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Abstract p90 Ribosomal S6 Kinase 2 (p90RSK2) is a substrate of ERK and plays an active role in anti-apoptosis signaling and cell cycle regulation. Our proteomics studies revealed that p90RSK2 is not only activated but also tyrosine-phosphorylated in hematopoietic cells expressing constitutively activated leukemogenic FGFR3 mutants. FGFR3 is a receptor-tyrosine kinase that responds to fibroblast growth factor (FGF). Recent studies have suggested that FGFR3 may play a significant role in the pathogenesis and disease progression of some hematopoietic malignancies including t(4;14)(p16.3;q32.3) multiple myeloma, which is associated with dysregulation of FGFR3. We found that the MEK1 inhibitor U0126 effectively inhibited RSK2 activation induced by FGFR3, suggesting that FGFR3 activates RSK2 through the MEK/ERK pathway. Interestingly, FGFR3-dependent tyrosine phosphorylation at Y529 was required for phosphorylation and activation of the Ser/Thr kinase RSK2 by ERK. By using a specific phospho-RSK2 (Y529) antibody, we identified FGFR3 as the direct tyrosine kinase that phosphorylates RSK2 at Y529 in an in vitro kinase assay, which only contains purified recombinant active FGFR3 and RSK2 proteins. Substitution of Y529 in RSK2 attenuates inactive ERK binding to RSK2, which is prior to and required for ERK-dependent phosphorylation and activation of RSK2. In addition, reconstitution of RSK2 phosphorylation at Y529 consequently enhanced inactive ERK binding by a GST pull down, suggesting that phosphorylation at Y529 by FGFR3 facilitates ERK binding to RSK2, which is a sequential event and precedes inactive ERK binding. These observations support a novel two-step model that FGFR3 activates RSK2 by both assisting inactive ERK binding via tyrosine phosphorylation of RSK2 at Y529 and activating the MEK/ERK pathway. Moreover, we observed that RSK2 is specifically phosphorylated at Y529 and activated in FGFR3 expressing various human t(4;14) positive MM cell lines. Targeted down-regulation of RSK2 by siRNA induced significant apoptosis in FGFR3-expressing human myeloma cell lines including OPM1 and KMS11. In addition, a specific RSK inhibitor fmk induced significant apoptosis in t(4;14) positive human myeloma cells including KMS11, OPM1, LP1 and KMS18, but not in control t(4;14) negative myeloma cell lines including ANBL6 and U266. Similar results were obtained by using another RSK specific and potent inhibitor BI-D1870. Furthermore, we observed that fmk induced significant apoptosis in primary CD138-positive, FGFR3-expressing myeloma cells from a t(4;14) positive multiple myeloma patient, but not in the control CD138-negative cells from the same patient, nor primary samples from a t(4;14)-negative patient as a control. Together, these data not only support our model that RSK2 is a critical signaling effector of FGFR3 in hematopoietic transformation, but also provide “proof of principle” for the therapeutic potential of RSK2 inhibitors such as fmk in treatment of t(4;14) positive, FGFR3-expressing multiple myeloma with minimal non-specific cytotoxicity.

Objective Bronchopulmonary dysplasia (BPD) is a condition of neonatal chronic lung disease due to disruption or dysregulation of pulmonary development. It is associated with decreased septation, alveolar hypoplasia, and abnormal disruption of pulmonary alveolar capillaries resulting in increased pulmonary resistance. However, the pathophysiology of BPD in the large conducting airways is not fully understood. The objective of our study was to determine if the luminal area of the central airways, measured by imaging, are altered in patients with a history of BPD. We hypothesized that compared to age‐ and sex‐matched controls, BPD patients would have a decreased luminal area of the central conducting airways and lung volume. Methods Fourteen BPD patients (8M/6F; Age=5.7±5.5 yrs) who had undergone a chest computed tomography (CT) scan were retrospectively identified. Age and sex matched controls (8M/6F, Age=5.6±5.3 yrs) who had undergone a CT scan for trauma were used for comparison. Measurement and analysis were performed by a single investigator using commercially available software (Aquarius, TeraRecon Inc.) that reconstructs the airways into 3D. Measurements of luminal airway area were conducted at three points based on anatomic bifurcations (corresponding to the beginning, middle and end of the airway) for each of the following structures: trachea, left main bronchus, left upper lobe, left lower lobe, right main bronchus, intermediate bronchus, and right upper lobe. Each airway’s luminal area was calculated based on the averages of the three measures obtained for each anatomic airway structure. Independent Student’s t‐test was used for statistical analysis. Results Airway luminal area was not different between BPD patients and matched controls (trachea: 89.8±57.1 vs 87.4±58.6 mm 2 ; right main bronchus: 58.8±40.8 vs 64.0±44.5 mm 2 ; intermediate bronchus: 33.6±21.2 vs 41.0±28.9 mm 2 ; left main bronchus: 39.3±27.1 vs 46.6±36.5 mm 2 ; left lower lobe: 22.7±14.6 vs 29.9±18.9 mm 2 ; p>0.05 for all comparisons; BPD and controls, respectively). Total lung volume detected in the CT scans was not different between BPD patients and matched controls (3069±1073 vs 3215±984 cm 3 , p>0.05). Conclusions Our results suggest the luminal areas of the large conducting airways in patients with BPD are not different from matched controls. Given the degree of respiratory complications and sequelae associated with BPD, our findings suggest future research should focus on alveolar and vascular impaired physiological and anatomic developments versus the large conducting airways. Support or Funding Information National Institutes of Health Grants R35‐HL139854 (MJJ), F32 HL‐131151 (SEB), T32 DK‐007352‐39 (CCW) and Mayo Clinic Internal Funding and Natural Science and Engineering Council of Canada RGPIN‐2019‐04615 (PBD)

