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In the evolving landscape of biomedical research, the convergence of molecular biology and translational medicine has ushered in a new era of pharmaceutical innovation. This paradigm shift, characterized by significant advances in targeted therapies and gene editing, emphasizes the critical role of integrating academic research - and academic researchers - within industry settings. Contemporary innovation models are moving beyond traditional, corporation-centered frameworks, adopting more open, collaborative approaches. Here, we discuss the challenges and solutions brought about by this new direction in pharma innovation and describe the BioMed X innovation model, a unique open innovation approach that has been growing continuously over the past ten years.
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BACKGROUND: The envelope protein of human endogenous retrovirus W (HERV-W-Env) is expressed by macrophages and microglia, mediating axonal damage in chronic active MS lesions. OBJECTIVE AND METHODS: This phase 2, double-blind, 48-week trial in relapsing-remitting MS with 48-week extension phase assessed the efficacy and safety of temelimab; a monoclonal antibody neutralizing HERV-W-Env. The primary endpoint was the reduction of cumulative gadolinium-enhancing T1-lesions in brain magnetic resonance imaging (MRI) scans at week 24. Additional endpoints included numbers of T2 and T1-hypointense lesions, magnetization transfer ratio, and brain atrophy. In total, 270 participants were randomized to receive monthly intravenous temelimab (6, 12, or 18 mg/kg) or placebo for 24 weeks; at week 24 placebo-treated participants were re-randomized to treatment groups. RESULTS: The primary endpoint was not met. At week 48, participants treated with 18 mg/kg temelimab had fewer new T1-hypointense lesions (p = 0.014) and showed consistent, however statistically non-significant, reductions in brain atrophy and magnetization transfer ratio decrease, as compared with the placebo/comparator group. These latter two trends were sustained over 96 weeks. No safety issues emerged. CONCLUSION: Temelimab failed to show an effect on features of acute inflammation but demonstrated preliminary radiological signs of possible anti-neurodegenerative effects. Current data support the development of temelimab for progressive MS. TRIAL REGISTRATION: CHANGE-MS: ClinicalTrials.gov: NCT02782858, EudraCT: 2015-004059-29; ANGEL-MS: ClinicalTrials.gov: NCT03239860, EudraCT: 2016-004935-18.
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Anticuerpos Monoclonales Humanizados , Esclerosis Múltiple Recurrente-Remitente , Esclerosis Múltiple , Anticuerpos Monoclonales Humanizados/efectos adversos , Anticuerpos Monoclonales Humanizados/uso terapéutico , Método Doble Ciego , Productos del Gen env/uso terapéutico , Humanos , Imagen por Resonancia Magnética , Esclerosis Múltiple/tratamiento farmacológico , Esclerosis Múltiple Recurrente-Remitente/diagnóstico por imagen , Esclerosis Múltiple Recurrente-Remitente/tratamiento farmacológico , Esclerosis Múltiple Recurrente-Remitente/patología , Resultado del TratamientoRESUMEN
Artefenomel and DSM265 are two new compounds that have been shown to be well tolerated and effective when administered as monotherapy malaria treatment. This study aimed to determine the safety, pharmacokinetics, and pharmacodynamics of artefenomel and DSM265 administered in combination to healthy subjects in a volunteer infection study using the Plasmodium falciparum-induced blood-stage malaria model. Thirteen subjects were inoculated with parasite-infected erythrocytes on day 0 and received a single oral dose of artefenomel and DSM265 on day 7. Cohort 1 (n = 8) received 200 mg artefenomel plus 100 mg DSM265, and cohort 2 (n = 5) received 200 mg artefenomel plus 50 mg DSM265. Blood samples were collected to measure parasitemia, gametocytemia, and artefenomel-DSM265 plasma concentrations. There were no treatment-related adverse events. The pharmacokinetic profiles of artefenomel and DSM265 were similar to those of the compounds when administered as monotherapy, suggesting no pharmacokinetic interactions. A reduction in parasitemia occurred in all subjects following treatment (log10 parasite reduction ratios over 48 h [PRR48] of 2.80 for cohort 1 and 2.71 for cohort 2; parasite clearance half-lives of 5.17 h for cohort 1 and 5.33 h for cohort 2). Recrudescence occurred in 5/8 subjects in cohort 1 between days 19 and 28 and in 5/5 subjects in cohort 2 between days 15 and 22. Low-level gametocytemia (1 to 330 female gametocytes/ml) was detected in all subjects from day 14. The results of this single-dosing combination study support the further clinical development of the use of artefenomel and DSM265 in combination as a treatment for falciparum malaria. (This study has been registered at ClinicalTrials.gov under identifier NCT02389348.).
