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1.
J Cancer Res Clin Oncol ; 147(12): 3639-3651, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34499223

RESUMO

Cancer immunotherapy is a major breakthrough in tumor therapy and has been used in monotherapy or combination therapy. However, it has been associated with poor immune tolerance in some patients or immune-related adverse events. Therefore, ideal and reliable tumor elimination strategies are urgently needed to overcome these shortcomings. Phosphatidylserine (PS) is a negatively charged phospholipid, usually present in the inner lobules of eukaryotic cell membranes. Under certain physiological or pathological conditions, PS may be exposed on the outer leaflets of apoptotic cells serving as recognition signals by phagocytes and modulating the immune response. On the contrary, increased exposure of PS in the tumor microenvironment can significantly antagonize the body's anti-tumor immunity, thereby promoting tumor growth and metastasis. During radiotherapy and chemotherapy, PS-mediated immunosuppression increases the PS levels in necrotic tissue in the tumor microenvironment, further suppressing tumor immunity. PS-targeted therapy is a promising strategy in cancer immunotherapy. It inhibits tumor growth and improves the anti-tumor activity of immune checkpoint inhibitors. A comprehensive understanding of the mechanism of PS-targeted therapy opens up a new perspective for future cancer immunotherapies.


Assuntos
Imunoterapia/métodos , Neoplasias , Fosfatidilserinas/antagonistas & inibidores , Animais , Humanos , Inibidores de Checkpoint Imunológico/farmacologia , Neoplasias/imunologia , Neoplasias/metabolismo , Fosfatidilserinas/imunologia , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/fisiologia
2.
Virology ; 560: 17-33, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34020328

RESUMO

Envelope phosphatidylserine (PtdSer) and phosphatidylethanolamine (PtdEtr) have been shown to mediate binding of enveloped viruses. However, commonly used PtdSer binding molecules such as Annexin V cannot block PtdSer-mediated viral infection. Lack of reagents that can conceal envelope PtdSer and PtdEtr and subsequently inhibit infection hinders elucidation of the roles of the envelope phospholipids in viral infection. Here, we developed sTIM1dMLDR801, a reagent capable of blocking PtdSer- and PtdEtr-dependent infection of enveloped viruses. Using sTIM1dMLDR801, we found that envelope PtdSer and/or PtdEtr can support ZIKV infection of not only human but also mosquito cells. In a mouse model for ZIKV infection, sTIM1dMLDR801 reduced ZIKV load in serum and the spleen, indicating envelope PtdSer and/or PtdEtr support in viral infection in vivo. sTIM1dMLDR801 will enable elucidation of the roles of envelope PtdSer and PtdEtr in infection of various virus species, thereby facilitating identification of their receptors and transmission mechanisms.


Assuntos
Antivirais/farmacologia , Fosfatidiletanolaminas/antagonistas & inibidores , Fosfatidilserinas/antagonistas & inibidores , Ligação Viral/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacos , Zika virus/efeitos dos fármacos , Células A549 , Animais , Linhagem Celular , Chlorocebus aethiops , Culicidae/virologia , Feminino , Células HEK293 , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Proto-Oncogênicas/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Receptor de Interferon alfa e beta/genética , Células Vero , Envelope Viral/metabolismo , Carga Viral/efeitos dos fármacos , Zika virus/crescimento & desenvolvimento , Infecção por Zika virus/tratamento farmacológico , Infecção por Zika virus/patologia , Infecção por Zika virus/transmissão , Receptor Tirosina Quinase Axl
3.
J Surg Res ; 245: 604-609, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31499368

RESUMO

BACKGROUND: Phosphatidylserine (PS) is a key cell membrane phospholipid normally maintained on the inner cell surface but externalizes to the outer surface in response to cellular stress. We hypothesized that PS exposure mediates organ dysfunction in hemorrhagic shock. Our aims were to evaluate PS blockade on (1) pulmonary, (2) renal, and (3) gut function, as well as (4) serum lysophosphatidic acid (LPA), an inflammatory mediator generated by PS externalization, as a possible mechanism mediating organ dysfunction. MATERIALS AND METHODS: Rats were either (1) monitored for 130 min (controls, n = 3), (2) hemorrhaged then resuscitated (hemorrhage only group, n = 3), or (3) treated with Diannexin (DA), a PS blocking agent, followed by hemorrhage and resuscitation (DA + hemorrhage group, n = 4). Pulmonary dysfunction was assessed by arterial partial pressure of oxygen, renal dysfunction by serum creatinine, and gut dysfunction by mesenteric endothelial permeability (LP). LPA levels were measured in all groups. RESULTS: Pulmonary: there was no difference in arterial partial pressure of oxygen between groups. Renal: after resuscitation, creatinine levels were lower after PS blockade with DA versus hemorrhage only group (P = 0.01). Gut: LP was decreased after PS blockade with DA versus hemorrhage only group (P < 0.01). Finally, LPA levels were also lower after PS blockade with DA versus the hemorrhage only group but higher than the control group (P < 0.01). CONCLUSIONS: PS blockade with DA decreased renal and gut dysfunction associated with hemorrhagic shock and attenuated the magnitude of LPA generation. Our findings suggest potential for therapeutic targets in the future that could prevent organ dysfunction associated with hemorrhagic shock.


