RESUMEN
BACKGROUND: Immune checkpoint inhibitors are approved for the treatment of various tumors, but the response rate is not satisfactory in certain malignancies. Inhibitor of apoptosis proteins (IAP) ubiquitin-E3 ligase activity is involved in the regulation of immune responses. APG-1387 is a novel second mitochondria-derived activator of caspase (Smac) mimetic IAP inhibitor. The aim of this study was to explore the synergistic effect of APG-1387 when combined with anti-PD-1 antibody in a preclinical setting. METHODS: We utilized syngeneic mouse models of ovarian cancer (ID8), colon cancer (MC38), malignant melanoma (B16), and liver cancer (Hepa1-6) to assess the combination effect of APG-1387 and anti-PD-1 antibody, including immune-related factors, tumor growth, and survival. MSD V-PLEX validated assays were used to measure in vitro and in vivo cytokine release. RESULTS: In ID8 ovarian cancer and MC38 colon cancer models, APG-1387 and anti-PD1 antibody had synergistic antitumor effects. In the MC38 model, the combination of APG-1387 and anti-PD-1 antibody significantly inhibited tumor growth (P < 0.0001) and increased the survival rate of tumor-bearing animals (P < 0.001). Moreover, we found that APG-1387 upregulated tumor-infiltrating CD3 + NK1.1 + cells by nearly 2-fold, by promoting tumor cell secretion of IL-12. Blocking IL-12 secretion abrogated the synergistic effects of APG-1387 and anti-PD-1 antibody in both MC38 and ID8 models. CONCLUSIONS: APG-1387 has the potential to turn "cold tumors" into hot ones by recruiting more CD3 + NK1.1 + cells into certain tumors. Based on these and other data, the safety and therapeutic effect of this combination will be investigated in a phase 1/2 trial in patients with advanced solid tumors or hematologic malignancies (NCT03386526).
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Hyperpolarized carbon-13 MRI has the advantage of allowing the study of glycolytic flow in vivo or in vitro dynamically in real-time. The apparent exchange rate constant of a metabolite dynamic signal reflects the metabolite changes of a disease. Downstream metabolites can have a low signal-to-noise ratio (SNR), causing apparent exchange rate constant inconsistencies. Thus, we developed a method that estimates a more accurate metabolite signal. This method utilizes a kinetic model and background noise to estimate metabolite signals. Simulations and in vitro studies with photon-irradiated and control groups were used to evaluate the procedure. Simulated and in vitro exchange rate constants estimated using our method were compared with the raw signal values. In vitro data were also compared to the Area-Under-Curve (AUC) of the cell medium in 13C Nuclear Magnetic Resonance (NMR). In the simulations and in vitro experiments, our technique minimized metabolite signal fluctuations and maintained reliable apparent exchange rate constants. In addition, the apparent exchange rate constants of the metabolites showed differences between the irradiation and control groups after using our method. Comparing the in vitro results obtained using our method and NMR, both solutions showed consistency when uncertainty was considered, demonstrating that our method can accurately measure metabolite signals and show how glycolytic flow changes. The method enhanced the signals of the metabolites and clarified the metabolic phenotyping of tumor cells, which could benefit personalized health care and patient stratification in the future.
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Imagen por Resonancia Magnética , Ácido Pirúvico , Humanos , Cinética , Espectroscopía de Resonancia Magnética/métodos , Relación Señal-RuidoRESUMEN
In mammals, ether lipids exert a wide spectrum of signaling and structural functions, such as stimulation of immune responses, anti-tumor activities, and enhancement of sperm functions. Abnormal accumulation of monoalkyl-diacylglycerol (MADAG) was found in Wolman's disease, a human genetic disorder defined by a deficiency in lysosomal acid lipase. In the current study, we found that among the nine recombinant human lipid acyltransferases examined, acyl-CoA:diacylglycerol acyltransferase (DGAT)1, DGAT2, acyl-CoA:monoacylglycerol acyltransferase (MGAT)2, MGAT3, acyl-CoA:wax-alcohol acyltransferase 2/MFAT, and DGAT candidate 3 were able to use 1-monoalkylglycerol (1-MAkG) as an acyl acceptor for the synthesis of monoalkyl-monoacylglycerol (MAMAG). These enzymes demonstrated different enzymatic turnover rates and relative efficiencies for the first and second acylation steps leading to the synthesis of MAMAG and MADAG, respectively. They also exhibited different degrees of substrate preference when presented with 1-monooleoylglycerol versus 1-MAkG. In CHO-K1 cells, treatment with DGAT1 selective inhibitor, XP-620, completely blocked the synthesis of MADAG, indicating that DGAT1 is the predominant enzyme responsible for the intracellular synthesis of MADAG in this model system. The levels of MADAG in the adrenal gland of DGAT1 KO mice were reduced as compared with those of the WT mice, suggesting that DGAT1 is a major enzyme for the synthesis of MADAG in this tissue. Our findings indicate that several of these lipid acyltransferases may be able to synthesize neutral ether lipids in mammals.
