ABSTRACT IMPACT: The impact of this study is that the results may lead to the development of effective educational programs and a comprehensive cancer control program while verifying patients and medical care providers adherence and compliance with cancer clinical guidelines. OBJECTIVES/GOALS: The objective of this study is to assess risk factors, preventive measures, and screening practices for human papilloma virus (HPV) associated cancers in a sub-population in Central-Eastern Puerto Rico (PR). METHODS/STUDY POPULATION: This is a sub-analysis from an annual descriptive cross-sectional questionnaire of risk factors, preventive measures, and screening practices for cancer in PR administered at a private hospital campus using a convenience sample of healthy and non-healthy adults. RESULTS/ANTICIPATED RESULTS: Out of 345 enrolled subjects in 2019 for the questionnaire, 67 were enrolled by the first author, from which 66 (19%) subjects qualified for this sub-analysis for completing the study: 79% females. When analyzing HPV risk factors, 5% of the participants were smokers. Eleven percent of the subjects received the preventive HPV vaccine. Among those non-vaccinated and eligible for vaccination, 95% were willing to get it. Seventy one percent of females 21-29 years old and 97% of 30-65 years olds had age-appropriate cervical cancer screening. DISCUSSION/SIGNIFICANCE OF FINDINGS: Despite the low prevalence of HPV vaccination, almost all of the subjects within the age range for HPV vaccination were willing to get it. Also, there was a lower prevalence of cervical cancer screening in females 21-29 years old when compared with 30-45 years old. In conclusion, there is a need for more education about HPV associated cancers and vaccine.

The availability of large healthcare datasets offers the opportunity for researchers to navigate the traditional clinical and translational science research stages in a nonlinear manner. In particular, data scientists can harness the power of large healthcare datasets to bridge from preclinical discoveries (T0) directly to assessing population-level health impact (T4). A successful bridge from T0 to T4 does not bypass the other stages entirely; rather, effective team science makes a direct progression from T0 to T4 impactful by incorporating the perspectives of researchers from every stage of the clinical and translational science research spectrum. In this exemplar, we demonstrate how effective team science overcame challenges and, ultimately, ensured success when a diverse team of researchers worked together, using healthcare big data to test population-level substance use disorder (SUD) hypotheses generated from preclinical rodent studies. This project, called Advancing Substance use disorder Knowledge using Big Data (ASK Big Data), highlights the critical roles that data science expertise and effective team science play in quickly translating preclinical research into public health impact.

OBJECTIVES/GOALS: Post-traumatic stress disorder (PTSD) develops in a subset of individuals (15-25%) exposed to trauma. We report our preliminary findings investigating underlying peripheral immune responses related to risk and resilience to PTSD. METHODS/STUDY POPULATION: Sprague-Dawley rats (half male) were exposed to predator scent stress in the form of the fox pheromone 2,5-dihydro-2,4,5-trimethylthiazoline (TMT) once for 10 minutes. Seven days later, rats were assessed for persistent anxiety-like behavior using the acoustic startle response task (ASR) and elevated plus maze (EPM). Tail blood was collected for later inflammation analysis at three time points: 1) prior to experiment start, 2) after TMT exposure, and 3) after ASR and EPM. Using a with-in subjects design, our experiment elucidates the connection between PTSD-like symptoms and baseline immune function, inflammatory stress responses, and chronic inflammatory state after stress exposure. RESULTS/ANTICIPATED RESULTS: In comparison to healthy controls, humans with PTSD show elevated blood levels of inflammatory markers. Human studies also show a relationship between baseline immune dysfunction and later PTSD development. In agreement with this literature, we anticipate PTSD-like rats will have increased blood levels of inflammation markers at all three time points compared to resilient and control rats. These findings will back-translate human findings in support of the predator scent stress preclinical model of PTSD. DISCUSSION/SIGNIFICANCE: Our findings will elucidate the temporal aspects of proinflammatory markers and PTSD development. This study will indicate if baseline peripheral immune dysfunction, inflammatory response immediately after stress exposure, and/or chronic inflammatory state predicts ptsd-like behavior in rats.

