Osteosarcoma tumors maintain intra-tumoral transcriptional heterogeneity during bone and lung colonization.

BACKGROUND: Tumors are complex tissues containing collections of phenotypically diverse malignant and nonmalignant cells. We know little of the mechanisms that govern heterogeneity of tumor cells nor of the role heterogeneity plays in overcoming stresses, such as adaptation to different microenvironments. Osteosarcoma is an ideal model for studying these mechanisms-it exhibits widespread inter- and intra-tumoral heterogeneity, predictable patterns of metastasis, and a lack of clear targetable driver mutations. Understanding the processes that facilitate adaptation to primary and metastatic microenvironments could inform the development of therapeutic targeting strategies. RESULTS: We investigated single-cell RNA-sequencing profiles of 47,977 cells obtained from cell line and patient-derived xenograft models as cells adapted to growth within primary bone and metastatic lung environments. Tumor cells maintained phenotypic heterogeneity as they responded to the selective pressures imposed during bone and lung colonization. Heterogenous subsets of cells defined by distinct transcriptional profiles were maintained within bone- and lung-colonizing tumors, despite high-level selection. One prominent heterogenous feature involving glucose metabolism was clearly validated using immunofluorescence staining. Finally, using concurrent lineage tracing and single-cell transcriptomics, we found that lung colonization enriches for multiple clones with distinct transcriptional profiles that are preserved across cellular generations. CONCLUSIONS: Response to environmental stressors occurs through complex and dynamic phenotypic adaptations. Heterogeneity is maintained, even in conditions that enforce clonal selection. These findings likely reflect the influences of developmental processes promoting diversification of tumor cell subpopulations, which are retained, even in the face of selective pressures.

BMI metrics and their association with adiposity, cardiometabolic risk factors, and biomarkers in children and adolescents.

BACKGROUND: There are limited data comparing the relative associations of various BMI metrics with adiposity and cardiometabolic risk factors in youth. OBJECTIVE: Examine correlations of 7 different BMI metrics with adiposity, cardiometabolic risk factors, and biomarkers (i.e. blood pressure, waist circumference, cholesterol, leptin, insulin, high molecular weight adiponectin, high-sensitivity c-reactive protein (hsCRP)). METHODS: This was a cross-sectional analysis of youth in all BMI categories. BMI metrics: BMI z-score (BMIz), extended BMIz (ext.BMIz), BMI percentile (BMIp), percent of the BMI 95th percentile (%BMIp95), percent of the BMI median (%BMIp50), triponderal mass index (TMI), and BMI (BMI). Correlations between these BMI metrics and adiposity, visceral adiposity, cardiometabolic risk factors and biomarkers were summarized using Pearson's correlations. RESULTS: Data from 371 children and adolescents ages 8-21 years old were included in our analysis: 52% were female; 20.2% with Class I obesity, 20.5% with Class II, and 14.3% with Class III obesity. BMIp consistently demonstrated lower correlations with adiposity, risk factors, and biomarkers (r = 0.190-0.768) than other BMI metrics. The %BMIp95 and %BMIp50 were marginally more strongly correlated with measures of adiposity as compared to other BMI metrics. The ext.BMIz did not meaningfully outperform BMIz. CONCLUSION: Out of all the BMI metrics evaluated, %BMIp95 and %BMIp50 were the most strongly correlated with measures of adiposity. %BMIp95 has the benefit of being used currently to define obesity and severe obesity in both clinical and research settings. BMIp consistently had the lowest correlations. Future research should evaluate the longitudinal stability of various BMI metrics and their relative associations with medium to long-term changes in adiposity and cardiometabolic outcomes in the context of intervention trials.

Hypoxia-Inducible Factor α Subunits Regulate Tie2-Expressing Macrophages That Influence Tumor Oxygen and Perfusion in Murine Breast Cancer.