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Aciltransferasas/metabolismo , Diglicéridos/biosíntesis , Diglicéridos/química , Éteres/química , Acilación , Animales , Células CHO , Chlorocebus aethiops , Cricetulus , Diglicéridos/metabolismo , HumanosRESUMEN
Homeostatic temperature regulation is fundamental to mammalian physiology and is controlled by acute and chronic responses of local, endocrine and nervous regulators. Here, we report that loss of the heparan sulfate proteoglycan, syndecan-1, causes a profoundly depleted intradermal fat layer, which provides crucial thermogenic insulation for mammals. Mice without syndecan-1 enter torpor upon fasting and show multiple indicators of cold stress, including activation of the stress checkpoint p38α in brown adipose tissue, liver and lung. The metabolic phenotype in mutant mice, including reduced liver glycogen, is rescued by housing at thermoneutrality, suggesting that reduced insulation in cool temperatures underlies the observed phenotypes. We find that syndecan-1, which functions as a facultative lipoprotein uptake receptor, is required for adipocyte differentiation in vitro. Intradermal fat shows highly dynamic differentiation, continuously expanding and involuting in response to hair cycle and ambient temperature. This physiology probably confers a unique role for Sdc1 in this adipocyte sub-type. The PPARγ agonist rosiglitazone rescues Sdc1-/- intradermal adipose tissue, placing PPARγ downstream of Sdc1 in triggering adipocyte differentiation. Our study indicates that disruption of intradermal adipose tissue development results in cold stress and complex metabolic pathology.
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Diferenciación Celular/genética , Proteína Quinasa 14 Activada por Mitógenos/genética , PPAR gamma/genética , Estrés Fisiológico/genética , Sindecano-1/genética , Adipocitos/efectos de los fármacos , Adipocitos/metabolismo , Tejido Adiposo Pardo/efectos de los fármacos , Tejido Adiposo Pardo/metabolismo , Animales , Frío , Ratones , Proteína Quinasa 14 Activada por Mitógenos/metabolismo , PPAR gamma/agonistas , PPAR gamma/metabolismo , Rosiglitazona , Sindecano-1/metabolismo , Tiazolidinedionas/administración & dosificaciónRESUMEN
Reducing triacylglycerol (TAG) in the liver continues to pose a challenge in states of nonalcoholic hepatic steatosis. MonoacylglycerolO-acyltransferase (MOGAT) enzymes convert monoacylglycerol (MAG) to diacylglycerol, a precursor for TAG synthesis, and are involved in a major pathway of TAG synthesis in selected tissues, such as small intestine. MOGAT1 possesses MGAT activity in in vitro assays, but its physiological function in TAG metabolism is unknown. Recent studies suggest a role for MOGAT1 in hepatic steatosis in lipodystrophic [1-acylglycerol-3-phosphateO-acyltransferase (Agpat)2(-/-)] and obese (ob/ob) mice. To test this, we deletedMogat1in theAgpat2(-/-)andob/obgenetic background to generateMogat1(-/-);Agpat2(-/-)andMogat1(-/-);ob/obdouble knockout (DKO) mice. Here we report that, despite the absence ofMogat1in either DKO mouse model, we did not find any decrease in liver TAG by 16 weeks of age. Additionally, there were no measureable changes in plasma glucose (diabetes) and insulin resistance. Our data indicate a minimal role, if any, of MOGAT1 in liver TAG synthesis, and that TAG synthesis in steatosis associated with lipodystrophy and obesity is independent of MOGAT1. Our findings suggest that MOGAT1 likely has an alternative function in vivo.