OBJECTIVES/GOALS: The aim of thispaper is to raise awareness of the limitations of the current pMRI training paradigm and to recommend a standardization of skills to expand diversity among field-based neuroimaging technicians. METHODS/STUDY POPULATION: Currently, there are seven international brain research initiatives. The goal is to establish and understand the cultural values a society holds and how the outcomes of research may be adopted into societal practice. We must also consider the benefits of early detection amongst minoritized communities in neuroscience research. Neuroimaging in remote settings can allow patients to advocate more accurately for timely medical care which can lead to better health outcomes. According to the Journal of Neurological Sciences, neuroscience accounts for 9% of total medical publications. RESULTS/ANTICIPATED RESULTS: Neuroimaging research continues to evolve in terms of resolution and portability. T By harnessing diverse data, we are able to utilize advanced neuroimaging techniques and software technology to recognize trends amongst subgroups of individuals who were previously considered geographically inaccessible. DISCUSSION/SIGNIFICANCE: Despite the excitement and promise of portable neuroimaging devices, there is a fundamental need to establish standardized training procedures that can be accessed by individuals of all socioeconomic backgrounds.

OBJECTIVES/GOALS: The goal of this study is to demonstrate growth in leadership skills in areas of milestone selection, conditions of success, and self-efficacy for females in the Capita3 Leader Launch (LL) Program. METHODS/STUDY POPULATION: Forty females were self-selected to participate in the LL Program, a 21 day program which is organized around a highly significant career goal that each participant selects. The program is designed to guide participants through a unique and transformative process to increase the probability that each participant will achieve their selected career goal. In the fall of 2022, seventy people participated in the LL program. Participants were asked to fill out a Leadership assessment prior to participation and then again following participation in the program. RESULTS/ANTICIPATED RESULTS: Immediately after the program, participants self-report 86% increased confidence, 85% being on track to achieving their selected milestone or an appropriate pivot, and 80% feel more equipped to navigate their external environment. DISCUSSION/SIGNIFICANCE: This study demonstrates that the LL Program addresses barriers that are neither fully understood nor addressed by existing programs and policies for women. Future programs can use the LL Program to help females take on high visibility assignments, larger organizational roles, and leadership positions within the current organization environment.

OBJECTIVES/SPECIFIC AIMS: Childhood obesity has become an issue of some concern worldwide. Some reviews and a recent study in adults have indicated that obesity-related inflammatory responses produce brain damage. However, studies exploring associations between inflammation and executive functions in children are overlooked. Therefore, the objective of this cross-sectional study is to determine whether difficulties in executive functions and emotional processing are associated with obesity and inflammation. METHODS/STUDY POPULATION: We have recruited 12 of a total of 60 children aged 6–8 years old. They have completed the NIH Toolbox Cognition Battery and the NEPSY II Affect Recognition tests. Samples of plasma and saliva were collected to quantify inflammatory biomarkers cytokines (IL-6 and TNF-α) assay by Luminex procedure. We performed descriptive analysis and Mann-Whitney U test to compare obese Versus nonobese groups. RESULTS/ANTICIPATED RESULTS: Obese children have lower scores in measures of affect recognition than healthy weight children. They also showed higher median scores in both salivary and plasma IL-6 and TNF-α. DISCUSSION/SIGNIFICANCE OF IMPACT: Although no statistical differences were found among groups in either measurement, these preliminary data based on the initial recruitment suggest that children with higher body mass index may have difficulties in emotional processing. More data will be available after completing recruitment to determine if the association between obesity and affect recognition is significant and if it is mediated by inflammation.

