Metabolic Burden Research Articles

18 F-FDG PET/CT for evaluation of metastases in nonsmall cell lung cancer on the efficacy of immunotherapy.

This study aimed to investigate the relationship between 18 F-fluorodeoxyglucose PET/computed tomography ( 18 F-FDG PET/CT) metabolic parameters and clinical benefit and prognosis in nonsmall cell lung cancer (NSCLC). In total, 34 advanced NSCLC patients who received 18 F-FDG PET/CT before immunotherapy were retrospectively included in this study. All patients were divided into two groups, the clinical benefit (CB) group and the no-clinical benefit (no-CB) group, based on the efficacy of evaluation after 6 months of treatment. Also clinical information, characteristics of metastases, survival, PD-L1 expression level and glucose metabolic parameters were evaluated. Finally, 24 patients were in the CB group, and 10 patients were in the no-CB group. There was a significant difference between the CB group and the no-CB group in TNM stages ( P = 0.005), visceral and bone metastasis ( P = 0.031), metabolic tumor volume of primary lesion (MTV-P; P = 0.003), the metabolic tumor volume of whole-body (MTVwb; P = 0.005) and total lesion glycolysis of whole-body (TLGwb, P = 0.015). However, for patient outcomes, the independent prognostic factors associated with progression free survival were TNM stage (HR = 0.113; 95% CI, 0.029-0.439; P = 0.002), TLG-P (HR = 0.085; 95% CI, 0.018-0.402; P = 0.002) and TLG-LN (HR = 0.068; 95% CI, 0.015-0.308; P = 0.000), and the TLG-LN (HR = 0.242; 95% CI, 0.066-0.879; P = 0.002) was the independent prognostic factor associated with overall survival. Metastatic lesion burden evaluated by 18 F-FDG PET/ CT can predict response to immunotherapy in advanced NSCLC patients, in which lymph node metastasis lesion metabolic burden is a meaningful predictor, but a large multicenter trial is still needed to validate this conclusion.

Uncovering the Gut-Liver Axis Biomarkers for Predicting Metabolic Burden in Mice.

Western diet (WD) intake, aging, and inactivation of farnesoid X receptor (FXR) are risk factors for metabolic and chronic inflammation-related health issues ranging from metabolic dysfunction-associated steatotic liver disease (MASLD) to dementia. The progression of MASLD can be escalated when those risks are combined. Inactivation of FXR, the receptor for bile acid (BA), is cancer prone in both humans and mice. The current study used multi-omics including hepatic transcripts, liver, serum, and urine metabolites, hepatic BAs, as well as gut microbiota from mouse models to classify those risks using machine learning. A linear support vector machine with K-fold cross-validation was used for classification and feature selection. We have identified that increased urine sucrose alone achieved 91% accuracy in predicting WD intake. Hepatic lithocholic acid and serum pyruvate had 100% and 95% accuracy, respectively, to classify age. Urine metabolites (decreased creatinine and taurine as well as increased succinate) or increased gut bacteria (Dorea, Dehalobacterium, and Oscillospira) could predict FXR deactivation with greater than 90% accuracy. Human disease relevance is partly revealed using the metabolite-disease interaction network. Transcriptomics data were also compared with the human liver disease datasets. WD-reduced hepatic Cyp39a1 (cytochrome P450 family 39 subfamily a member 1) and increased Gramd1b (GRAM domain containing 1B) were also changed in human liver cancer and metabolic liver disease, respectively. Together, our data contribute to the identification of noninvasive biomarkers within the gut-liver axis to predict metabolic status.

Comparison of Baseline 68Ga-FAPI and 18F-FDG PET/CT for Prediction of Response and Clinical Outcome in Patients with Unresectable Hepatocellular Carcinoma Treated with PD-1 Inhibitor and Lenvatinib.

