Macrophage fumarate hydratase restrains mtRNA-mediated interferon production.

Metabolic rewiring underlies the effector functions of macrophages1-3, but the mechanisms involved remain incompletely defined. Here, using unbiased metabolomics and stable isotope-assisted tracing, we show that an inflammatory aspartate-argininosuccinate shunt is induced following lipopolysaccharide stimulation. The shunt, supported by increased argininosuccinate synthase (ASS1) expression, also leads to increased cytosolic fumarate levels and fumarate-mediated protein succination. Pharmacological inhibition and genetic ablation of the tricarboxylic acid cycle enzyme fumarate hydratase (FH) further increases intracellular fumarate levels. Mitochondrial respiration is also suppressed and mitochondrial membrane potential increased. RNA sequencing and proteomics analyses demonstrate that there are strong inflammatory effects resulting from FH inhibition. Notably, acute FH inhibition suppresses interleukin-10 expression, which leads to increased tumour necrosis factor secretion, an effect recapitulated by fumarate esters. Moreover, FH inhibition, but not fumarate esters, increases interferon-ß production through mechanisms that are driven by mitochondrial RNA (mtRNA) release and activation of the RNA sensors TLR7, RIG-I and MDA5. This effect is recapitulated endogenously when FH is suppressed following prolonged lipopolysaccharide stimulation. Furthermore, cells from patients with systemic lupus erythematosus also exhibit FH suppression, which indicates a potential pathogenic role for this process in human disease. We therefore identify a protective role for FH in maintaining appropriate macrophage cytokine and interferon responses.

4-Octyl Itaconate and Dimethyl Fumarate Induce Secretion of the Anti-Inflammatory Protein Annexin A1 via NRF2.

Annexin A1 is a key anti-inflammatory effector protein that is involved in the anti-inflammatory effects of glucocorticoids. 4-Octyl itaconate (4-OI), a derivative of the endogenous metabolite itaconate, which is abundantly produced by LPS-activated macrophages, has recently been identified as a potent anti-inflammatory agent. The anti-inflammatory effects of 4-OI share a significant overlap with those of dimethyl fumarate (DMF), a derivate of another Krebs cycle metabolite fumarate, which is already in use clinically for the treatment of inflammatory diseases. In this study we show that both 4-OI and DMF induce secretion of the 33-kDa form of annexin A1 from murine bone marrow-derived macrophages, an effect that is much more pronounced in LPS-stimulated cells. We also show that this 4-OI- and DMF-driven annexin A1 secretion is NRF2-dependent and that other means of activating NRF2 give rise to the same response. Lastly, we demonstrate that the cholesterol transporter ABCA1, which has previously been implicated in annexin A1 secretion, is required for this process in macrophages. Our findings contribute to the growing body of knowledge on the anti-inflammatory effects of the Krebs cycle metabolite derivatives 4-OI and DMF.

The phosphorylation of AMPKß1 is critical for increasing autophagy and maintaining mitochondrial homeostasis in response to fatty acids.

Fatty acids are vital for the survival of eukaryotes, but when present in excess can have deleterious consequences. The AMP-activated protein kinase (AMPK) is an important regulator of multiple branches of metabolism. Studies in purified enzyme preparations and cultured cells have shown that AMPK is allosterically activated by small molecules as well as fatty acyl-CoAs through a mechanism involving Ser108 within the regulatory AMPK ß1 isoform. However, the in vivo physiological significance of this residue has not been evaluated. In the current study, we generated mice with a targeted germline knock-in (KI) mutation of AMPKß1 Ser108 to Ala (S108A-KI), which renders the site phospho-deficient. S108A-KI mice had reduced AMPK activity (50 to 75%) in the liver but not in the skeletal muscle. On a chow diet, S108A-KI mice had impairments in exogenous lipid-induced fatty acid oxidation. Studies in mice fed a high-fat diet found that S108A-KI mice had a tendency for greater glucose intolerance and elevated liver triglycerides. Consistent with increased liver triglycerides, livers of S108A-KI mice had reductions in mitochondrial content and respiration that were accompanied by enlarged mitochondria, suggestive of impairments in mitophagy. Subsequent studies in primary hepatocytes found that S108A-KI mice had reductions in palmitate- stimulated Cpt1a and Ppargc1a mRNA, ULK1 phosphorylation and autophagic/mitophagic flux. These data demonstrate an important physiological role of AMPKß1 Ser108 phosphorylation in promoting fatty acid oxidation, mitochondrial biogenesis and autophagy under conditions of high lipid availability. As both ketogenic diets and intermittent fasting increase circulating free fatty acid levels, AMPK activity, mitochondrial biogenesis, and mitophagy, these data suggest a potential unifying mechanism which may be important in mediating these effects.