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Aciltransferasas/deficiencia , Aciltransferasas/genética , Eliminación de Gen , Lipodistrofia/genética , Enfermedad del Hígado Graso no Alcohólico/enzimología , Enfermedad del Hígado Graso no Alcohólico/genética , Animales , Femenino , Insulina/sangre , Resistencia a la Insulina , Lipodistrofia/complicaciones , Lipodistrofia/metabolismo , Hígado/metabolismo , Masculino , Ratones , Ratones Obesos , Enfermedad del Hígado Graso no Alcohólico/sangre , Enfermedad del Hígado Graso no Alcohólico/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
The intestine plays a prominent role in the biosynthesis of triacylglycerol (triglyceride; TAG). Digested dietary TAG is repackaged in the intestine to form the hydrophobic core of chylomicrons, which deliver metabolic fuels, essential fatty acids, and other lipid-soluble nutrients to the peripheral tissues. By controlling the flux of dietary fat into the circulation, intestinal TAG synthesis can greatly impact systemic metabolism. Genes encoding many of the enzymes involved in TAG synthesis have been identified. Among TAG synthesis enzymes, acyl-CoA:monoacylglycerol acyltransferase 2 and acyl-CoA:diacylglycerol acyltransferase (DGAT)1 are highly expressed in the intestine. Their physiological functions have been examined in the context of whole organisms using genetically engineered mice and, in the case of DGAT1, specific inhibitors. An emerging theme from recent findings is that limiting the rate of TAG synthesis in the intestine can modulate gut hormone secretion, lipid metabolism, and systemic energy balance. The underlying mechanisms and their implications for humans are yet to be explored. Pharmacological inhibition of TAG hydrolysis in the intestinal lumen has been employed to combat obesity and associated disorders with modest efficacy and unwanted side effects. The therapeutic potential of inhibiting specific enzymes involved in intestinal TAG synthesis warrants further investigation.
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Metabolismo Energético/fisiología , Absorción Intestinal/fisiología , Mucosa Intestinal/metabolismo , Metabolismo de los Lípidos/fisiología , Triglicéridos/biosíntesis , Animales , Quilomicrones/genética , Quilomicrones/metabolismo , Diacilglicerol O-Acetiltransferasa/biosíntesis , Diacilglicerol O-Acetiltransferasa/genética , Regulación Enzimológica de la Expresión Génica/fisiología , Humanos , Ratones , Triglicéridos/genéticaRESUMEN
The absorption of dietary fat involves the re-esterification of digested triacylglycerol in the enterocytes, a process catalyzed by acyl-CoA:monoacylglycerol acyltransferase (MGAT) 2. Mice without a functional gene encoding MGAT2 (Mogat2(-/-)) are protected from diet-induced obesity. Surprisingly, these mice absorb normal amounts of dietary fat but increase their energy expenditure. MGAT2 is expressed in tissues besides intestine, including adipose tissue in both mice and humans. To test the hypothesis that intestinal MGAT2 regulates systemic energy balance, we generated and characterized mice deficient in MGAT2 specifically in the small intestine (Mogat2(IKO)). We found that, like Mogat2(-/-) mice, Mogat2(IKO) mice also showed a delay in fat absorption, a decrease in food intake, and a propensity to use fatty acids as fuel when first exposed to a high fat diet. Mogat2(IKO) mice increased energy expenditure although to a lesser degree than Mogat2(-/-) mice and were protected against diet-induced weight gain and associated comorbidities, including hepatic steatosis, hypercholesterolemia, and glucose intolerance. These findings illustrate that intestinal lipid metabolism plays a crucial role in the regulation of systemic energy balance and may be a feasible intervention target. In addition, they suggest that MGAT activity in extraintestinal tissues may also modulate energy metabolism.
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Grasas de la Dieta/efectos adversos , Intolerancia a la Glucosa/enzimología , Intestinos/enzimología , N-Acetilglucosaminiltransferasas/metabolismo , Obesidad/enzimología , Animales , Grasas de la Dieta/farmacología , Ingestión de Alimentos/genética , Metabolismo Energético/efectos de los fármacos , Metabolismo Energético/genética , Hígado Graso/enzimología , Hígado Graso/genética , Hígado Graso/patología , Hígado Graso/prevención & control , Eliminación de Gen , Intolerancia a la Glucosa/genética , Intolerancia a la Glucosa/patología , Intolerancia a la Glucosa/prevención & control , Humanos , Hipercolesterolemia/enzimología , Hipercolesterolemia/genética , Hipercolesterolemia/patología , Hipercolesterolemia/prevención & control , Absorción Intestinal/efectos de los fármacos , Absorción Intestinal/genética , Intestinos/patología , Ratones , Ratones Noqueados , N-Acetilglucosaminiltransferasas/genética , Obesidad/inducido químicamente , Obesidad/genética , Obesidad/patología , Obesidad/prevención & controlRESUMEN
PURPOSE: B-cell lymphoma-extra-large (BCL-xL) regulates apoptosis and is an attractive anticancer therapeutic target. However, BCL-xL inhibition also kills mature platelets, hampering clinical development. Using an innovative prodrug strategy, we have developed pelcitoclax (APG-1252), a potent, dual BCL-2 and BCL-xL inhibitor. Aims of this study were to characterize the antitumor activity and safety of pelcitoclax and explore its underlying mechanisms of action (MOA). PATIENTS AND METHODS: Cell line-derived xenograft and patient-derived xenograft (PDX) models were tested to evaluate antitumor activity and elucidate MOA. Subjects (N = 50) with metastatic small-cell lung cancer and other solid tumors received intravenous pelcitoclax once or twice weekly. Primary outcome measures were safety and tolerability; preliminary efficacy (responses every 2 cycles per RECIST version 1.1) represented a secondary endpoint. RESULTS: Pelcitoclax exhibited strong BAX/BAKâdependent and caspase-mediated antiproliferative and apoptogenic activity in various cancer cell lines. Consistent with cell-based apoptogenic activity, pelcitoclax disrupted BCL-xL:BIM and BCL-xL:PUMA complexes in lung and gastric cancer PDX models. Levels of BCL-xL complexes correlated with tumor growth inhibition by pelcitoclax. Combined with taxanes, pelcitoclax enhanced antitumor activity by downregulating antiapoptotic protein myeloid cell leukemia-1 (MCL-1). Importantly, pelcitoclax was well tolerated and demonstrated preliminary therapeutic efficacy, with overall response and disease control rates of 6.5% and 30.4%, respectively. Most common treatment-related adverse events included transaminase elevations and reduced platelets that were less frequent with a once-weekly schedule. CONCLUSIONS: Our data demonstrate that pelcitoclax has antitumor activity and is well tolerated, supporting its further clinical development for human solid tumors, particularly combined with agents that downregulate MCL-1.