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Adamantano/análogos & derivados , Antimaláricos/administración & dosificación , Malaria Falciparum/tratamiento farmacológico , Parasitemia/tratamiento farmacológico , Peróxidos/administración & dosificación , Pirimidinas/administración & dosificación , Triazoles/administración & dosificación , Adamantano/administración & dosificación , Adamantano/farmacocinética , Administración Oral , Adulto , Antimaláricos/farmacocinética , Combinación de Medicamentos , Femenino , Voluntarios Sanos , Humanos , Malaria Falciparum/metabolismo , Malaria Falciparum/parasitología , Masculino , Persona de Mediana Edad , Parasitemia/metabolismo , Parasitemia/parasitología , Peróxidos/farmacocinética , Plasmodium falciparum/efectos de los fármacos , Pirimidinas/farmacocinética , Triazoles/farmacocinética , Adulto JovenRESUMEN
DSM265 is a novel antimalarial drug in clinical development that acts as a selective inhibitor of Plasmodium dihydroorotate dehydrogenase. In a previous phase 1b study, a single 150-mg dose of DSM265 showed partial efficacy against experimentally induced blood-stage Plasmodium falciparum malaria (IBSM). Pharmacokinetic/pharmacodynamic modeling predicted a human efficacious dose of 340 mg. The primary objectives of the current study were to determine the safety and efficacy of a single oral 400-mg dose of DSM265 against P. falciparum in the IBSM model. Eight healthy participants were inoculated intravenously with 2,800 parasites and treated with DSM265 7 days later. Unexpectedly, one participant did not develop parasitemia during the study. All other participants developed parasitemia, with the complete clearance of asexual parasites occurring following DSM265 treatment. All seven subjects also became gametocytemic. The secondary objectives were to investigate the gametocytocidal and transmission-blocking activity of a second 400-mg dose of DSM265, which was administered 23 days after inoculation. Gametocytes were not cleared by the second dose of DSM265, and transmission-blocking activity could not be determined due to low gametocyte densities. Three DSM265-related adverse events occurred, including a cutaneous rash in one subject on the day of the second DSM265 dose. The results obtained in this study support the prediction of the efficacious dose of DSM265 and provide further evidence that DSM265 is generally safe and well tolerated. In addition, this study confirms preclinical data indicating that DSM265 permits the development and maturation of gametocytes and does not clear mature circulating gametocytes. (This study has been registered at ClinicalTrials.gov under identifier NCT02573857.).
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Antimaláricos/farmacología , Malaria Falciparum/tratamiento farmacológico , Plasmodium falciparum/fisiología , Pirimidinas/farmacología , Triazoles/farmacología , Animales , Anopheles , Antimaláricos/administración & dosificación , Antimaláricos/efectos adversos , Antimaláricos/farmacocinética , Femenino , Semivida , Voluntarios Sanos , Humanos , Malaria Falciparum/parasitología , Malaria Falciparum/transmisión , Masculino , Mosquitos Vectores , Parasitemia/tratamiento farmacológico , Parasitemia/parasitología , Plasmodium falciparum/efectos de los fármacos , Pirimidinas/administración & dosificación , Pirimidinas/efectos adversos , Pirimidinas/farmacocinética , Triazoles/administración & dosificación , Triazoles/efectos adversos , Triazoles/farmacocinética , Adulto JovenRESUMEN
BACKGROUND: DSM265 is a novel, long-duration inhibitor of plasmodium dihydroorotate dehydrogenase (DHODH) with excellent selectivity over human DHODH and activity against blood and liver stages of Plasmodium falciparum. This study aimed to assess the efficacy of DSM265 in patients with P falciparum or Plasmodium vivax malaria infection. METHODS: This proof-of-concept, open-label, phase 2a study was conducted at the Asociación Civil Selva Amazónica in Iquitos, Peru. Patients aged 18-70 years, weighing 45-90 kg, who had clinical malaria (P falciparum or P vivax monoinfection) and fever within the previous 24 h were eligible. Exclusion criteria were clinical or laboratory signs of severe malaria, inability to take oral medicine, and use of other antimalarial treatment in the preceding 14 days. Patients were divided into cohorts of those with P falciparum (cohort a) or P vivax (cohort b) infection. Two initial cohorts received single oral doses of 400 mg DSM265. Patients were followed up for efficacy for 28 days and safety for 35 days. Further cohorts received escalated or de-escalated doses of DSM265, after safety and efficacy assessment of the initial dose. The primary endpoints were the proportion of patients achieving PCR-adjusted adequate clinical and parasitological response (ACPR) by day 14 for patients infected with P falciparum and the proportion of patients achieving a crude cure by day 14 for those infected with P vivax. Cohort success, the criteria for dose escalation, was defined as ACPR (P falciparum) or crude cure (P vivax) in at least 80% of patients in the cohort. The primary analysis was done in the intention-to-treat population (ITT) and the per-protocol population, and safety analyses were done in all patients who received the study drug. This study is registered at ClinicalTrials.gov (NCT02123290). FINDINGS: Between Jan 12, 2015, and Dec 2, 2015, 45 Peruvian patients (24 with P falciparum [cohort a] and 21 with P vivax [cohort b] infection) were sequentially enrolled. For patients with P falciparum malaria in the per-protocol population, all 11 (100%) in the 400 mg group and eight (80%) of ten in the 250 mg group achieved ACPR on day 14. In the ITT analysis, 11 (85%) of 13 in the 400 mg group and eight (73%) of 11 in the 250 mg group achieved ACPR at day 14. For the patients with P vivax malaria, the primary endpoint was not met. In the per-protocol analysis, none of four patients who had 400 mg, three (50%) of six who had 600 mg, and one (25%) of four who had 800 mg DSM265 achieved crude cure at day 14. In the ITT analysis, none of five in the 400 mg group, three (33%) of nine in the 600 mg group, and one (14%) of seven in the 800 mg group achieved crude cure at day 14. During the 28-day extended observation of P falciparum patients, a resistance-associated mutation in the gene encoding the DSM265 target DHODH was observed in two of four recurring patients. DSM265 was well tolerated. The most common adverse events were pyrexia (20 [44%] of 45) and headache (18 [40%] of 45), which are both common symptoms of malaria, and no patients had any treatment-related serious adverse events or adverse events leading to study discontinuation. INTERPRETATION: After a single dose of DSM265, P falciparum parasitaemia was rapidly cleared, whereas against P vivax, DSM265 showed less effective clearance kinetics. Its long duration of action provides the potential to prevent recurrence of P falciparum after treatment with a single dose, which should be further assessed in future combination studies. FUNDING: The Global Health Innovative Technology Fund, the Bill & Melinda Gates Foundation, the National Institutes of Health (R01 AI103058), the Wellcome Trust, and the UK Department of International Development.