Tie2-expressing monocytes/macrophages (TEMs) are a distinct subset of proangiogenic monocytes selectively recruited to tumors in breast cancer. Because of the hypoxic nature of solid tumors, we investigated if oxygen, via hypoxia-inducible transcription factors HIF-1α and HIF-2α, regulates TEM function in the hypoxic tumor microenvironment. We orthotopically implanted PyMT breast tumor cells into the mammary fat pads of syngeneic LysMcre, HIF-1α fl/fl /LysMcre, or HIF-2α fl/fl /LysMcre mice and evaluated the tumor TEM population. There was no difference in the percentage of tumor macrophages among the mouse groups. In contrast, HIF-1α fl/fl /LysMcre mice had a significantly smaller percentage of tumor TEMs compared with control and HIF-2α fl/fl /LysMcre mice. Proangiogenic TEMs in macrophage HIF-2α-deficient tumors presented significantly more CD31+ microvessel density but exacerbated hypoxia and tissue necrosis. Reduced numbers of proangiogenic TEMs in macrophage HIF-1α-deficient tumors presented significantly less microvessel density but tumor vessels that were more functional as lectin injection revealed more perfusion, and functional electron paramagnetic resonance analysis revealed more oxygen in those tumors. Macrophage HIF-1α-deficient tumors also responded significantly to chemotherapy. These data introduce a previously undescribed and counterintuitive prohypoxia role for proangiogenic TEMs in breast cancer which is, in part, suppressed by HIF-2α.

Validity of the Adult Eating Behavior Questionnaire for adolescents treated in a weight management clinic.

BACKGROUND: The Child and Adult Eating Behavior Questionnaires (CEBQ, AEBQ) are established measures of eating behaviors. However, no similar measure is available for adolescents. Prior research has validated the AEBQ in adult samples, and one study has explored using the measure with adolescents. However, no studies to date have examined the validity of the AEBQ in adolescent clinical populations. Furthermore, no studies have examined associations between the AEBQ and indicators of health status in adolescents. METHODS: A total of 280 adolescents (12-17 years old, 60% female) seen in a pediatric weight management clinic completed the AEBQ at intake. Confirmatory factor analysis (CFA) was conducted with AEBQ items to evaluate the model fit of one-, two-, seven-, and eight-factor structures. Intercorrelations between scale scores from AEBQ Food Approach and Food Avoidance domains were calculated. Associations of AEBQ scales with body mass index (BMI) and binge-eating behaviors were examined using Spearman Rho correlations and independent t-tests. RESULTS: CFAs revealed that the best fitting model was a seven-factor structure excluding the Hunger scale, although overall model fit was only marginally acceptable (X2 = 980.94, CFI = 0.925, TLI = 0.915, RMSEA = 0.074). Intercorrelation analyses indicated that all Food Approach scales were significantly associated with one another (r = 0.243-0.654); Food Avoidance scales were inconsistently correlated (r = 0.034-0.439). No AEBQ scales were correlated with BMI (r = -0.101-0.082). Stronger links were found with binge eating; higher frequency binge-related behaviors were associated with higher Food Approach scores. CONCLUSIONS: The seven-factor structure of AEBQ demonstrates a marginally acceptable fit for treatment-seeking adolescents with obesity. The Food Approach scales demonstrated more convergent validity than the Food Avoidance scales. The Food Approach scales also exhibited some clinical utility for identifying patients with increased risk for binge eating, which is a common target for behavioral intervention. Implications for maximizing the AEBQ's potential for assessing eating behaviors in adolescents with obesity are discussed.

Predictive Value of Developmental Assessment in a Neonatal Intensive Care Unit (NICU) Follow-Up Clinic.

