, are generated in the lymph nodes (LNs) and require IL-7R signaling to maintain their longevity in the liver. Hapten sensitization initiates CXCR3-dependent recruitment of IL-7Rα+ ILC1s into skin-draining LNs, where they are primed and acquire hapten-specific memory potential. Memory IL-7Rα+ ILC1s then exit draining LNs and are preferentially recruited, via CXCR6, to reside in the liver. Moreover, long-term blockade of IL-7R signaling significantly reduces ILC1-mediated memory responses. Thus, our results identify a memory IL-7Rα+ ILC1 population and reveal a LN-liver axis that is essential for ILC1 memory generation and long-term maintenance.


Assuntos
Memória Imunológica , Células Matadoras Naturais/imunologia , Fígado/imunologia , Linfonodos/imunologia , Receptores de Interleucina-7/imunologia , Baço/imunologia , Animais , Linhagem da Célula/imunologia , Cloridrato de Fingolimode/farmacologia , Expressão Gênica , Haptenos/administração & dosagem , Imunidade Inata/efeitos dos fármacos , Imunização , Imunossupressores/farmacologia , Integrina alfa1/genética , Integrina alfa1/imunologia , Células Matadoras Naturais/citologia , Células Matadoras Naturais/efeitos dos fármacos , Fígado/citologia , Fígado/efeitos dos fármacos , Linfonodos/citologia , Linfonodos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Parabiose/métodos , Cultura Primária de Células , Receptores CXCR3/deficiência , Receptores CXCR3/genética , Receptores CXCR3/imunologia , Receptores CXCR6/genética , Receptores CXCR6/imunologia , Receptores de Interleucina-7/genética , Baço/citologia , Baço/efeitos dos fármacos
14.
Chem Pharm Bull (Tokyo) ; 66(10): 1015-1018, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30270236

RESUMO

FTY720 is employed for the treatment of multiple sclerosis and exerts apoptotic effects on various cancers through protein phosphatase 2A (PP2A) activation. In compound 4, the dihydroxy head group of FTY720 was modified into dihydroxy phenyl group. The cell survival in compound 4 treated colorectal and gastric cancer cells was significantly reduced as compared with control, 34.6 and 25.1%, respectively. The docking study of compound 4 showed that the aromatic head group effectively binds to PP2A.


Assuntos
Antineoplásicos/farmacologia , Cloridrato de Fingolimode/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Cloridrato de Fingolimode/síntese química , Cloridrato de Fingolimode/química , Humanos , Simulação de Acoplamento Molecular , Estrutura Molecular , Relação Estrutura-Atividade , Células Tumorais Cultivadas
15.
Molecules ; 23(11)2018 Oct 24.
Artigo em Inglês | MEDLINE | ID: mdl-30355990

RESUMO

FTY720 inhibits various cancers through PP2A activation. The structure of FTY720 is also used as a basic structure for the design of sphingosine kinase (SK) inhibitors. We have synthesized derivatives using an amide chain in FTY720 with a phenyl backbone, and then compounds were screened by an MTT cell viability assay. The PP2A activity of compound 7 was examined. The phosphorylation levels of AKT and ERK, downstream targets of PP2A, in the presence of compound 7, were determined. Compound 7 may exhibit anticancer effects through PP2A activation rather than the mechanism by inhibition of SK1 in cancer cells. In the docking study of compound 7 and PP2A, the amide chain of compound 7 showed an interaction with Asn61 that was different from FTY720, which is expected to affect the activity of the compound.


Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Cloridrato de Fingolimode/síntese química , Cloridrato de Fingolimode/farmacologia , Proteína Fosfatase 2/antagonistas & inibidores , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Cloridrato de Fingolimode/análogos & derivados , Humanos , Concentração Inibidora 50 , Modelos Moleculares , Conformação Molecular , Estrutura Molecular , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Relação Estrutura-Atividade
16.
Biochem Biophys Res Commun ; 505(4): 1032-1037, 2018 11 10.
Artigo em Inglês | MEDLINE | ID: mdl-30314693

RESUMO

Necrotizing enterocolitis (NEC) remains one of the leading causes of death in neonatal infants and new therapeutic strategies for NEC are urgently required. The immunomodulatory agent FTY720 has been shown to have protective effects in various inflammatory diseases. In this study, we hypothesized that treatment with FTY720 confers protection against experimental NEC. Experimental NEC was induced in five-day-old C57BL/6 neonatal mice by hyperosmolar formula feeding plus hypoxia and lipopolysaccharide (LPS) challenges. Induction of NEC resulted in substantial weight loss and high mortality compared to the control group, whereas FTY720 treatment significantly attenuated weight loss and improved survival in NEC-challenged neonatal mice. FTY720 treatment strongly ameliorated NEC-induced intestinal injury with reduced apoptosis and up-regulation of intestinal barrier proteins in the ileal tissues. Furthermore, FTY720 treatment abrogated NEC-initiated intestinal and systemic inflammation with markedly diminished inflammatory cytokines and chemokines. Moreover, FTY720 treatment suppressed NEC-activated CXCL5/CXCR2 axis with down-regulated expression of CXCL5 and CXCR2 at both mRNA and protein levels. Thus, we demonstrate that FTY720 protects neonatal mice against NEC-associated lethality by ameliorating intestinal injury and attenuating inflammation, possibly via its down-regulation of NEC-induced activation of intestinal CXCL5/CXCR2 axis.


