bluuebunny/crossref_metadata_2025 huggingface dataset:
299 Serum Aldosterone and Urine Electrolytes Dynamics in Response to DASH Diet Intervention
Authors: Bielopolski Dana , Qureshi Adam , Ronning Andrea , Tobin Jonathan N. , Kost Rhonda G.
Published: April 2022

OBJECTIVES/GOALS: Dietary approach to stop hypertension (DASH) is a proven intervention to treat hypertension. Despite years of research the immediate physiologic response to its implementation was never characterized.This translational trial describes the biological pathway from nutrition through hormons, urine electrolytes and blood pressure reduction. METHODS/STUDY POPULATION: A single center interventional trial. Stage 1 hypertensive otherwise healthy volunteers were admitted for 14-days, transitioning from American style diet to DASH diet. Nutritional habits were assessed with food frequency questionnaires, and menus designed according to the guidelines of the National Heart Blood and Lung Institute (NHBLI) of the National Institute of Health (NIH). Data were collected daily for vital signs, blood and urine. Participants completed two 24-hour ABPM on days 1,10, and two 24-hour urine collections in parallel. We conducted a follow up visit two weeks after discharge. RESULTS/ANTICIPATED RESULTS: 9 volunteers (78% male, 89% Black individuals) completed the protocol. During an inpatient stay, they consumed a mean daily potassium intake of 5.6 g (Â±0.7g) and 2.6 g (Â±0.3g) of Sodium. Serum Aldosterone increased from day 0 (mean 8.3, range 2.8-18.9) to day 5 (mean 17.8, range 10.2-27.2) after intervention, and decreased on day 11 (mean 11.5, range 4.8-18.2) despite continuous exposure (p-value=0.002). The urine electrolyte ratio of ([Na]/[K]) decreased from a mean of 3.5 before intervention to 1.16 on day 4, creating a statistically significant slope (p-value<0.001). Blood pressure by 24-hour ABPM decreased by 3.7 mmHg systolic BP and 2.3 mmHg diastolic BP from day 1 to 10 for the entire period, and for measures taken during sleep or awake time, assessed separately. DISCUSSION/SIGNIFICANCE: Shifting from a high-sodium low-potassium diet to the opposite composition leads to serial physiological changes that are governed by aldosterone and result in blood pressure reduction. Urine electrolyte ratio reflects nutritional changes within 4 days of transition and should guide clinicians in assessing lifestyle modification adherence.
303 Providers Assessment of Nutritional Practices for the Duchenne and Becker Muscular Dystrophy Population
Authors: Fawcett Kindann , Veerapandiyan Aravindhan
Published: April 2022

OBJECTIVES/GOALS: Nutrition plays an imperative role in the management of Duchenne and Becker Muscular Dystrophy, but guidelines for nutrition counseling are absent. This study was designed to gain insight into provider experiences with nutritional services to find solutions for future counseling. METHODS/STUDY POPULATION: In this prospective, observational exploratory study, semi structured interviews were conducted at Certified Duchenne Care Centers (CDCCs) to gain insight on providers experiences with nutritional services and to identify barriers and solutions to nutrition education/care/counseling at CDCCs. Interviews were video recorded and then transcribed for themes. Overarching themes gave insight for a quantitative survey to be sent out, assessing all members on the multidisciplinary team perceptions, confidence, barriers, and solutions to providing nutritional care to Duchenne and Becker patients. RESULTS/ANTICIPATED RESULTS: We anticipate this study will provide novel data and key information from providers regarding nutrition education /care/counseling efforts in the multidisciplinary care of neuromuscular diseases. DISCUSSION/SIGNIFICANCE: Results will demonstrate the need for higher standards and more specific recommendations in nutritional services at CDCCs, while providing a framework for referrals, continuing education opportunities, and increasing providers’ confidence and abilities to provide sound nutritional advice.
314 Exoskeletons increase paretic limb use in stroke survivors during a bimanual virtual reality reaching task
Authors: Brunfeldt Alexander , Lum Peter , Bregman Barbara
Published: April 2022

OBJECTIVES/GOALS: Almost 8 million Americans live with disability caused by stroke. However, recent advances in stroke rehabilitation are costly and lack resemblance to activities of daily living. The goal of this study was to develop a rehabilitation platform to increase stroke patients paretic limb use using inexpensive virtual reality and exoskeleton devices. METHODS/STUDY POPULATION: We conducted a feasibility study with 2 hemiplegic stroke participants. They reached for targets in a virtual reality environment using both hands. They completed 162 reaches divided into 3 blocks. Following baseline, we used an exoskeleton to provide 50% arm weight compensation to the paretic limb and used wrist weights to provide 50% arm weight resistance to the non-paretic limb. We removed the exoskeleton and wrist weights during the retention block. We used electromyography to approximate muscle activity in the biceps brachii. Relative contribution (RC) was calculated as the displacement of the paretic arm divided by the sum of displacements for both arms. Muscle contribution (MC) was calculated as the root mean square of paretic arm muscle activity divided by the sum of activity for both arms. RESULTS/ANTICIPATED RESULTS: During baseline, RC of the impaired limb was 44% and 48%, and MC of the impaired bicep was 43% and 35% in two mild to moderately impaired patients (Fugl-Meyer Upper Extremity scores of 43 and 37, respectively). During loading, RC increased by 5.6% and 1.1% and MC decreased by 8.3% and 11.8%. These data suggest hemiplegic stroke participants alter limb coordination when our device normalizes muscular output asymmetries between limbs. Importantly, these results closely match data from our previous work in 12 healthy controls, where we found a 2% increase in RC is significantly predicted by a 11% decrease in MC. By collecting more data on stroke patients, we will quantify this tradeoff between coordination and muscle activity modulation, allowing us to optimize the exoskeleton mechanics to maximize paretic limb use. DISCUSSION/SIGNIFICANCE: We demonstrate our platform is well tolerated by mild to moderately impaired stroke patients; this feasibility study forms the basis for low cost at-home technologies for stroke rehabilitation. With further development, clinicians can use our platform to fine-tune the level of limb constraint based on the individual needs of the patient.
326 Integrated Analysis of Genetic Databases Identifies miRNA Associated With Poor Survival In Melanoma
Authors: McCallister Reid , Subramanian Chitra , Mantena Nikhil , Cohen Mark
Published: April 2022