Fibroblast activation protein contributes to immunosuppression and resistance to immunotherapies. This study aimed to compare baseline 68Ga-labeled fibroblast activation protein inhibitor (68Ga-FAPI) PET/CT and 18F-FDG PET/CT in response and survival prediction in unresectable hepatocellular carcinoma (uHCC) patients treated with the combination of programmed cell death 1 (PD-1) inhibitor and lenvatinib. Methods: In this prospective cohort study, 22 patients with uHCC who underwent baseline 18F-FDG and 68Ga-FAPI PET/CT and soon began taking a combination of PD-1 inhibitor and lenvatinib were recruited. Semiquantitative indices of baseline PET/CT were measured as 18F-FDG SUVmax, metabolic tumor volume, total lesion glycolysis, 68Ga-FAPI SUVmax, 68Ga-FAPI-avid tumor volume (FTV), and total lesion fibroblast activation protein expression (TLF). The primary endpoint was durable or nondurable clinical benefit after treatment, and the secondary endpoints were progression-free survival (PFS) and overall survival (OS). Results: The overall response rate of the combination therapy was 41% (9/22). Fifty percent of patients had durable clinical benefit. Median PFS and OS were 4.8 and 14.4 mo, respectively. Patients with nondurable clinical benefit showed a significantly higher FTV and TLF than those with durable clinical benefit, whereas 18F-FDG parameters overlapped. A higher 68Ga-FAPI-avid tumor burden (FTV > 230.46 cm3 or TLF > 961.74 SUVbody weight⋅cm3) predicted both shorter PFS (4.0 vs. 13.5 mo, P = 0.016) and shorter OS (7.8 mo vs. not reached, P = 0.030). Patients with a higher metabolic tumor burden (metabolic tumor volume > 206.80 cm3 or total lesion glycolysis > 693.53 SUVbody weight⋅cm3) showed a shorter OS although the difference did not reach statistical significance (P = 0.085). In multivariate analysis, a higher 68Ga-FAPI-avid tumor burden (hazard ratio [HR], 3.88 [95% CI, 1.26-12.01]; P = 0.020) and macrovascular invasion (HR, 4.00 [95% CI, 1.06-15.14]; P = 0.039) independently predicted a shorter PFS, whereas a higher 68Ga-FAPI-avid tumor burden (HR, 5.92 [95% CI, 1.19-29.42]; P = 0.035) and bone metastases (HR, 5.88 [95% CI, 1.33-25.93]; P = 0.022) independently predicted a shorter OS. Conclusion: Volumetric indices on baseline 68Ga-FAPI PET/CT were potentially independent prognostic factors to predict durable clinical benefit, PFS, and OS in uHCC patients treated with a combination of PD-1 and lenvatinib. Baseline 68Ga-FAPI PET/CT may facilitate uHCC patient selection before combination therapy.

Association between daily aspirin therapy and risk of hepatocellular carcinoma according to metabolic risk factor burden in non-cirrhotic patients with chronic hepatitis B.

Several studies have demonstrated chemopreventive effects of aspirin against hepatocellular carcinoma (HCC) in patients with chronic hepatitis B (CHB). To investigate the associations of aspirin use with risks of HCC, liver-related mortality, and major bleeding according to metabolic risk factor burden among non-cirrhotic patients with CHB METHODS: Using the Korean National Health Insurance Service database, we identified 282,611 non-cirrhotic adults with CHB. Data on obesity, diabetes, high blood pressure, and hypercholesterolemia were collected. Subjects were stratified into lower and higher metabolic risk groups (≤2 and ≥3 risk factors, respectively). Propensity score-matched cohorts of aspirin users and non-users were generated. Risks of HCC, liver-related death and major bleeding were analyzed. During the median follow-up of 7.4 years, positive associations between metabolic risk factor burden and outcomes were verified (all ptrend < 0.001). In the lower metabolic risk group (13,104 pairs), the association between aspirin use and HCC risk was not significant after multivariable adjustment (adjusted subdistribution hazard ratio [aSHR]: 0.93; 95% CI: 0.84-1.03); however, aspirin use was associated with elevated major bleeding risk (aSHR: 1.22; 95% CI: 1.08-1.39). In the higher metabolic risk group (2984 pairs), aspirin use was associated with reduced risks of HCC (aSHR: 0.72; 95% CI: 0.57-0.91) and liver-related mortality (aSHR: 0.69; 95% CI: 0.50-0.96) without an increase in risk of major bleeding (aSHR: 1.02; 95% CI: 0.79-1.32). Aspirin therapy was associated with reduced risks of HCC and liver-related death without excess risk of major bleeding, in non-cirrhotic patients with CHB who had a higher metabolic risk factor burden.