Assuntos
Anexina A5/administração & dosagem , Fosfatidilserinas/antagonistas & inibidores , Ressuscitação/métodos , Choque Hemorrágico/terapia , Animais , Modelos Animais de Doenças , Feminino , Humanos , Infusões Intravenosas , Mucosa Intestinal/fisiopatologia , Rim/fisiopatologia , Pulmão/fisiopatologia , Lisofosfolipídeos/sangue , Escores de Disfunção Orgânica , Ratos , Choque Hemorrágico/sangue , Choque Hemorrágico/diagnóstico , Resultado do Tratamento
4.
Target Oncol ; 14(5): 541-550, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31429027

RESUMO

BACKGROUND: Bavituximab, an immunomodulator, targets phosphatidylserine (PS), a membrane lipid externalized on tumor and endothelial cells in response to sorafenib. OBJECTIVE: The objective of this phase II study was to assess the efficacy of combination bavituximab and sorafenib in advanced hepatocellular carcinoma (HCC). METHODS: In this single-arm phase II study, patients with HCC determined to be unresectable with Eastern Cooperative Oncology Group (ECOG) score ≤ 2, Child-Pugh score A/B7 received intravenous bavituximab 3 mg/kg weekly and oral sorafenib 400 mg twice daily until disease progression or intolerable toxicity. We investigated time to progression (TTP) for patients receiving combination bavituximab and sorafenib compared with that for sorafenib-only historical controls. RESULTS: In total, 38 patients were accrued. The median follow-up was 6.1 months. Patient characteristics were as follows: median age 61 years; male 82%; hepatitis C virus 79%; Black 39%, Hispanic 26%, White 29%; previous treatment 39%; macrovascular invasion 84%; and extrahepatic metastases 24%. The median TTP was 6.7 months (95% confidence interval [CI] 4-17). The median overall survival was 6.1 months (95% CI 5-8), and the median disease-specific survival was 8.6 months (95% CI 6-14). Two patients experienced partial responses; none had a complete response. The disease control rate was 58%. Treatment-related adverse events were observed in 63% of patients, with the most commonly reported therapy-related symptoms being diarrhea (32%), fatigue (26%), and anorexia (24%). CONCLUSIONS: The efficacy of adding bavituximab to sorafenib for the treatment of advanced HCC was inconclusive; however, the combination regimen did not exacerbate toxicities associated with single-agent sorafenib. CLINICALTRIALS. GOV IDENTIFIER: NCT01264705.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma Hepatocelular/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Sorafenibe/uso terapêutico , Carcinoma Hepatocelular/mortalidade , Feminino , Seguimentos , Humanos , Imunomodulação , Neoplasias Hepáticas/mortalidade , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Fosfatidilserinas/antagonistas & inibidores , Análise de Sobrevida , Resultado do Tratamento
5.
Surgery ; 166(5): 844-848, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31285044

RESUMO

BACKGROUND: Phosphatidylserine is usually an intracellularly oriented cell membrane phospholipid. Externalized phosphatidylserine on activated cells is a signal for phagocytosis. In sepsis, persistent phosphatidylserine exposure is also a signal for activation of the coagulation and inflammatory cascades. As such, phosphatidylserine may be a key molecule in sepsis induced cellular and organ injury. We hypothesize that phosphatidylserine blockade provides a protective effect in sepsis induced organ dysfunction. METHODS: Sepsis was induced in adult female rats using an endotoxin model. Diannexin, a homodimer of annexin A5, was administered for phosphatidylserine blockade. Rats were allocated to control (n = 5), sepsis (n = 6), or sepsis and phosphatidylserine blockade (n = 9) groups. Gut, pulmonary, renal, and hematologic dysfunctions were evaluated by mesenteric microvascular fluid leak, partial pressure of oxygen, serum creatinine, activated clotting time, and glomerular fibrin deposition, respectively. RESULTS: Rats in the sepsis group demonstrated gut, renal, and hematologic dysfunction. Phosphatidylserine blockade reversed signs of gut dysfunction and mesenteric microvascular leak (P < .01). In addition, phosphatidylserine blockade corrected systemic coagulopathy, as measured by activated clotting time (P = .03) and glomerular fibrin deposition (P = .008). There was no difference in renal dysfunction (P = .1) or pulmonary dysfunction in any of the groups (P = .6). CONCLUSION: In sepsis, phosphatidylserine blockade had a protective effect on gut dysfunction and coagulopathy. Increased phosphatidylserine exposure may be a key mediator of organ dysfunction and coagulopathy during sepsis. These data may provide insights into novel treatment options for septic patients.


