Nonlinear Mixed-Effects Model of Z-Endoxifen Concentrations in Tamoxifen-Treated Patients from the CEPAM Cohort.

Tamoxifen is widely used in patients with hormone receptor-positive breast cancer. The polymorphic enzyme CYP2D6 is primarily responsible for metabolic activation of tamoxifen, resulting in substantial interindividual variability of plasma concentrations of its most important metabolite, Z-endoxifen. The Z-endoxifen concentration thresholds below which tamoxifen treatment is less efficacious have been proposed but not validated, and prospective trials of individualized tamoxifen treatment to achieve Z-endoxifen concentration thresholds are considered infeasible. Therefore, we aim to validate the association between Z-endoxifen concentration and tamoxifen treatment outcomes, and identify a Z-endoxifen concentration threshold of tamoxifen efficacy, using pharmacometric modeling and simulation. As a first step, the CYP2D6 Endoxifen Percentage Activity Model (CEPAM) cohort was created by pooling data from 28 clinical studies (> 7,000 patients) with measured endoxifen plasma concentrations. After cleaning, data from 6,083 patients were used to develop a nonlinear mixed-effect (NLME) model for tamoxifen and Z-endoxifen pharmacokinetics that includes a conversion factor to allow inclusion of studies that measured total endoxifen but not Z-endoxifen. The final parent-metabolite NLME model confirmed the primary role of CYP2D6, and contributions from body weight, CYP2C9 phenotype, and co-medication with CYP2D6 inhibitors, on Z-endoxifen pharmacokinetics. Future work will use the model to simulate Z-endoxifen concentrations in patients receiving single agent tamoxifen treatment within large prospective clinical trials with long-term survival to identify the Z-endoxifen concentration threshold below which tamoxifen is less efficacious. Identification of this concentration threshold would allow personalized tamoxifen treatment to improve outcomes in patients with hormone receptor-positive breast cancer.

Effect of metformin and lifestyle intervention on adipokines and hormones in breast cancer survivors: a pooled analysis from two randomized controlled trials.

PURPOSE: We investigated the effect of metformin and lifestyle intervention on metabolic, inflammatory, and steroid biomarkers of breast cancer (BC) recurrence risk in two intervention trials among BC survivors with overweight or obesity. METHODS: Baseline and follow-up serum samples collected during the two trials were analyzed and data pooled. The USA trial (Reach for Health) included postmenopausal BC survivors (n = 333) randomly assigned to 6-month metformin vs placebo and lifestyle intervention (LSI) vs control (2 × 2 factorial design). The Italian trial (MetBreCS) included BC survivors (n = 40) randomized to 12-month metformin vs placebo. Insulin resistance (HOMA-IR), adipokines, cytokines, and steroids were measured. RESULTS: Metformin compared to placebo showed a favorable decrease in leptin (- 8.8 vs - 3.5 ng/mL; p < 0.01) and HOMA-IR (- 0.48 vs - 0.25; p = 0.03), and an increase in SHBG (2.80 vs 1.45 nmol/L; p < 0.01). Excluding women taking aromatase inhibitors, metformin (n = 84) compared to placebo (n = 99) decreased estradiol (- 4 vs 0 pmol/L; p < 0.01), estrone (- 8 vs 2 pmol/L; p < 0.01) and testosterone (- 0.1 vs 0 nmol/L-; p = 0.02). LSI favorably affected adiponectin (0.45 vs - 0.06 ug/mL; p < 0.01), leptin (- 10.5 vs - 4.4 ng/mL; p < 0.01), HOMA-IR (- 0.6 vs 0.2; p = 0.03), and SHBG (2.7 vs 1.1 nMol/L; p = 0.04) compared to controls. The strongest impact was observed combining metformin with LSI on adipokines, CRP, SHBG, and estrogens. CONCLUSIONS: Supportive healthy lifestyle programs combined with metformin to achieve maximal risk reduction among BC cancer survivors are recommended, especially for those with obesity in menopause.

Alternative dosing regimen of exemestane in a randomized presurgical trial: the role of obesity in biomarker modulation.