Prostagram magnetic resonance imaging in a screening population: Prostate Imaging-Reporting and Data System or Likert?

OBJECTIVE: To compare biopsy recommendation rates and accuracy of the Prostate Imaging-Reporting and Data System, version 2 (PI-RADSv2) with the Likert scale for detection of clinically significant and insignificant prostate cancer in men screened within the Imperial Prostate 1 Prostate Cancer Screening Trial Using Imaging (IP1-PROSTAGRAM). PATIENTS AND METHODS: Men aged 50-69 years were screened with Prostagram MRI. Scans were prospectively reported using both PI-RADSv2 (excluding dynamic contrast-enhanced sequence score) and 5-point Likert scores by expert uro-radiologists. Systematic and targeted transperineal biopsy was recommended if the scan was scored ≥ 3, based on either reporting system. The proportion of patients recommended for biopsy and detection rates for Grade Groups (GGs) 1 and ≥ 2 were compared. Receiver operating characteristic (ROC) analysis was performed to compare performance. RESULTS: A total of 406 men underwent Prostagram MRI. The median (interquartile range) age and prostate-specific antigen level were 57 (53-61) years and 0.91 (0.56-1.74) ng/mL, respectively. At MRI score ≥ 3, more patients were recommended for biopsy based on Likert criteria (94/406; 23%, 95% confidence interval [CI] 19.2%-27.6%) compared to PI-RADSv2 (72/406; 18%, 95% CI 14.2%-21.9%; P = 0.03). For MRI scores ≥ 4, PI-RADSv2 and Likert scales led to 43/406 (11%, 95% CI 7.9%-14.1%) and 35/406 (9%, 95% CI 6.2%-11.9%) men recommended for biopsy (P = 0.40). For GG ≥ 2 detection, PIRADSv2 and Likert detected 22% (95% CI 11.4%-30.8%, 14/72) and 16% (95% CI 9.5%-25.3%, 15/94), respectively (P = 0.56). For GG1 cancers detection these were 11% (95% CI 4.3%-19.6%, seven of 72) vs 11% (95% CI 4.7%-17.8%, nine of 94; P = 1.00). The accuracy of PI-RADSv2 and Likert scale was similar (area under the ROC curve 0.64 vs 0.65, P = 0.95). CONCLUSIONS: In reporting non-contrast-enhanced Prostagram MRI in a screening population, the PI-RADSv2 and Likert scoring systems were equally accurate; however, Likert scale use led to more men undergoing biopsy without a subsequent increase in significant cancer detection rates. To improve reporting of Prostagram MRI, either the PI-RADSv2 or a modified Likert scale or a standalone scoring system should be developed.

Posterior cerebral territory ischemia in pediatric moyamoya: Surgical techniques and long-term clinical and radiographic outcomes.

PURPOSE: To describe a surgical technique for posterior cerebral revascularization in pediatric patients with moyamoya arteriopathy. Here, we describe the clinical characteristics, surgical indications, operative techniques, and clinical and radiographic outcomes in a series of pediatric patients with moyamoya disease affecting the posterior cerebral artery (PCA) territory. METHODS: A retrospective single-center series of all pediatric patients with moyamoya disease who presented to our institute between July 2009 through August 2019 were reviewed. The clinical characteristics, surgical indications, operative techniques, and long-term clinical and radiographic outcomes of pediatric moyamoya patients with PCA territory ischemia were collected and analyzed. RESULTS: A total of 10 PCA revascularization procedures were performed in 9 patients, 5 female, ages 1 to 11.1 years (average 5.2 years). Complications included 1 stroke, with no infections, hemorrhages, seizures, or deaths. One patient had less than 1 year of radiographic and clinical follow-up. In 8 of 9 patients with at least 1 year of radiographic follow-up, there was engraftment of surgical vessels present in all cases. No new strokes were identified on long-term follow-up despite the radiographic progression of the disease. In the 8 cases available for analysis, the average follow-up was 50.8 months with a range of 12 to 117 months. CONCLUSIONS: PCA territory ischemia in patients with progressive moyamoya disease can be surgically treated with indirect revascularization. Here, we describe our experience with PCA revascularization procedures for moyamoya disease, including pial pericranial dural (PiPeD) revascularization and pial synangiosis utilizing the occipital artery. These surgical options may be useful for decreasing the risk of stroke in pediatric moyamoya patients with severe posterior circulation disease.