Assuntos
Anexina A5/administração & dosagem , Insuficiência de Múltiplos Órgãos/prevenção & controle , Fosfatidilserinas/antagonistas & inibidores , Sepse/tratamento farmacológico , Animais , Modelos Animais de Doenças , Feminino , Humanos , Infusões Intravenosas , Lipopolissacarídeos/toxicidade , Insuficiência de Múltiplos Órgãos/etiologia , Insuficiência de Múltiplos Órgãos/patologia , Fosfatidilserinas/metabolismo , Ratos , Sepse/complicações , Sepse/patologia , Resultado do Tratamento
6.
FEBS Open Bio ; 9(3): 446-456, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30868053

RESUMO

One of the major roles of professional phagocytes is the removal of dead cells in the body. We know less about the clearance of necrotic cells than apoptotic cell phagocytosis, despite the fact that both types of dead cells need to be cleared together and necrotic cells appear often in pathological settings. In the present study, we examined phagocytosis of heat- or H2O2-killed necrotic and apoptotic thymocytes by mouse bone marrow-derived macrophages (BMDMs) in vitro and found that the two cell types are engulfed at equal efficiency and compete with each other when added together to BMDMs. Phagocytosis of both apoptotic and necrotic thymocytes was decreased by (a) blocking phosphatidylserine on the surface of dying cells; (b) inhibition of Mer tyrosine kinase, Tim-4, integrin ß3 receptor signaling, or Ras-related C3 botulinum toxin substrate 1 activity; or (c) using BMDMs deficient for transglutaminase 2. Stimulation of liver X, retinoid X, retinoic acid or glucocorticoid nuclear receptors in BMDMs enhanced not only apoptotic, but also necrotic cell uptake. Electron microscopic analysis of the engulfment process revealed that the morphology of phagosomes and the phagocytic cup formed during the uptake of dying thymocytes is similar for apoptotic and necrotic cells. Our data indicate that apoptotic and necrotic cells are cleared via the same mechanisms, and removal of necrotic cells in vivo can be facilitated by molecules known to enhance the uptake of apoptotic cells.


Assuntos
Apoptose , Macrófagos/metabolismo , Necrose/metabolismo , Fosfatidilserinas/metabolismo , Timócitos/metabolismo , Animais , Apoptose/efeitos dos fármacos , Células Cultivadas , Temperatura Alta , Peróxido de Hidrogênio/farmacologia , Macrófagos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fosfatidilserinas/antagonistas & inibidores , Timócitos/efeitos dos fármacos
7.
Am J Case Rep ; 19: 1042-1046, 2018 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-30166503

RESUMO

BACKGROUND Antiphospholipid syndrome (APS) is an autoimmune disease characterized by antibodies directed against phospholipids on plasma membranes. Through unclear mechanisms, APS confers hypercoagulability. APS may cause recurrent thromboses in the arterial and venous vasculature. We report a case of primary APS resulting in cerebral venous thrombosis and ST-elevation myocardial infarction (STEMI) for which only antiphosphatidylserine (aPS) IgM antibody was positive after extensive investigation. CASE REPORT A 48-year-old male was admitted after a witnessed generalized seizure with subsequent confusion. Imaging demonstrated thrombosis of multiple central nervous system (CNS) sinuses, including the superior sagittal sinus and bilateral transverse sinuses. The patient was heparinized with aggressive hydration, which proved inadequate, prompting endovascular thrombectomy. Three months later, despite anticoagulation therapy, the patient developed a STEMI when International Normalized Ratio (INR) was 1.8. Echocardiogram (ECHO) and PAN CT scan were normal. Initial coagulation studies demonstrated normal anticardiolipin antibody, prothrombin time, partial thromboplastin time, and platelet count. Outpatient coagulation studies revealed normal antithrombin III, protein C/S, hemoglobin electrophoresis, homocysteine, anti-ß2 glycoprotein 1 antibodies, and D-Dimer. Factor V Leiden, JAK 2 mutation, prothrombin gene mutation, and tests for paroxysmal nocturnal hemoglobinuria (PNH) were negative. A positive phosphatidylserine IgM was detected. The patient was continued on warfarin (10 mg daily) with a target INR of 3.0-3.5 and clopidogrel (75 mg daily). CONCLUSIONS Despite extensive investigation, this patient only showed evidence of elevated aPS IgM antibodies, likely contributing to his CNS venous sinus thromboses and STEMI. It is important to screen for antiphosphatidylserine antibodies in cases of unprovoked thrombosis when standard thrombophilia analysis is unrevealing. This will assist in identifying pathogenicity and help prevent recurrence of subsequent thromboses.


Assuntos
Anticorpos Antifosfolipídeos/imunologia , Síndrome Antifosfolipídica/diagnóstico , Infarto do Miocárdio/imunologia , Fosfatidilserinas/imunologia , Trombose dos Seios Intracranianos/diagnóstico , Anticorpos Antifosfolipídeos/efeitos adversos , Síndrome Antifosfolipídica/complicações , Síndrome Antifosfolipídica/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Infarto do Miocárdio/diagnóstico , Infarto do Miocárdio/etiologia , Fosfatidilserinas/antagonistas & inibidores , Trombose dos Seios Intracranianos/etiologia , Trombose dos Seios Intracranianos/imunologia
9.
Biochim Biophys Acta Mol Cell Biol Lipids ; 1863(9): 980-990, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29787912

RESUMO

Given their important role in neuronal function, there has been an increasing focus on altered lipid levels in brain disorders. The effect of a high-fat (HF) diet on the lipid profiles of the cortex, hippocampus, hypothalamus, and olfactory bulb of the mouse brain was investigated using nanoflow ultrahigh pressure liquid chromatography-electrospray ionization-tandem mass spectrometry in the current study. For 8 weeks, two groups of 5-week-old mice were fed either an HF or normal diet (6 mice from each group analyzed as the F and N groups, respectively). The remaining mice in both groups then received a 4-week normal diet. Each group was then subdivided into two groups for another 4-week HF or normal diet. Quantitative analysis of 270 of the 359 lipids identified from brain tissue revealed that an HF diet significantly affected the brain lipidome in all brain regions that were analyzed. The HF diet significantly increased diacylglycerols, which play a role in insulin resistance in all regions that were analyzed. Although the HF diet increased most lipid species, the majority of phosphatidylserine species were decreased, while lysophosphatidylserine species, with the same acyl chain, were substantially increased. This result can be attributed to increased oxidative stress due to the HF diet. Further, weight-cycling (yo-yo effect) was found more critical for the perturbation of brain lipid profiles than weight gain without a preliminary experience of an HF diet. The present study reveals systematic alterations in brain lipid levels upon HF diet analyzed either by lipid class and molecular levels.