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Compuestos de Anilina , Antineoplásicos , Neoplasias Pulmonares , Linfoma de Células B , Piperidinas , Humanos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteína bcl-X/metabolismo , Línea Celular Tumoral , Ensayos Antitumor por Modelo de Xenoinjerto , Proteínas Proto-Oncogénicas c-bcl-2 , Antineoplásicos/efectos adversos , Apoptosis , Neoplasias Pulmonares/tratamiento farmacológico , Linfoma de Células B/tratamiento farmacológicoRESUMEN
Acyl CoA:monoacylglycerol acyltransferase (MGAT) catalyzes the resynthesis of triacylglycerol, a crucial step in the absorption of dietary fat. Mice lacking the gene Mogat2, which codes for an MGAT highly expressed in the small intestine, are resistant to obesity and other metabolic disorders induced by high-fat feeding. Interestingly, these Mogat2â»/â» mice absorb normal amounts of dietary fat but exhibit a reduced rate of fat absorption, increased energy expenditure, decreased respiratory exchange ratio, and impaired metabolic efficiency. MGAT2 is expressed in tissues besides intestine. To test the hypothesis that intestinal MGAT2 enhances metabolic efficiency and promotes the storage of metabolic fuels, we introduced the human MOGAT2 gene driven by the intestine-specific villin promoter into Mogat2â»/â» mice. We found that the expression of MOGAT2 in the intestine increased intestinal MGAT activity, restored fat absorption rate, partially corrected energy expenditure, and promoted weight gain upon high-fat feeding. However, the changes in respiratory exchange ratio were not reverted, and the recoveries in metabolic efficiency and weight gain were incomplete. These data indicate that MGAT2 in the intestine plays an indispensable role in enhancing metabolic efficiency but also raise the possibility that MGAT2 in other tissues may contribute to the regulation of energy metabolism.
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Aciltransferasas/biosíntesis , Metabolismo Energético , Regulación Enzimológica de la Expresión Génica , Absorción Intestinal/fisiología , Intestinos/enzimología , Aciltransferasas/genética , Animales , Grasas de la Dieta/metabolismo , Grasas de la Dieta/farmacología , Humanos , Ratones , Ratones Noqueados , Especificidad de Órganos/fisiologíaRESUMEN
OBJECTIVE: Pedestrian accidents contribute significantly to the high number of annual traffic casualties. It is therefore crucial for pedestrians to use safety measures, such as a crosswalk, and to activate pedestrian signals. However, people often fail to activate the signal or are unable to do so - those who are visually impaired or have occupied hands may be unable to activate the system. Failure to activate the signal can result in an accident. This paper proposes an improvement to crosswalk safety by designing a system that can detect pedestrians and trigger the pedestrian signal automatically when necessary. METHODS: In this study, a dataset of images was collected in order to train a Convolutional Neural Network (CNN) to distinguish between pedestrians (including bicycle riders) when crossing the street. The resulting system can capture and evaluate images in real-time, and the result can be used to automatically activate a system such as a pedestrian signal. A threshold system is also implemented that triggers the crosswalk only when the positive predictions pass the threshold. This system was tested by deploying it at three real-world environments and comparing the results with a recorded video of the camera's view. RESULTS: The CNN prediction model is able to correctly predict pedestrian and cyclist intentions with an average accuracy of 84.96% and had an absence trigger rate of 0.037%. The prediction accuracy varies based on the location and whether a cyclist or pedestrian is in front of the camera. Pedestrians crossing the street were correctly predicted more accurately than cyclists crossing the street by up to 11.61%, while passing (i.e., non-crossing) cyclists were correctly ignored more than passing pedestrians, by up to 18.75%. CONCLUSION: Based on the testing of the system in real-world environments, the authors conclude that it is feasible as a back-up system that can complement existing pedestrian signal buttons, and thereby improve the overall safety of crossing the street. Further improvements to the accuracy can be achieved with a more comprehensive dataset for a specific location where the system is deployed. Implementing different computer vision techniques optimized for tracking objects should also increase the accuracy.