Natural (dihydro)phenanthrene plant compounds are direct activators of AMPK through its allosteric drug and metabolite-binding site.

AMP-activated protein kinase (AMPK) is a central energy sensor that coordinates the response to energy challenges to maintain cellular ATP levels. AMPK is a potential therapeutic target for treating metabolic disorders, and several direct synthetic activators of AMPK have been developed that show promise in preclinical models of type 2 diabetes. These compounds have been shown to regulate AMPK through binding to a novel allosteric drug and metabolite (ADaM)-binding site on AMPK, and it is possible that other molecules might similarly bind this site. Here, we performed a high-throughput screen with natural plant compounds to identify such direct allosteric activators of AMPK. We identified a natural plant dihydrophenathrene, Lusianthridin, which allosterically activates and protects AMPK from dephosphorylation by binding to the ADaM site. Similar to other ADaM site activators, Lusianthridin showed preferential activation of AMPKß1-containing complexes in intact cells and was unable to activate an AMPKß1 S108A mutant. Lusianthridin dose-dependently increased phosphorylation of acetyl-CoA carboxylase in mouse primary hepatocytes, which led to a corresponding decrease in de novo lipogenesis. This ability of Lusianthridin to inhibit lipogenesis was impaired in hepatocytes from ß1 S108A knock-in mice and mice bearing a mutation at the AMPK phosphorylation site of acetyl-CoA carboxylase 1/2. Finally, we show that activation of AMPK by natural compounds extends to several analogs of Lusianthridin and the related chemical series, phenanthrenes. The emergence of natural plant compounds that regulate AMPK through the ADaM site raises the distinct possibility that other natural compounds share a common mechanism of regulation.

Cancer Cell Membrane Wrapped Nanoparticles for the Delivery of a Bcl-2 Inhibitor to Triple-Negative Breast Cancer.

Overexpression of the antiapoptotic protein B-cell lymphoma 2 (Bcl-2) is correlated with poor survival outcomes in triple-negative breast cancer (TNBC), making Bcl-2 inhibition a promising strategy to treat this aggressive disease. Unfortunately, Bcl-2 inhibitors developed to date have limited clinical success against solid tumors, owing to poor bioavailability, insufficient tumor delivery, and off-target toxicity. To circumvent these problems, we loaded the Bcl-2 inhibitor ABT-737 in poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) that were wrapped with phospholipid membranes derived from 4T1 murine mammary cancer cells, which mimic the growth and metastasis of human TNBC. We show that the biomimetic cancer cell membrane coating enabled the NPs to preferentially target 4T1 TNBC cells over noncancerous mammary epithelial cells in vitro and significantly increased NP accumulation in orthotopic 4T1 tumors in mice after intravenous injection by over 2-fold compared to poly(ethylene glycol)-poly(lactide-co-glycolic) (PEG-PLGA) copolymer NPs. Congruently, the ABT-737 loaded, cancer cell membrane-wrapped PLGA NPs (ABT CCNPs) induced higher levels of apoptosis in TNBC cells in vitro than ABT-737 delivered freely or in PEG-PLGA NPs. When tested in a syngeneic spontaneous metastasis model, the ABT CCNPs significantly increased apoptosis (evidenced by elevated active caspase-3 and decreased Bcl-2 staining) and decreased proliferation (denoted by reduced Ki67 staining) throughout tumors compared with saline or ABT-loaded PEG-PLGA NP controls. Moreover, the ABT CCNPs did not alter animal weight or blood composition, suggesting that the specificity afforded by the TNBC cell membrane coating mitigated the off-target adverse effects typically associated with ABT-737. Despite these promising results, the low dose of ABT CCNPs administered only modestly reduced primary tumor growth and metastatic nodule formation in the lungs relative to controls. We posit that increasing the dose of ABT CCNPs, altering the treatment schedule, or encapsulating a more potent Bcl-2 inhibitor may yield more robust effects on tumor growth and metastasis. With further development, drug-loaded biomimetic NPs may safely treat solid tumors such as TNBC that are characterized by Bcl-2 overexpression.