OBJECTIVE: Neonatal Intensive Care Unit (NICU) Follow-Up programs vary in the duration for which they monitor child development and neurocognitive outcomes. This study explores the early predictive value of a widely used developmental measure for intellectual functioning during early childhood to better inform whether there is value added in continued monitoring. METHODS: Participants were 209 children who had at least two assessments between the ages of 1 and 6 years old as part of NICU Follow-Up clinic. The Bayley Scales of Infant and Toddler Development, Third Edition (Bayley-III) was administered when children were 1 and 2 years old and the Wechsler Preschool and Primary Scale of Intelligence, Fourth Edition (WPPSI-IV) was administered when children were 3 years and older. RESULTS: The Bayley-III at 1 year of age was a significant predictor of Bayley-III performance at age 2. Similarly, Bayley-III at ages 1 year and 2 years were significant predictors of WPPSI-IV performance. Strength of prediction was moderate with the majority of variance unexplained. Exploratory analyses examining whether early developmental abilities as assessed on the Bayley-III could identify patients at risk for poorer WPPSI-IV performance indicated appropriate specificity but inadequate sensitivity. CONCLUSIONS: This study supports ongoing assessment of children who were born with perinatal complications into at least early childhood. Assessing development only during the infant and toddler years did not sufficiently identify children who went on to have lower cognitive functioning in preschool and the early school years.

GD2-directed CAR-T cells in combination with HGF-targeted neutralizing antibody (AMG102) prevent primary tumor growth and metastasis in Ewing sarcoma.

Ewing sarcoma (EWS) is the second most common and aggressive type of metastatic bone tumor in adolescents and young adults. There is unmet medical need to develop and test novel pharmacological targets and novel therapies to treat EWS. Here, we found that EWS expresses high levels of a p53 isoform, delta133p53. We further determined that aberrant expression of delta133p53 induced HGF secretion resulting in tumor growth and metastasis. Thereafter, we evaluated targeting EWS tumors with HGF receptor neutralizing antibody (AMG102) in preclinical studies. Surprisingly, we found that targeting EWS tumors with HGF receptor neutralizing antibody (AMG102) in combination with GD2-specific, CAR-reengineered T-cell therapy synergistically inhibited primary tumor growth and establishment of metastatic disease in preclinical models. Furthermore, our data suggested that AMG102 treatment alone might increase leukocyte infiltration including efficient CAR-T access into tumor mass and thereby improves its antitumor activity. Together, our findings warrant the development of novel CAR-T-cell therapies that incorporate HGF receptor neutralizing antibody to improve therapeutic potency, not only in EWS but also in tumors with aberrant activation of the HGF/c-MET pathway.

Adverse Childhood Experiences and Weight Status among Adolescents.

OBJECTIVE: To investigate the relationship between adverse childhood experiences (ACEs) and weight status among adolescents. STUDY DESIGN: Data were drawn from the Minnesota Student Survey, a large (n = 105 759), statewide, anonymous survey of public school students in eighth, ninth, and eleventh grades. Self-reported height and weight were used to calculate body mass index. Multinomial logistic regression was used to examine associations between self-reported ACEs and weight status, controlling for key sociodemographic characteristics. RESULTS: ACEs were positively associated with weight status; adolescents with more ACEs were more likely to have overweight, obesity, and severe obesity than adolescents with no ACEs. Adolescents who reported an ACE were 1.2, 1.4, and 1.5 times as likely to have overweight, obesity, and severe obesity, respectively, compared with their peers with no ACEs. There was no relationship between ACEs and underweight. CONCLUSIONS: The results of this large sample of adolescents with anonymous data support the hypothesis that ACEs and obesity are strongly associated. The directionality of this relationship needs to be understood. Moreover, these findings suggest that child health professionals may need to screen for ACEs as an important aspect of clinical weight management.

Relationships of Anxiety and Depression with Cardiovascular Health in Youth with Normal Weight to Severe Obesity.

OBJECTIVE: To evaluate the relationships of depression and anxiety symptoms with cardiovascular disease (CVD) risk factors and measures of vascular health in youth. STUDY DESIGN: Participants (n = 202) were 8- to 18-year-olds from a cross-sectional study evaluating cardiovascular health across a wide range of body mass index values (normal weight to severe obesity). CVD risk measurement included blood pressure, fasting lipids, glucose, insulin, carotid artery intima-media thickness, compliance and distensibility, brachial artery flow-mediated dilation, carotid-radial artery pulse wave velocity, body fat percentage, and a metabolic syndrome cluster score. Anxiety and depression symptoms were self-reported on the Screen for Child Anxiety Related Disorders and Center for Epidemiological Studies Depression Scale for Children. Two sets of adjustment variables were used in evaluation of differences between those with and without anxiety or depression symptomatology for the CVD risk factor and vascular outcomes. The first set included adjustment for Tanner stage, sex, and race; the second was additionally adjusted for percent body fat. RESULTS: Anxiety was not significantly associated with CVD risk factors or vascular health in either model. Depression was associated with high-density lipoprotein cholesterol, triglycerides, and metabolic syndrome cluster score; these relationships were attenuated when accounting for percent body fat. CONCLUSIONS: When accounting for body fat, we found no clear relationship of self-reported depression or anxiety symptoms with CVD risk factors or vascular health in youth.