Assuntos
Córtex Cerebral/efeitos dos fármacos , Dieta Hiperlipídica , Gorduras na Dieta/administração & dosagem , Hipocampo/efeitos dos fármacos , Hipotálamo/efeitos dos fármacos , Metaboloma , Bulbo Olfatório/efeitos dos fármacos , Animais , Química Encefálica , Córtex Cerebral/metabolismo , Cromatografia Líquida de Alta Pressão , Diglicerídeos/agonistas , Diglicerídeos/metabolismo , Hipocampo/metabolismo , Hipotálamo/metabolismo , Resistência à Insulina , Metabolismo dos Lipídeos/efeitos dos fármacos , Lisofosfolipídeos/agonistas , Lisofosfolipídeos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Bulbo Olfatório/metabolismo , Estresse Oxidativo , Fosfatidilserinas/antagonistas & inibidores , Fosfatidilserinas/metabolismo , Espectrometria de Massas por Ionização por Electrospray
10.
J Trauma Acute Care Surg ; 84(6): 838-846, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29538229

RESUMO

BACKGROUND: Translocation of phosphatidylserine from the inner leaflet to the outer leaflet of the endothelial membrane via phospholipid scramblase-1 (PLSCR1) is an apoptotic signal responsible for the loss of endothelial barrier integrity after ischemia-reperfusion injury (IRI). We hypothesized that inhibiting phosphatidylserine expression on endothelial cells would attenuate IRI induced increases in hydraulic permeability (Lp). METHODS: Mesenteric Lp was measured in rat post-capillary mesenteric venules subjected to IRI via superior mesenteric artery (SMA) occlusion (45 minutes) and release (300 minutes) in conjunction with several inhibitors of phosphatidylserine exposure as follows: (1) inhibition of PLSCR1 translocation (dithioerythritol, n = 3), (2) inhibition of PLSCR1 membrane trafficking (2-bromopalmitate [2-BP], n = 3), and (3) inhibition of ion exchange necessary for PLSCR1 function (4,4'-Diisothiocyano-2,2'-stilbenedisulfonic acid [DIDS], n = 3). Under the same IRI conditions, rats were also administered targeted inhibitors of phosphatidylserine exposure including knockdown of PLSCR1 (n = 3) using RNA interference (RNAi), and as a potential therapeutic tool Diannexin, a selective phosphatidylserine blocker (n = 3). RESULTS: During IRI net Lp increased by 80% (p < 0.01). Net reductions of Lp were accomplished by 2-BP (46% reduction, p = 0.005), combined DET + 2-BP + DIDS (32% reduction, p = 0.04), RNAi (55% reduction, p = 0.002), Diannexin administered pre-SMA artery occlusion (73% reduction, p = 0.001), and post-SMA occlusion (70% reduction, p = 0.002). CONCLUSION: Phosphatidylserine exposure is a key event in the pathogenesis of microvascular dysfunction during IRI. Clinically, inhibition of phosphatidylserine exposure is a promising strategy that may 1 day be used to mitigate the effects of IRI.


Assuntos
Endotélio Vascular/metabolismo , Fosfatidilserinas/metabolismo , Traumatismo por Reperfusão/metabolismo , Ácido 4,4'-Di-Isotiocianoestilbeno-2,2'-Dissulfônico/farmacologia , Animais , Anexina A5/farmacologia , Permeabilidade Capilar , Ditioeritritol/farmacologia , Feminino , Fosfatidilserinas/antagonistas & inibidores , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Traumatismo por Reperfusão/prevenção & controle , Vênulas
11.
Semin Cancer Biol ; 52(Pt 1): 17-25, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-28870843

RESUMO

Cancer is a leading cause of mortality and morbidity globally. Many prominent cancer-associated molecules have been identified over the recent years which include EGFR, CD44, TGFbRII, HER2, miR-497, NMP22, BTA, Fibrin/FDP etc. These biomarkers are often used for screening, detection, diagnosis, prognosis, prediction and monitoring of cancer development. Phosphatidylserine (PS) is an essential component in all human cells which is present on the inner leaflet of the cell membrane. The oxidative stress causes exposure of PS on the surface of the vascular endothelium in the cancer cells (lung, breast, pancreatic, bladder, skin, brain metastasis, rectal adenocarcinoma etc.) but not on the normal cells. The external PS is regulated by calcium-dependent flippase activity. Cancer cell lines with high surface PS have low flippase activity and high intracellular calcium content. Human Annexin-V, PS targeting antibodies (PGN635 and bavituximab and mch1N11), lysosomal protein, phospholipid Saposin C dioleoylphosphatidylserine (SapC-DOPS), peptide-peptoid hybrid PPS1, PS-binding 14-mer peptide (PSBP-6) and hexapeptide (E3) have been reported to target PS present on cancer cell surface. High expression of CD47 inhibits tumor cell phagocytosis by macrophages. The PS cancer biomarker has also been used to target the drugs to cancer cells specifically without affecting other healthy cells. Currently, the fusion protein (FP) consisting of L-methionase linked to human Annexin-V has been reported to target the cancer cells. The FP catalyzes the conversion of non-toxic prodrug selenomethionine into toxic methyl selenol which thus also prevents the methionine (essential amino acid) supplementation to the cancer cells.