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Antimaláricos/administración & dosificación , Malaria Falciparum/tratamiento farmacológico , Malaria Vivax/tratamiento farmacológico , Plasmodium falciparum/inmunología , Pirimidinas/administración & dosificación , Triazoles/administración & dosificación , Adulto , Estudios de Cohortes , Dihidroorotato Deshidrogenasa , Femenino , Humanos , Malaria Falciparum/inmunología , Malaria Falciparum/parasitología , Malaria Vivax/inmunología , Malaria Vivax/parasitología , Masculino , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH , PerúRESUMEN
BACKGROUND: Combination medicines including an artemisinin are the mainstay of antimalarial therapy. Artemisinins are potent embryotoxicants in animal species due to their trioxane moiety. METHODS: As part of its development, the new synthetic trioxolane antimalarial artefenomel (OZ439) was tested in rat whole embryo culture and in rat embryo-fetal toxicity studies with dosing throughout organogenesis or with a single dose on Gestational Day (GD) 12. The single-dose studies included groups treated with artesunate to allow a direct comparison of the embryotoxicity of the two antimalarials and included toxicokinetics hematology and histological examination of embryos. In addition, the distribution of artefenomel-related material in plasma was determined after the administration of 14 C-artefenomel. RESULTS: Artefenomel and artesunate showed similar patterns of embryotoxicity including cardiovascular defects and resorption with a steep dose-response. They both also caused a depletion of circulating embryonic erythroblasts both in vitro and in vivo and decreases in maternal reticulocyte count. However, artefenomel was â¼250-fold less potent than the active metabolite of artesunate (dihydroartemisinin) as an embryotoxicant in vitro. The safety margin (based on AUC) for artefenomel administered on GD 12 was approximately 100-fold greater than that for artesunate. Also, unlike artesunate, artefenomel was not a selective developmental toxicant. CONCLUSIONS: The lesser embryotoxicity of artefenomel is likely linked to its original design which included two blocking side groups that had been introduced to lower the reactivity with ferrous iron. Our data support the hypothesis that artefenomel's improved safety margin is linked to a lower potential for inhibiting heme biosynthesis in embryonic erythroblasts.
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Adamantano/análogos & derivados , Antimaláricos/toxicidad , Artesunato/toxicidad , Embrión de Mamíferos/efectos de los fármacos , Peróxidos/toxicidad , Adamantano/farmacocinética , Adamantano/toxicidad , Animales , Artemisininas/toxicidad , Benzoxazinas/toxicidad , Relación Dosis-Respuesta a Droga , Femenino , Desarrollo Fetal/efectos de los fármacos , Edad Gestacional , Hemo/biosíntesis , Técnicas de Cultivo de Órganos , Organogénesis/efectos de los fármacos , Peróxidos/farmacocinética , Ftalimidas/toxicidad , RatasRESUMEN
BACKGROUND: DSM265 is a novel antimalarial that inhibits plasmodial dihydroorotate dehydrogenase, an enzyme essential for pyrimidine biosynthesis. We investigated the safety, tolerability, and pharmacokinetics of DSM265, and tested its antimalarial activity. METHODS: Healthy participants aged 18-55 years were enrolled in a two-part study: part 1, a single ascending dose (25-1200 mg), double-blind, randomised, placebo-controlled study, and part 2, an open-label, randomised, active-comparator controlled study, in which participants were inoculated with Plasmodium falciparum induced blood-stage malaria (IBSM) and treated with DSM265 (150 mg) or mefloquine (10 mg/kg). Primary endpoints were DSM265 safety, tolerability, and pharmacokinetics. Randomisation lists were created using a validated, automated system. Both parts were registered with the Australian New Zealand Clinical Trials Registry, number ACTRN12613000522718 (part 1) and number ACTRN12613000527763 (part 2). FINDINGS: In part 1, 73 participants were enrolled between April 12, 2013, and July 14, 2015 (DSM265, n=55; placebo, n=18). In part 2, nine participants were enrolled between Sept 30 and Nov 25, 2013 (150 mg DSM265, n=7; 10 mg/kg mefloquine, n=2). In part 1, 117 adverse events were reported; no drug-related serious or severe events were reported. The most common drug-related adverse event was headache. The mean DSM265 peak plasma concentration (Cmax) ranged between 1310 ng/mL and 34â800 ng/mL and was reached in a median time (tmax) between 1·5 h and 4 h, with a mean elimination half-life between 86 h and 118 h. In part 2, the log10 parasite reduction ratio at 48 h in the DSM265 (150 mg) group was 1·55 (95% CI 1·42-1·67) and in the mefloquine (10 mg/kg) group was 2·34 (2·17-2·52), corresponding to a parasite clearance half-life of 9·4 h (8·7-10·2) and 6·2 h (5·7-6·7), respectively. The median minimum inhibitory concentration of DSM265 in blood was estimated as 1040 ng/mL (range 552-1500), resulting in a predicted single efficacious dose of 340 mg. Parasite clearance was significantly faster in participants who received mefloquine than in participants who received DSM265 (p<0·0001). INTERPRETATION: The good safety profile, long elimination half-life, and antimalarial effect of DSM265 supports its development as a partner drug in a single-dose antimalarial combination treatment. FUNDING: Wellcome Trust, UK Department for International Development, Global Health Innovative Technology Fund, Bill & Melinda Gates Foundation.