The Effect of Platform Switching on Periimplant Crevicular Fluid Content During Early Wound Healing.

PURPOSE: The objective of this study was to investigate the soft tissue response and periimplant crevicular fluid (PICF) content around platform-switched (PS) and platform-matched (PM) implants during early healing. MATERIALS AND METHODS: Nonsmokers treatment planned to receive a single implant in 2 quadrants were recruited. Two-stage implant placement protocol with 1 PM and 1 PS implant was implemented. Periimplant probing depths (PDs), modified sulcus bleeding index, and plaque indices were recorded, and PICF was collected at 1, 2, 4, and 6 weeks after abutment connection. RESULTS: PD readings were higher at week 1 than at week 6 for both groups (P = 0.0005). PD was statistically deeper in PM than in PS at week 1 (P = 0.03). There was a time-dependent decrease in total PICF volume for both groups. This decrease was statistically significant for PS (P = 0.0005), with no differences between the 2 groups at any time (P > 0.05). The decrease observed in both PM and PS for PICF interleukin 6 and macrophage inflammatory protein-1ß, and in PS for tumor necrosis factor-α (TNF-α) was statistically significant (P ≤ 0.03). TNF-α was statistically higher in PS than in PM at week 1 (P = 0.005). CONCLUSION: Within the limits of this study, it seems that periimplant soft tissue response around PM and PS implants is mostly similar during the early healing period.

New class of 8-aryl-7-deazaguanine cell permeable fluorescent probes.

A one step synthesis of fluorescent 8-aryl-(7-deazaguanines) has been accomplished. Probes exhibit blue to green high quantum yield fluorescence in a variety of organic and aqueous solutions, high extinction coefficients, and large Stokes shifts often above 100 nm. The probes are highly cell permeable, and exhibit stable bright fluorescence once intracellular; therefore are suited to the design of biosensors.

Preferential, enhanced breast cancer cell migration on biomimetic electrospun nanofiber 'cell highways'.

BACKGROUND: Aggressive metastatic breast cancer cells seemingly evade surgical resection and current therapies, leading to colonization in distant organs and tissues and poor patient prognosis. Therefore, high-throughput in vitro tools allowing rapid, accurate, and novel anti-metastatic drug screening are grossly overdue. Conversely, aligned nanofiber constitutes a prominent component of the late-stage breast tumor margin extracellular matrix. This parallel suggests that the use of a synthetic ECM in the form of a nanoscale model could provide a convenient means of testing the migration potentials of cancer cells to achieve a long-term goal of providing clinicians an in vitro platform technology to test the efficacy of novel experimental anti-metastatic compounds. METHODS: Electrospinning produces highly aligned, cell-adhesive nanofiber matrices by applying a strong electric field to a polymer-containing solution. The resulting fibrous microstructure and morphology closely resembles in vivo tumor microenvironments suggesting their use in analysis of migratory potentials of metastatic cancer cells. Additionally, a novel interface with a gel-based delivery system creates CXCL12 chemotactic gradients to enhance CXCR4-expressing cell migration. RESULTS: Cellular dispersions of MCF-10A normal mammary epithelial cells or human breast cancer cells (MCF-7 and MDA-MB-231) seeded on randomly-oriented nanofiber exhibited no significant differences in total or net distance traveled as a result of the underlying topography. Cells traveled ~2-5 fold greater distances on aligned fiber. Highly-sensitive MDA-MB-231 cells displayed an 82% increase in net distance traversed in the presence of a CXCL12 gradient. In contrast, MCF-7 cells exhibited only 31% increase and MCF-10A cells showed no statistical difference versus control or vehicle conditions. MCF-10A cells displayed little sensitivity to CXCL12 gradients, while MCF-7 cells displayed early sensitivity when CXCL12 concentrations were higher. MDA-MB-231 cells displayed low relative expression levels of CXCR4, but high sensitivity resulting in 55-fold increase at late time points due to CXCL12 gradient dissipation. CONCLUSIONS: This model could create clinical impact as an in vitro diagnostic tool for rapid assessment of tumor needle biopsies to confirm metastatic tumors, their invasiveness, and allow high-throughput drug screening providing rapid development of personalized therapies.