Combination cancer imaging and phototherapy mediated by membrane-wrapped nanoparticles.

Cancer is a devastating health problem with inadequate treatment options. Many conventional treatments for solid-tumor cancers lack tumor specificity, which results in low efficacy and off-target damage to healthy tissues. Nanoparticle (NP)-mediated photothermal therapy (PTT) is a promising minimally invasive treatment for solid-tumor cancers that has entered clinical trials. Traditionally, NPs used for PTT are coated with passivating agents and/or targeting ligands, but alternative coatings are being explored to enhance tumor specific delivery. In particular, cell-derived membranes have emerged as promising coatings that improve the biointerfacing of photoactive NPs, which reduces their immune recognition, prolongs their systemic circulation and increases their tumor accumulation, allowing for more effective PTT. To maximize treatment success, membrane-wrapped nanoparticles (MWNPs) that enable dual tumor imaging and PTT are being explored. These multifunctional theranostic NPs can be used to enhance tumor detection and/or ensure a sufficient quantity of NPs that have arrived in the tumor prior to laser irradiation. This review summarizes the current state-of-the-art in engineering MWNPs for combination cancer imaging and PTT and discusses considerations for the path toward clinical translation.

Protein targeting by the itaconate family in immunity and inflammation.

Immune cells are metabolically plastic and respond to inflammatory stimuli with large shifts in metabolism. Itaconate is one of the most up-regulated metabolites in macrophages in response to the gram negative bacterial product LPS. As such, itaconate has recently been the subject of intense research interest. The artificial derivatives, including 4-Octyl Itaconate (4-OI) and Dimethyl Itaconate (DI) and naturally produced isomers, mesaconate and citraconate, have been tested in relation to itaconate biology with similarities and differences in the biochemistry and immunomodulatory properties of this family of compounds emerging. Both itaconate and 4-OI have been shown to modify cysteines on a range of target proteins, with the modification being linked to a functional change. Targets include KEAP1 (the NRF2 inhibitor), GAPDH, NLRP3, JAK1, and the lysosomal regulator, TFEB. 4-OI and DI are more electrophilic, and are therefore stronger NRF2 activators, and inhibit the production of Type I IFNs, while itaconate inhibits SDH and the dioxygenase, TET2. Additionally, both itaconate and derivates have been shown to be protective across a wide range of mouse models of inflammatory and infectious diseases, through both distinct and overlapping mechanisms. As such, continued research involving the comparison of itaconate and related molecules holds exciting prospects for the study of cysteine modification and pathways for immunomodulation and the potential for new anti-inflammatory therapeutics.

miR-182 integrates apoptosis, growth, and differentiation programs in glioblastoma.

Glioblastoma multiforme (GBM) is a lethal, therapy-resistant brain cancer consisting of numerous tumor cell subpopulations, including stem-like glioma-initiating cells (GICs), which contribute to tumor recurrence following initial response to therapy. Here, we identified miR-182 as a regulator of apoptosis, growth, and differentiation programs whose expression level is correlated with GBM patient survival. Repression of Bcl2-like12 (Bcl2L12), c-Met, and hypoxia-inducible factor 2α (HIF2A) is of central importance to miR-182 anti-tumor activity, as it results in enhanced therapy susceptibility, decreased GIC sphere size, expansion, and stemness in vitro. To evaluate the tumor-suppressive function of miR-182 in vivo, we synthesized miR-182-based spherical nucleic acids (182-SNAs); i.e., gold nanoparticles covalently functionalized with mature miR-182 duplexes. Intravenously administered 182-SNAs penetrated the blood-brain/blood-tumor barriers (BBB/BTB) in orthotopic GBM xenografts and selectively disseminated throughout extravascular glioma parenchyma, causing reduced tumor burden and increased animal survival. Our results indicate that harnessing the anti-tumor activities of miR-182 via safe and robust delivery of 182-SNAs represents a novel strategy for therapeutic intervention in GBM.

AMPK mediates energetic stress-induced liver GDF15.