Metabolomics of Escherichia coli for Disclosing Novel Metabolic Engineering Strategies for Enhancing Hydrogen and Ethanol Production.

The biological production of hydrogen is an appealing approach to mitigating the environmental problems caused by the diminishing supply of fossil fuels and the need for greener energy. Escherichia coli is one of the best-characterized microorganisms capable of consuming glycerol-a waste product of the biodiesel industry-and producing H2 and ethanol. However, the natural capacity of E. coli to generate these compounds is insufficient for commercial or industrial purposes. Metabolic engineering allows for the rewiring of the carbon source towards H2 production, although the strategies for achieving this aim are difficult to foresee. In this work, we use metabolomics platforms through GC-MS and FT-IR techniques to detect metabolic bottlenecks in the engineered ΔldhΔgndΔfrdBC::kan (M4) and ΔldhΔgndΔfrdBCΔtdcE::kan (M5) E. coli strains, previously reported as improved H2 and ethanol producers. In the M5 strain, increased intracellular citrate and malate were detected by GC-MS. These metabolites can be redirected towards acetyl-CoA and formate by the overexpression of the citrate lyase (CIT) enzyme and by co-overexpressing the anaplerotic human phosphoenol pyruvate carboxykinase (hPEPCK) or malic (MaeA) enzymes using inducible promoter vectors. These strategies enhanced specific H2 production by up to 1.25- and 1.49-fold, respectively, compared to the reference strains. Other parameters, such as ethanol and H2 yields, were also enhanced. However, these vectors may provoke metabolic burden in anaerobic conditions. Therefore, alternative strategies for a tighter control of protein expression should be addressed in order to avoid undesirable effects in the metabolic network.

Impact of Liver Transplantation on Adipose Tissue Compartments and Its Association With Metabolic Sequela.

Loss of skeletal muscle can be accompanied by an increase in adipose tissue leading to sarcopenic obesity. There are limited data on how liver transplantation (LT) might impact adipose tissue compartments, particularly among patients with metabolically active disease, such as nonalcoholic steatohepatitis (NASH) and subsequent metabolic sequela. Skeletal muscle, visceral adipose tissue (VAT), and subcutaneous adipose tissue (SAT) were measured using cross-sectional imaging performed in 190 patients pre-LT, 6 mo post-LT and 12 mo post-LT. Changes in adipose tissue and their impact on metabolic diseases were determined in patients transplanted for NASH versus non-NASH. Skeletal muscle, VAT, and SAT were similar in patients with NASH and non-NASH pre-LT despite a higher burden of metabolic diseases in patients with NASH. Following LT, no significant differences between skeletal muscle and SAT were observed in the entire cohort and among patients with NASH (versus non-NASH). LT recipients with the highest muscle mass pre-LT were at the greatest risk for muscle loss post-LT. A time-dependent increase in VAT was noted post-LT, which was more robust among patients with a history of NASH cirrhosis. In adjusted multivariate analysis, NASH versus non-NASH was a strong predictor of post-LT increase in VAT (β-coefficient 3.00, P = 0.04). Pre-LT VAT was an independent predictor of post-LT serum triglycerides (β-coefficient 5.49 ± 2.78, P = 0.05) and low-density lipoprotein cholesterol (β-coefficient 1.80 ± 0.75, P = 0.02). A trend between pre-LT VAT and diabetes was noted but did not reach statistical significance. VAT but not SAT increases rapidly after LT, especially among patients transplanted for NASH cirrhosis and predicts future metabolic burden.

Burdens of non-communicable disease attributable to metabolic risk factors in Australia, 1990–2019: joinpoint regression analysis of the Global Burden of Disease Study

BackgroundLong-term and comparative assessments of trends in non-communicable disease (NCD) burden attributable to metabolic risk are sparse. This study aimed to assess burdens and trends of NCD attributable to metabolic...