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Accidentes de Tránsito , Peatones , Humanos , Accidentes de Tránsito/prevención & control , Seguridad , Intención , Redes Neurales de la Computación , CaminataRESUMEN
PURPOSE: This global phase I trial investigated the safety, efficacy, pharmacokinetics, and pharmacodynamics of lisaftoclax (APG-2575), a novel, orally active, potent selective B-cell lymphoma 2 (BCL-2) inhibitor, in patients with relapsed or refractory chronic lymphocytic leukemia or small lymphocytic lymphoma (R/R CLL/SLL) and other hematologic malignancies (HMs). PATIENTS AND METHODS: Maximum tolerated dose (MTD) and recommended phase II dose were evaluated. Outcome measures were safety and tolerability (primary) and pharmacokinetic variables and antitumor effects (secondary). Pharmacodynamics in patient tumor cells were explored. RESULTS: Among 52 patients receiving lisaftoclax, MTD was not reached. Treatment-emergent adverse events (TEAEs) included diarrhea (48.1%), fatigue (34.6%), nausea (30.8%), anemia and thrombocytopenia (28.8% each), neutropenia (26.9%), constipation (25.0%), vomiting (23.1%), headache (21.2%), peripheral edema and hypokalemia (17.3% each), and arthralgia (15.4%). Grade ≥ 3 hematologic TEAEs included neutropenia (21.2%), thrombocytopenia (13.5%), and anemia (9.6%), none resulting in treatment discontinuation. Clinical pharmacokinetic and pharmacodynamic results demonstrated that lisaftoclax had a limited plasma residence and systemic exposure and elicited rapid clearance of malignant cells. With a median treatment of 15 (range, 6-43) cycles, 14 of 22 efficacy-evaluable patients with R/R CLL/SLL experienced partial responses, for an objective response rate of 63.6% and median time to response of 2 (range, 2-8) cycles. CONCLUSIONS: Lisaftoclax was well tolerated, with no evidence of tumor lysis syndrome. Dose-limiting toxicity was not reached at the highest dose level. Lisaftoclax has a unique pharmacokinetic profile compatible with a potentially more convenient daily (vs. weekly) dose ramp-up schedule and induced rapid clinical responses in patients with CLL/SLL, warranting continued clinical investigation.
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Anemia , Antineoplásicos , Neoplasias Hematológicas , Leucemia Linfocítica Crónica de Células B , Linfoma de Células B , Neutropenia , Trombocitopenia , Humanos , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucemia Linfocítica Crónica de Células B/patología , Antineoplásicos/efectos adversos , Linfoma de Células B/patología , Neoplasias Hematológicas/tratamiento farmacológico , Neutropenia/inducido químicamente , Anemia/inducido químicamente , Anemia/tratamiento farmacológico , Trombocitopenia/inducido químicamente , Proteínas Proto-Oncogénicas c-bcl-2RESUMEN
Transmembrane protein 135 (TMEM135) is thought to participate in the cellular response to increased intracellular lipids yet no defined molecular function for TMEM135 in lipid metabolism has been identified. In this study, we performed a lipid analysis of tissues from Tmem135 mutant mice and found striking reductions of docosahexaenoic acid (DHA) across all Tmem135 mutant tissues, indicating a role of TMEM135 in the production of DHA. Since all enzymes required for DHA synthesis remain intact in Tmem135 mutant mice, we hypothesized that TMEM135 is involved in the export of DHA from peroxisomes. The Tmem135 mutation likely leads to the retention of DHA in peroxisomes, causing DHA to be degraded within peroxisomes by their beta-oxidation machinery. This may lead to generation or alteration of ligands required for the activation of peroxisome proliferator-activated receptor a (PPARa) signaling, which in turn could result in increased peroxisomal number and beta-oxidation enzymes observed in Tmem135 mutant mice. We confirmed this effect of PPARa signaling by detecting decreased peroxisomes and their proteins upon genetic ablation of Ppara in Tmem135 mutant mice. Using Tmem135 mutant mice, we also validated the protective effect of increased peroxisomes and peroxisomal beta-oxidation on the metabolic disease phenotypes of leptin mutant mice which has been observed in previous studies. Thus, we conclude that TMEM135 has a role in lipid homeostasis through its function in peroxisomes.