In a 3-arm presurgical trial, four-six weeks exemestane 25 mg three times/week (TIW) was non-inferior to 25 mg/day (QD) in suppressing circulating estradiol in postmenopausal women with ER-positive breast cancer. Since obesity may decrease exemestane efficacy, we analyzed changes in sex steroids, adipokines, Ki-67, and drug levels in relation to obesity. Postmenopausal women with early-stage ER-positive breast cancer were randomized to either exemestane 25 mg QD (n = 57), 25 mg TIW (n = 57), or 25 mg/week (QW, n = 62) for 4-6 weeks before breast surgery. Serum and tissue pre- and post-treatment biomarkers were stratified by body mass index (BMI)< or ≥30 kg/m2. Post-treatment median exemestane and 17-OH exemestane levels were 5-6 times higher in the QD arm compared to the TIW arm. For obese women, TIW maintained comparable reductions to QD in systemic estradiol levels, although the reduction in estrone was less with the TIW regimen. There was less suppression of SHBG with the TIW versus the QD dose schedule in obese women which should result in less systemic bioavailable estrogens. Metabolically, the effect of the TIW regimen was similar to the QD regimen for obese women in terms of leptin suppression and increase in the adiponectin-leptin ratio. Reduction in tissue Ki-67 was less for obese women on the TIW regimen than QD, although changes were similar for non-obese women. Our findings suggest that TIW exemestane should be explored further for primary cancer prevention in both normal weight and obese cohorts.

A randomized Phase I pre-operative window trial of transdermal endoxifen in women planning mastectomy: Evaluation of dermal safety, intra-mammary drug distribution, and biologic effects.

Breast cancer prevention only requires local exposure of the breast to active drug. However, oral preventive agents entail systemic exposure, causing adverse effects that limit acceptance by high-risk women. Drug-delivery through the breast skin is an attractive option, but requires demonstration of dermal safety and drug distribution throughout the breast. We formulated the tamoxifen metabolite (E/Z)-endoxifen for transdermal delivery and tested it in a placebo-controlled, double-blinded Phase I trial with dose escalation from 10 to 20 mg daily. The primary endpoint was dermal toxicity. Thirty-two women planning mastectomy were randomized (2:1) to endoxifen-gel or placebo-gel applied to both breasts for 3-5 weeks. Both doses of endoxifen-gel incurred no dermal or systemic toxicity compared to placebo. All endoxifen-treated breasts contained the drug at each of five sampling locations; the median per-person tissue concentration in the treated participants was 0.6 ng/g (IQR 0.4-1.6), significantly higher (p < 0.001) than the median plasma concentration (0.2 ng/mL, IQR 0.2-0.2). The median ratio of the more potent (Z)-isomer to (E)-isomer at each breast location was 1.50 (IQR 0.96-2.54, p < 0.05). No discernible effects of breast size or adiposity on tissue concentrations were observed. At the endoxifen doses and duration used, and the tissue concentration achieved, we observed a non-significant overall reduction of tumor proliferation (Ki67 LI) and significant downregulation of gene signatures known to promote cancer invasion (FN1, SERPINH1, PLOD2, PDGFA, ITGAV) (p = 0.03). Transdermal endoxifen is an important potential breast cancer prevention agent but formulations with better dermal penetration are needed.

Preoperative risk factors associated with left ventricular dysfunction after bariatric surgery.

A large proportion of patients with severe obesity remain with left ventricular (LV) dysfunction after bariatric surgery. We assessed whether preoperative evaluation by echocardiography and inflammatory proteins can identify this high-risk group. In the Bariatric Surgery on the West Coast of Norway study, 75 patients (44 ± 10 years, body mass index [BMI] 41.5 ± 4.7 kg/m2) were prospectively evaluated by echocardiography and inflammatory proteins (high-sensitivity C-reactive protein [hsCRP], serum amyloid A [SAA] and calprotectin) before and one year after Roux-en-Y gastric bypass surgery. LV mechanics was assessed by the midwall shortening (MWS) and global longitudinal strain (GLS). Bariatric surgery improved BMI and GLS, and lowered hsCRP, calprotectin and SAA (p < 0.05). MWS remained unchanged and 35% of patients had impaired MWS at 1-year follow-up. A preoperative risk index including sex, hypertension, ejection fraction (EF) and high hsCRP (index 1) or SAA (index 2) predicted low 1-year MWS with 81% sensitivity/71% specificity (index 1), and 77% sensitivity/77% specificity (index 2) in ROC analyses (AUC 0.80 and 0.79, p < 0.001). Among individuals with severe obesity, women and patients with hypertension, increased serum levels of inflammatory proteins and reduced EF are at high risk of impaired LV midwall mechanics 1 year after bariatric surgery.ClinicalTrials.gov identifier NCT01533142 February 15, 2012.

Presurgical Oral Tamoxifen vs Transdermal 4-Hydroxytamoxifen in Women With Ductal Carcinoma In Situ: A Randomized Clinical Trial.