Growth differentiating factor-15 (GDF15) is an emerging target for the treatment of obesity and metabolic disease partly due to its ability to suppress food intake. GDF15 expression and secretion are thought to be regulated by a cellular integrated stress response, which involves endoplasmic reticulum (ER) stress. AMPK is another cellular stress sensor, but the relationship between AMPK, ER stress, and GDF15 has not been assessed in vivo. Wildtype (WT), AMPK ß1 deficient (AMPKß1-/- ), and CHOP-/- mice were treated with three distinct AMPK activators; AICAR, which is converted to ZMP mimicking the effects of AMP on the AMPKγ isoform, R419, which indirectly activates AMPK through inhibition of mitochondrial respiration, or A769662, a direct AMPK activator which binds the AMPKß1 isoform ADaM site causing allosteric activation. Following treatments, liver Gdf15, markers of ER-stress, AMPK activity, adenine nucleotides, circulating GDF15, and food intake were assessed. AICAR and R419 caused ER and energetic stress, increased GDF15 expression and secretion, and suppressed food intake. Direct activation of AMPK ß1 containing complexes by A769662 increased hepatic Gdf15 expression, circulating GDF15, and suppressed food intake, independent of ER stress. The effects of AICAR, R419, and A769662 on GDF15 were attenuated in AMPKß1-/- mice. AICAR and A769662 increased GDF15 to a similar extent in WT and CHOP-/- mice. Herein, we provide evidence that AMPK plays a role in mediating the induction of GDF15 under conditions of energetic stress in mouse liver in vivo.

Adjusting for verification bias in diagnostic accuracy measures when comparing multiple screening tests - an application to the IP1-PROSTAGRAM study.

INTRODUCTION: Novel screening tests used to detect a target condition are compared against either a reference standard or other existing screening methods. However, as it is not always possible to apply the reference standard on the whole population under study, verification bias is introduced. Statistical methods exist to adjust estimates to account for this bias. We extend common methods to adjust for verification bias when multiple tests are compared to a reference standard using data from a prospective double blind screening study for prostate cancer. METHODS: Begg and Greenes method and multiple imputation are extended to include the results of multiple screening tests which determine condition verification status. These two methods are compared to the complete case analysis using the IP1-PROSTAGRAM study data. IP1-PROSTAGRAM used a paired-cohort double-blind design to evaluate the use of imaging as alternative tests to screen for prostate cancer, compared to a blood test called prostate specific antigen (PSA). Participants with positive imaging (index) and/or PSA (control) underwent a prostate biopsy (reference standard). RESULTS: When comparing complete case results to Begg and Greenes and methods of multiple imputation there is a statistically significant increase in the specificity estimates for all screening tests. Sensitivity estimates remained similar across the methods, with completely overlapping 95% confidence intervals. Negative predictive value (NPV) estimates were higher when adjusting for verification bias, compared to complete case analysis, even though the 95% confidence intervals overlap. Positive predictive value (PPV) estimates were similar across all methods. CONCLUSION: Statistical methods are required to adjust for verification bias in accuracy estimates of screening tests. Expanding Begg and Greenes method to include multiple screening tests can be computationally intensive, hence multiple imputation is recommended, especially as it can be modified for low prevalence of the target condition.

Photoresponsive miR-34a/Nanoshell Conjugates Enable Light-Triggered Gene Regulation to Impair the Function of Triple-Negative Breast Cancer Cells.

Triple-negative breast cancer (TNBC) is an aggressive disease that requires new interventions. A promising approach to improve patient prognosis is to introduce tumor suppressive miR-34a into TNBC cells. Unfortunately, naked miR-34a is not effective therapeutically because it is degraded by nucleases and cannot passively enter cells. Nanocarriers designed to increase miR-34a stability and cellular entry have lacked specificity and potency. To overcome these limitations, we conjugated miR-34a to photoresponsive gold nanoshells (NS), which can release tethered miR-34a upon excitation with continuous wave (CW) or nanosecond (ns) pulsed near-infrared light to facilitate on-demand gene regulation. We demonstrate that miR-34a/NS can regulate downstream miR-34a targets following irradiation to reduce TNBC cell viability, proliferation, and migration. Further, we show ns pulsed light releases miRNA more effectively than CW light, and that released miR-34a is as potent as transfected miR-34a. These findings signify miR-34a/NS as promising tools for precisely controlled gene regulation of TNBC.

Nerve transfer for restoration of lower motor neuron-lesioned bladder function. Part 2: correlation between histological changes and nerve evoked contractions.