534 Background: mTOR plays a critical role in promoting tumor cell growth. In preclinical studies, the anti-tumor activity of mTOR inhibitors is attenuated by feedback up-regulation of AKT mediated by IGF-1R. We designed a phase I trial to determine the maximum-tolerated dose (MTD), dose-limiting toxicities (DLT) and pharmacodynamic effects of the IGF-1R antibody IMC-A12 in combination with temsirolimus (tem) in patients (pts) with metastatic breast cancer (MBC) where mTOR is frequently activated. Methods: A 3+3 phase I design was chosen. Tem and IMC-A12 were administered IV days (d) 1, 8, 15, and 22 of a 4-week cycle in pts with MBC refractory to standard therapies. Tumor response was evaluated by RECIST. Adverse events (AE) were reported using CTC v3.0. Serum IGF 1 and C-peptide levels on d2 (24h post infusion) and d8 prior to drug infusion were compared to baseline (BL) using paired t-test. Results: Of 26 pts enrolled, 4 did not complete cycle 1 due to progression (3) or co-morbid condition (1). MTD was determined from remaining 22 pts aged 34-72 (median 48) years with ECOG PS 0 (55%) or 1 (45%). 86% had ER+ cancer. Median number of regimens for MBC was 4. Two DLTs at the starting DL (DL 1) necessitated dose de-escalation of tem to 20mg (DL-1), then to 15 mg (DL-2) which was tolerable (Table). Subsequent dose escalation of IMC-A12 led to DLTs in 0 of 6 in DL-2A and 2 of 3 pts in DL-2B. The MTD was defined as DL-2A. Other AEs included gr 1/2 fatigue, neutropenia, anemia, and hyperglycemia. No CR or PR, but 4 SD lasting ≥ 4 months were observed. At DL-2, -2A and -2B, serum IGF 1 levels were significantly elevated on d2 (p <0.002) and d8 compared to BL (p <0.001), but C-peptide levels were not found to differ from BL. Conclusions: The MTD for the combination of IMC-A12 and tem in pts with MBC is lower than that observed for single agents alone. A phase II study is ongoing in MBC. The study is supported in part by ASCO CDA, Komen Craft to CXM, N01-CM62205 and N01-CM-2011-00071. [Table: see text]

Background: Tumoral melanosis clinically resembles metastatic melanoma, occurs in the context of regressed disease, and requires evaluation to rule out underlying melanoma and metastatic disease. Histopathology demonstrates a nodular infiltrate of melanophages in the dermis, subcutaneous tissue, deep soft tissue, or lymph nodes in the absence of viable melanocytes. Recent limited reports of tumoral melanosis in the context of immunotherapy with ipilimumab (monoclonal antibody targeting CTLA-4) as well as nivolumab and pembrolizumab (humanized monoclonal antibodies against programmed death 1 receptor) highlight a unique presentation representative of treatment-related tumor regression and an association with a favorable clinical response. Objective: To describe our experience with tumoral melanosis in the setting of immunotherapy for metastatic melanoma and elucidate the clinical and histopathological features. Methods: Retrospective case series from a single tertiary care institution. Results: We describe 10 cases of patients with metastatic melanoma who received treatment with immunotherapy before the development of tumoral melanosis. Length of time between the initiation of therapy and the onset of tumoral melanosis ranged from 2 to 20 months with a mean time of 10 months. At the end of the follow-up period, 8 patients were classified as having a complete or partial response to treatment with immunotherapy. One patient had progression of visceral and cutaneous disease on ipilimumab despite developing tumoral melanosis, and 1 patient had yet to undergo repeat imaging. Furthermore, at the end of follow-up, 3 patients were alive with no evidence of active disease, 5 patients were alive with disease, and 1 patient was deceased, although this patient died of a cardiovascular event unrelated to his underlying melanoma. Of the patients who were classified as alive with disease, 2 patients had minimal remaining disease, and 2 patients had an almost complete response on immunotherapy with recurrence of visceral metastases after immunotherapy was discontinued. One patient developed new peritoneal and cutaneous metastases on pembrolizumab despite development of tumoral melanosis. Conclusions: The underlying biologic mechanisms and prognostic implications of tumoral melanosis in the setting of immunotherapy remain to be elucidated. Further prospective studies with a larger cohort and prolonged follow-up are necessary to better understand the incidence, prevalence, and oncologic outcomes in patients with tumoral melanosis who receive immunotherapy.