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Antimaláricos/administración & dosificación , Mefloquina/uso terapéutico , Pirimidinas/administración & dosificación , Pirimidinas/farmacocinética , Triazoles/administración & dosificación , Triazoles/farmacocinética , Adolescente , Adulto , Antimaláricos/farmacocinética , Antimaláricos/uso terapéutico , Australia , Dihidroorotato Deshidrogenasa , Método Doble Ciego , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacocinética , Semivida , Humanos , Malaria Falciparum/tratamiento farmacológico , Persona de Mediana Edad , Nueva Zelanda , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH , Plasmodium falciparum , Pirimidinas/uso terapéutico , Triazoles/uso terapéuticoRESUMEN
BACKGROUND: A drug for causal (ie, pre-erythrocytic) prophylaxis of Plasmodium falciparum malaria with prolonged activity would substantially advance malaria control. DSM265 is an experimental antimalarial that selectively inhibits the parasite dihydroorotate dehydrogenase. DSM265 shows in vitro activity against liver and blood stages of P falciparum. We assessed the prophylactic activity of DSM265 against controlled human malaria infection (CHMI). METHODS: At the Institute of Tropical Medicine, Eberhard Karls University (Tübingen, Germany), healthy, malaria-naive adults were allocated to receive 400 mg DSM265 or placebo either 1 day (cohort 1A) or 7 days (cohort 2) before CHMI by direct venous inoculation (DVI) of 3200 aseptic, purified, cryopreserved P falciparum sporozoites (PfSPZ Challenge; Sanaria Inc, Rockville, MD, USA). An additional group received daily atovaquone-proguanil (250-100 mg) for 9 days, starting 1 day before CHMI (cohort 1B). Allocation to DSM265, atovaquone-proguanil, or placebo was randomised by an interactive web response system. Allocation to cohort 1A and 1B was open-label, within cohorts 1A and 2, allocation to DSM265 and placebo was double-blinded. All treatments were given orally. Volunteers were treated with an antimalarial on day 28, or when parasitaemic, as detected by thick blood smear (TBS) microscopy. The primary efficacy endpoint was time-to-parasitaemia, assessed by TBS. All participants receiving at least one dose of chemoprophylaxis or placebo were considered for safety, those receiving PfSPZ Challenge for efficacy analyses. Log-rank test was used to compare time-to-parasitemia between interventions. The trial was registered with ClinicalTrials.gov, number NCT02450578. FINDINGS: 22 participants were enrolled between Oct 23, 2015, and Jan 18, 2016. Five participants received 400 mg DSM265 and two participants received placebo 1 day before CHMI (cohort 1A), six participants received daily atovaquone-proguanil 1 day before CHMI (cohort 1B), and six participants received 400 mg DSM265 and two participants received placebo 7 days before CHMI (cohort 2). Five of five participants receiving DSM265 1 day before CHMI and six of six in the atovaquone-proguanil cohort were protected, whereas placebo recipients (two of two) developed malaria on days 11 and 14. When given 7 days before CHMI, three of six volunteers receiving DSM265 became TBS positive on days 11, 13, and 24. The remaining three DSM265-treated, TBS-negative participants of cohort 2 developed transient submicroscopic parasitaemia. Both participants receiving placebo 7 days before CHMI became TBS positive on day 11. The only possible DSM265-related adverse event was a moderate transient elevation in serum bilirubin in one participant. INTERPRETATION: A single dose of 400 mg DSM265 was well tolerated and had causal prophylactic activity when given 1 day before CHMI. Future trials are needed to investigate further the use of DSM265 for the prophylaxis of malaria. FUNDING: Global Health Innovative Technology Fund, Wellcome Trust, Bill & Melinda Gates Foundation through Medicines for Malaria Venture, and the German Center for Infection Research.