Importance: Oral tamoxifen citrate benefits women with ductal carcinoma in situ (DCIS), but concern about toxic effects has limited acceptance. Previous pilot studies have suggested transdermal 4-hydroxytamoxifen gel has equivalent antiproliferative efficacy to oral tamoxifen, with low systemic exposure. Objective: To demonstrate that 4-hydroxytamoxifen gel applied to the breast skin is noninferior to oral tamoxifen in its antiproliferative effect in DCIS lesions. Design, Setting, and Participants: This randomized, double-blind, phase 2 preoperative window trial was performed at multicenter breast surgery referral practices from May 31, 2017, to January 27, 2021. Among 408 women with estrogen receptor-positive DCIS who were approached, 120 consented and 100 initiated study treatment. The most common reasons for nonparticipation were surgical delay, disinterest in research, and concerns about toxic effects. Data were analyzed from January 26, 2021, to October 5, 2022. Intervention: Random assignment to oral tamoxifen citrate, 20 mg/d, and gel placebo or 4-hydroxytamoxifen gel, 2 mg/d per breast, and oral placebo, for 4 to 10 weeks, followed by DCIS resection. Main Outcomes and Measures: The primary end point was absolute change in DCIS Ki-67 labeling index (Ki67-LI). Secondary end points included 12-gene DCIS Score, breast tissue tamoxifen metabolite concentrations, tamoxifen-responsive plasma protein levels, and patient-reported symptoms. Noninferiority of Ki67-LI reduction by 4-hydroxytamoxifen gel was tested using analysis of covariance; within- and between-arm comparisons were performed with paired t tests for mean values or the Wilcoxon rank sum test for medians. Results: Of 90 participants completing treatment (mean [SD] age, 55 [11] years; 8 [8.9%] Asian, 16 [17.8%] Black, 8 [8.9%] Latina, and 53 [58.9%] White), 15 lacked residual DCIS in the surgical sample, leaving 75 evaluable for the primary end point analysis (40 in the oral tamoxifen group and 35 in the 4-hydroxytamoxifen gel group). Posttreatment Ki67-LI was 3.3% higher (80% CI, 2.1%-4.6%) in the 4-hydroxytamoxifen gel group compared with the oral tamoxifen group, exceeding the noninferiority margin (2.6%). The DCIS Score decreased more with oral tamoxifen treatment (-16 [95% CI, -22 to -9.4]) than with 4-hydroxytamoxifen gel (-1.8 [95% CI, -5.8 to 2.3]). The median 4-hydroxytamoxifen concentrations deep in the breast were nonsignificantly higher in the oral tamoxifen group (5.7 [IQR, 4.0-7.9] vs 3.8 [IQR, 1.3-7.9] ng/g), whereas endoxifen was abundant in the oral tamoxifen group and minimal in the 4-hydroxytamoxifen gel group (median, 13.0 [IQR, 8.9-20.6] vs 0.3 [IQR, 0-0.3] ng/g; P < .001). Oral tamoxifen caused expected adverse changes in plasma protein levels and vasomotor symptoms, with minimal changes in the transdermal group. Conclusions and Relevance: In this randomized clinical trial, antiproliferative noninferiority of 4-hydroxytamoxifen gel to oral tamoxifen was not confirmed, potentially owing to endoxifen exposure differences. New transdermal approaches must deliver higher drug quantities and/or include the most potent metabolites. Trial Registration: ClinicalTrials.gov Identifier: NCT02993159.

High-fat diet impact on intestinal cholesterol conversion by the microbiota and serum cholesterol levels.

Cholesterol-to-coprostanol conversion by the intestinal microbiota has been suggested to reduce intestinal and serum cholesterol availability, but the relationship between intestinal cholesterol conversion and the gut microbiota, dietary habits, and serum lipids has not been characterized in detail. We measured conserved proportions of cholesterol high and low-converter types in individuals with and without obesity from two distinct, independent low-carbohydrate high-fat (LCHF) dietary intervention studies. Across both cohorts, cholesterol conversion increased in previous low-converters after LCHF diet and was positively correlated with the fecal relative abundance of Eubacterium coprostanoligenes. Lean cholesterol high-converters had increased serum triacylglycerides and decreased HDL-C levels before LCHF diet and responded to the intervention with increased LDL-C, independently of fat, cholesterol, and saturated fatty acid intake. Our findings identify the cholesterol high-converter type as a microbiome marker, which in metabolically healthy lean individuals is associated with increased LDL-C in response to LCHF.

Long-term impact of gastropexy on use of acid-reducing medication, second operations for gastroesophageal reflux and subjective reflux symptoms after sleeve gastrectomy.