Abstract Eating disorders and substance use disorders frequently co‐occur. Twin studies reveal shared genetic variance between liabilities to eating disorders and substance use, with the strongest associations between symptoms of bulimia nervosa and problem alcohol use (genetic correlation [ r g ], twin‐based = 0.23‐0.53). We estimated the genetic correlation between eating disorder and substance use and disorder phenotypes using data from genome‐wide association studies (GWAS). Four eating disorder phenotypes (anorexia nervosa [AN], AN with binge eating, AN without binge eating, and a bulimia nervosa factor score), and eight substance‐use‐related phenotypes (drinks per week, alcohol use disorder [AUD], smoking initiation, current smoking, cigarettes per day, nicotine dependence, cannabis initiation, and cannabis use disorder) from eight studies were included. Significant genetic correlations were adjusted for variants associated with major depressive disorder and schizophrenia. Total study sample sizes per phenotype ranged from ~2400 to ~537 000 individuals. We used linkage disequilibrium score regression to calculate single nucleotide polymorphism‐based genetic correlations between eating disorder‐ and substance‐use‐related phenotypes. Significant positive genetic associations emerged between AUD and AN ( r g = 0.18; false discovery rate q = 0.0006), cannabis initiation and AN ( r g = 0.23; q < 0.0001), and cannabis initiation and AN with binge eating ( r g = 0.27; q = 0.0016). Conversely, significant negative genetic correlations were observed between three nondiagnostic smoking phenotypes (smoking initiation, current smoking, and cigarettes per day) and AN without binge eating ( r gs = −0.19 to −0.23; qs < 0.04). The genetic correlation between AUD and AN was no longer significant after co‐varying for major depressive disorder loci. The patterns of association between eating disorder‐ and substance‐use‐related phenotypes highlights the potentially complex and substance‐specific relationships among these behaviors.

New Findings What is the central question of this study? Are sex difference in the central airways present in healthy paediatric patients? What is the main finding and its importance? In patients ≤12 years we found no sex differences in central airway luminal area. After 14 years, the males had significantly larger central airway luminal areas than the females. The sex differences were minimized, but preserved when correcting for height. Luminal area is the main determinant of airway resistance and our finding could help explain sex differences in pulmonary system limitations to exercise in paediatric patients. Abstract Cross‐sectional airway area is the main determinant of resistance to airflow in the respiratory system. In paediatric patients (<18 years), previous evidence for sex differences in cross‐sectional airway area was limited to patients with history of pulmonary disease or cadaveric studies with small numbers of subjects. These studies either only report tracheal data and do not include a range of ages or correct for height. Therefore, we sought to assess sex differences in airway luminal area utilizing paediatric patients of varying ages and no history of respiratory disease. Using three‐dimensional reconstructions from high‐resolution computed tomography scans, we retrospectively assessed the cross‐sectional airway area in healthy paediatric females ( n = 97) and males ( n = 128) over a range of ages (1–17 years). The areas of the trachea, left main bronchus, left upper lobe, left lower lobe, right main bronchus, intermediate bronchus and right upper lobe were measured at three discrete points by a blinded investigator. No differences between the sexes were noted in the cross‐sectional areas of the youngest (ages 1–12 years) patients ( P > 0.05). However, in patients ≥14 years the cross‐sectional areas were larger in the males compared to females in most airway sites. For instance, the cross‐sectional size of the trachea was 25% (218 ± 44 vs . 163 ± 24 mm 2 , P < 0.01) larger in males vs . females among ages 13–17 years. When accounting for height, these sex differences in airway areas were attenuated, but persisted. Our results indicate that sex differences in paediatric airway cross‐sectional area manifest after age ≥14 years and are independent of height.