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Ácidos Docosahexaenoicos , Metabolismo de los Lípidos , Proteínas de la Membrana , Peroxisomas , Animales , Ratones , Ácidos Docosahexaenoicos/metabolismo , Homeostasis , Oxidación-Reducción , Receptores Activados del Proliferador del Peroxisoma/metabolismo , Peroxisomas/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas Mitocondriales/metabolismoRESUMEN
Phenylketonuria (PKU) is caused by a mutation in the phenylalanine (phe) hydroxylase gene and requires a low-phe diet plus amino acid (AA) formula to prevent cognitive impairment. Glycomacropeptide (GMP) contains minimal phe and provides a palatable alternative to AA formula. Our objective was to compare growth, body composition, and energy balance in Pah(enu2) (PKU) and wild-type mice fed low-phe GMP, low-phe AA, or high-phe casein diets from 3-23 wk of age. The 2 × 2 × 3 design included main effects of genotype, sex, and diet. Fat and lean mass were assessed by dual-energy X-ray absorptiometry, and acute energy balance was assessed by indirect calorimetry. PKU mice showed growth and lean mass similar to wild-type littermates fed the GMP or AA diets; however, they exhibited a 3-15% increase in energy expenditure, as reflected in oxygen consumption, and a 3-30% increase in food intake. The GMP diet significantly reduced energy expenditure, food intake, and plasma phe concentration in PKU mice compared with the casein diet. The high-phe casein diet or the low-phe AA diet induced metabolic stress in PKU mice, as reflected in increased energy expenditure and intake of food and water, increased renal and spleen mass, and elevated plasma cytokine concentrations consistent with systemic inflammation. The low-phe GMP diet significantly attenuated these adverse effects. Moreover, total fat mass, %body fat, and the respiratory exchange ratio (CO(2) produced/O(2) consumed) were significantly lower in PKU mice fed GMP compared with AA diets. In summary, GMP provides a physiological source of low-phe dietary protein that promotes growth and attenuates the metabolic stress induced by a high-phe casein or low-phe AA diet in PKU mice.
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Caseínas/farmacología , Queso/análisis , Fragmentos de Péptidos/farmacología , Fenilcetonurias/tratamiento farmacológico , Fenilcetonurias/metabolismo , Absorciometría de Fotón , Aminoácidos/sangre , Aminoácidos/farmacología , Animales , Composición Corporal/efectos de los fármacos , Peso Corporal/efectos de los fármacos , Citocinas/sangre , Dieta , Dieta con Restricción de Proteínas , Metabolismo Energético/efectos de los fármacos , Crecimiento/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Tamaño de los Órganos/efectos de los fármacos , Consumo de Oxígeno/efectos de los fármacos , Fenotipo , Fenilcetonurias/genética , Bazo/efectos de los fármacos , Bazo/crecimiento & desarrollo , Bazo/patología , Esplenomegalia/inducido químicamente , Esplenomegalia/patologíaRESUMEN
Biotherapeutics are highly efficacious, but the pain and inconvenience of chronic injections lead to poor patient compliance and compromise effective disease management. Despite innumerable attempts, oral delivery of biotherapeutics remains unsuccessful due to their degradation in the gastrointestinal (GI) environment and poor intestinal absorption. We have developed an orally ingestible robotic pill (RP) for drug delivery, which protects the biotherapeutic drug payload from digestion in the GI tract and auto-injects it into the wall of the small intestine as a safe, pain-free injection since the intestines are insensate to sharp stimuli. The payload is delivered upon inflation of a balloon folded within the RP, which deflates immediately after drug delivery. Here we present results from two clinical studies demonstrating the safety, tolerability and performance of the RP in healthy humans. In the first study, three versions of the RP (A, B and C) were evaluated, which were identical in all respects except for the diameter of the balloon. The RP successfully delivered a biotherapeutic (octreotide) in 3 out of 12 subjects in group A, 10 out of 20 subjects in group B and 16 out of 20 subjects in group C, with a mean bioavailability of 65 ± 9% (based on successful drug deliveries in groups A and B). Thus, reliability of drug delivery with the RP ranged from 25 to 80%, with success rate directly related to balloon size. In a separate study, the deployment of the RP was unaffected by fed or fasting conditions suggesting that the RP may be taken with or without food. These promising clinical data suggest that biotherapeutics currently administered parenterally may be safely and reliably delivered via this versatile, orally ingestible drug delivery platform.
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Procedimientos Quirúrgicos Robotizados , Administración Oral , Disponibilidad Biológica , Sistemas de Liberación de Medicamentos , Voluntarios Sanos , Humanos , Reproducibilidad de los ResultadosRESUMEN
The hydrolysis of fat stored in adipose tissues is crucial for providing energy during fasting and exercise, and dysregulation of fat breakdown may contribute to metabolic disease. In this issue of Cell Metabolism, report that CGI-58/ABHD5, a lipid-droplet-associated protein that is mutated in a rare disease characterized by excess lipid storage, activates adipose triglyceride lipase and thus may regulate fat mobilization.