We investigated whether adding gastropexy to sleeve gastrectomy (SG) reduced gastroesophageal reflux disease (GERD) in patients operated for severe obesity, assessed mainly by use of anti-reflux medication (ARM) and second operations due to GERD worsening. In a prospective non-randomized study, patients undergoing SG at two Norwegian hospitals were included from 2011 to 2015 and followed for 7 years. GERD was defined by regular use of ARM, and epigastric pain and heartburn were measured by the Rome II questionnaire. Gastropexy was done by suturing the gastrocolic ligament to the staple line. Patients undergoing SG only, mainly before gastropexia was introduced in 2013, were compared to those with additional gastropexy from 2013 onwards. Of 376 included patients (75% females, mean age 42.6 years and BMI 42.9 kg/m2 ), 350 (93%) and 232 (62%) were available for evaluation after 1 and 7 years, respectively. Baseline characteristics in the no-gastropexy (n = 235) and gastropexy groups (n = 141) were similar. In patients without ARM use before surgery, the use increased and in those that used ARM at baseline, the proportion decreased, with no difference in the no-gastropexy and gastropexy groups. With a combined endpoint of ARM use and/or second operation for GERD, there was no difference during follow-up between the two groups. With time, adding gastropexy did not reduce symptoms of GERD significantly. In this population, adding gastropexy to SG did not reduce use of ARM and/or second operation for uncontrolled GERD, epigastric pain or heartburn during the first 7 postoperative years.

Importance of endocrine treatment adherence and persistence in breast cancer survivorship: a systematic review.

PURPOSE: Adjuvant endocrine treatment is essential for treating luminal subtypes of breast cancer, which constitute 75% of all breast malignancies. However, the detrimental side effects of treatment make it difficult for many patients to complete the guideline-required treatment. Such non-adherence may jeopardize the lifesaving ability of anti-estrogen therapy. In this systematic review, we aimed to assess the consequences of non-adherence and non-persistence from available studies meeting strict statistical and clinical criteria. METHODS: A systematic literature search was performed using several databases, yielding identification of 2,026 studies. After strict selection, 14 studies were eligible for systematic review. The review included studies that examined endocrine treatment non-adherence (patients not taking treatment as prescribed) or non-persistence (patients stopping treatment prematurely), in terms of the effects on event-free survival or overall survival among women with non-metastatic breast cancer. RESULTS: We identified 10 studies measuring the effects of endocrine treatment non-adherence and non-persistence on event-free survival. Of these studies, seven showed significantly poorer survival for the non-adherent or non-persistent patient groups, with hazard ratios (HRs) ranging from 1.39 (95% CI, 1.07 to 1.53) to 2.44 (95% CI, 1.89 to 3.14). We identified nine studies measuring the effects of endocrine treatment non-adherence and non-persistence on overall survival. Of these studies, seven demonstrated significantly reduced overall survival in the groups with non-adherence and non-persistence, with HRs ranging from 1.26 (95% CI, 1.11 to 1.43) to 2.18 (95% CI, 1.99 to 2.39). CONCLUSION: The present systematic review demonstrates that non-adherence and non-persistence to endocrine treatment negatively affect event-free and overall survival. Improved follow-up, with focus on adherence and persistence, is vital for improving health outcomes among patients with non-metastatic breast cancer.

Sex-specific responses in glucose-insulin homeostasis and lipoprotein-lipid components after high-dose supplementation with marine n-3 PUFAs in abdominal obesity: a randomized double-blind crossover study.

Background: Clinical studies on effects of marine-derived omega-3 (n-3) polyunsaturated fatty acids (PUFAs), mainly eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), and the plant-derived omega-6 (n-6) PUFA linoleic acid (LA) on lipoprotein-lipid components and glucose-insulin homeostasis have shown conflicting results, which may partly be explained by differential responses in females and males. However, we have lacked data on sexual dimorphism in the response of cardiometabolic risk markers following increased consumption of n-3 or n-6 PUFAs. Objective: To explore sex-specific responses after n-3 (EPA + DHA) or n-6 (LA) PUFA supplementation on circulating lipoprotein subfractions, standard lipids, apolipoproteins, fatty acids in red blood cell membranes, and markers of glycemic control/insulin sensitivity among people with abdominal obesity. Methods: This was a randomized double-blind crossover study with two 7-week intervention periods separated by a 9-week washout phase. Females (n = 16) were supplemented with 3 g/d of EPA + DHA (fish oil) or 15 g/d of LA (safflower oil), while males (n = 23) received a dose of 4 g/d of EPA + DHA or 20 g/d of LA. In fasting blood samples, we measured lipoprotein particle subclasses, standard lipids, apolipoproteins, fatty acid profiles, and markers of glycemic control/insulin sensitivity. Results: The between-sex difference in relative change scores was significant after n-3 for total high-density lipoproteins (females/males: -11%*/-3.3%, p = 0.036; *: significant within-sex change), high-density lipoprotein particle size (+2.1%*/-0.1%, p = 0.045), and arachidonic acid (-8.3%*/-12%*, p = 0.012), and after n-6 for total (+37%*/+2.1%, p = 0.041) and small very-low-density lipoproteins (+97%*/+14%, p = 0.021), and lipoprotein (a) (-16%*/+0.1%, p = 0.028). Circulating markers of glucose-insulin homeostasis differed significantly after n-3 for glucose (females/males: -2.1%/+3.9%*, p = 0.029), insulin (-31%*/+16%, p < 0.001), insulin C-peptide (-12%*/+13%*, p = 0.001), homeostasis model assessment of insulin resistance index 2 (-12%*/+14%*, p = 0.001) and insulin sensitivity index 2 (+14%*/-12%*, p = 0.001), and quantitative insulin sensitivity check index (+4.9%*/-3.4%*, p < 0.001). Conclusion: We found sex-specific responses after high-dose n-3 (but not n-6) supplementation in circulating markers of glycemic control/insulin sensitivity, which improved in females but worsened in males. This may partly be related to the sex differences we observed in several components of the lipoprotein-lipid profile following the n-3 intervention. Clinical trial registration: https://clinicaltrials.gov/, identifier [NCT02647333].