We determined the effect of pelvic organ decentralization and reinnervation 1 yr later on urinary bladder histology and function. Nineteen canines underwent decentralization by bilateral transection of all coccygeal and sacral (S) spinal roots, dorsal roots of lumbar (L)7, and hypogastric nerves. After exclusions, eight were reinnervated 12 mo postdecentralization with obturator-to-pelvic and sciatic-to-pudendal nerve transfers, then euthanized 8-12 mo later. Four served as long-term decentralized only animals. Before euthanasia, pelvic or transferred nerves and L1-S3 spinal roots were stimulated and maximum detrusor pressure (MDP) recorded. Bladder specimens were collected for histological and ex vivo smooth muscle contractility studies. Both reinnervated and decentralized animals showed less or denuded urothelium, fewer intramural ganglia, and more inflammation and collagen, than controls, although percent muscle was maintained. In reinnervated animals, pgp9.5+ axon density was higher compared with decentralized animals. Ex vivo smooth muscle contractions in response to KCl correlated positively with submucosal inflammation, detrusor muscle thickness, and pgp9.5+ axon density. In vivo, reinnervated animals showed higher MDP after stimulation of L1-L6 roots compared with their transected L7-S3 roots, and reinnervated and decentralized animals showed lower MDP than controls after stimulation of nerves (due likely to fibrotic nerve encapsulation). MDP correlated negatively with detrusor collagen and inflammation, and positively with pgp9.5+ axon density and intramural ganglia numbers. These results demonstrate that bladder function can be improved by transfer of obturator nerves to pelvic nerves at 1 yr after decentralization, although the fibrosis and inflammation that developed were associated with decreased contractile function.

Critical Evaluation of Different Lysosomal Labeling Methods Used to Analyze RNA Nanocarrier Trafficking in Cells.

The use of nucleic acids to regulate gene expression is a rapidly developing field with immense clinical potential. Nanomaterials are frequently used to deliver nucleic acids into cells as they can overcome the poor cellular uptake and endo/lysosomal degradation of bare nucleic acids. For these nanocarriers to be effective, they must escape endo/lysosomal compartments to deliver their nucleic acid cargo into the cytosol (for ribonucleic acid (RNA)) or nucleus (for deoxyribonucleic acid (DNA)). This process is poorly understood and remains an area of active research toward the goal of developing effective delivery strategies. Fluorescent endo/lysosomal markers are among the most widely employed tools used to evaluate the endosomal escape of nucleic acid nanocarriers. However, the endo/lysosomal labeling method may alter the extent of and route of nanocarrier uptake by cells. The impact of these markers on cellular function and cell-nanocarrier interactions has not been probed in a systematic manner. To investigate this, we compared the effects of several common lysosomal labeling methods, namely, LysoTracker Red (LT Red), transient lysosomal-associated membrane protein 1-mutant green fluorescent protein (LAMP1-mGFP) transfection (Transient GFP), and stable lentiviral LAMP1-mGFP transfection (Stable GFP), on cellular metabolic activity, nanocarrier uptake, nanocarrier/lysosomal label colocalization, and gene silencing potency in U87 glioblastoma and MDA-MB-231 breast cancer cells using polyethyleneimine (PEI)/ribonucleic acid (RNA) polyplexes as a model nanocarrier. In both U87s and MDA-MB-231s, Transient GFP and LT Red labeling reduced metabolic activity relative to untransfected (Parental) cells, while Stable GFP labeling increased metabolic activity. Congruently, flow cytometry indicates Stable GFP cells have greater polyplex uptake than LT Red-labeled cells in both cell lines. Despite these similar trends in uptake, polyplex intracellular trafficking differs in the two cell lines, as confocal imaging revealed greater polyplex/lysosome colocalization in Stable GFP U87 cells than LT Red-labeled U87 cells, while the trend was reversed in MBA-MB-231s. The level of RNA-mediated gene silencing achieved in Parental versus Stable GFP U87 and MDA-MB-231 cells agreed with the observed levels of polyplex/lysosome colocalization, supporting the established concept that endosomal escape is the rate-limiting step for RNA interference. These findings indicate that lysosomal labels can profoundly alter cellular function and cell-nanocarrier interactions, presenting critical new considerations for researchers investigating nanoparticle trafficking.

Safety and efficacy of an extended-release peptide YY analogue for obesity: A randomized, placebo-controlled, phase 1 trial.