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Antimaláricos/administración & dosificación , Quimioprevención , Malaria Falciparum/tratamiento farmacológico , Plasmodium falciparum/inmunología , Pirimidinas/administración & dosificación , Triazoles/administración & dosificación , Administración Intravenosa , Adolescente , Adulto , Antimaláricos/uso terapéutico , Método Doble Ciego , Femenino , Humanos , Malaria Falciparum/inmunología , Malaria Falciparum/parasitología , Masculino , Persona de Mediana Edad , Parasitemia/inmunología , Parasitemia/parasitología , Pirimidinas/uso terapéutico , Esporozoítos/inmunología , Triazoles/uso terapéutico , VoluntariosRESUMEN
BACKGROUND: Ferroquine (SSR97193) is a candidate anti-malarial currently undergoing clinical trials for malaria. To better understand its pharmacokinetic (PK) and pharmacodynamic (PD) parameters the compound was tested in the experimentally induced blood stage malaria infection model in volunteers. METHODS: Male and non-pregnant female aged 18-50 years were screened for this phase II, controlled, single-centre clinical trial. Subjects were inoculated with ~1800 viable Plasmodium falciparum 3D7A-infected human erythrocytes, and treated with a single-dose of 800 mg ferroquine. Blood samples were taken at defined time-points to measure PK and PD parameters. The blood concentration of ferroquine and its active metabolite, SSR97213, were measured on dry blood spot samples by ultra-performance liquid chromatography with tandem mass spectrometry (LC-MS/MS). Parasitaemia and emergence of gametocytes were monitored by quantitative PCR. Safety was determined by recording adverse events and monitoring clinical laboratory assessments during the course of the study. RESULTS: Eight subjects were enrolled into the study, inoculated with infected erythrocytes and treated with 800 mg ferroquine. Ferroquine was rapidly absorbed with maximal exposure after 4-8 and 4-12 h exposure for SSR97213. Non-compartmental PK analysis resulted in estimates for half-lives of 10.9 and 23.8 days for ferroquine and SSR97213, respectively. Parasite clearance as reported by parasite reduction ratio was 162.9 (95 % CI 141-188) corresponding to a parasite clearance half-life of 6.5 h (95 % CI: 6.4-6.7 h). PK/PD modelling resulted in a predicted minimal parasiticidal concentration of 20 ng/mL, and the single dosing tested in this study was predicted to maintain an exposure above this threshold for 454 h (37.8 days). Although ferroquine was overall well tolerated, transient elevated transaminase levels were observed in three subjects. Paracetamol was the only concomitant treatment among the two out of these three subjects that may have played a role in the elevated transaminases levels. No clinically significant ECG abnormalities were observed. CONCLUSIONS: The parameters and PK/PD model derived from this study pave the way to the further rational development of ferroquine as an anti-malarial partner drug. The safety of ferroquine has to be further explored in controlled human trials. Trial registration anzctr.org.au (registration number: ACTRN12613001040752), registered 18/09/2013.
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Aminoquinolinas/administración & dosificación , Antimaláricos/administración & dosificación , Compuestos Ferrosos/administración & dosificación , Malaria Falciparum/tratamiento farmacológico , Plasmodium falciparum/efectos de los fármacos , Adolescente , Adulto , Aminoquinolinas/farmacocinética , Aminoquinolinas/farmacología , Antimaláricos/farmacocinética , Antimaláricos/farmacología , Sangre/parasitología , Análisis Químico de la Sangre , Cromatografía Liquida , Femenino , Compuestos Ferrosos/farmacocinética , Compuestos Ferrosos/farmacología , Voluntarios Sanos , Humanos , Masculino , Metalocenos , Persona de Mediana Edad , Carga de Parásitos , Plasmodium falciparum/aislamiento & purificación , Reacción en Cadena en Tiempo Real de la Polimerasa , Espectrometría de Masas en Tándem , Resultado del Tratamiento , Adulto JovenRESUMEN
Type 1 diabetes (T1D) remains a major health problem worldwide, with a steadily rising incidence yet no cure. Phosphoinositide 3-kinase-γ (PI3Kγ), a member of a family of lipid kinases expressed primarily in leukocytes, has been the subject of substantial research for its role in inflammatory diseases. However, the role of PI3Kγ inhibition in suppressing autoimmune T1D remains to be explored. We tested the role of the PI3Kγ inhibitor AS605240 in preventing and reversing diabetes in NOD mice and assessed the mechanisms by which this inhibition abrogates T1D. Our data indicate that the PI3Kγ pathway is highly activated in T1D. In NOD mice, we found upregulated expression of phosphorylated Akt (PAkt) in splenocytes. Notably, T regulatory cells (Tregs) showed significantly lower expression of PAkt compared with effector T cells. Inhibition of the PI3Kγ pathway by AS605240 efficiently suppressed effector T cells and induced Treg expansion through the cAMP response element-binding pathway. AS605240 effectively prevented and reversed autoimmune diabetes in NOD mice and suppressed T-cell activation and the production of inflammatory cytokines by autoreactive T cells in vitro and in vivo. These studies demonstrate the key role of the PI3Kγ pathway in determining the balance of Tregs and autoreactive cells regulating autoimmune diabetes.