Metabolic role of the hepatic valine/3-hydroxyisobutyrate (3-HIB) pathway in fatty liver disease.

BACKGROUND: The valine (branched-chain amino acid) metabolite 3-hydroxyisobutyrate (3-HIB), produced by 3-Hydroxyisobutyryl-CoA Hydrolase (HIBCH), is associated with insulin resistance and type 2 diabetes, but implicated tissues and cellular mechanisms are poorly understood. We hypothesized that HIBCH and 3-HIB regulate hepatic lipid accumulation. METHODS: HIBCH mRNA in human liver biopsies ("Liver cohort") and plasma 3-HIB ("CARBFUNC" cohort) were correlated with fatty liver and metabolic markers. Human Huh7 hepatocytes were supplemented with fatty acids (FAs) to induce lipid accumulation. Following HIBCH overexpression, siRNA knockdown, inhibition of PDK4 (a marker of FA ß-oxidation) or 3-HIB supplementation, we performed RNA-seq, Western blotting, targeted metabolite analyses and functional assays. FINDINGS: We identify a regulatory feedback loop between the valine/3-HIB pathway and PDK4 that shapes hepatic FA metabolism and metabolic health and responds to 3-HIB treatment of hepatocytes. HIBCH overexpression increased 3-HIB release and FA uptake, while knockdown increased cellular respiration and decreased reactive oxygen species (ROS) associated with metabolic shifts via PDK4 upregulation. Treatment with PDK4 inhibitor lowered 3-HIB release and increased FA uptake, while increasing HIBCH mRNA. Implicating this regulatory loop in fatty liver, human cohorts show positive correlations of liver fat with hepatic HIBCH and PDK4 expression (Liver cohort) and plasma 3-HIB (CARBFUNC cohort). Hepatocyte 3-HIB supplementation lowered HIBCH expression and FA uptake and increased cellular respiration and ROS. INTERPRETATION: These data implicate the hepatic valine/3-HIB pathway in mechanisms of fatty liver, reflected in increased plasma 3-HIB concentrations, and present possible targets for therapeutic intervention. FUNDING: Funding was provided by the Research Council of Norway (263124/F20), the University of Bergen, the Western Norway Health Authorities, Novo Nordisk Scandinavia AS, the Trond Mohn Foundation and the Norwegian Diabetes Association.

Obesity Is Associated with Distorted Proteoglycan Expression in Adipose Tissue.

Proteoglycans are central components of the extracellular matrix (ECM) and binding partners for inflammatory chemokines. Morphological differences in the ECM and increased inflammation are prominent features of the white adipose tissues in patients with obesity. The impact of obesity and weight loss on the expression of specific proteoglycans in adipose tissue is not well known. This study aimed to investigate the relationship between adiposity and proteoglycan expression. We analyzed transcriptomic data from two human bariatric surgery cohorts. In addition, RT-qPCR was performed on adipose tissues from female and male mice fed a high-fat diet. Both visceral and subcutaneous adipose tissue depots were analyzed. Adipose mRNA expression of specific proteoglycans, proteoglycan biosynthetic enzymes, proteoglycan partner molecules, and other ECM-related proteins were altered in both human cohorts. We consistently observed more profound alterations in gene expression of ECM targets in the visceral adipose tissues after surgery (among others VCAN (p = 0.000309), OGN (p = 0.000976), GPC4 (p = 0.00525), COL1A1 (p = 0.00221)). Further, gene analyses in mice revealed sex differences in these two tissue compartments in obese mice. We suggest that adipose tissue repair is still in progress long after surgery, which may reflect challenges in remodeling increased adipose tissues. This study can provide the basis for more mechanistic studies on the role of proteoglycans in adipose tissues in obesity.

Relationship between Ketones, Ghrelin, and, Appetite on Isocaloric Diets with Varying Carbohydrate Quality and Amount: Results from a Randomized Controlled Trial in People with Obesity (CARBFUNC).