P16-004-23 Folate Malnutrition as a Metabolic Burden To Promote Advanced Hyperglycemia by Switching Bioenergetic Fluxes and Metabolic Flexibility

P16-004-23 Folate Malnutrition as a Metabolic Burden To Promote Advanced Hyperglycemia by Switching Bioenergetic Fluxes and Metabolic Flexibility

Growth feedback confers cooperativity in resource-competing synthetic gene circuits

Modularity is a key concept in designing synthetic gene circuits, as it allows for constructing complex molecular systems using well-characterized building blocks. One of the major challenges in this field is that these modular components often do not function as expected when assembled into larger circuits. One of the major issues is caused by resource competition, where multiple genes in the circuit compete for the same limited cellular resources, such as transcription factors and ribosomes. In addition, the mutual inhibition between synthetic gene circuits and cell growth results in growth feedback that significantly impacts its host-circuit dynamics. However, the complexity of the gene circuit dynamics under intertwined resource competition and growth feedback is not fully understood. This study developed a theoretical framework to examine the dynamics of synthetic gene circuits by considering both growth feedback and resource competition. Our results suggest a cooperative behavior between resource-competing gene circuits under growth feedback. Cooperation or competition is non-monotonically determined by the metabolic burden threshold. These two diverse effects could lead to the activation or deactivation of one circuit by the other. Lastly, the cooperativity mediated by growth feedback can attenuate the winner-takes-all resource competition. These findings show that coupling growth feedback and resource competition plays a crucial role in the dynamics of the host-circuit system, and understanding its effects helps control unexpected gene expression behaviors.

More than skin-deep: visceral fat is strongly associated with disease activity, function and metabolic indices in psoriatic disease

ObjectiveTo compare body composition between patients with psoriatic disease (PsD), including cutaneous psoriasis (PsO) and psoriatic arthritis (PsA), and controls, and to explore associations between disease activity and measures of function and metabolic derangement.MethodsBody composition was assessed by air displacement plethysmography (ADP) and MRI-derived fat segmentation using an automated pipeline (FatSegNet). Function was assessed by Health Assessment Questionnaire (HAQ) and metabolic status by fasting lipid profile, insulin and adiponectin. Active and inactive PsO and PsA were defined by body surface area (BSA) and Psoriasis Area Severity Index (PASI) and minimal disease activity (MDA), respectively.ResultsThirty patients (median disease duration 15 years; median age 52 years) and 30 BMI-matched controls were enrolled. Compared with controls, all MRI-derived body composition parameters—whole-body volume, subcutaneous adipose tissue (SAT), visceral adipose tissue (VAT), abdominal adipose tissue (AAT), VAT/AAT and VAT/SAT—were higher in the PsD group, specifically, those with active disease. Body mass, body fat, whole-body volume and whole-body VAT were correlated with higher triglycerides, cholesterol:HDL (high-density lipoprotein), insulin resistance and lower adiponectin as well as higher HAQ and lower MDA.ConclusionsIn this pilot study, patients with PsD revealed excessive total adipose tissue and a greater volume of metabolically unfavourable ectopic fat, including VAT, compared with BMI-matched controls, which also correlated with HAQ, disease activity and overall dysmetabolism. We also provide the first evidence in patients with PsD for the clinical application of FatSegNet: a novel, automated and rapid deep learning pipeline for providing accurate MRI-based measurement of fat segmentation. Our findings suggest the need for a more integrated approach to the management of PsD, which considers both the metabolic and inflammatory burden of disease. More specifically, visceral fat is a surrogate marker of uncontrolled PsD and may be an important future target for both pharmacological and lifestyle interventions.

Current state of molecular and metabolic strategies for the improvement of L-asparaginase expression in heterologous systems.

Heterologous expression of L-asparaginase (L-ASNase) has become an important area of research due to its clinical and food industry applications. This review provides a comprehensive overview of the molecular and metabolic strategies that can be used to optimize the expression of L-ASNase in heterologous systems. This article describes various approaches that have been employed to increase enzyme production, including the use of molecular tools, strain engineering, and in silico optimization. The review article highlights the critical role that rational design plays in achieving successful heterologous expression and underscores the challenges of large-scale production of L-ASNase, such as inadequate protein folding and the metabolic burden on host cells. Improved gene expression is shown to be achievable through the optimization of codon usage, synthetic promoters, transcription and translation regulation, and host strain improvement, among others. Additionally, this review provides a deep understanding of the enzymatic properties of L-ASNase and how this knowledge has been employed to enhance its properties and production. Finally, future trends in L-ASNase production, including the integration of CRISPR and machine learning tools are discussed. This work serves as a valuable resource for researchers looking to design effective heterologous expression systems for L-ASNase production as well as for enzymes production in general.