Assuntos
Quimiocina CXCL5/biossíntese , Enterocolite Necrosante/tratamento farmacológico , Cloridrato de Fingolimode/farmacologia , Inflamação/tratamento farmacológico , Intestinos/efeitos dos fármacos , Intestinos/lesões , Receptores de Interleucina-8B/biossíntese , Animais , Quimiocina CXCL5/metabolismo , Modelos Animais de Doenças , Enterocolite Necrosante/metabolismo , Feminino , Inflamação/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Receptores de Interleucina-8B/metabolismo
17.
Eur J Med Chem ; 159: 217-242, 2018 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-30292898

RESUMO

A series of compounds containing pyrrolidine and pyrrolizidine cores with appended hydrophobic substituents were prepared as constrained analogs of FTY720 and phytosphingosine. The effect of these compounds on the viability of cancer cells, on downregulation of the nutrient transport systems, and on their ability to cause vacuolation was studied. An attempt to inhibit HDACs with some phosphate esters of our analogs was thwarted by our failure to reproduce the reported inhibitory action of FTY720-phosphate.


Assuntos
Antineoplásicos/farmacologia , Epigênese Genética/efeitos dos fármacos , Cloridrato de Fingolimode/farmacologia , Esfingosina/análogos & derivados , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Cloridrato de Fingolimode/análogos & derivados , Cloridrato de Fingolimode/química , Camundongos , Estrutura Molecular , Esfingosina/síntese química , Esfingosina/química , Esfingosina/farmacologia , Relação Estrutura-Atividade
18.
Front Immunol ; 9: 2189, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30319641

RESUMO

About 40% of patients with systemic lupus erythematosus experience diffuse neuropsychiatric manifestations, including impaired cognition and depression. Although the pathogenesis of diffuse neuropsychiatric SLE (NPSLE) is not fully understood, loss of brain barrier integrity, autoreactive antibodies, and pro-inflammatory cytokines are major contributors to disease development. Fingolimod, a sphingosine-1-phosphate (S1P) receptor modulator, prevents lymphocyte egress from lymphoid organs through functional antagonism of S1P receptors. In addition to reducing the circulation of autoreactive lymphocytes, fingolimod has direct neuroprotective effects such as preserving brain barrier integrity and decreasing pro-inflammatory cytokine secretion by astrocytes and microglia. Given these effects, we hypothesized that fingolimod would attenuate neurobehavioral deficits in MRL-lpr/lpr (MRL/lpr) mice, a validated neuropsychiatric lupus model. Fingolimod treatment was initiated after the onset of disease, and mice were assessed for alterations in cognitive function and emotionality. We found that fingolimod significantly attenuated spatial memory deficits and depression-like behavior in MRL/lpr mice. Immunofluorescent staining demonstrated a dramatic lessening of brain T cell and macrophage infiltration, and a significant reduction in cortical leakage of serum albumin, in fingolimod treated mice. Astrocytes and endothelial cells from treated mice exhibited reduced expression of inflammatory genes, while microglia showed differential regulation of key immune pathways. Notably, cytokine levels within the cortex and hippocampus were not appreciably decreased with fingolimod despite the improved neurobehavioral profile. Furthermore, despite a reduction in splenomegaly, lymphadenopathy, and circulating autoantibody titers, IgG deposition within the brain was unaffected by treatment. These findings suggest that fingolimod mediates attenuation of NPSLE through a mechanism that is not dependent on reduction of autoantibodies or cytokines, and highlight modulation of the S1P signaling pathway as a novel therapeutic target in lupus involving the central nervous system.