BACKGROUND: Low-carbohydrate high-fat (LCHF) diets may suppress the increase in appetite otherwise seen after diet-induced fat loss. However, studies of diets without severe energy restriction are lacking, and the effects of carbohydrate quality relative to quantity have not been directly compared. OBJECTIVES: To evaluated short- (3 mo) and long-term (12 mo) changes in fasting plasma concentrations of total ghrelin, ß-hydroxybutyrate (ßHB), and subjective feelings of appetite on 3 isocaloric eating patterns within a moderate caloric range (2000-2500 kcal/d) and with varying carbohydrate quality or quantity. METHODS: We performed a randomized controlled trial of 193 adults with obesity, comparing eating patterns based on "acellular" carbohydrate sources (e.g., flour-based whole-grain products; comparator arm), "cellular" carbohydrate sources (minimally processed foods with intact cellular structures), or LCHF principles. Outcomes were compared by an intention-to-treat analysis using constrained linear mixed modeling. This trial was registered at clinicaltrials.gov as NCT03401970. RESULTS: Of the 193 adults, 118 (61%) and 57 (30%) completed 3 and 12 mo of follow-up. Throughout the intervention, intakes of protein and energy were similar with all 3 eating patterns, with comparable reductions in body weight (5%-7%) and visceral fat volume (12%-17%) after 12 mo. After 3 mo, ghrelin increased significantly with the acellular (mean: 46 pg/mL; 95% CI: 11, 81) and cellular (mean: 54 pg/mL; 95% CI: 21, 88) diets but not with the LCHF diet (mean: 11 pg/mL; 95% CI: -16, 38). Although ßHB increased significantly more with the LCHF diet than with the acellular diet after 3 m (mean: 0.16 mmol/L; 95% CI: 0.09, 0.24), this did not correspond to a significant group difference in ghrelin (unless the 2 high-carbohydrate groups were combined [mean: -39.6 pg/mL; 95% CI: -76, -3.3]). No significant between-group differences were seen in feelings of hunger. CONCLUSIONS: Modestly energy-restricted isocaloric diets differing in carbohydrate cellularity and amount showed no significant differences in fasting total ghrelin or subjective hunger feelings. An increase in ketones with the LCHF diet to 0.3-0.4 mmol/L was insufficient to substantially curb increases in fasting ghrelin during fat loss.

Dietary fish intake increased the concentration of soluble ACE2 in rats: can fish consumption reduce the risk of COVID-19 infection through interception of SARS-CoV-2 by soluble ACE2?

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) enters the cells after binding to the membrane-bound receptor angiotensin-converting enzyme 2 (ACE2), but this may be prevented through interception by soluble ACE2 (sACE2) or by inhibition of the ACE2 receptor, thus obstructing cell entry and replication. The main objective of this study was to investigate if fish intake affected the concentration of sACE2 in rats. The secondary aim was to evaluate the in vitro ACE2-inhibiting activity of fish proteins. Rats were fed cod muscle as 25 % of dietary protein, and blood was collected after 4 weeks of intervention. Muscle, backbone, skin, head, stomach, stomach content, intestine and swim bladder from haddock, saithe, cod and redfish were hydrolysed with trypsin before ACE2-inhibiting activity was measured in vitro. In vivo data were compared using unpaired Student's t test, and in vitro data were compared using one-way ANOVA followed by the Tukey HSD post hoc test. The mean sACE2 concentration was 47 % higher in rats fed cod when compared with control rats (P 0·034), whereas serum concentrations of angiotensin II and TNF-α were similar between the two experimental groups. Muscle, backbone, skin and head from all four fish species inhibited ACE2 activity in vitro, whereas the remaining fractions had no effect. To conclude, our novel data demonstrate that fish intake increased the sACE2 concentration in rats and that the hydrolysed fish proteins inhibited ACE2 activity in vitro.

Impaired Adipocyte SLC7A10 Promotes Lipid Storage in Association With Insulin Resistance and Altered BCAA Metabolism.