Effectiveness and predictors of weight loss response to phentermine plus lifestyle modifications among youth in a paediatric weight management clinical setting.

BACKGROUND: Anti-obesity medications (AOMs) are promising lifestyle modification (LSM) adjuncts for obesity treatment, and phentermine is commonly prescribed in paediatric weight management clinics. Determining 'real-world' AOM effectiveness and characteristics predicting response is important. OBJECTIVES: We sought to describe phentermine plus LSM effectiveness and identify baseline characteristics predicting response. METHODS: This was a retrospective cohort study among youth seen in a US academic-based weight management clinic from 2012 to 2020. Baseline characteristics (e.g., body mass index (BMI), liver transaminases, eating-related behaviours) and outcomes (%BMI of 95th percentile (%BMIp95), BMI, %BMI change, weight) were determined through electronic health records and intake surveys. RESULTS: Among 91 youth prescribed phentermine plus LSM over 8 years (mean %BMIp95 150%), %BMIp95 was statistically significantly reduced at 1.5, 3, 6 and 12 months (peak reduction 10.9 percentage points at 6 months; p < 0.001). Considering multiple comparisons, the presence of baseline elevated alanine aminotransferase was associated with statistically significant smaller 1.5-month %BMIp95 reductions (p = 0.001) and higher food responsiveness with smaller 3- (p = 0.001) and 6-month (p < 0.001) reductions. CONCLUSIONS: Phentermine plus LSM reduced %BMIp95 among youth in a weight management clinic, and baseline characteristics may help determine those more or less likely to respond. Prospective studies are needed to further characterize effectiveness and confirm response predictors.

Clinical effectiveness and predictors of response to topiramate plus lifestyle modification in youth with obesity seen in a weight management clinical setting.

Introduction: Obesity affects approximately 20% of U.S. youth. Anti-obesity medications (AOMs) are promising lifestyle modification adjuncts for obesity treatment, and topiramate is commonly prescribed in pediatric weight management clinics. It is important to determine "real-world" effectiveness of AOMs and, given shifts towards personalized approaches, characteristics potentially predicting better or worse response. We therefore sought to describe clinical effectiveness from topiramate plus lifestyle modification, and to determine if baseline phenotypic characteristics are associated with better or worse response. Methods: We performed a retrospective cohort study (2012-2020) among youth (<18 years old) followed in a U.S. academic-based weight management clinic. Baseline characteristics (i.e., body mass index (BMI), liver function tests, eating-related behaviors) and outcomes (%BMI of 95th percentile (%BMIp95), BMI, percent %BMI change, weight) were determined through review of electronic health records and clinic intake survey data. Results: Among 282 youth prescribed topiramate plus lifestyle modifications (mean baseline age 12.7 years, %BMIp95 144%), %BMIp95 and percent BMI change were statistically significantly reduced at each time point (1.5-, 3-, 6-, and 12-month %BMIp95 reductions: -2.2, -3.9, -6.6, and -9.3 percentage points, respectively; percent BMI reduction: -1.2%, -1.9%, -3.2%, and -3.4%, respectively; all p<0.01). Considering multiple comparisons, no baseline characteristics statistically significantly predicted response at any time point. Conclusions: We found that topiramate plus lifestyle modification reduced %BMIp95 and BMI among youth in a weight management clinical setting, and that no baseline characteristics evaluated were associated with response. These results should be considered preliminary given the observational nature of this study, and prospective studies are needed to further characterize clinical effectiveness and identify and confirm potential predictors of response.