OBJECTIVES/GOALS: Despite advances in precision medicine and understanding of the molecular pathways, melanoma remains the deadliest skin cancer, warranting identification of novel biomarkers. In this study, we performed bioinformatic analysis of melanoma patient tumors to identify novel dysregulated gene and micro-RNA (miRNA) targets responsible for survival. METHODS/STUDY POPULATION: Genetic sequencing data for 594 patient samples of melanoma and normal skin tissue from 10 databases were accessed using the NCBI Gene Expression Omnibus. Genes and miRNA that were significantly dysregulated (adjusted p-value < 0.05, log fold change > Â± 2) in melanoma compared to normal skin were identified using the GEO2R program. Dataset expression profiles were cross-referenced to identify genetic elements dysregulated in at least 50% of datasets and filtered for association with poor survival using R2 Genomics Analysis and Visualization Platform. DAVID 6.8 provided pathway analysis of dysregulated genes. miRTarBase linked genes associated with poor survival and dysregulated miRNA from our database analysis. RESULTS/ANTICIPATED RESULTS: Bioinformatic analysis revealed consistent differential regulation of 205 genes (down=177 and up=28) and 38 miRNA across datasets with fold change >2 (bonf. p<0.05). Pathway analysis indicated that PPAR, phosphatidylinositol signaling, Rap1 signaling, and p53 signaling pathways were enriched by downregulated genes while the NF-kB pathway was enriched up regulated genes. Survival analysis of the differentially regulated genes identified 11 downregulated (ACSL1, CEBPA, CES4A, CRIP1, GATA3, HLADQB2, PTGS1, PYCARD, PPARG, PKP3, RSSF6) and 3 upregulated (DUXAP10, SLC2A3 and PRAME) hub genes to be associated with poor overall survival. Out of the 13 miRNA associated with hub genes, five miRNA (hsa-miR-125b-5p, hsa-miR-130b-3p, hsa-miR-26n-5p, hsa-miR-30b, hsa-miR-30c) were linked to multiple hub genes. DISCUSSION/SIGNIFICANCE: Our analysis identified 14 hub genes (regulators of PPARA, adipocyte differentiation, and transcriptional pathways) as well as miRNAs hsa-miR-30c (regulator of PTGS1 and SLC2A3) and hsa-let-7i-5p (regulator of ASCL1) as potential therapeutic targets. Further studies for validation of the targets are needed for clinical translation in melanoma.
327 Nociplastic pain and early discontinuation of aromatase inhibitor therapy in breast cancer patients over age 65
Authors: Joyce Elizabeth , Kidwell Kelley M. , Henry N. Lynn
Published: April 2022

OBJECTIVES/GOALS: Side effects are why up to 50% of women with hormone-positive breast cancer prematurely discontinue aromatase inhibitor (AI) therapy. Pain from altered nociception without clear tissue/nerve damage (nociplastic) is a hypothesized contributing factor to this. Our objective was to evaluate the relationship between nociplastic pain and AI duration. METHODS/STUDY POPULATION: Patients with breast cancer diagnosed between 2012-19 were identified from the University of Michigan Genomics Initiative (MGI). Patients who were female, >65 years old at time of breast cancer diagnosis, and had hormone receptor-positive disease met inclusion criteria. Prior to undergoing surgery, patients completed validated surveys about overall worst pain (Brief Pain Inventory [BPI]), nociplastic pain (2011 Fibromyalgia Survey [FS]), and life satisfaction, with higher scores representing more of the factor. Breast cancer history, treatment, and patient demographics were abstracted from the medical record. Univariate analysis was conducted to evaluate the relationship between age, body-mass index (BMI), chemotherapy, BPI, FS, life satisfaction, and time to discontinuation of initial AI. RESULTS/ANTICIPATED RESULTS: 207 patients were eligible, 133 of whom initiated AI therapy. Of the 133 analyzed patients, mean age was 70.7 years and mean BMI was 30.3. 28 (21%) underwent adjuvant chemotherapy and 79 (59%) received adjuvant radiation prior to initiation of AI therapy. Average nociplastic pain score was 4.0/31 (standard deviation [SD] 4.6), worst pain was 1.5/10 (SD 1.9), and life satisfaction score was 7.3/10 (SD 2.8). The initial AI for 94% of patients (125 patients) was anastrozole. On univariate analysis, only higher nociplastic pain score was statistically associated with premature discontinuation of AI with a HR 1.07 (95%CI 1.00-1.13, p = 0.036). On multivariable analysis, no factors remained statistically significant, although there was a trend for nociplastic pain (HR 1.06, 95%CI 1.00-1.13, p = 0.068). DISCUSSION/SIGNIFICANCE: It is important to identify variables predicting tolerance to therapy so patients can be optimally counseled. Our study suggests that patients with pre-existing baseline pain disorders may be more likely to be nonpersistent with AI therapy. Future study should be conducted to determine if treatments for nocipastic pain improve AI persistence.
328 Urine Biomarker Predictors of Incident Hospitalization in People Living with HIV
Authors: Lai Mason , Estrella Michelle
Published: April 2022