AbstractAdipose‐derived mesenchymal stem cell (ADSC)‐based regenerative therapies have shown potential for use in many chronic diseases. Aging diminishes stem cell regenerative potential, yet it is unknown whether stem cells from aged donors cause adverse effects in recipients. ADSCs can be obtained using minimally invasive approaches and possess low immunogenicity. Nevertheless, we found that transplanting ADSCs from old donors, but not those from young donors, induces physical dysfunction in older recipient mice. Using single‐cell transcriptomic analysis, we identified a naturally occurring senescent cell‐like population in ADSCs primarily from old donors that resembles in vitro‐generated senescent cells with regard to a number of key pathways. Our study reveals a previously unrecognized health concern due to ADSCs from old donors and lays the foundation for a new avenue of research to devise interventions to reduce harmful effects of ADSCs from old donors.

Alcoholic hepatitis (AH) is a severe inflammatory liver disease that develops in some heavy drinkers. The immune system in patients with AH is hyperactive and yet dysfunctional. Here, we investigated whether this immune‐dysregulated state is related to the alcoholic impact on immune checkpoints (ICPs). We used multiplex immunoassays and enzyme‐linked immunosorbent assay to quantify plasma levels of 18 soluble ICPs (sICPs) from 81 patients with AH, 65 heavy drinkers without liver diseases (HDCs), and 39 healthy controls (HCs) at baseline, 33 patients with AH and 32 HDCs at 6‐month follow‐up, and 18 patients with AH and 29 HDCs at 12‐month follow‐up. We demonstrated that baseline levels of 6 sICPs (soluble T‐cell immunoglobulin and mucin domain 3 [sTIM‐3], soluble cluster of differentiation [sCD]27, sCD40, soluble Toll‐like receptor‐2 [sTLR‐2], soluble herpesvirus entry mediator [sHVEM], and soluble lymphotoxin‐like inducible protein that competes with glycoprotein D for herpes virus entry on T cells [sLIGHT]) were up‐regulated, while 11 sICPs (soluble B‐ and T‐lymphocyte attenuator [sBTLA], sCD160, soluble cytotoxic T‐lymphocyte‐associated protein 4 [sCTLA‐4], soluble lymphocyte‐activation gene 3 [sLAG‐3], soluble programmed death 1 [sPD‐1], sPD ligand 1 [sPD‐L1], sCD28, soluble glucocorticoid‐induced tumor necrosis factor receptor‐related protein [sGITR], sGITR ligand [sGITRL], sCD80, and inducible T‐cell costimulator [sICOS]) were down‐regulated in patients with AH compared to HDCs. The up‐regulated sICPs except sLIGHT and down‐regulated sCD80, sCD160, sCTLA‐4, and sLAG‐3 correlated positively or negatively with AH disease severity, bacterial translocation, and inflammatory factors. At follow‐up, abstinent patients with AH still had higher levels of several sICPs compared to HDCs. We also compared expression of 10 membrane‐bound ICPs (mICPs) on peripheral blood mononuclear cells (PBMCs) from patients with AH and HCs by flow cytometry and found that several mICPs were dysregulated on blood cells from patients with AH. The function and regulation of sICPs and mICPs were studied using PBMCs from patients with AH and HCs. Recombinant sHVEM affected tumor necrosis factor (TNF)‐α and interferon‐γ production by T cells from patients with AH and HCs. Conclusion: Both sICPs and mICPs were dysregulated in patients with AH, and alcohol abstinence did not fully reverse these abnormalities. The HVEM axis plays a role in regulating T‐cell function in patients with AH.

Background The epidemiological transition of non-communicable diseases replacing infectious diseases as the main contributors to disease burden has been well documented in global health literature. Less focus, however, has been given to the relationship between sociodemographic changes and injury. The aim of this study was to examine the association between disability-adjusted life years (DALYs) from injury for 195 countries and territories at different levels along the development spectrum between 1990 and 2017 based on the Global Burden of Disease (GBD) 2017 estimates. Methods Injury mortality was estimated using the GBD mortality database, corrections for garbage coding and CODEm—the cause of death ensemble modelling tool. Morbidity estimation was based on surveys and inpatient and outpatient data sets for 30 cause-of-injury with 47 nature-of-injury categories each. The Socio-demographic Index (SDI) is a composite indicator that includes lagged income per capita, average educational attainment over age 15 years and total fertility rate. Results For many causes of injury, age-standardised DALY rates declined with increasing SDI, although road injury, interpersonal violence and self-harm did not follow this pattern. Particularly for self-harm opposing patterns were observed in regions with similar SDI levels. For road injuries, this effect was less pronounced. Conclusions The overall global pattern is that of declining injury burden with increasing SDI. However, not all injuries follow this pattern, which suggests multiple underlying mechanisms influencing injury DALYs. There is a need for a detailed understanding of these patterns to help to inform national and global efforts to address injury-related health outcomes across the development spectrum.