Aberrant activation of wound healing programs within the metastatic niche facilitates lung colonization by osteosarcoma cells.

Purpose: Lung metastasis is responsible for nearly all deaths caused by osteosarcoma, the most common pediatric bone tumor. How malignant bone cells coerce the lung microenvironment to support metastatic growth is unclear. This study delineates how osteosarcoma cells educate the lung microenvironment during metastatic progression. Experimental design: Using single-cell transcriptomics (scRNA-seq), we characterized genome- and tissue-wide molecular changes induced within lung tissues by disseminated osteosarcoma cells in both immunocompetent murine models of metastasis and patient samples. We confirmed transcriptomic findings at the protein level and determined spatial relationships with multi-parameter immunofluorescence. We evaluated the ability of nintedanib to impair metastatic colonization and prevent osteosarcoma-induced education of the lung microenvironment in both immunocompetent murine osteosarcoma and immunodeficient human xenograft models. Results: Osteosarcoma cells induced acute alveolar epithelial injury upon lung dissemination. scRNA-seq demonstrated that the surrounding lung stroma adopts a chronic, non-resolving wound-healing phenotype similar to that seen in other models of lung injury. Accordingly, metastasis-associated lung demonstrated marked fibrosis, likely due to the accumulation of pathogenic, pro-fibrotic, partially-differentiated epithelial intermediates. Inhibition of fibrotic pathways with nintedanib prevented metastatic progression in multiple murine and human xenograft models. Conclusions: Our work demonstrates that osteosarcoma cells co-opt fibrosis to promote metastatic outgrowth. When harmonized with data from adult epithelial cancers, our results support a generalized model wherein aberrant mesenchymal-epithelial interactions are critical for promoting lung metastasis. Adult epithelial carcinomas induce fibrotic pathways in normal lung fibroblasts, whereas osteosarcoma, a pediatric mesenchymal tumor, exhibits fibrotic reprogramming in response to the aberrant wound-healing behaviors of an otherwise normal lung epithelium, which are induced by tumor cell interactions. Statement of translational relevance: Therapies that block metastasis have the potential to save the majority of lives lost due to solid tumors. Disseminated tumor cells must educate the foreign, inhospitable microenvironments they encounter within secondary organs to facilitate metastatic colonization. Our study elucidated that disseminated osteosarcoma cells survive within the lung by co-opting and amplifying the lung's endogenous wound healing response program. More broadly, our results support a model wherein mesenchymal-epithelial cooperation is a key driver of lung metastasis. Osteosarcoma, a pediatric mesenchymal tumor, undergoes lung epithelial induced fibrotic activation while also transforming normal lung epithelial cells towards a fibrosis promoting phenotype. Conversely, adult epithelial carcinomas activate fibrotic signaling in normal lung mesenchymal fibroblasts. Our data implicates fibrosis and abnormal wound healing as key drivers of lung metastasis across multiple tumor types that can be targeted therapeutically to disrupt metastasis progression.

Appetitive and psychological phenotypes of pediatric patients with obesity.

BACKGROUND: Obesity is a heterogeneous disease with variable treatment response. Identification of the unique constellation of contributors to obesity may allow for targeted interventions and improved outcomes. OBJECTIVE: Identify empirically derived phenotypes of pediatric patients with obesity based on appetitive and psychological correlates of obesity. METHODS: This cross-sectional study included patients aged 5-12 years who were treated in a weight management clinic and completed standard intake questionnaires including Child Eating Behavior Questionnaire (CEBQ), Vanderbilt ADHD Scale and Pediatric Symptom Checklist. Phenotypes were elicited using latent profile analysis of 12 indicators: eight CEBQ subscales, inattention, hyperactivity/impulsivity, internalizing and externalizing symptoms. RESULTS: Parents/guardians of 384 patients (mean age 9.8 years, mean BMI 30.3 kg/m2 ) completed the intake questionnaires. A 4-phenotype model best fits the data. Hedonic Impulsive phenotype (42.5%) exhibited high food enjoyment and hyperactivity/impulsivity. Inattentive Impulsive phenotype (27.4%) exhibited overall low food approach and high food avoid behaviours, and highest inattention. Hedonic Emotional phenotype (20.8%) scored the highest on food enjoyment, internalizing and externalizing symptoms. Picky Eating phenotype (9.3%) scored the lowest on food approach, inattention, hyperactivity/impulsivity, internalizing and externalizing symptoms. CONCLUSION: Appetitive traits and psychological symptoms appear to cluster in distinct patterns, giving rise to four unique phenotypic profiles, which, if replicated, may help inform the development of tailored treatment plans.