OBJECTIVES/GOALS: This study aims to determine 1) which urine biomarkers of kidney health measured in the ambulatory setting predict incident hospitalization; 2) to determine whether time-updated measures of ambulatory urine biomarkers and their changes/trajectories have added value over baseline alone. METHODS/STUDY POPULATION: Participants in the Predictors of Acute Renal Injury Study (PARIS), a prospective cohort of 478 HIV+ patients followed at the Johns Hopkins HIV Clinic, had sociodemographic, clinical data, and biosamples taken until hospitalization or up to 3 years annually. Among those hospitalized, clinical data and biosamples were collected serially during hospitalization and at 3 and 12 months post-discharge. For each of the 10 biomarkers measured, we will evaluate the association of the biomarker and risk of incident hospitalization using Cox hazards regression, adjusting for sociodemographics, comorbidities, HIV history, and medications. Biomarkers will be evaluated at baseline and as time-updated and change over time. RESULTS/ANTICIPATED RESULTS: We anticipate that higher baseline levels and increasing levels of urine albumin, Î±1M, Î²2M, NGAL, IL-18, KIM-1, MCP-1, YKL-40 will be independently associated with increased risk of incident hospitalization whereas higher and increasing levels of uromodulin and EGF will be associated with lower risk of incident hospitalization. These biomarkers collectively capture the following dimensions of kidney health: endothelial injury, tubular injury and function, inflammation, and fibrosis. We anticipate increased risk of incident hospitalization in HIV+ persons in the highest tertile of baseline, time-updated, and change over time biomarkers, relative to those within the lowest tertile. DISCUSSION/SIGNIFICANCE: This study will improve our understanding of the evolution of biomarkers of kidney health from the ambulatory to the hospitalized setting and will quantify the clinical implications of subclinical kidney damage among people living with HIV, a high-risk patient population with unique kidney pathophysiology.
338 Diffusion Basis Spectrum Imaging (DBSI) Prognosticates Outcomes for Cervical Spondylotic Myelopathy after Surgery
Authors: Zhang Justin , Javeed Saad , Greenberg Jacob K. , Jayasekera Dinal , Dibble Christopher F. , Blum Jacob , Jakes Rachel , Sun Peng , Song Sheng-Kwei , Ray Wilson Z.
Published: April 2022

OBJECTIVES/GOALS: Diffusion basis spectrum imaging (DBSI) allows for detailed evaluation of white matter microstructural changes present in cervical spondylotic myelopathy (CSM). Our goal is to utilize multidimensional clinical and quantitative imaging data to characterize disease severity and predict long-term outcomes in CSM patients undergoing surgery. METHODS/STUDY POPULATION: A single-center prospective cohort study enrolled fifty CSM patients who underwent surgical decompression and twenty healthy controls from 2018-2021. All patients underwent diffusion tensor imaging (DTI), DBSI, and complete clinical evaluations at baseline and 2-years follow-up. Primary outcome measures were the modified Japanese Orthopedic Association score (mild [mJOA 15-17], moderate [mJOA 12-14], severe [mJOA 0-11]) and SF-36 Physical and Mental Component Summaries (PCS and MCS). At 2-years follow-up, improvement was assessed via established MCID thresholds. A supervised machine learning classification model was used to predict treatment outcomes. The highest-performing algorithm was a linear support vector machine. Leave-one-out cross-validation was utilized to test model performance. RESULTS/ANTICIPATED RESULTS: A total of 70 patients – 20 controls, 25 mild, and 25 moderate/severe CSM patients – were enrolled. Baseline clinical and DTI/DBSI measures were significantly different between groups. DBSI Axial and Radial Diffusivity were significantly correlated with baseline mJOA and mJOA recovery, respectively (r=-0.33, p<0.01; r=-0.36, p=0.02). When predicting baseline disease severity (mJOA classification), DTI metrics alone performed with 38.7% accuracy (AUC: 72.2), compared to 95.2% accuracy (AUC: 98.9) with DBSI metrics alone. When predicting improvement after surgery (change in mJOA), clinical variables alone performed with 33.3% accuracy (AUC: 0.40). When combining DTI or DBSI parameters with key clinical covariates, model accuracy improved to 66.7% (AUC: 0.65) and 88.1% (AUC: 0.95) accuracy, respectively. DISCUSSION/SIGNIFICANCE: DBSI metrics correlate with baseline disease severity and outcome measures at 2-years follow-up. Our results suggest that DBSI may serve as a valid non-invasive imaging biomarker for CSM disease severity and potential for postoperative improvement.
339 A serum exocrine enzyme as a biomarker of response to immunotherapy in type 1 diabetes.
Authors: Bruggeman Brittany S , McGrail Kieran , Gonzalez Nicole , Wasserfall Clive , Campbell-Thompson Martha , Atkinson Mark , Haller Michael , Schatz Desmond
Published: April 2022