AIM: To report the results from a Phase 1 trial of an extended-release peptide YY analogue, Y14, developed for the treatment of obesity. METHODS: Y14 was evaluated in overweight/obese volunteers in a Phase 1 randomized placebo-controlled trial, conducted in a clinical trial unit in the United Kingdom. Part A was a blinded single-ascending-dose study evaluating doses up to 36 mg. Part B was double-blinded and tested multiple ascending doses between 9 and 36 mg, given at 7- to 14-day intervals, over the course of 28 days, with up to five doses given per participant. The primary outcome was safety and tolerability; the secondary outcome was assessment of pharmacokinetic (PK) characteristics. Exploratory outcomes included food intake, body weight change and glucose tolerance after multiple doses. RESULTS: Between April 11, 2017 and December 24, 2018, 53 participants were enrolled into Part A and 24 into Part B of the trial. The PK characteristics were compatible with administration every 7 to 14 days. The most common adverse events (AEs) were nausea, vomiting or administration site reactions, which were mild in most cases and settled with time. No serious AE occurred. Participants given multiple doses of Y14 lost between -2.87 and -3.58 kg body weight compared with placebo (P <0.0001) at 31 days from the first dose, with profound reductions in food intake of 38% to 55% (P <0.0001, compared to placebo) and there was no evidence of tachyphylaxis. CONCLUSIONS: Our results support the continued development of Y14 as a novel treatment for obesity.

The SGLT2 inhibitor canagliflozin suppresses lipid synthesis and interleukin-1 beta in ApoE deficient mice.

Sodium-glucose cotransporter 2 inhibitors such as canagliflozin lower blood glucose and reduce cardiovascular events in people with type 2 diabetes through mechanisms that are not fully understood. Canagliflozin has been shown to increase the activity of the AMP-activated protein kinase (AMPK), a metabolic energy sensor important for increasing fatty acid oxidation and energy expenditure and suppressing lipogenesis and inflammation, but whether AMPK activation is important for mediating some of the beneficial metabolic effects of canagliflozin has not been determined. We, therefore, evaluated the effects of canagliflozin in female ApoE-/- and ApoE-/-AMPK ß1-/- mice fed a western diet. Canagliflozin increased fatty acid oxidation and energy expenditure and lowered adiposity, blood glucose and the respiratory exchange ratio independently of AMPK ß1. Canagliflozin also suppressed liver lipid synthesis and the expression of ATP-citrate lyase, acetyl-CoA carboxylase and sterol response element-binding protein 1c independently of AMPK ß1. Canagliflozin lowered circulating IL-1ß and studies in bone marrow-derived macrophages indicated that in contrast with the metabolic adaptations, this effect required AMPK ß1. Canagliflozin had no effect on the size of atherosclerotic plaques in either ApoE-/- and ApoE-/-AMPK ß1-/- mice. Future studies investigating whether reductions in liver lipid synthesis and macrophage IL-1ß are important for the cardioprotective effects of canagliflozin warrant further investigation.

Postoperative MR imaging surveillance of pediatric craniopharyngioma: new institutional guidelines.

PURPOSE: To develop postoperative surveillance protocols that yield efficient detection rates of tumor recurrence or progression using fewer imaging studies and less cost. METHOD: This is a retrospective cohort study of all pediatric craniopharyngioma patients who have been diagnosed and treated at Boston Children's Hospital (BCH) between 1990 and 2017. All statistical analyses were performed using Stata. RESULTS: Eighty patients (43 males and 37 females) fulfilled the inclusion criteria. The mean age at time of diagnosis was 8.6 ± 4.4 years. The mean follow-up period was 10.9 ± 6.5 years. Overall 30/80 (37.5%) patients experienced tumor recurrence/progression. The median latency to recurrence/progression was 12.75 months (range 3 to 108 months), with 76.6% of the recurrences/progressions taking place within the first 2 years postoperatively. Given the lack of any clinical symptoms/signs associated with the vast majority of the recurrent/progressed cases, we propose postoperative MR imaging surveillance protocols that are substantially less intensive than the current practice. Therefore, we recommend the following postoperative MR imaging surveillance protocols, stratified by management strategies; 0, 9, 15, 36, 48, and 60 months for patients who underwent GTR, 0, 3, 6,12, 18, and 24 months for patients who underwent STR alone and 0, 3, 12, 72, 96, and 120 months for patients who underwent STR followed by subsequent XRT. CONCLUSION: The proposed postoperative MR imaging surveillance protocols would provide a potential 50% decrement of healthcare costs. It may also minify the psychological burden of frequent MR scanning for these patients and their families.