The tigecycline resistance gene tetX has an expensive fitness cost based on increased outer membrane permeability and metabolic burden in Escherichia coli

Livestock-derived tetX-positive Escherichia coli with tigecycline resistance poses a serious risk to public health. Fitness costs, antibiotic residues, and other tetracycline resistance genes (TRGs) are fundamental in determining the spread of tetX in the environment, but there is a lack of relevant studies. The results of this study showed that both tetO and tetX resulted in reduction in growth and an increased in the metabolic burden of E. coli, but the presence of doxycycline reversed this phenomenon. Moreover, the protection of E. coli growth and metabolism by tetO was superior to that of tetX in the presence of doxycycline, resulting in a much lower competitiveness of tetX-carrying E. coli than tetO-carrying E. coli. The results of RNA-seq showed that the increase in outer membrane proteins (ompC, ompF and ompT) of tetX-carrying E. coli resulted in increased membrane permeability and biofilm formation, which is an important reason for fitness costs. Overall, the increased membrane permeability and metabolic burden of E. coli is the mechanistic basis for the high fitness cost of tetX, and the spread of tetO may limit the spread of tetX. This study provides new insights into the rational use of tetracycline antibiotics to control the spread of tetX.

Impact of the longitudinal changes of myocardial remodeling on cardiovascular outcomes of kidney transplantation recipients

Abstract Funding Acknowledgements Type of funding sources: None. Background Kidney transplantation (KT) improves the hemodynamic burden associated with volume overload, and anemia in patients with chronic kidney disease, but cardiovascular disease remains the leading cause of death after KT. This study evaluated the longitudinal change of eccentric and concentric myocardial remodeling and its impact on cardiovascular outcomes after KT. Methods A total of 600 patients who underwent echocardiography before and 3 years after KT were included from a multi-center observational cohort (KNOW-KT). Conventional echocardiograph evaluated left ventricular ejection fraction (LV EF), LV mass index (LVMI) LV end-diastolic dimension (LVEDD), and relative wall thickness (RWT) at baseline and after KT, and the changes of these parameters and their effect on the cardiovascular events were investigated. Results After KT, LV EF, LVMI, and the myocardial eccentricity represented by LVEDD were significantly improved for 3 years (LV EF; 61.4±7.9 to 64.8±6.0 %, LVMI; 113.4 ± 31.8 to 94.4 ± 23.4 g/m2, LVEDD; 51.2 ± 5.6 to 47.1 ± 4.8 mm), however, the myocardial concentricity represented by RWT were increased from 0.39 ± 0.07 to 0.41 ± 0.07. The proportion of LV concentric remodeling significantly increased after KT compared to the baseline (Figure 1). In the correlation analysis between the change of echocardiographic and clinical parameters, the change of LVEDD was associated with change of hemodynamic stress such as hemoglobin or systolic blood pressure, whereas the change of RWT showed significant association with change of metabolic stress such as HbA1c and triglycerides. There were 30 major adverse cardiovascular events (MACE, myocardial infarction, angina, or strokes) after KT, and the incidence of MACE significantly increased in patients with the increased RWT group, but not in the increased LVEDD group after KT (Figure 2). In multivariate analysis corrected for baseline echocardiographic parameters, age, sex, diabetes, etc., an increase in RWT, independently predicted the occurrence of MACE (HR 2.20 CI 1.21–3.99, p&amp;lt;0.01), but other echocardiographic parameters including LVEF, LVMI, and LVEDD did not. Conclusions Longitudinal changes in myocardial concentricity is associated with the metabolic stress and have a significant impact on the new onset cardiovascular disease in KT patients. Improving metabolic burden after KT should be considered as an important strategy for amelioration of cardiac remodeling and adverse event.