OBJECTIVES/GOALS: We assessed the relationship between C-peptide preservation and a serum exocrine pancreatic enzyme (trypsin) in a recently concluded clinical trial. We hypothesized that immunomodulatory treatment resulting in improved beta-cell function would be associated with improved trypsin levels in subjects with recent-onset type 1 diabetes (T1D). METHODS/STUDY POPULATION: In a three-arm, randomized, double-masked, placebo-controlled trial 'Antithymocyte Globulin (ATG) and pegylated granulocyte colony stimulating factor (GCSF) in New Onset Type 1 Diabetes’ 89 subjects with recent-onset T1D (duration <100 days) were enrolled and randomized to 3 groups: low-dose ATG (2.5 mg/kg IV) followed by pegylated GCSF (6 mg subcutaneously every 2 weeks for 6 doses), low-dose ATG alone, and placebo. We compared longitudinal serum levels of an exocrine enzyme (trypsin) in a subset of responders to therapy (defined as subjects with at least 60% of baseline area under the curve (AUC) C-peptide levels at 96 weeks, n=4) versus placebo 'responders’ (n=2) and non-responders (n=25), and treated (n=19) versus placebo (n=12) subjects at baseline, 2 weeks, and 6 months after treatment. RESULTS/ANTICIPATED RESULTS: There was no observed difference in treated (n=20) versus placebo (n=12) longitudinal trends in trypsin levels when compared to baseline levels. However, responders to immunotherapy (n=4) had 6 month trypsin levels that were 114% of baseline whereas placebo subject 'responders’ (n=2), placebo subjects (n=10), and non-responders to immunotherapy (n=15) had trypsin levels that were 81-93% of baseline (unpaired T test p=0.05). Overall, we found that serum trypsin, a marker of exocrine pancreatic function, had a normal upward trend in new-onset T1D subjects who responded clinically to immunotherapy but declined in subjects who did not respond or who were not treated. These results were bordering on statistical significance but did not reach significance, likely due to the small sample size. DISCUSSION/SIGNIFICANCE: An improvement in trypsin, a marker of exocrine function, after response to immunotherapy in new-onset T1D may be due to a direct impact on exocrine function versus an indirect effect from improved beta cell function. Future studies will be needed to confirm our findings in a larger sample and evaluate the mechanism for improved exocrine function.
343 Brain-derived Extracellular Vesicles: A Novel Biomarker of CNS Metals Load with Applications in Identifying Neurodegenerative Diseases
Authors: Saxena Roheeni , Saglimbeni Brianna , Strait Madeleine , Alayyoub Mohammad , Comfort Nicole , Re Diane
Published: April 2022

OBJECTIVES/GOALS: This study aims to develop a method to examine whether blood-borne CNS-EV metal cargoes can serve as reliable biomarkers of CNS metal load and reveal a link between metal load and ALS development (i.e., neurodegenerative disease development). METHODS/STUDY POPULATION: CNS-EVs were isolated from human blood and plasma samples via direct immunoprecipitation using biotinylated antibodies for proteins known to be expressed in neurons (Contactin-2, i.e.) and astrocytes (glial-glutamate-aspartate-transporter, i.e., GLAST). Once isolated, protein concentrations in the EV samples were analyzed via ELISA assay, EV abundance was measured using ViewSizer Nanoparticle Tracking analysis, and EVs were visualized via Transmission Electron Microscopy. EVs were then analyzed for metal contents using a Perkin-Elmer NexION 350S via an ICP-MS/MS dynamic reaction cell method. RESULTS/ANTICIPATED RESULTS: Preliminary results demonstrate that it is feasible to quantify the metal contents of these CNS-derived EVs, particularly in terms of toxic metals known to be associated with neurodegenerative disorders, including copper, zinc, lead, aluminum, manganese, and iron. DISCUSSION/SIGNIFICANCE: CNS-derived EVs isolated from peripheral blood draws show promise as a potential biomarker of real-time metal load in the brain and spinal cord, with promising applications in predicting future development of neurodegenerative disorders (i.e., ALS) among patients with relevant elevated CNS metal loads.
363 School-age Morphological And Neurodevelopmental Outcomes In Patients With Sagittal Craniosynos
Authors: Fotouhi Annahita R , Said Abdullah M , Skolnick Gary B , Cradock Mary M , Naidoo Sybill D , Patel Kamlesh B
Published: April 2022

OBJECTIVES/GOALS: Several studies compare perioperative parameters and postoperative morphology of open cranial vault remodeling (CVR) and endoscopic-assisted craniectomy (EAC) in the treatment of sagittal craniosynostosis. However, data on neurodevelopmental outcomes comparing these techniques is lacking. METHODS/STUDY POPULATION: Patients with repaired sagittal synostosis were enrolled from a single tertiary care center between five and fifteen years of age. Patients with any craniofacial syndrome (e.g., 22q11.2 deletion) or other medical conditions that could affect neurodevelopment were excluded. Neurodevelopment was measured using the Differential Ability Scale-II (DAS-II). The General Cognitive Ability (GCA) score was the primary outcome measure, describing overall intellectual ability. Secondary outcome measures were measurements of cephalic index (CI) from pre- and postoperative imaging studies. Family socioeconomic status (SES) was estimated using the Hollingshead index (total score, continuous). RESULTS/ANTICIPATED RESULTS: 26 patients [10 CVR (2 Female:8 Male), 16 EAC (6 F:10 M)] were studied. Mean age at repair was greater with CVR than EAC (15.5 Â± 8.5 months vs 3.5 Â± 1.3 months, respectively, p < 0.001). Mean age at DAS-II testing was 9.0 Â± 2.2 years. Patients GCA scores were within the average range relative to test norms (CVR 95.7 Â± 15.4 vs EAC 102.6 Â± 17.4). After adjusting for SES and preoperative CI, group differences in GCA were modest and statistically nonsignificant (p = 0.646). GCA scores were associated with SES (p = 0.054) but not preop CI (p = 0.479). Differences between CVR and EAC were not statistically significant for any of the imaging parameters (pre and postop CI, pre to postop change in CI, or age at postop imaging; 0.131 <= p <= 0.867). None of the independent variables were significantly associated with postop CI (0.140 <= p <= 0.689). DISCUSSION/SIGNIFICANCE: Interim analysis of the preliminary data suggests no association between surgical procedure and cognitive and morphological outcomes at school-age in patients with sagittal craniosynostosis who underwent CVR versus EAC. Collection of data continues, with the goal to enroll 50 participants in each group by 2022.
371 Decreased Contraction Rate, Altered Calcium Transients, and Increased Proliferation seen in Patient-specific iPSC-CMs Modeling Ebsteins Anomaly and Left Ventricular Noncompaction
Authors: Samudrala Sai Suma , Anfinson Melissa , Cavanaugh Matthew , Kim Min-Su , Lamberton Peter , Radandt Jackson , Brown Ryan , Liang Huan Ling , Stamm Karl , Zeeshan Afzal , Strande Jennifer , Frommelt Michele , Lough John W. , Fitts Robert , Mitchell Michael E. , Tomita-Mitchel Aoy
Published: April 2022