Host-derived growth factors drive ERK phosphorylation and MCL1 expression to promote osteosarcoma cell survival during metastatic lung colonization.

PURPOSE: For patients with osteosarcoma, disease-related mortality most often results from lung metastasis-a phenomenon shared with many solid tumors. While established metastatic lesions behave aggressively, very few of the tumor cells that reach the lung will survive. By identifying mechanisms that facilitate survival of disseminated tumor cells, we can develop therapeutic strategies that prevent and treat metastasis. METHODS: We analyzed single cell RNA-sequencing (scRNAseq) data from murine metastasis-bearing lungs to interrogate changes in both host and tumor cells during colonization. We used these data to elucidate pathways that become activated in cells that survive dissemination and identify candidate host-derived signals that drive activation. We validated these findings through live cell reporter systems, immunocytochemistry, and fluorescent immunohistochemistry. We then validated the functional relevance of key candidates using pharmacologic inhibition in models of metastatic osteosarcoma. RESULTS: Expression patterns suggest that the MAPK pathway is significantly elevated in early and established metastases. MAPK activity correlates with expression of anti-apoptotic genes, especially MCL1. Niche cells produce growth factors that increase ERK phosphorylation and MCL1 expression in tumor cells. Both early and established metastases are vulnerable to MCL1 inhibition, but not MEK inhibition in vivo. Combining MCL1 inhibition with chemotherapy both prevented colonization and eliminated established metastases in murine models of osteosarcoma. CONCLUSION: Niche-derived growth factors drive MAPK activity and MCL1 expression in osteosarcoma, promoting metastatic colonization. Although later metastases produce less MCL1, they remain dependent on it. MCL1 is a promising target for clinical trials in both human and canine patients.

Financial Incentives and Treatment Outcomes in Adolescents With Severe Obesity: A Randomized Clinical Trial.

Importance: Adolescent severe obesity is usually not effectively treated with traditional lifestyle modification therapy. Meal replacement therapy (MRT) shows short-term efficacy for body mass index (BMI; calculated as weight in kilograms divided by height in meters squared) reduction in adolescents, and financial incentives (FIs) may be an appropriate adjunct intervention to enhance long-term efficacy. Objective: To evaluate the effect of MRT plus FIs vs MRT alone on BMI, body fat, and cardiometabolic risk factors in adolescents with severe obesity. Design, Setting, and Participants: This was a randomized clinical trial of MRT plus FIs vs MRT alone at a large academic health center in the Midwest conducted from 2018 to 2022. Participants were adolescents (ages 13-17 y) with severe obesity (≥120% of the 95th BMI percentile based on sex and age or ≥35 BMI, whichever was lower) who were unaware of the FI component of the trial until they were randomized to MRT plus FIs or until the end of the trial. Study staff members collecting clinical measures were blinded to treatment condition. Data were analyzed from March 2022 to February 2024. Interventions: MRT included provision of preportioned, calorie-controlled meals (~1200 kcals/d). In the MRT plus FI group, incentives were provided based on reduction in body weight from baseline. Main Outcomes and Measures: The primary end point was mean BMI percentage change from randomization to 52 weeks. Secondary end points included total body fat and cardiometabolic risk factors: blood pressure, triglyceride to high-density lipoprotein ratio, heart rate variability, and arterial stiffness. Cost-effectiveness was additionally evaluated. Safety was assessed through monthly adverse event monitoring and frequent assessment of unhealthy weight-control behaviors. Results: Among 126 adolescents with severe obesity (73 female [57.9%]; mean [SD] age, 15.3 [1.2] years), 63 participants received MRT plus FIs and 63 participants received only MRT. At 52 weeks, the mean BMI reduction was greater by -5.9 percentage points (95% CI, -9.9 to -1.9 percentage points; P = .004) in the MRT plus FI compared with the MRT group. The MRT plus FI group had a greater reduction in mean total body fat mass by -4.8 kg (95% CI, -9.1 to -0.6 kg; P = .03) and was cost-effective (incremental cost-effectiveness ratio, $39 178 per quality-adjusted life year) compared with MRT alone. There were no significant differences in cardiometabolic risk factors or unhealthy weight-control behaviors between groups. Conclusions and Relevance: In this study, adding FIs to MRT resulted in greater reductions in BMI and total body fat in adolescents with severe obesity without increased unhealthy weight-control behaviors. FIs were cost-effective and possibly promoted adherence to health behaviors. Trial Registration: ClinicalTrials.gov Identifier: NCT03137433.