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Diabetes Mellitus Tipo 1/tratamiento farmacológico , Inhibidores de las Quinasa Fosfoinosítidos-3 , Quinoxalinas/uso terapéutico , Linfocitos T Reguladores/efectos de los fármacos , Tiazolidinedionas/uso terapéutico , Animales , Diabetes Mellitus Tipo 1/metabolismo , Femenino , Hiperglucemia/tratamiento farmacológico , Hiperglucemia/metabolismo , Ratones , Ratones Endogámicos NOD , Fosforilación/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Quinoxalinas/farmacología , Bazo/efectos de los fármacos , Bazo/metabolismo , Linfocitos T Reguladores/metabolismo , Tiazolidinedionas/farmacologíaRESUMEN
Obligate intracellular pathogens such as Leishmania specifically target host phagocytes for survival and replication. Phosphoinositide 3-kinase γ (PI3Kγ), a member of the class I PI3Ks that is highly expressed by leukocytes, controls cell migration by initiating actin polymerization and cytoskeletal reorganization, which are processes also critical for phagocytosis. In this study, we demonstrate that class IB PI3K, PI3Kγ, plays a critical role in pathogenesis of chronic cutaneous leishmaniasis caused by L. mexicana. Using the isoform-selective PI3Kγ inhibitor, AS-605240 and PI3Kγ gene-deficient mice, we show that selective blockade or deficiency of PI3Kγ significantly enhances resistance against L. mexicana that is associated with a significant suppression of parasite entry into phagocytes and reduction in recruitment of host phagocytes as well as regulatory T cells to the site of infection. Furthermore, we demonstrate that AS-605240 is as effective as the standard antileishmanial drug sodium stibogluconate in treatment of cutaneous leishmaniasis caused by L. mexicana. These findings reveal a unique role for PI3Kγ in Leishmania invasion and establishment of chronic infection, and demonstrate that therapeutic targeting of host pathways involved in establishment of infection may be a viable strategy for treating infections caused by obligate intracellular pathogens such as Leishmania.
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Resistencia a la Enfermedad/efectos de los fármacos , Leishmania mexicana , Leishmaniasis Cutánea/parasitología , Fosfatidilinositol 3-Quinasas/metabolismo , Quinoxalinas/farmacología , Tiazolidinedionas/farmacología , Animales , Gluconato de Sodio Antimonio/uso terapéutico , Citometría de Flujo , Interacciones Huésped-Parásitos/efectos de los fármacos , Humanos , Leishmaniasis Cutánea/fisiopatología , Macrófagos , Ratones , Ratones Endogámicos C57BL , Microscopía Fluorescente , Neutrófilos , Fagocitos/efectos de los fármacos , Inhibidores de las Quinasa Fosfoinosítidos-3 , Quinoxalinas/uso terapéutico , Tiazolidinedionas/uso terapéuticoRESUMEN
Apoptosis signal-regulating kinase 1 (ASK1) is an evolutionarily conserved mitogen-activated protein kinase (MAPK) kinase kinase which plays important roles in stress and immune responses. Here, we show that ASK1 deficiency attenuates neuroinflammation in experimental autoimmune encephalomyelitis (EAE), without affecting the proliferation capability of T cells. Moreover, we found that EAE upregulates expression of Toll-like receptors (TLRs) in activated astrocytes and microglia, and that TLRs can synergize with ASK1-p38 MAPK signalling in the release of key chemokines from astrocytes. Consequently, oral treatment with a specific small molecular weight inhibitor of ASK1 suppressed EAE-induced autoimmune inflammation in both spinal cords and optic nerves. These results suggest that the TLR-ASK1-p38 pathway in glial cells may serve as a valid therapeutic target for autoimmune demyelinating disorders including multiple sclerosis.
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Enfermedades Desmielinizantes/metabolismo , MAP Quinasa Quinasa Quinasa 5/metabolismo , Transducción de Señal , Receptores Toll-Like/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Animales , Encefalopatías/tratamiento farmacológico , Encefalopatías/genética , Encefalopatías/inmunología , Encefalopatías/metabolismo , Quimiocinas/inmunología , Enfermedades Desmielinizantes/tratamiento farmacológico , Enfermedades Desmielinizantes/genética , Enfermedades Desmielinizantes/inmunología , Modelos Animales de Enfermedad , Encefalitis , Inhibidores Enzimáticos/administración & dosificación , Femenino , Regulación de la Expresión Génica , Enfermedad de Hashimoto/tratamiento farmacológico , Enfermedad de Hashimoto/genética , Enfermedad de Hashimoto/inmunología , Enfermedad de Hashimoto/metabolismo , Humanos , MAP Quinasa Quinasa Quinasa 5/antagonistas & inhibidores , MAP Quinasa Quinasa Quinasa 5/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Neuroglía/metabolismo , Ratas , Ratas Sprague-Dawley , Receptores Toll-Like/genética , Proteínas Quinasas p38 Activadas por Mitógenos/genéticaRESUMEN
RATIONALE: Sepsis is a leading cause of death in the intensive care unit, characterized by a systemic inflammatory response (SIRS) and bacterial infection, which can often induce multiorgan damage and failure. Leukocyte recruitment, required to limit bacterial spread, depends on phosphoinositide-3 kinase γ (PI3Kγ) signaling in vitro; however, the role of this enzyme in polymicrobial sepsis has remained unclear. OBJECTIVES: This study aimed to determine the specific role of the kinase activity of PI3Kγ in the pathogenesis of sepsis and multiorgan damage. METHODS: PI3Kγ wild-type, knockout, and kinase-dead mice were exposed to cecal ligation and perforation-induced sepsis and assessed for survival; pulmonary, hepatic, and cardiovascular damage; coagulation derangements; systemic inflammation; bacterial spread; and neutrophil recruitment. Additionally, wild-type mice were treated either before or after the onset of sepsis with a PI3Kγ inhibitor and assessed for survival, neutrophil recruitment, and bacterial spread. MEASUREMENTS AND MAIN RESULTS: Both genetic and pharmaceutical PI3Kγ kinase inhibition significantly improved survival, reduced multiorgan damage, and limited bacterial decompartmentalization, while modestly affecting SIRS. Protection resulted from both neutrophil-independent mechanisms, involving improved cardiovascular function, and neutrophil-dependent mechanisms, through reduced susceptibility to neutrophil migration failure during severe sepsis by maintaining neutrophil surface expression of the chemokine receptor, CXCR2. Furthermore, PI3Kγ pharmacological inhibition significantly decreased mortality and improved neutrophil migration and bacterial control, even when administered during established septic shock. CONCLUSIONS: This study establishes PI3Kγ as a key molecule in the pathogenesis of septic infection and the transition from SIRS to organ damage and identifies it as a novel possible therapeutic target.