OBJECTIVES/GOALS: In a familial case where 10 of 17 members inherited EA/LVNC in an autosomal dominant pattern, we discovered a novel, damaging missense variant in the gene KLHL26 that segregates with disease and comprises an altered electrostatic surface profile, likely decoupling the CUL3-interactome. We hypothesize that this KLHL26 variant is etiologic of EA/LVNC. METHODS/STUDY POPULATION: We differentiated a family trio (a heart-healthy daughter and EA/LVNC-affected mother and daughter) of induced pluripotent stem cells into cardiomyocytes (iPSC-CMs) in a blinded manner on three iPSC clones per subject. Using flow cytometry, immunofluorescence, and biomechanical, electrophysiological, and automated contraction methods, we investigated iPSC-CM differentiation efficiency between D10-20, contractility analysis and cell cycle regulation at D20, and sarcomere organization at D60. We further conducted differential analyses following label-free protein and RNA-Seq quantification at D20. Via CRISPR-Cas9 gene editing, we plan to characterize KLHL26 variant-specific iPSC-CM alterations and connect findings to discoveries from patient-specific studies. RESULTS/ANTICIPATED RESULTS: All iPSC lines differentiated into CMs with an increased percentage of cTnT+ cells in the affected daughter line. In comparison to the unaffected, affected iPSC-CMs had fewer contractions per minute and altered calcium transients, mainly a higher amount of total calcium release, faster rate of rise and faster rate of fall. The affected daughter line further had shorter shortening and relaxation times, higher proliferation, lower apoptosis, and a smaller cell surface area per cardiac nucleus. The affected mother line trended in a similar direction to the affected daughter line. There were no gross differences in sarcomere organization between the lines. We also discovered differential expression of candidate proteins such as kinase VRK1 and collagen COL5A1 from proteomic profiling. DISCUSSION/SIGNIFICANCE: These discoveries suggest that EA/LVNC characteristics or pathogenesis may result from decreased contractile ability, altered calcium transients, and cell cycle dysregulation. Through the KLHL26 variant correction and introduction in the daughter lines, we will build upon this understanding to inform exploration of critical clinical targets.
382 Strategies for commercializing non-patentable innovations developed at CTSA hubs
Authors: Quanbeck Andrew , Saha-Muldowney Mondira , Mahoney Jane , Garza-Hennessy Rose , Meline Rachael , Hirschfield Sheena
Published: April 2022

OBJECTIVES/GOALS: This presentation reports activities of a NCATS-funded collaborative working group created to promote dissemination and implementation (D&I) research within the CTSA landscape. Our working group seeks to meet both the conceptual as well as practical challenges to advancing the utilization of D&I across the translational science spectrum. METHODS/STUDY POPULATION: A fundamental focus of D&I is supporting the movement of effective health interventions into real-world use so that they benefit population health. Yet, this process remains unpredictable, with some interventions receiving widespread uptake in practice and others (of similar potential benefit) failing to translate. The value of research efforts is wasted when directed toward the “wrong” interventions. Recent discussion and experience amongst investigators in our collaborative working group has resulted in new ways of addressing this problem. Specifically, tools borrowed from business and management have shown promise in predicting which health interventions have the highest potential for commercialization and dissemination. RESULTS/ANTICIPATED RESULTS: We will conduct an environmental scan of CTSA hubs to understand their approaches to supporting commercialization and business development around research products, identifying the most promising and effective methods and processes. We will compile various tools for identifying and supporting interventions with the highest potential for commercialization, including how to form the multidisciplinary and stakeholder-engaged teams necessary to make these determinations. Finally, we will further explore the differences between patentable and non-patentable innovations and make recommendations for CTSAs in supporting the latter. DISCUSSION/SIGNIFICANCE: Commercialization of non-patentable interventions is an essential and underexplored element of the translational science spectrum. The perspectives and methods of D&I should not be relegated to late-stage translational steps, but rather inform the conduct of translational science writ large.
383 Balancing science policy and patient advocacy in medical education: the case of differences of sex development.
Authors: Jorge Juan Carlos , Trinidad-Piñeiro Jennyvette , Rodríguez-Cintrón Gisette , Ortiz-López Frances , Valerio-Perez Leidy E.
Published: April 2022