Assuntos
Biomarcadores Tumorais/metabolismo , Membrana Celular/metabolismo , Terapia de Alvo Molecular/métodos , Neoplasias/metabolismo , Fosfatidilserinas/metabolismo , Biomarcadores Tumorais/antagonistas & inibidores , Membrana Celular/efeitos dos fármacos , Humanos , Modelos Biológicos , Terapia de Alvo Molecular/tendências , Neoplasias/diagnóstico , Neoplasias/tratamento farmacológico , Fosfatidilserinas/antagonistas & inibidores , Ligação Proteica , Saposinas/metabolismo , Saposinas/uso terapêutico
12.
Biochem Biophys Res Commun ; 487(3): 560-566, 2017 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-28427942

RESUMO

Ethyl pyruvate (EP) is a stable lipophilic pyruvate derivative. Studies demonstrated that EP shows potent anti-oxidation, anti-inflammatory and anti-coagulant effects. Inflammation and coagulation are closely interacted with platelet activation. However, it is unclear whether EP has anti-platelet effects. Therefore, we investigated the anti-platelet effect of EP in this study in vitro. We found that EP inhibited agonists induced platelets aggregation, ATP release and adhesion to collagen. Flow cytometric analysis revealed that EP inhibited agonist induced platelets PAC-1 binding, as well as P-selectin and CD40L expression. The underlying mechanism of action may involve the inhibition of platelet PI3K/Akt and Protein Kinase C (PKC) signaling pathways. Additionally, EP dose dependently inhibited platelet PS exposure induced by high concentration thrombin. Lactate dehydrogenase (LDH) activity assay and mice platelet count implied that EP may have no toxic effect on platelets. Therefore, we are the first to report that EP has potent anti-platelet activity and attenuates platelet PS exposure in vitro, suggesting that the inhibitory effects of EP on platelets may also play important roles in improvement of inflammation and coagulation disorder in related animal models.


Assuntos
Proteínas Sanguíneas/farmacologia , Fosfatidilserinas/antagonistas & inibidores , Agregação Plaquetária/efeitos dos fármacos , Piruvatos/farmacologia , Trifosfato de Adenosina/antagonistas & inibidores , Trifosfato de Adenosina/metabolismo , Animais , Colágeno/antagonistas & inibidores , Colágeno/farmacologia , Relação Dose-Resposta a Droga , Feminino , Voluntários Saudáveis , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Fosfatidilserinas/administração & dosagem , Fosfatidilserinas/farmacologia , Adesividade Plaquetária/efeitos dos fármacos , Relação Estrutura-Atividade , Trombina/antagonistas & inibidores , Trombina/farmacologia
13.
Breast Cancer Res ; 18(1): 50, 2016 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-27169467

RESUMO

BACKGROUND: The purpose of this study was to investigate the potential of antibody-directed immunotherapy targeting the aminophospholipid phosphatidylserine, which promotes immunosuppression when exposed in the tumor microenvironment, alone and in combination with antibody treatment towards the T-cell checkpoint inhibitor PD-1 in breast carcinomas, including triple-negative breast cancers. METHODS: Immune-competent mice bearing syngeneic EMT-6 or E0771 tumors were subjected to treatments comprising of a phosphatidylserine-targeting and an anti-PD-1 antibody either as single or combinational treatments. Anti-tumor effects were determined by tumor growth inhibition and changes in overall survival accompanying each treatment. The generation of a tumor-specific immune response in animals undergoing complete tumor regression was assessed by secondary tumor cell challenge and splenocyte-produced IFNγ in the presence or absence of irradiated tumor cells. Changes in the presence of tumor-infiltrating lymphocytes were assessed by flow cytometry, while mRNA-based immune profiling was determined using NanoString PanCancer Immune Profiling Panel analysis. RESULTS: Treatment by a phosphatidylserine-targeting antibody inhibits in-vivo growth and significantly enhances the anti-tumor activity of antibody-mediated PD-1 therapy, including providing a distinct survival advantage over treatment by either single agent. Animals in which complete tumor regression occurred with combination treatments were resistant to secondary tumor challenge and presented heightened expression levels of splenocyte-produced IFNγ. Combinational treatment by a phosphatidylserine-targeting antibody with anti-PD-1 therapy increased the number of tumor-infiltrating lymphocytes more than that observed with single-arm therapies. Finally, immunoprofiling analysis revealed that the combination of anti-phosphatidylserine targeting antibody and anti-PD-1 therapy enhanced tumor-infiltrating lymphocytes, and increased expression of pro-immunosurveillance-associated cytokines while significantly decreasing expression of pro-tumorigenic cytokines that were induced by single anti-PD-1 therapy. CONCLUSIONS: Our data suggest that antibody therapy targeting phosphatidylserine-associated immunosuppression, which has activity as a single agent, can significantly enhance immunotherapies targeting the PD-1 pathway in murine breast neoplasms, including triple-negative breast cancers.