Assuntos
Depressão/imunologia , Cloridrato de Fingolimode/farmacologia , Vasculite Associada ao Lúpus do Sistema Nervoso Central/psicologia , Lisofosfolipídeos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Esfingosina/análogos & derivados , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/imunologia , Autoanticorpos/imunologia , Técnicas de Observação do Comportamento , Comportamento Animal/efeitos dos fármacos , Encéfalo/citologia , Encéfalo/imunologia , Encéfalo/fisiologia , Cognição/efeitos dos fármacos , Cognição/fisiologia , Citocinas/imunologia , Depressão/tratamento farmacológico , Depressão/psicologia , Modelos Animais de Doenças , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/imunologia , Feminino , Cloridrato de Fingolimode/uso terapêutico , Humanos , Vasculite Associada ao Lúpus do Sistema Nervoso Central/tratamento farmacológico , Vasculite Associada ao Lúpus do Sistema Nervoso Central/genética , Vasculite Associada ao Lúpus do Sistema Nervoso Central/imunologia , Lisofosfolipídeos/imunologia , Camundongos , Camundongos Endogâmicos MRL lpr , Microglia/efeitos dos fármacos , Microglia/imunologia , Receptores de Lisoesfingolipídeo/antagonistas & inibidores , Receptores de Lisoesfingolipídeo/imunologia , Receptores de Lisoesfingolipídeo/metabolismo , Transdução de Sinais/imunologia , Esfingosina/imunologia , Esfingosina/metabolismo , Resultado do Tratamento
19.
Sci Rep ; 8(1): 15371, 2018 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-30337577

RESUMO

There are no data on the effects of fingolimod, an immunomodulatory drug used in treatment of multiple sclerosis (MS), on circulating tight-junction (TJ) protein levels as well as on peripheral blood mononuclear cells (PBMC) migration. Serum TJ protein [occludin (OCLN), claudin-5 (CLN-5) and zonula occludens-1 (ZO-1)] levels, sphingosine-1 phosphate 1 (S1P1) receptor expression on circulating leukocyte populations as well as in vitro PBMC migration were longitudinally assessed in 20 MS patients under 12-months fingolimod treatment and correlated with clinical and magnetic resonance imaging (MRI) parameters. After 12 months of treatment, a significant reduction of mean relapse rate as well as number of active lesions at MRI was found. TJ protein levels significantly decreased and were associated with reduction of S1P1 expression as well as of PBMC in vitro migratory activity. A significant correlation of CLN-5/OCLN ratio with new T2 MRI lesions and a significant inverse correlation of CLN-5/ZO-1 ratio with disability scores were found. These findings support possible in vivo effects of fingolimod on the blood-brain barrier (BBB) functional activity as well as on peripheral cell trafficking that could result in avoiding passage of circulating autoreactive cells into brain parenchyma. Circulating TJ protein levels and respective ratios could be further studied as a novel candidate biomarker of BBB functional status to be monitored in course of fingolimod as well as of other immunomodulatory treatments in MS.


Assuntos
Biomarcadores/sangue , Movimento Celular , Cloridrato de Fingolimode/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Leucócitos Mononucleares/efeitos dos fármacos , Esclerose Múltipla/patologia , Proteínas de Junções Íntimas/sangue , Adulto , Quimiotaxia , Feminino , Humanos , Imunossupressores/farmacologia , Técnicas In Vitro , Estudos Longitudinais , Masculino , Esclerose Múltipla/sangue , Esclerose Múltipla/tratamento farmacológico , Estudos Prospectivos , Receptores de Lisoesfingolipídeo/sangue
20.
Mol Med Rep ; 18(6): 5151-5158, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30320355

RESUMO

Vemurafenib, a selective inhibitor of mutated BRAF, is used to treat late­stage melanoma. However, resistance to vemurafenib is urgently required as it can have fatal consequences. Fingolimod (FTY720), a sphingosine­1­phosphate receptor modulator, has been used for the treatment of several malignant neoplasms in clinical trials. The present study investigated the effects of FTY720 and vemurafenib combination treatment on cell death induction, and defined the molecular mechanisms in vemurafenib­resistant melanoma cells. The combination treatment with FTY720 and vemurafenib reduced cell viability, and the expression of apoptosis­associated cleaved poly (adenosine diphosphate­ribose) polymerase (PARP) was increased when compared with treatment with vemurafenib alone in WM­115 cells, a vemurafenib­resistant human melanoma cell line. In addition, the protein expression of phosphorylated extracellular signal­related kinase (ERK) in WM­115 cells was decreased by this combination treatment. Vemurafenib­resistant SK­Mel­28 cells (R­SK­Mel) were established by culturing SK­Mel­28 cells, which are the most sensitive to vemurafenib, in the presence of vemurafenib. Similar to WM­155 cells, the viability of R­SK­Mel cells was reduced and the expression of cleaved PARP was increased by the combination treatment with FTY720 and vemurafenib. In addition, the expression of phosphorylated ERK and Akt was also reduced by this treatment. These results suggested that FTY720 and vemurafenib synergistically induced cell death by downregulating proliferation and survival signalling pathways in vemurafenib­resistant melanoma cells.


Assuntos
Antineoplásicos/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Cloridrato de Fingolimode/farmacologia , Vemurafenib/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Sinergismo Farmacológico , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Melanoma/genética , Melanoma/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos
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