OBJECTIVES/GOALS: The clinical management of differences of sex development (DSDs) aims to guarantee best practices in medical care while addressing concerns related to non-reversible surgeries. Rhetorical analysis was conducted to study the balance between science policy and patient advocacy related to DSD surgeries as depicted in medical education materials METHODS/STUDY POPULATION: Unrestricted transcripts of two educational videos and text from all chapters of a handbook addressed to medical learners and faculty by the Association of American Medical Colleges (AAMC) were submitted to automated word cloud analysis (NVivo, QSR InternationalÂ®). Words with a weighted percentage > 0.19% from total words of a given source were defined as words of frequent use and were selected for further analysis after exclusion of words as conjunctions, prepositions, pronouns, or conversational fillers. Words sharing noun, adjective and adverb forms were coded and weighed as a single word following the Oxford dictionary. Discrepancies on word selection, exclusion or coding were resolved between four raters. The rhetorical context of most frequent words was identified. RESULTS/ANTICIPATED RESULTS: The word cloud analysis of the video resource intended for medical learners (n=104 words of frequent use) and the video intended for medical faculty (n= 94 words of frequent use) depicts a patient-centered approach (word people’) that is based on expert opinion (word [I] think’). The handbook (n= 998 words of frequent use) makes reiterated reference to patients’; lgbt’; gender’; health’; and caring’ while underscoring health concerns that are unrelated to genital variance (health’; caring’ and medical’). The noun surgery’ did not figure among the most frequent words in spoken language nor in written text even when summing its adjective and adverb forms. DISCUSSION/SIGNIFICANCE: Educational materials by the AAMC on DSDs accentuate patient-centered care within a medical humanism framework. However, the lack of discussion of DSD surgeries is an educational gap that should be addressed by key science policy and patient advocacy stakeholders.
384 Motives for kratom self-medication: contents of public comments solicited by the FDA
Authors: Tobacyk Julia , Parks Brian , Brents Lisa
Published: April 2022

OBJECTIVES/GOALS: The present study sought to investigate kratom use motives among the U.S population with the goal of discovering previously unknown health-related reasons for kratom use by the general public. METHODS/STUDY POPULATION: To guide decisions regarding kratom regulation, the FDA solicited comments from the public regarding the abuse potential of these substances, medical usefulness, and impact of scheduling changes from July 2021 until August 2021. Comment participation was open to the public. The first 6,353 consecutive comments posted on the Federal Register website were retrieved and analyzed. Duplicate comments and comments not pertaining to kratom were excluded from the analysis. The comment submissions were reviewed and categorized using an inductive approach via thematic content analysis. RESULTS/ANTICIPATED RESULTS: Respondents reported over 108 independent health-related reasons for kratom self-medication. Most often fell under the categories of mental health (1911 counts), pain management (1873 counts), substance use disorder (1635 counts), rheumatic diseases (613 counts), and degenerative spine diseases (247 counts). Many comments (701 counts) reported use for miscellaneous purposes, which included to increase focus (212 counts), treat insomnia (127 counts), and decrease fatigue (99 counts). Neurological diseases (e.g., migraines, restless legs syndrome, and multiple sclerosis) and digestive disorders (e.g., irritable bowel syndrome and Crohns Disease) were also reported (147 and 96 counts, respectively). Respondents also reported on the abuse potential and adverse effects of Kratom (122 counts). DISCUSSION/SIGNIFICANCE: This is the first study to delineate and classify motives for kratom use among Americans. Individuals reported using kratom for a wide spectrum of health-related reasons. Though these results may be influenced by the placebo effect, they suggest that kratom alkaloids may possess therapeutic activity for previously unknown applications.
396 Brain pathophysiology in SARS-CoV-2 disease
Authors: Stanic Branka Milicic , de Souza Aline M.A. , Ji Hong , Wu Xie , Speth Robert C. , Duggal Nisha K. , Ecelbarger Carolyn A. , Saavedra Juan M. , Lee Dexter L. , Sandberg Kathryn
Published: April 2022

OBJECTIVES/GOALS: The SARS-CoV-2 (Severe Acute Respiratory Syndrome CoronaVirus-2), which underlies the current COVID-19 pandemic, among other tissues, also targets the central nervous system (CNS). The goal of this study is to investigate mechanisms of neuroinflammation in Lipopolysaccharides (LPS)-treated mouse model and SARS-CoV-2-infected hamsters. METHODS/STUDY POPULATION: In this research I will assay vascular reactivity of cerebral vessels to assess vascular dysfunction within the microcirculation. I will determine expression of proinflammatory cytokines, coagulation factors and AT1 receptors (AT1R) in isolated microvessels from the circle of Willis to assess inflammation, thrombosis and RAS activity in the microvasculature. LPS and SARS-CoV-2, are both associated with coagulopathies and because of that I will measure concentration of PAI-1, von Willebrand Factor, thrombin and D-dimer to assess the thrombotic pathway in the circulation. Histology and immunohistochemistry will assess immune cell type infiltration into the brain parenchyma, microglia activation and severity of neuroinflammation and neural injury. RESULTS/ANTICIPATED RESULTS: We hypothesize that under conditions of reduced ACE2 (e.g., SARS-CoV-2 infection), AT1R activity is upregulated in the microvasculature. In the presence of an inflammatory insult, these AT1Rs promote endothelialitis and immunothrombosis through pro-thrombotic pathways and pro-inflammatory cytokine production leading to endothelial dysfunction in the microvasculature, blood brain barrier (BBB) injury, deficits in cognition and increased anxiety. We will test this hypothesis through 2 aims: Aim 1: Determine the role of the pro-injury arm of the RAS in the pathophysiology of the brain in animal models of neuroinflammation and COVID-19. Aim 1: Determine the role of the protective arm of the RAS in the pathophysiology of the brain in animal models of neuroinflammation and COVID-19. DISCUSSION/SIGNIFICANCE: This study will provide insights that will complement on-going clinical trials on angiotensin type 1 receptor (AT1R) blockers (ARBs) in COVID-19. This research is a necessary first step in understanding mechanisms of brain pathogenesis that can set the groundwork for future studies of more complex models of disease.
400 Potential Drug Therapy for Fragile X Tremor/Ataxia Syndrome
Authors: Brown Collis , Sobrian Sonya , Hudson Tamaro
Published: April 2022