Assuntos
Anticorpos Monoclonais/farmacologia , Antineoplásicos/farmacologia , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismo , Neoplasias de Mama Triplo Negativas/imunologia , Neoplasias de Mama Triplo Negativas/metabolismo , Animais , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Biomarcadores Tumorais , Linhagem Celular Tumoral , Citocinas/genética , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Expressão Gênica , Perfilação da Expressão Gênica , Humanos , Imunoterapia , Interferon gama/biossíntese , Estimativa de Kaplan-Meier , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Camundongos , Terapia de Alvo Molecular , Fosfatidilserinas/antagonistas & inibidores , Receptor de Morte Celular Programada 1/genética , Baço/citologia , Baço/imunologia , Baço/metabolismo , Linfócitos T/efeitos dos fármacos , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/patologia , Ensaios Antitumorais Modelo de Xenoenxerto
14.
Cancer Immunol Res ; 4(6): 531-40, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-27045021

RESUMO

In tumor-bearing animals, the membrane phospholipid phosphatidylserine (PS) suppresses immune responses, suggesting that PS signaling could counteract the antitumor effect of antibody-driven immune checkpoint blockade. Here, we show that treating melanoma-bearing mice with a PS-targeting antibody enhances the antitumor activity of downstream checkpoint inhibition. Combining PS-targeting antibodies with CTLA-4 or PD-1 blockade resulted in significantly greater inhibition of tumor growth than did single-agent therapy. Moreover, combination therapy enhanced CD4(+) and CD8(+) tumor-infiltrating lymphocyte numbers; elevated the fraction of cells expressing the proinflammatory cytokines IL2, IFNγ, and TNFα; and increased the ratio of CD8 T cells to myeloid-derived suppressor cells and regulatory T cells in tumors. Similar changes in immune cell profiles were observed in splenocytes. Taken together, these data show that antibody-mediated PS blockade enhances the antitumor efficacy of immune checkpoint inhibition. Cancer Immunol Res; 4(6); 531-40. ©2016 AACR.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Antígeno CTLA-4/imunologia , Melanoma Experimental/tratamento farmacológico , Fosfatidilserinas/antagonistas & inibidores , Receptor de Morte Celular Programada 1/imunologia , Animais , Anticorpos Monoclonais/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Citocinas/biossíntese , Feminino , Imunofenotipagem , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/imunologia , Masculino , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Transplante de Neoplasias , Fosfatidilserinas/imunologia , Baço/imunologia
15.
Oncotarget ; 7(21): 30678-90, 2016 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-27120792

RESUMO

Phosphatidylserine (PS) is an anionic phospholipid maintained on the inner-leaflet of the cell membrane and is externalized in malignant cells. We previously launched a careful unbiased selection targeting biomolecules (e.g. protein, lipid or carbohydrate) distinct to cancer cells by exploiting HCC4017 lung cancer and HBEC30KT normal epithelial cells derived from the same patient, identifying HCC4017 specific peptide-peptoid hybrid PPS1. In this current study, we identified PS as the target of PPS1. We validated direct PPS1 binding to PS using ELISA-like assays, lipid dot blot and liposome based binding assays. In addition, PPS1 recognized other negatively charged and cancer specific lipids such as phosphatidic acid, phosphatidylinositol and phosphatidylglycerol. PPS1 did not bind to neutral lipids such as phosphatidylethanolamine found in cancer and phosphatidylcholine and sphingomyelin found in normal cells. Further we found that the dimeric version of PPS1 (PPS1D1) displayed strong cytotoxicity towards lung cancer cell lines that externalize PS, but not normal cells. PPS1D1 showed potent single agent anti-tumor activity and enhanced the efficacy of docetaxel in mice bearing H460 lung cancer xenografts. Since PS and anionic phospholipid externalization is common across many cancer types, PPS1 may be an alternative to overcome limitations of protein targeted agents.


Assuntos
Neoplasias Pulmonares/tratamento farmacológico , Oligopeptídeos/farmacologia , Fosfatidilserinas/antagonistas & inibidores , Ensaios Antitumorais Modelo de Xenoenxerto , Animais , Linhagem Celular , Linhagem Celular Tumoral , Feminino , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Lipídeos de Membrana/antagonistas & inibidores , Lipídeos de Membrana/metabolismo , Camundongos Endogâmicos NOD , Camundongos SCID , Terapia de Alvo Molecular , Oligopeptídeos/química , Oligopeptídeos/metabolismo , Peptídeos/química , Peptoides/química , Fosfatidilserinas/metabolismo , Ligação Proteica , Carga Tumoral/efeitos dos fármacos
16.
Ann Surg Oncol ; 23(Suppl 5): 583-591, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-26847681