Asunto(s)
Insuficiencia Multiorgánica/enzimología , Infiltración Neutrófila/fisiología , Fosfatidilinositol 3-Quinasas/fisiología , Sepsis/enzimología , Animales , Quimiotaxis de Leucocito , Fosfatidilinositol 3-Quinasa Clase Ib , Modelos Animales de Enfermedad , Inhibidores Enzimáticos/farmacología , Isoenzimas/antagonistas & inhibidores , Isoenzimas/fisiología , Estimación de Kaplan-Meier , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Insuficiencia Multiorgánica/prevención & control , Inhibidores de las Quinasa Fosfoinosítidos-3 , Sepsis/prevención & control , Transducción de Señal/efectos de los fármacos , Síndrome de Respuesta Inflamatoria Sistémica/enzimología , Síndrome de Respuesta Inflamatoria Sistémica/prevención & controlAsunto(s)
1-Fosfatidilinositol 4-Quinasa/antagonistas & inhibidores , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Leucemia Linfocítica Crónica de Células B/patología , Relación Dosis-Respuesta a Droga , Humanos , Isoenzimas/antagonistas & inhibidores , Células Tumorales CultivadasRESUMEN
Deregulation of the phosphoinositide-3-OH kinase (PI(3)K) pathway has been implicated in numerous pathologies including cancer, diabetes, thrombosis, rheumatoid arthritis and asthma. Recently, small-molecule and ATP-competitive PI(3)K inhibitors with a wide range of selectivities have entered clinical development. In order to understand the mechanisms underlying the isoform selectivity of these inhibitors, we developed a new expression strategy that enabled us to determine to our knowledge the first crystal structure of the catalytic subunit of the class IA PI(3)K p110 delta. Structures of this enzyme in complex with a broad panel of isoform- and pan-selective class I PI(3)K inhibitors reveal that selectivity toward p110 delta can be achieved by exploiting its conformational flexibility and the sequence diversity of active site residues that do not contact ATP. We have used these observations to rationalize and synthesize highly selective inhibitors for p110 delta with greatly improved potencies.
Asunto(s)
Dominio Catalítico , Fosfatidilinositol 3-Quinasas/química , Inhibidores de Proteínas Quinasas/química , Adenosina Trifosfato/química , Adenosina Trifosfato/metabolismo , Animales , Línea Celular , Simulación por Computador , Cristalografía por Rayos X , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Modelos Moleculares , Fosfatidilinositol 3-Quinasas/metabolismo , Dominios y Motivos de Interacción de Proteínas , Spodoptera , Relación Estructura-Actividad , Especificidad por SustratoRESUMEN
The gamma isoform of phosphoinositide 3-kinase (PI3Kgamma) has been viewed as restricted to leukocytes mediating the regulation of chemokine-induced migration and recruitment of neutrophils, monocytes, and macrophages. In line with the observation that PI3Kgamma-deficient mice display defects in adaptive immunity, inhibition of PI3Kgamma reduces synovial inflammation in the collagen-induced arthritis mouse model of inflammatory arthritis [rheumatoid arthritis (RA)], which has been attributed to reduced influx of inflammatory cells. Challenging the concept of leukocyte-restricted PI3Kgamma function, we report here a novel, nonredundant function of PI3Kgamma as an important regulator of fibroblast-induced cartilage destruction during chronic destructive arthritis. We show that in human tumor necrosis factor transgenic mice, the loss of PI3Kgamma leads to a milder inflammatory arthritis. Interestingly, PI3Kgamma deficiency does not alter the recruitment of inflammatory cells, but significantly reduces cartilage damage through reduced expression of matrix metalloproteinases in fibroblasts and chondrocytes. In vitro analyses demonstrate that the decreased invasiveness of fibroblasts is mediated by reduced phosphorylation of Akt and extracellular signal-regulated kinase. Using a PI3Kgamma specific inhibitor, these data are confirmed in human synovial fibroblasts from patients with RA who exhibit a disease-specific up-regulation of PI3Kgamma. Our data indicate that in addition to mediating the recruitment of inflammatory cells, PI3Kgamma is an important regulator of fibroblast-mediated joint destruction in RA and suggest that specific inhibitors of PI3Kgamma will interfere with the activation of RA synovial fibroblasts and reduce cartilage destruction in RA.