OBJECTIVES/GOALS: Fragile X-associated tremor/ataxia syndrome (FXTAS) is a devastating rare neurological disorder that negatively impacts movement and cognition. To date, there are no effective pharmacological treatments for FXTAS. Our goal was to develop a cell culture model of FXTAS to investigate promising therapeutics. METHODS/STUDY POPULATION: To establish mitochondrial dysfunction, normal human cell lines and human-induced pluripotent cells were treated with multiple concentrations of glucose/ glucose oxidase (GluOx) at 2,12, and 24 hour time points to induce varying intensities of oxidative stress. The degrees of oxidative stress were measured by apoptosis and mitochondrial reactive oxygen species (ROS) production. Curcumin and MSKE compounds effective against oxidative damage in mitochondria were used to rescue glucose oxidase-induced oxidative damage in both cell lines. To test the ability of these drugs to restore mitochondrial health, cell viability and cellular superoxide production were assessed by propidium iodide and the MitoSox fluorescence assay, respectively. RESULTS/ANTICIPATED RESULTS: We anticipated that GluOx at varying concentrations and time points would proportionally increase levels of apoptosis and mitochondrial ROS, reflective of mitochondrial damage, with the most severe dysfunction occurring at a dose of 25 nM and the longest duration of 24-hr exposure. Administration of MSKE in concentrations ranging from 10-8 to 10-5 M in half log increments, did not reverse the oxidative defects induced in the cell lines. However, curcumin concentrations increased cell viability at the 2, 12, and 24 hour time period. Results indicate that the research design should be modified by increasing the concentration of both glucose and MSKE to provide a reliable test of the hypothesis. DISCUSSION/SIGNIFICANCE: These studies illustrate the usefulness of this in vitro model to test novel therapeutics in neuronal FXTAS models and expand the discovery of mitochondrial markers for the syndrome.
408 In vivo calcium imaging in the medial prefrontal cortex reveals novel site of action for therapeutic effects of Neuromedin U
Authors: Miller Sierra N. , Chapman Holly , Kasper James , Smith Ashley E. , Anastasio Noelle , Hommel Jonathan D.
Published: April 2022

OBJECTIVES/GOALS: The primary goals of this study are 1) expand our understanding of the neural circuitry influenced by the neuropeptide Neuromedin U (NMU) via its receptor Neuromedin U Receptor 2 (NMUR2), and 2) provide alternative top-down mechanisms for how NMU regulates high fat food intake and cocaine taking. METHODS/STUDY POPULATION: Immunohistochemistry (IHC) for NMUR2 and cell markers was performed on rat brain tissue containing the medial prefrontal cortex (mPFC). To identify the source of the presynaptic NMUR2, anterograde tracing from the paraventricular nucleus or dorsal raphe nucleus to the mPFC utilizing an AAV2- dsRed-synaptobrevin fusion protein were performed (n=3) and will be followed by IHC. Using in vivo calcium imaging technology (InScopix nVista), neuronal activity (calcium transients) was recorded from the mPFC of two awake, freely behaving rats. Each animal underwent a single session of 30 minutes baseline activity, intraperitoneal NMU administration, and 30 minutes of post-treatment activity. Activity was then processed and recorded as distinct events using the InScopix data acquisition software. RESULTS/ANTICIPATED RESULTS: Medial prefrontal cortex staining for NMUR2 revealed a characteristic “beads on a string” motif, consistent with presynaptic receptor expression. Furthermore, we expect the anterograde tracing experiment will show colocalization of the dsRed-synaptobrevin fusion protein with NMUR2 on synaptic inputs into the medial prefrontal cortex. Following quantification of pre- and post- treatment events using the InScopix data acquisition software, total events during the pre- and post-treatment time periods were calculated. In these studies, both animals demonstrated a clear increase in calcium transient activity between pre- and post- treatment evaluations, suggesting that NMU administration increases the neuronal activity of neurons in the prefrontal cortex. DISCUSSION/SIGNIFICANCE: This research provides a new site of action for the known therapeutic effects of NMU. We demonstrate the presence of presynaptic NMUR2 in the mPFC and show that systemic administration of NMU increases mPFC neuronal activity. This illustrates NMU may act as a top-down mediator for substance use disorders and binge eating behaviors.
416 LRP1 as a modulator of hippocampal neurogenesis and neurodegeneration
Authors: Dietert Kristi , Mahesula Swetha , Reed Pamela , Sprague Shane , Kokovay Erzsebet , Sayre Naomi
Published: April 2022