CONTEXT: The neutral amino acid transporter SLC7A10/ASC-1 is an adipocyte-expressed gene with reduced expression in insulin resistance and obesity. Inhibition of SLC7A10 in adipocytes was shown to increase lipid accumulation despite decreasing insulin-stimulated uptake of glucose, a key substrate for de novo lipogenesis. These data imply that alternative lipogenic substrates to glucose fuel continued lipid accumulation during insulin resistance in obesity. OBJECTIVE: We examined whether increased lipid accumulation during insulin resistance in adipocytes may involve alter flux of lipogenic amino acids dependent on SLC7A10 expression and activity, and whether this is reflected by extracellular and circulating concentrations of marker metabolites. METHODS: In adipocyte cultures with impaired SLC7A10, we performed RNA sequencing and relevant functional assays. By targeted metabolite analyses (GC-MS/MS), flux of all amino acids and selected metabolites were measured in human and mouse adipose cultures. Additionally, SLC7A10 mRNA levels in human subcutaneous adipose tissue (SAT) were correlated to candidate metabolites and adiposity phenotypes in 2 independent cohorts. RESULTS: SLC7A10 impairment altered expression of genes related to metabolic processes, including branched-chain amino acid (BCAA) catabolism, lipogenesis, and glyceroneogenesis. In 3T3-L1 adipocytes, SLC7A10 inhibition increased fatty acid uptake and cellular content of glycerol and cholesterol. SLC7A10 impairment in SAT cultures altered uptake of aspartate and glutamate, and increased net uptake of BCAAs, while increasing the net release of the valine catabolite 3- hydroxyisobutyrate (3-HIB). In human cohorts, SLC7A10 mRNA correlated inversely with total fat mass, circulating triacylglycerols, BCAAs, and 3-HIB. CONCLUSION: Reduced SLC7A10 activity strongly affects flux of BCAAs in adipocytes, which may fuel continued lipogenesis during insulin resistance, and be reflected in increased circulating levels of the valine-derived catabolite 3-HIB.

Phenotypic diversity of human adipose tissue-resident NK cells in obesity.

Natural killer (NK) cells have emerged as key mediators of obesity-related adipose tissue inflammation. However, the phenotype of NK cell subsets residing in human adipose tissue are poorly defined, preventing a detailed understanding of their role in metabolic disorders. In this study, we applied multicolor flow cytometry to characterize CD56bright and CD56dim NK cells in blood and adipose tissue depots in individuals with obesity and identified surface proteins enriched on adipose tissue-resident CD56bright NK cells. Particularly, we found that adipose tissue harbored clusters of tissue-resident CD56bright NK cells signatured by the expression of CD26, CCR5 and CD63, possibly reflecting an adaptation to the microenvironment. Together, our findings provide broad insights into the identity of NK cells in blood and adipose tissue in relation to obesity.

Efficacy of Alternative Dose Regimens of Exemestane in Postmenopausal Women With Stage 0 to II Estrogen Receptor-Positive Breast Cancer: A Randomized Clinical Trial.

Importance: Successful therapeutic cancer prevention requires definition of the minimal effective dose. Aromatase inhibitors decrease breast cancer incidence in high-risk women, but use in prevention and compliance in adjuvant settings are hampered by adverse events. Objective: To compare the noninferiority percentage change of estradiol in postmenopausal women with estrogen receptor-positive breast cancer given exemestane, 25 mg, 3 times weekly or once weekly vs a standard daily dose with a noninferiority margin of -6%. Design, Setting, and Participants: This multicenter, presurgical, double-blind phase 2b randomized clinical trial evaluated 2 alternative dosing schedules of exemestane. Postmenopausal women with estrogen receptor-positive breast cancer who were candidates for breast surgery were screened from February 1, 2017, to August 31, 2019. Blood samples were collected at baseline and final visit; tissue biomarker changes were assessed from diagnostic biopsy and surgical specimen. Biomarkers were measured in different laboratories between April 2020 and December 2021. Interventions: Exemestane, 25 mg, once daily, 3 times weekly, or once weekly for 4 to 6 weeks before surgery. Main Outcomes and Measures: Serum estradiol concentrations were measured by solid-phase extraction followed by liquid chromatography-tandem mass spectrometry detection. Toxic effects were evaluated using the National Cancer Institute terminology criteria, and Ki-67 was assessed by immunohistochemistry. Results: A total of 180 women were randomized into 1 of the 3 arms; median (IQR) age was 66 (60-71) years, 63 (60-69) years, and 65 (61-70) years in the once-daily, 3-times-weekly, and once-weekly arms, respectively. In the intention-to-treat population (n = 171), the least square mean percentage change of serum estradiol was -89%, -85%, and -60% for exemestane once daily (n = 55), 3 times weekly (n = 56), and once weekly (n = 60), respectively. The difference in estradiol percentage change between the once-daily and 3-times-weekly arms was -3.6% (P for noninferiority = .37), whereas in compliant participants (n = 153), it was 2.0% (97.5% lower confidence limit, -5.6%; P for noninferiority = .02). Among secondary end points, Ki-67 and progesterone receptor were reduced in all arms, with median absolute percentage changes of -7.5%, -5.0%, and -4.0% for Ki-67 in the once-daily, 3-times-weekly, and once-weekly arms, respectively (once daily vs 3 times weekly, P = .31; once daily vs once weekly, P = .06), and -17.0%, -9.0%, and -7.0% for progesterone receptor, respectively. Sex hormone-binding globulin and high-density lipoprotein cholesterol had a better profile among participants in the 3-times-weekly arm compared with once-daily arm. Adverse events were similar in all arms. Conclusions and Relevance: In this randomized clinical trial, exemestane, 25 mg, given 3 times weekly in compliant patients was noninferior to the once-daily dosage in decreasing serum estradiol. This new schedule should be further studied in prevention studies and in women who do not tolerate the daily dose in the adjuvant setting. Trial Registration: ClinicalTrials.gov Identifier: NCT02598557; EudraCT: 2015-005063-16.