#3940 RISK OF MAJOR ADVERSE CARDIOVASCULAR EVENTS ACCORDING TO DIALYSIS MODALITY

Abstract Background and Aims Among dialysis patients, cardiovascular events are the leading cause of death. Little is known about difference in the frequency of cardiovascular events between various dialysis modalities. Glucose load may contribute to a metabolic burden in peritoneal dialysis (PD). On the other hand, hemodialysis can cause intradialytic hypotension, cardiac stunning, and arrhythmias, which are associated with increased risk of death. We compared risk of major cardiovascular events in patients who started continuous ambulatory PD (CAPD), automated PD (APD) and home HD with in-center HD patients. Method We included 968 patients who entered dialysis in the Helsinki-Uusimaa healthcare district in Finland from 2004 to 2017, of whom 162 were on CAPD, 229 on APD, 145 on home HD and 432 on in-center HD at day 90 from the start of dialysis. Patients were followed up for 5 years or until the end of 2019. Major adverse cardiovascular event (MACE) was defined as acute myocardial infarction, stroke or death due to cardiovascular disease. The cumulative incidence of the first MACE was calculated by taking other causes of death into account as competing risk events and censoring at time of kidney transplantation. Cox regression was used to compare risk of MACE between dialysis modalities with adjustment for age, gender, primary renal disease, prior comorbidities (coronary heart disease, left ventricular hypertrophy, heart failure and stroke), and laboratory data (plasma albumin, phosphate and ionized calcium). Imputation was performed to replace missing values of comorbidities and laboratory data. Results Of all 968 patients, 195 (20%) experienced a MACE during the entire follow-up and 62 (6.4%) during the first year of follow-up. The cumulative incidence of first MACE was similar in in-center HD and CAPD patients and higher than that in APD and home HD patients (Figure 1). Without adjustments, the hazard ratio of MACE was 0.83 [95% CI 0.56–1.2] for CAPD, 0.49 [95% CI 0.31–0.77] for APD and 0.42 [95% CI 0.23–0.78] for home HD in comparison to in-center HD. After adjustment for possible confounders, the hazard ratio of MACE was 1.1 [95% CI 0.70–1.6] for CAPD, 0.88 [95% CI 0.53–1.5] for APD and 0.80 [95% CI 0.41–1.6] for home HD and not statistically significantly different in comparison to in-center HD. Conclusion We observed a lower risk of MACE among patients who entered APD or home hemodialysis compared to in-center hemodialysis, but after adjusting for potential confounding factors, the difference diminished and was no longer statistically significant.

Obesity-induced metabolic imbalance allosterically modulates CtBP2 to inhibit PPAR-alpha transcriptional activity

Maintenance of metabolic homeostasis is secured by metabolite-sensing systems, which can be overwhelmed by constant macronutrient surplus in obesity. Not only the uptake processes but also the consumption of energy substrates determine the cellular metabolic burden. We herein describe a novel transcriptional system in this context comprised of peroxisome proliferator-activated receptor alpha (PPARα), a master regulator for fatty acid oxidation, and C-terminal binding protein 2 (CtBP2), a metabolite-sensing transcriptional co-repressor. CtBP2 interacts with PPARα to repress its activity, and the interaction is enhanced upon binding to malonyl-CoA, a metabolic intermediate increased in tissues in obesity and reported to suppress fatty acid oxidation through inhibition of carnitine palmitoyltransferase 1 (CPT1). In line with our preceding observations that CtBP2 adopts a monomeric configuration upon binding to acyl-CoAs, we determined that mutations in CtBP2 that shift the conformational equilibrium toward monomers increase the interaction between CtBP2 and PPARα. In contrast, metabolic manipulations that reduce malonyl-CoA decreased the formation of the CtBP2/PPARα complex. Consistent with these in vitro findings, we found that the CtBP2/PPARα interaction is accelerated in obese livers while genetic deletion of CtBP2 in the liver causes derepression of PPARα target genes. These findings support our model where CtBP2 exists primarily as a monomer in the metabolic milieu of obesity to repress PPARα, representing a liability in metabolic diseases that can be exploited to develop therapeutic approaches.

MOSUNETUZUMAB DEMONSTRATES DURABLE RESPONSES IN PATIENTS WITH RELAPSED/REFRACTORY FOLLICULAR LYMPHOMA AND ≥2 PRIOR THERAPIES: UPDATED ANALYSIS OF A PIVOTAL PHASE II STUDY

MOSUNETUZUMAB DEMONSTRATES DURABLE RESPONSES IN PATIENTS WITH RELAPSED/REFRACTORY FOLLICULAR LYMPHOMA AND ≥2 PRIOR THERAPIES: UPDATED ANALYSIS OF A PIVOTAL PHASE II STUDY