OBJECTIVES/GOALS: This project aims to elucidate the mechanism by which LRP1 governs hippocampal neurogenesis with a particular focus on its relevance in brain aging and memory loss. We are specifically interested in further discerning the intricacies of a novel relationship we have discovered between LRP1 and CXCR4 in adult neural stem cells. METHODS/STUDY POPULATION: For the in vivo studies, we are using a triple-transgenic mouse model in which we knockout LRP1 in adult neural stem cells. This is accomplished using a nestin-driven Cre-ER system in animals with floxed LRP1 and a floxed stop codon preceding a td-tomato reporter. The reporter allows for visualization of cells with the knockout and for trafficking and differentiation assays to be easily accomplished. We study stroke recovery using the middle cerebral artery occlusion model and brain aging by inducing the knockout and allowing the mice to age. We perform behavioral batteries and histological analysis on these mice to elucidate functional changes in neurogenesis. We also incorporate in vitro studies using primary neural stem cell cultures to mechanistically test the role of LRP1 in neural stem cell function. RESULTS/ANTICIPATED RESULTS: We have discovered that neural stem cell LRP1 knockout caused a 10-fold loss of CXCR4 expression in conjunction with deficits in ischemia-stimulated migration from the subventricular zone. We also found that uninjured aged mice lacking neural stem cell LRP1 displayed spatial memory deficits at 9 months of age (6 months after knockout), suggesting dysregulated hippocampal function. Given this, we hypothesize that LRP1 regulates CXCR4 in the subgranular zone NSCs to enhance hippocampal memory function. Ongoing research is testing our hypothesis via hippocampal functional tests and in vitro trafficking/expression assays. We expect our research to elucidate a previously unknown link between three independently identified effectors of neurodegenerative disease: LRP1, CXCR4, and neurogenesis. DISCUSSION/SIGNIFICANCE: The role of LRP1 in Alzheimers disease has long eluded clarity despite its known role in trafficking many major disease players – ApoE, amyloid beta and tau. Elucidating its role in hippocampal neurogenesis, a potential disease-modifying process, could lead to novel therapeutic approaches in diseases that cause the death of 1/3 of senior citizens.
420 Comparison of Statin Use to Non-Use on Cerebral Blood Flow Velocity in Older Adults at Risk for Alzheimers Disease: Data from a Phase II Multisite Clinical Trial
Authors: Aaron Stacey E. , Tomoto Tsubasa , Zhang Rong , Billinger Sandra A.
Published: April 2022

OBJECTIVES/GOALS: Reduced cerebral blood flow (CBF) along with vascular risk factors (e.g., dyslipidemia) are prevalent in Alzheimers disease (AD) and related dementias. Statins are one of the most effective pharmacologic treatments for vascular risk reduction, which may contribute to CBF in individuals with an increased risk for AD. METHODS/STUDY POPULATION: Cross-sectional analysis of 212 older adults with a family history of dementia. Heart rate via electrocardiogram, mean arterial pressure (MAP) via brachial sphygmomanometers, end-tidal CO2 via capnograph, and CBF velocity at the middle cerebral artery (MCAv) via transcranial Doppler ultrasound were collected following 20-minutes of supine rest. Mean MCAv (cm/s) was measured within each cardiac cycle and averaged over an 8-minute duration. Cerebrovascular conductance was calculated by dividing mean MCAv by MAP. Pulsatility Index was calculated by subtracting systolic MCAv from diastolic MCAv and then dividing by mean MCAv. RESULTS/ANTICIPATED RESULTS: 125 females (68 Â± 6 years; 49 statin) and 87 males (70 Â± 6 years; 47 statin) were included in analyses. There were no significant differences between heart rate, MAP, or end-tidal CO2 between statin and non-statin users. After controlling for age, sex, and low-density and high-density lipoprotein, statin use did not significantly contribute to MCAv (p = 0.09). However, statin use did significantly contribute to cerebrovascular conductance (MCAv/MAP; p = 0.03) as well as Pulsatility Index (assessment of cerebral health, p < 0.01). DISCUSSION/SIGNIFICANCE: Our findings suggest statin use significantly and positively contributes to resting cerebral blood flow velocity and cerebrovascular health. Further investigation is warranted into statin interventions with other components of cerebrovascular function, as differences may have implications for brain health and disease pathogenesis.
421 Specific cephalosporin antibiotics deplete tumor PD-L1 to inhibit DNA damage sensing and sensitize to Chk1 inhibitors in vivo
Authors: Murray Clare , Kornepati Anand , Padron Alvaro , Deng Yilun , Garcia Myrna , Bai Haiyan , Curiel Tyler
Published: April 2022

OBJECTIVES/GOALS: Tumor PDL1 signals to immune cells for tumor immune evasion but has cell-intrinsic signals that promote tumor virulence. We identify novel tumor PDL1 depleting drugs (PDDs) to interrupt tumor-intrinsic PDL1 signals and sensitize tumors to targeted therapy in vitro and in vivo. METHODS/STUDY POPULATION: We screened the Prestwick and LOPAC libraries for FDA-approved drugs reducing B16 melanoma PDL1 > 2.6-fold. ?-lactam antibiotics were used at 80 ?M and Chk1 inhibitor rabusertib as indicated in T24 human bladder cancer, and murine ID8agg ovarian cancer, 4T1 breast cancer and B16. Genetic PDL1 KO was by CRISPR or shRNA and re-expression by lentivirus. Viability was by MTT and protein by immunoblot. We challenged 5 NSG mice/group with 2x106 T24 (SQ) cells and 5 BALB/c mice/group with 5x105 4T1 cells (mammary fat pad) and treated with cefepime (200 mg/kg), rabusertib (2.5 mg/kg), vehicle, or combo daily from day 3. RESULTS/ANTICIPATED RESULTS: Structurally-related ?-lactam antibiotics cefepime and ceftazidime are tumor PDDs. Cefepime or ceftazidime reduced tumor PD-L1 and thus its cell-intrinsic signals to deplete the DNA damage sensing Chk2 protein and promote rabusertib synthetic lethality in vitro and in vivo in a tumor PDL1-dependent manner independent of immunity. Structurally distinct ?-lactam antibiotics did not sensitize tumor cells to rabusertib, suggesting ?-lactam antimicrobial functions did not promote PDL1 depletion or rabusertib treatment effects in vivo. Although rabusertib effects were immune-independent, both PDDs induced immunogenic tumor STING signaling, suggesting they can improve tumor immunotherapy. DISCUSSION/SIGNIFICANCE: We show a rapidly translatable way to deplete detrimental tumor-intrinsic PDL1 signals, and sensitize tumors to rabusertib. We are testing PDD structure activity relationships to improve PDD effects and testing PDD effects on other treatments, e.g., PARP inhibitors, immunotherapy.