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Tejido Adiposo/enzimología , Esterasas/metabolismo , Lipasa/metabolismo , Errores Innatos del Metabolismo Lipídico/enzimología , Fosfolipasas A/metabolismo , Triglicéridos/metabolismo , 1-Acilglicerol-3-Fosfato O-Aciltransferasa , Animales , Activación Enzimática , Esterasas/genética , Humanos , Lipasa/genética , Errores Innatos del Metabolismo Lipídico/genética , Lipólisis , Mutación , SíndromeRESUMEN
Acyl CoA:monoacylglycerol acyltransferase 2 (MGAT2) is thought to be crucial for dietary fat absorption. Indeed, mice lacking the enzyme (Mogat2(-/-)) are resistant to obesity and other metabolic disorders induced by high-fat feeding. However, these mice absorb normal quantities of fat. To explore whether a high level of dietary fat is an essential part of the underlying mechanism(s), we examined metabolic responses of Mogat2(-/-) mice to diets containing varying levels of fat. Mogat2(-/-) mice exhibited 10-15% increases in energy expenditure compared with wild-type littermates; although high levels of dietary fat exacerbated the effect, this phenotype was expressed even on a fat-free diet. When deprived of food, Mogat2(-/-) mice expended energy and lost weight like wild-type controls. To determine whether MGAT2 deficiency protects against obesity in the absence of high-fat feeding, we crossed Mogat2(-/-) mice with genetically obese Agouti mice. MGAT2 deficiency increased energy expenditure and prevented these mice from gaining excess weight. Our results suggest that MGAT2 modulates energy expenditure through multiple mechanisms, including one independent of dietary fat; these findings also raise the prospect of inhibiting MGAT2 as a strategy for combating obesity and related metabolic disorders resulting from excessive calorie intake.
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Aciltransferasas/deficiencia , Grasas de la Dieta/metabolismo , Metabolismo Energético , Ratones Obesos/metabolismo , Aumento de Peso , Aciltransferasas/genética , Animales , Ingestión de Alimentos/fisiología , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Consumo de OxígenoRESUMEN
Calorie restriction (CR) promotes healthy ageing in diverse species. Recently, it has been shown that fasting for a portion of each day has metabolic benefits and promotes lifespan. These findings complicate the interpretation of rodent CR studies, in which animals typically eat only once per day and rapidly consume their food, which collaterally imposes fasting. Here we show that a prolonged fast is necessary for key metabolic, molecular and geroprotective effects of a CR diet. Using a series of feeding regimens, we dissect the effects of calories and fasting, and proceed to demonstrate that fasting alone recapitulates many of the physiological and molecular effects of CR. Our results shed new light on how both when and how much we eat regulate metabolic health and longevity, and demonstrate that daily prolonged fasting, and not solely reduced caloric intake, is likely responsible for the metabolic and geroprotective benefits of a CR diet.
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Envejecimiento/metabolismo , Restricción Calórica , Animales , Longevidad/fisiología , RatonesRESUMEN
The incidence of metabolic syndrome continues to rise globally. In mice, intravenous administration of interleukin-22 (IL-22) ameliorates various disease phenotypes associated with diet-induced metabolic syndrome. In patients, oral treatment is favored over intravenous treatment, but methodologies to deliver IL-22 via the oral route are nonexistent. The goal of this study was to assess to what extent engineered Lactobacillus reuteri secreting IL-22 could ameliorate nonalcoholic fatty liver disease. We used a mouse model of diet-induced obesity and assessed various markers of metabolic syndrome following treatment with L. reuteri and a recombinant derivative. Mice that received an 8-week treatment of wild-type probiotic gained less weight and had a smaller fat pad than the control group, but these phenotypes were not further enhanced by recombinant L. reuteri However, L. reuteri secreting IL-22 significantly reduced liver weight and triglycerides at levels that exceeded those of the probiotic wild-type treatment group. Our findings are interesting in light of the observed phenotypes associated with reduced nonalcoholic liver disease, in humans the most prevalent chronic liver disease, following treatment of a next-generation probiotic that is administered orally. Once biological and environmental containment strategies are in place, therapeutic applications of recombinant Lactobacillus reuteri are on the horizon.IMPORTANCE In humans, nonalcoholic fatty liver disease (NAFLD) is the most prevalent liver disease due to the increased prevalence of obesity. While treatment of NAFLD is often geared toward lifestyle changes, such as diet and exercise, the use of dietary supplements such as probiotics is underinvestigated. Here, we report that probiotic Lactobacillus reuteri reduces fatty liver in a mouse model of diet-induced obesity. This phenotype was further enhanced upon delivery of recombinant interleukin-22 by engineered Lactobacillus reuteri These observations pave the road to a better understanding of probiotic mechanisms driving the reduction of diet-induced steatosis and to development of next-generation probiotics for use in the clinic. Ultimately, these studies may lead to rational selection of (engineered) probiotics to ameliorate fatty liver disease.