Differential dose-response patterns of intracellular and extracellular antibiotic resistance genes under sub-lethal antibiotic exposure

Although antibiotics are one of the most significant factors contributing to the propagation of antibiotic resistance genes (ARGs), studies on the dose-response relationship at sub-lethal concentrations of antibiotics remain scarce, despite their importance for assessing the risks of antibiotics in the environment. In this study, we constructed a series of microcosms to investigate the propagation of intracellular (iARGs) and extracellular (eARGs) ARGs in both water and biofilms when exposed to antibiotics at various concentrations (1–100 μg/L) and frequencies. Results showed that eARGs were more abundant than iARGs in water, while iARGs were the dominant ARGs form in biofilms. eARGs showed differentiated dose-response relationships from iARGs. The abundance of iARGs increased with the concentration of antibiotics as enhanced selective pressure overcame the metabolic burden of antibiotic-resistant bacteria carrying ARGs. However, the abundance of eARGs decreased with increasing antibiotic concentrations because less ARGs were secreted from bacterial hosts at higher concentrations (100 μg/L). Furthermore, combined exposure to two antibiotics (tetracycline & imipenem) showed a synergistic effect on the propagation of iARGs, but an antagonistic effect on the propagation of eARGs compared to exposure to a single antibiotic. When exposed to antibiotic at a fixed total dose, one-time dosing (1 time/10 d) favored the propagation of iARGs, while fractional dosing (5 times /10 d) favored the propagation of eARGs. This study sheds light on the propagation of antibiotic resistance in the environment and can help in assessing the risks associated with the use of antibiotics.

0006 Circadian Misalignment Increases the Impact of Visceral Adiposity on Metabolic Syndrome Burden in Adolescents

Abstract Introduction Although insufficient sleep is a known risk factor for metabolic syndrome (MetS), the circadian timing of sleep is also involved in cardiac and metabolic regulation. As a result, circadian misalignment of the sleep-wake cycle, which is highly prevalent in adolescents, may independently impact the burden of MetS in youth. Methods We analyzed data from 277 population-based randomly-selected adolescents from the Penn State Child Cohort (median 16 years; 47% female; 21% racial/ethnic minority) who had at least 5 nights of at-home actigraphy (ACT), an in-lab 9-h polysomnography (PSG) and an in-lab dual-energy X-ray absorptiometry (DEXA) scan. Two ACT-measured metrics of circadian misalignment were examined as effect modifiers: sleep midpoint (SM), the intra-individual mean midpoint of the sleep period, and sleep regularity (SR), the intra-individual standard deviation of sleep midpoint. DEXA-measured visceral adipose tissue (VAT) was the primary predictor. A continuous MetS score (i.e., sum of the age and sex adjusted z-scores of waist circumference, blood pressure, HOMA-IR, triglycerides, and HDL cholesterol) was the primary outcome. Linear regression models tested SM and SR as effect modifiers of the association of VAT with MetS, while accounting for sex, race/ethnicity, age, ACT-sleep duration, ACT-sleep variability, and PSG-apnea/hypopnea index. Results A significant interaction was found between VAT and SR on MetS (p-interaction=0.05), while not between VAT and SM (p-interaction=0.15). Among adolescents with high sleep irregularity (≥ 45 minutes; n=164), each standard deviation increase in VAT was associated with 2.4 (0.2) standard deviations increase in MetS (p&amp;lt; 0.01), while this association was weaker among adolescents with low sleep irregularity [&amp;lt; 45 minutes; n=113; β=1.7 (0.3), p&amp;lt; 0.01]. Conclusion An irregular circadian timing of sleep may further increase the impact of visceral adiposity on MetS burden in adolescents. Sleep irregularity, independent of sleep apnea and insufficient sleep, may contribute to the development of cardiometabolic sequelae associated with central obesity. Support (if any) National Institutes of Health (R01HL136587, UL1TR000127) and American Heart Association (23PRE1011962)

Predictive Value of Total Metabolic Tumor Burden Prior to Treatment in NSCLC Patients Treated with Immune Checkpoint Inhibition.