Asunto(s)
Artritis/metabolismo , Cartílago/patología , Fosfatidilinositol 3-Quinasas/metabolismo , Animales , Artritis/genética , Artritis/patología , Artritis Reumatoide/metabolismo , Condrocitos/metabolismo , Fosfatidilinositol 3-Quinasa Clase Ib , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Femenino , Fibroblastos/citología , Fibroblastos/metabolismo , Expresión Génica/fisiología , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , Masculino , Metaloproteasas/metabolismo , Ratones , Ratones Noqueados , Fosfatidilinositol 3-Quinasas/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Membrana Sinovial/citología , Factores de Tiempo , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
OBJECTIVE: Phosphatidylinositol 3-OH kinase (PI3K) has a long-recognized role in beta-cell mass regulation and gene transcription and is implicated in the modulation of insulin secretion. The role of nontyrosine kinase receptor-activated PI3K isoforms is largely unexplored. We therefore investigated the role of the G-protein-coupled PI3Kgamma and its catalytic subunit p110gamma in the regulation of insulin granule recruitment and exocytosis. RESEARCH DESIGN AND METHODS: The expression of p110gamma was knocked down by small-interfering RNA, and p110gamma activity was selectively inhibited with AS605240 (40 nmol/l). Exocytosis and granule recruitment was monitored by islet perifusion, whole-cell capacitance, total internal reflection fluorescence microscopy, and electron microscopy in INS-1 and human beta-cells. Cortical F-actin was examined in INS-1 cells and human islets and in mouse beta-cells lacking the phosphatase and tensin homolog (PTEN). RESULTS: Knockdown or inhibition of p110gamma markedly blunted depolarization-induced insulin secretion and exocytosis and ablated the exocytotic response to direct Ca(2+) infusion. This resulted from reduced granule localization to the plasma membrane and was associated with increased cortical F-actin. Inhibition of p110gamma had no effect on F-actin in beta-cells lacking PTEN. Finally, the effect of p110gamma inhibition on granule localization and exocytosis could be rapidly reversed by agents that promote actin depolymerization. CONCLUSIONS: The G-protein-coupled PI3Kgamma is an important determinant of secretory granule trafficking to the plasma membrane, at least in part through the negative regulation of cortical F-actin. Thus, p110gamma activity plays an important role in maintaining a membrane-docked, readily releasable pool of secretory granules in insulinoma and human beta-cells.
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Células Secretoras de Insulina/citología , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Vesículas Secretoras/metabolismo , Actinas/metabolismo , Animales , Canales de Calcio/fisiología , Línea Celular Tumoral , Fosfatidilinositol 3-Quinasa Clase Ib , Exocitosis/fisiología , Humanos , Secreción de Insulina , Insulinoma , Isoenzimas/genética , Isoenzimas/metabolismo , Potenciales de la Membrana/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Neoplasias Pancreáticas , Técnicas de Placa-Clamp , Fosfatidilinositol 3-Quinasas/genética , Inhibidores de las Quinasa Fosfoinosítidos-3 , Quinoxalinas/farmacología , ARN Interferente Pequeño , Tiazolidinedionas/farmacologíaRESUMEN
Bone morphogenetic proteins (BMPs) are potent regulators of several cellular events. We report that exposure of C2C12 cells to BMP2 leads to an increase in cell migration and a rapid rearrangement of the actin filaments into cortical protrusions. These effects required independent and parallel activation of the Cdc42 small GTPase and the alpha-isoform of the phosphoinositide 3-kinase (PI3Kalpha), because ectopic expression of a dominant-negative form of Cdc42 or distinct pharmacological PI3K inhibitors abrogated these responses. Furthermore, we demonstrate that BMP2 activates different group I and group II PAK isoforms as well as LIMK1 with similar kinetics to Cdc42 or PI3K activation. BMP2 activation of PAK and LIMK1, measured by either kinase activity or with antibodies raised against phosphorylated residues at their activation loops, were abolished by blocking PI3K-signaling pathways. Together, these findings suggest that Cdc42 and PI3K signals emanating from BMP receptors are involved in specific regulation of actin assembly and cell migration.
Asunto(s)
Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Proteína Morfogenética Ósea 2/metabolismo , Movimiento Celular/fisiología , Fosfatidilinositol 3-Quinasas/metabolismo , Proteína de Unión al GTP cdc42/metabolismo , Células 3T3 , Animales , Células Cultivadas , Quinasas Lim/metabolismo , Ratones , Transducción de SeñalRESUMEN
The phosphoinositide 3-kinase (PI3K) pathway crucially controls metabolism and cell growth. Although different PI3K catalytic subunits are known to play distinct roles, the specific in vivo function of p110beta (the product of the PIK3CB gene) is not clear. Here, we show that mouse mutants expressing a catalytically inactive PIK3CB(K805R) mutant survived to adulthood but showed growth retardation and developed mild insulin resistance with age. Pharmacological and genetic analyses of p110beta function revealed that p110beta catalytic activity is required for PI3K signaling downstream of heterotrimeric guanine nucleotide-binding protein (G protein)-coupled receptors as well as to sustain long-term insulin signaling. In addition, PIK3CB(K805R) mice were protected in a model of ERBB2-driven tumor development. These findings indicate an unexpected role for p110beta catalytic activity in diabetes and cancer, opening potential avenues for therapeutic intervention.