Serum concentrations of amino acids and tryptophan metabolites are affected by consumption of a light breakfast: a clinical intervention study in adults with overweight or obesity.

BACKGROUND: Epidemiological studies often investigate amino acids and their metabolites as biomarkers, but do not always consistently use fasting or non-fasting blood samples, or may lack information on the prandial status of the study participants. Since little information is available on the effects of the prandial status on many biomarkers, and since blood is typically sampled early in the day with participants in a fasting state or after having consumed a light meal in many trials, the main purpose of this study was to investigate the short-term effects of a light breakfast on serum concentrations of amino acids and related metabolites. METHODS: Blood was collected from sixty-three healthy adults (36 women) in the fasting state and at set times for 120 min after intake of a light breakfast with low protein content (14 g protein, 2218 kJ). Relative changes in serum biomarker concentrations from fasting to postprandial serum concentrations were tested using T test. RESULTS: The serum concentrations of 13 of the 20 measured amino acids were significantly changed 60 min following breakfast intake, with the most marked effects seen as increases in alanine (34%) and proline (45%) concentrations. The response did not reflect the amino acid composition of the breakfast. The concentrations of seven kynurenine metabolites were significantly decreased after breakfast. CONCLUSION: Consumption of a light breakfast affected serum concentrations of several amino acids and related metabolites, underlining the importance of having information regarding the participants' prandial state at the time of blood sampling in studies including these biomarkers. TRIAL REGISTRATION: This trial was registered at clinicaltrials.gov as NCT02350595 (registered January 2015).

Plasma methylmalonic acid predicts risk of acute myocardial infarction and mortality in patients with coronary heart disease: A prospective 2-cohort study.

BACKGROUND: Elevated plasma methylmalonic acid (MMA) is reported in patients with established coronary heart disease (CHD) and is considered a marker of vitamin B12 deficiency. Moreover, MMA-dependent reactions have been linked to alterations in mitochondrial energy metabolism and oxidative stress, key features in the pathophysiology of cardiovascular diseases (CVDs). OBJECTIVES: We examined whether plasma MMA prospectively predicted the long-term risk of acute myocardial infarction (AMI) and mortality. METHODS AND RESULTS: Using Cox modeling, we estimated hazard ratios (HRs) for endpoints according to per 1-SD increment of log-transformed plasma MMA in two independent populations: the Western Norway Coronary Angiography Cohort (WECAC) (patients evaluated for CHD; n = 4137) and the Norwegian Vitamin Trial (NORVIT) (patients hospitalized with AMI; n = 3525). In WECAC and NORVIT, 12.8% and 18.0% experienced an AMI, whereas 21.8% and 19.9% died, of whom 45.5% and 60.3% from CVD-related causes during follow-up (range 3-11 years), respectively. In WECAC, age- and gender-adjusted HRs (95% confidence interval) were 1.18 (1.09-1.28), 1.25 (1.18-1.33), and 1.28 (1.17-1.40) for future AMI, total mortality, and CVD mortality, respectively. Corresponding risk estimates were 1.19 (1.10-1.28), 1.22 (1.14-1.31), and 1.30 (1.19-1.42) in NORVIT. These estimates were only slightly attenuated after multivariable adjustments. Across both cohorts, the MMA-risk association was stronger in older adults, women, and non-smokers. CONCLUSIONS: Elevated MMA was associated with an increased risk of AMI and mortality in patients with suspected or verified CHD.

Distinct T cell subsets in adipose tissue are associated with obesity.

Adipose tissue inflammation is a driving factor for the development of obesity-associated metabolic disturbances, and a role of adipose tissue T cells in initiating the pro-inflammatory signaling is emerging. However, data on human adipose tissue T cells in obesity are limited, reflected by the lack of phenotypic markers to define tissue-resident T cell subsets. In this study, we performed a deep characterization of T cells in blood and adipose tissue depots using multicolor flow cytometry and RNA sequencing. We identified distinct subsets of T cells associated with obesity expressing the activation markers, CD26 and CCR5, and obesity-specific genes that are potentially engaged in activating pro-inflammatory pathway, including ceramide signaling, autophagy, and IL-6 signaling. These findings increase our knowledge on the heterogeneity of T cells in adipose tissue and on subsets that may play a role in obesity-related pathogenesis.