We aimed to assess the predictive value of the total metabolic tumor burden prior to treatment in patients with advanced non-small-cell lung cancer (NSCLC) receiving immune checkpoint inhibitors (ICIs). Pre-treatment 2-deoxy-2-[18F]fluoro-D-glucose positron emission tomography/computed tomography (PET/CT) scans performed in two consecutive years for staging in adult patients with confirmed NSCLC were considered. Volume, maximum/mean standardized uptake value (SUVmax/SUVmean), metabolic tumor volume (MTV) and total lesion glycolysis (TLG) were assessed per delineated malignant lesion (including primary tumor, regional lymph nodes and distant metastases) in addition to the morphology of the primary tumor and clinical data. Total metabolic tumor burden was captured by totalMTV and totalTLG. Overall survival (OS), progression-free survival (PFS) and clinical benefit (CB) were used as endpoints for response to treatment. A total of 125 NSCLC patients were included. Osseous metastases were the most frequent distant metastases (n = 17), followed by thoracal distant metastases (pulmonal = 14 and pleural = 13). Total metabolic tumor burden prior to treatment was significantly higher in patients treated with ICIs (mean totalMTV ± standard deviation (SD) 72.2 ± 78.7; mean totalTLG ± SD 462.2 ± 538.9) compared to those without ICI treatment (mean totalMTV ± SD 58.1 ± 233.8; mean totalTLG ± SD 290.0 ± 784.2). Among the patients who received ICIs, a solid morphology of the primary tumor on imaging prior to treatment was the strongest outcome predictor for OS (Hazard ratio HR 28.04, p < 0.01), PFS (HR 30.89, p < 0.01) and CB (parameter estimation PE 3.46, p < 0.01), followed by the metabolic features of the primary tumor. Interestingly, total metabolic tumor burden prior to immunotherapy showed a negligible impact on OS (p = 0.04) and PFS (p = 0.01) after treatment given the hazard ratios of 1.00, but also on CB (p = 0.01) given the PE < 0.01. Overall, biomarkers on pre-treatment PET/CT scans showed greater predictive power in patients receiving ICIs, compared to patients without ICI treatment. Morphological and metabolic properties of the primary tumors prior to treatment in advanced NSCLC patients treated with ICI showed great outcome prediction performances, as opposed to the pre-treatment total metabolic tumor burdens, captured by totalMTV and totalTLG, both with negligible impact on OS, PFS and CB. However, the outcome prediction performance of the total metabolic tumor burden might be influenced by the value itself (e.g., poorer prediction performance at very high or very low values of total metabolic tumor burden). Further studies including subgroup analysis with regards to different values of total metabolic tumor burden and their respective outcome prediction performances might be needed.

Repressing expression of difficult-to-express recombinant proteins during the selection process increases productivity of CHO stable pools.

More than half of licensed therapeutic recombinant proteins (r-proteins) are manufactured using constitutively-expressing, stably-transfected Chinese hamster ovary (CHO) clones. While constitutive CHO expression systems have proven their efficacy for the manufacturing of monoclonal antibodies, many next-generation therapeutics such as cytokines and bispecific antibodies as well as biological targets such as ectodomains of transmembrane receptors remain intrinsically challenging to produce. Herein, we exploited a cumate-inducible CHO platform allowing reduced expression of various classes of r-proteins during selection of stable pools. Following stable pool generation, fed-batch productions showed that pools generated without cumate (OFF-pools) were significantly more productive than pools selected in the presence of cumate (ON-pools) for 8out of the 10r-proteins tested, including cytokines, G-protein coupled receptors (GPCRs), the HVEM membrane receptor ectodomain, the multifunctional protein High Mobility Group protein B1 (HMGB1), as well as monoclonal and bispecific T-cell engager antibodies. We showed that OFF-pools contain a significantly larger proportion of cells producing high levels of r-proteins and that these cells tend to proliferate faster when expression is turned off, suggesting that r-protein overexpression imposes a metabolic burden on the cells. Cell viability was lower and pool recovery was delayed during selection of ON-pools (mimicking constitutive expression), suggesting that high producers were likely lost or overgrown by faster-growing, low-producing cells. We also observed a correlation between the expression levels of the GPCRs with Binding immunoglobulin Protein, an endoplasmic reticulum (ER) stress marker. Taken together, these data suggest that using an inducible system to minimize r-protein expression during stable CHO pool selection reduces cellular stresses, including ER stress and metabolic burden, leading to pools with greater frequency of high-expressing cells, resulting in improved volumetric productivity.