SMART use of medications for the treatment of adolescent severe obesity: A sequential multiple assignment randomized trial protocol.

BACKGROUND: Severe obesity is a complex, chronic disease affecting nearly 9% of adolescents in the U.S. Although the current mainstay of treatment is lifestyle therapy, pediatric clinical practice guidelines recommend the addition of adjunct anti-obesity medication (AOM), such as phentermine and topiramate. However, guidance regarding when adjunct AOM should be started and how AOM should be used is unclear. Furthermore, an inherent limitation of current treatment guidelines is their "one-size-fits-all" approach, which does not account for the heterogeneous nature of obesity and high degree of patient variability in response to all interventions. METHODS: This paper describes the study design and methods of a sequential multiple assignment randomized trial (SMART), "SMART Use of Medications for the Treatment of Adolescent Severe Obesity." The trial will examine 1) when to start AOM (specifically phentermine) in adolescents who are not responding to lifestyle therapy and 2) how to modify AOM when there is a sub-optimal response to the initial pharmacological intervention (specifically, for phentermine non-responders, is it better to add topiramate to phentermine or switch to topiramate monotherapy). Critically, participant characteristics that may differentially affect response to treatment will be assessed and evaluated as potential moderators of intervention efficacy. CONCLUSION: Data from this study will be used to inform the development of an adaptive intervention for the treatment of adolescent severe obesity that includes empirically-derived decision rules regarding when and how to use AOM. Future research will test this adaptive intervention against standard "one-size-fits-all" treatments.

Evaluating appetite/satiety hormones and eating behaviours as predictors of weight loss maintenance with GLP-1RA therapy in adolescents with severe obesity.

INTRODUCTION: Whilst glucagon-like peptide-1 receptor agonists (GLP1-RAs) are effective for treating adolescent obesity, weight loss maintenance (WLM; preventing weight regain) remains a challenge. Our goal was to investigate appetite/satiety hormones and eating behaviours that may predict WLM with exenatide (a GLP1-RA) versus placebo in adolescents with severe obesity. METHODS: Adolescents who had ≥5% body mass index (BMI) reduction with meal replacement therapy were randomized to 52 weeks of once-weekly exenatide extended release or placebo. In this secondary analysis, eating behaviours and appetite/satiety regulation hormones post-meal replacement therapy (pre-randomization to exenatide or placebo) were evaluated as possible predictors of WLM. Percent change in BMI from randomization to 52 weeks served as the primary measure of WLM. RESULTS: The analysis included 66 adolescents (mean age 16.0 years; 47% female). Lower leptin response to meal testing was associated with greater WLM in terms of BMI percent change in those receiving exenatide compared to placebo (p = 0.007) after adjusting for sex, age and BMI. There were no other significant predictors of WLM. CONCLUSIONS: Prior to exenatide, lower leptin response to meals was associated with improved WLM with exenatide compared to placebo. The mostly null findings of this study suggest that GLP1-RA treatment may produce similar WLM for adolescents with obesity regardless of age, BMI, sex and eating behaviours.