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Hígado Graso/prevención & control , Interleucinas/administración & dosificación , Limosilactobacillus reuteri/genética , Obesidad/terapia , Probióticos/uso terapéutico , Animales , Biomarcadores , Dieta , Modelos Animales de Enfermedad , Interleucinas/genética , Masculino , Síndrome Metabólico/terapia , Ratones , Ratones Endogámicos C57BL , Obesidad/etiología , Proteínas Recombinantes/administración & dosificación , Proteínas Recombinantes/genética , Interleucina-22RESUMEN
Although prenatal alcohol exposure (PAE) reduces offspring growth, it may increase obesity risk at adolescence. Animal models of PAE display glucose intolerance and increased adiposity, suggesting that PAE causes metabolic reprogramming. We tested this hypothesis in a mouse model of binge PAE, wherein pregnant C57Bl/6J females received 3 g/kg alcohol (ETOH) daily from gestational day 12.5 to 17.5; maltodextrin (MD) and medium chain triglycerides (MCT) served as isocaloric nutritional controls, and sham (H2O) treatment controlled for gavage stress. Our comprehensive assessment quantified body composition, energy expenditure, glucose tolerance, and cardiovascular function in offspring at age 17 weeks. Although ETOH pups were initially lighter than all other groups, they did not have a unique obesogenic phenotype. Instead, a similar obesogenic phenotype emerged in all three caloric groups (MCT, MD, ETOH), such that caloric groups had greater post-weaning weight gain (both sexes), reduced gonadal fat weight (males), and reduced glucose clearance (males) compared against H2O offspring. PAE did not affect body composition, respiratory exchange ratio, metabolic adaption to high-fat or low-fat diet, eating behavior, and blood pressure, and ETOH values did not differ from those obtained from isocaloric controls. Exposure to a higher alcohol dose (4.5 g/kg) or a high-fat (60%) diet did not exacerbate differences in body composition or glucose tolerance. "PAE-specific" effects on postnatal growth, glucose tolerance, adiposity, or hypertension only emerged when PAE offspring were compared just against H2O controls, or against MD controls. We conclude that prior reports of obesity and glucose intolerance in adult PAE offspring reflect the contribution of added gestational calories, and not alcohol's pharmacologic action. Results suggest that the increased adiposity risk in FASD is not caused by metabolic reprogramming, and instead originates from behavioral, medication, and/or dietary practices. This study highlights the importance of appropriate dietary controls in nutritional studies of PAE.
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Consumo de Bebidas Alcohólicas/efectos adversos , Trastornos del Espectro Alcohólico Fetal/etiología , Exposición Materna , Síndrome Metabólico/etiología , Síndrome Metabólico/metabolismo , Efectos Tardíos de la Exposición Prenatal , Adiposidad , Animales , Biomarcadores , Peso Corporal , Dieta Alta en Grasa/efectos adversos , Modelos Animales de Enfermedad , Metabolismo Energético , Conducta Alimentaria , Femenino , Desarrollo Fetal/efectos de los fármacos , Intolerancia a la Glucosa , Hemodinámica , Masculino , Ratones , EmbarazoRESUMEN
Next-generation sequencing (NGS) is quickly revolutionizing how research into the genetic determinants of constitutional disease is performed. The technique is highly efficient with millions of sequencing reads being produced in a short time span and at relatively low cost. Specifically, targeted NGS is able to focus investigations to genomic regions of particular interest based on the disease of study. Not only does this further reduce costs and increase the speed of the process, but it lessens the computational burden that often accompanies NGS. Although targeted NGS is restricted to certain regions of the genome, preventing identification of potential novel loci of interest, it can be an excellent technique when faced with a phenotypically and genetically heterogeneous disease, for which there are previously known genetic associations. Because of the complex nature of the sequencing technique, it is important to closely adhere to protocols and methodologies in order to achieve sequencing reads of high coverage and quality. Further, once sequencing reads are obtained, a sophisticated bioinformatics workflow is utilized to accurately map reads to a reference genome, to call variants, and to ensure the variants pass quality metrics. Variants must also be annotated and curated based on their clinical significance, which can be standardized by applying the American College of Medical Genetics and Genomics Pathogenicity Guidelines. The methods presented herein will display the steps involved in generating and analyzing NGS data from a targeted sequencing panel, using the ONDRISeq neurodegenerative disease panel as a model, to identify variants that may be of clinical significance.