RESUMO

BACKGROUND: Currently, the only FDA-approved systemic therapy for hepatocellular carcinoma (HCC) is the multi-receptor tyrosine kinase inhibitor, sorafenib, which provides only modest clinical benefit. We recently showed that treatment with a phosphatidylserine (PS) targeting agent suppresses tumor growth by targeting tumor vasculature and reactivating antitumor immunity. METHODS: We tested the hypothesis that sorafenib increases PS exposure on tumor vasculature, thereby enhancing the antitumor efficacy of PS targeting. We evaluated the efficacy of combining a PS targeting agent (2aG4) with sorafenib in murine xenograft models of human HCC. RESULTS: Our results demonstrate that combination of 2aG4 and sorafenib had a superior therapeutic effect over single agent therapy. Mechanistic studies showed that sorafenib significantly increased PS exposure on tumor vasculature; the percentage of PS-positive vessels increased from 19 to 52, 23 to 68, and 30 to 55 % in PLC/PRF/5, C3A, and Huh7 tumors, respectively. Combination therapy significantly decreased tumor microvessel density and the level of M2 macrophages, while increasing the apoptotic index of tumor endothelial cells and the frequency of M1 macrophages. Furthermore, we report the findings of a Phase I clinical study of bavituximab, a chimeric version of 2aG4, combined with sorafenib in HCC patients. The Phase I results demonstrate the appropriate dose of bavituximab to be given with sorafenib in future clinical trials. CONCLUSIONS: Overall, these results strongly support the combination of bavituximab with sorafenib as a promising systemic therapeutic strategy for the treatment for advanced HCC patients.


Assuntos
Anticorpos Monoclonais/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma Hepatocelular/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Fosfatidilserinas/antagonistas & inibidores , Adulto , Idoso , Animais , Anticorpos Monoclonais/efeitos adversos , Anticorpos Monoclonais/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Apoptose/efeitos dos fármacos , Carcinoma Hepatocelular/irrigação sanguínea , Endotélio/efeitos dos fármacos , Endotélio/metabolismo , Feminino , Células Endoteliais da Veia Umbilical Humana , Humanos , Neoplasias Hepáticas/irrigação sanguínea , Masculino , Dose Máxima Tolerável , Camundongos , Pessoa de Meia-Idade , Transplante de Neoplasias , Niacinamida/administração & dosagem , Niacinamida/análogos & derivados , Fenótipo , Compostos de Fenilureia/administração & dosagem , Fosfatidilserinas/metabolismo , Rituximab/farmacologia , Sorafenibe , Ensaios Antitumorais Modelo de Xenoenxerto
17.
Food Chem Toxicol ; 89: 47-53, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26774912

RESUMO

Hydroxytyrosol (HT) is a phenolic antioxidant naturally occurring in virgin olive oil. In this study, we investigated the possible protective effects of HT on programmed suicidal death (eryptosis) induced by mercury (Hg) treatment in intact human erythrocytes (RBC). Our study confirms that the Hg-eryptosis is characterized by phosphatidylserine (PS) exposure at the cell surface, with cell shrinkage and ATP and glutathione depletion; calcium influx is also a key event that triggers eryptosis. Here we report that cell preconditioning with an optimal dose (1-5 µM) of HT prior to exposure to 2.5 µM HgCl2 causes a noteworthy decrease in PS-exposing RBC, almost restoring ATP and GSH content. Conversely, HT shows no effect against decrease in cell volume nor against influx of extracellular calcium. Taken together our data provide the first experimental evidence of the efficacy of HT in modulating the programmed suicidal death in non nucleated cells; the reported findings also confirm that the prevention of Hg toxicity should be regarded as an additional mechanism responsible for the health-promoting potential of this dietary phenol. Finally, virgin olive oil would appear to be a promising healthy food to reduce the adverse effects of chronic mercury exposure in humans.


Assuntos
Apoptose/efeitos dos fármacos , Exposição Ambiental , Eritrócitos/efeitos dos fármacos , Glutationa/metabolismo , Mercúrio/toxicidade , Álcool Feniletílico/análogos & derivados , Fosfatidilserinas/antagonistas & inibidores , Cálcio/metabolismo , Eritrócitos/metabolismo , Humanos , Técnicas In Vitro , Álcool Feniletílico/farmacologia
18.
Int Urol Nephrol ; 48(2): 175-81, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26560472

RESUMO

OBJECTIVES: We investigated the possible involvement of multidrug resistance protein 1 P-glycoprotein (MDR1 P-gp) in the oxalate-induced redistribution of phosphatidylserine in renal epithelial cell membranes. METHODS: Real-time PCR and western blotting were used to examine MDR1 expression in Madin-Darby canine kidney cells at the mRNA and protein levels, respectively, whereas surface-expressed phosphatidylserine was detected by the annexin V-binding assay. RESULTS: Oxalate treatment resulted in increased synthesis of MDR1, which resulted in phosphatidylserine (PS) externalization in the renal epithelial cell membrane. Treatment with the MDR1 inhibitor PSC833 significantly attenuated phosphatidylserine externalization. Transfection of the human MDR1 gene into renal epithelial cells significantly increased PS externalization. CONCLUSIONS: To our knowledge, this study is the first to show that oxalate increases the synthesis of MDR1 P-gp, which plays a key role in hyperoxaluria-promoted calcium oxalate urolithiasis by facilitating phosphatidylserine redistribution in renal epithelial cells.


Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Regulação da Expressão Gênica , Nefrolitíase/genética , Oxalatos/efeitos adversos , Fosfatidilserinas/metabolismo , RNA Mensageiro/genética , Urotélio/metabolismo , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Animais , Western Blotting , Membrana Celular/metabolismo , Membrana Celular/patologia , Células Cultivadas , Ciclosporinas/farmacologia , Cães , Resistência a Múltiplos Medicamentos , Citometria de Fluxo , Humanos , Nefrolitíase/tratamento farmacológico , Nefrolitíase/metabolismo , Fosfatidilserinas/antagonistas & inibidores , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Urotélio/efeitos dos fármacos , Urotélio/patologia
19.
Toxins (Basel) ; 7(11): 4390-403, 2015 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-26516916

RESUMO

Increased accumulation of indolic uremic solutes in the blood of uremic patients contributes to the risk of thrombotic events. Red blood cells (RBCs), the most abundant blood cells in circulation, may be a privileged target of these solutes. However, the effect of uremic solutes indoxyl sulfate (IS) and indole-3-acetic acid (IAA) on procoagulant activity (PCA) of erythrocyte is unclear. Here, RBCs from healthy adults were treated with IS and IAA (mean and maximal concentrations reported in uremic patients). Phosphatidylserine (PS) exposure of RBCs and their microparticles (MPs) release were labeled with Alexa Fluor 488-lactadherin and detected by flow cytometer. Cytosolic Ca(2+) ([Ca(2+)]) with Fluo 3/AM was analyzed by flow cytometer. PCA was assessed by clotting time and purified coagulation complex assays. We found that PS exposure, MPs generation, and consequent PCA of RBCs at mean concentrations of IS and IAA enhanced and peaked in maximal uremic concentrations. Moreover, 128 nM lactadherin, a PS inhibitor, inhibited over 90% PCA of RBCs and RMPs. Eryptosis or damage, by indolic uremic solutes was due to, at least partially, the increase of cytosolic [Ca(2+)]. Our results suggest that RBC eryptosis in uremic solutes IS and IAA plays an important role in thrombus formation through releasing RMPs and exposing PS. Lactadherin acts as an efficient anticoagulant in this process.


Assuntos
Coagulação Sanguínea/efeitos dos fármacos , Micropartículas Derivadas de Células/metabolismo , Eritrócitos/efeitos dos fármacos , Indóis/farmacologia , Fosfatidilserinas/farmacologia , Uremia/metabolismo , Anticoagulantes/farmacologia , Cálcio/sangue , Citosol/metabolismo , Fator Xa/análise , Feminino , Citometria de Fluxo , Humanos , Indicã/farmacologia , Indicã/urina , Ácidos Indolacéticos/farmacologia , Ácidos Indolacéticos/urina , Indóis/urina , Masculino , Fosfatidilserinas/antagonistas & inibidores , Tromboplastina/análise , Trombose/sangue , Trombose/urina , Adulto Jovem
20.
Biochim Biophys Acta ; 1848(11 Pt A): 2918-31, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26239537

RESUMO

Host defense-derived peptides have emerged as a novel strategy for the development of alternative anticancer therapies. In this study we report on characteristic features of human lactoferricin (hLFcin) derivatives which facilitate specific killing of cancer cells of melanoma, glioblastoma and rhabdomyosarcoma compared with non-specific derivatives and the synthetic peptide RW-AH. Changes in amino acid sequence of hLFcin providing 9-11 amino acids stretched derivatives LF11-316, -318 and -322 only yielded low antitumor activity. However, the addition of the repeat (di-peptide) and the retro-repeat (di-retro-peptide) sequences highly improved cancer cell toxicity up to 100% at 20 µM peptide concentration. Compared to the complete parent sequence hLFcin the derivatives showed toxicity on the melanoma cell line A375 increased by 10-fold and on the glioblastoma cell line U-87mg by 2-3-fold. Reduced killing velocity, apoptotic blebbing, activation of caspase 3/7 and formation of apoptotic DNA fragments proved that the active and cancer selective peptides, e.g. R-DIM-P-LF11-322, trigger apoptosis, whereas highly active, though non-selective peptides, such as DIM-LF11-318 and RW-AH seem to kill rapidly via necrosis inducing membrane lyses. Structural studies revealed specific toxicity on cancer cells by peptide derivatives with loop structures, whereas non-specific peptides comprised α-helical structures without loop. Model studies with the cancer membrane mimic phosphatidylserine (PS) gave strong evidence that PS only exposed by cancer cells is an important target for specific hLFcin derivatives. Other negatively charged membrane exposed molecules as sialic acid, heparan and chondroitin sulfate were shown to have minor impact on peptide activity.


Assuntos
Apoptose/efeitos dos fármacos , Dipeptídeos/farmacologia , Lactoferrina/farmacologia , Fosfatidilserinas/antagonistas & inibidores , Sequência de Aminoácidos , Varredura Diferencial de Calorimetria , Linhagem Celular Tumoral , Membrana Celular/química , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Dicroísmo Circular , Dipeptídeos/química , Relação Dose-Resposta a Droga , Humanos , Lactoferrina/química , Bicamadas Lipídicas/química , Bicamadas Lipídicas/metabolismo , Lipídeos de Membrana/antagonistas & inibidores , Lipídeos de Membrana/química , Lipídeos de Membrana/metabolismo , Microscopia de Fluorescência , Modelos Moleculares , Dados de Sequência Molecular , Fosfatidilserinas/química , Fosfatidilserinas/metabolismo , Estrutura Secundária de Proteína , Fatores de Tempo
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