OBJECTIVES/SPECIFIC AIMS: The goal of this study is to test the hypothesis that the adherens junctions of colon epithelial cells regulate lncRNAs levels and function via the microprocessor and RISC complexes to suppress expression of pro-tumorigenic markers and aberrant cell behavior. METHODS/STUDY POPULATION: To test this hypothesis, we used colon epithelial cancer cell lines. We performed RNA-seq following knockdown of PLEKHA7, a key component of the adherens junctions, to identify changes in lncRNA expression and downstream mRNA levels. We confirmed junctional localization of affected lncRNAs from the RNA-seq and those that we found in our preliminary study by using in situ hybridization (ISH). RESULTS/ANTICIPATED RESULTS: RNA-seq identified junction-associated lncRNAs whose expression levels are regulated by PLEKHA7. The top upregulated lncRNA upon PLEKHA7 depletion was MIR17HG, an oncogenic host transcript of a cluster of miRNAs. These mature miRNAs also co-precipitate with PLEKHA7. PLEKHA7 knockdown results in increased levels of MIR17HG, but only a subset of its hosted miRNAs (miR-19a,b). Notably, miR-19a and mir-19b are highly upregulated in colon cancer. Our data suggest that 2 PLEKHA7-associated miRNAs, miR-203a and miR-372, mediate suppression MIR17HG. Re-expression of PLEKHA7 in aggressive colon cancer cells that lack PLEKHA7 suppressed expression of MIR17HG, as well as anchorage independent growth of these cells. DISCUSSION/SIGNIFICANCE OF IMPACT: Our data point towards a novel mechanism of lncRNA regulation that tethers epithelial tissue integrity with pro-tumorigenic cell transformation. Reducing elevated MIR17HG levels, is a potential therapeutic approach to suppress the tumorigenic behavior of cells that have lost their junctional integrity and homeostasis. identify a network of miRNA-mRNA-lncRNA interactions that could be exploited for further mechanistic studies, as well as for diagnostic and therapeutic purposes in the future.

Objectives/Goals: This study looks to investigate the relationship between body mass index (BMI) and the treatment effect of cancer immunotherapies. Specifically, we will assess whether there is a significant difference in survival curves associated with varying BMI levels and track trends in BMI reporting over the last decade. Methods/Study Population: An individual patient meta-analysis will be conducted by reanalyzing raw data of phase 3 cancer immunotherapy trials (2013–2023) accessed via the database Vivli. Prior to making a formal data request, an exploratory search will be first done through clinicaltrials.gov to assess viability. Studies that report baseline BMI and treatment efficacy will be included. BMI will be analyzed as a continuous variable, with survival curves compared across different BMI ranges using restricted mean survival time and log-rank tests. Trials will be stratified by drug class and adjusted for race, age, and gender to account for potential sources of confounding/bias. Results/Anticipated Results: Results are currently still a work in progress as I am in the process of getting the dataset from Vivli. I anticipate that treatment effects in cancer immunotherapies will vary significantly by BMI. Furthermore, I expect to see significant disparities in survival outcomes between patients assigned to a low and high BMI category. Lastly, trends in the reporting of BMI across immunotherapy trials are expected to be inconsistent which highlights the need for more standardization in clinical trial datasets. Discussion/Significance of Impact: This study should address critical knowledge gaps in how BMI level is associated with immunotherapy outcomes. These findings could potentially guide personalized treatment strategies and highlight the importance of standardizing the variables clinical trials chose to report.

ABSTRACT IMPACT: Despite their clinical benefits reported in patients with Parkinson’s, statin use is not associated with cost savings. OBJECTIVES/GOALS: Statins have unique lipid-lowering, anti-inflammatory and anti-oxidant benefits. Their pleiotropic benefits were shown to decrease risk of occurrene and progression of Parkinson’s disease (PD). In this study we explored whether or not statin use reflects medical or prescription cost savings. METHODS/STUDY POPULATION: Records from the Medical Panels Expenditure Survey (MEPS) database made available by the Agency for Healthcare Research and Quality were used to identify all PD subjects (n=613). Demographics and PD ICD9/10 codes, 332/G20, were abstracted from the medical condition files for all the subjects (1996-2018). Prescribed cholesterol drugs were identified based on generic and brand names following a manual review to detect any misspellings. Total medical expenses and prescription expenses were abstracted for all identified PD subjects. Subject were surveyed for two consecutive years, thus expenses were assessed for each of the two surveyed years. Costs were adjusted for inflation and expressed in 2018 dollars. The relationship between cholesterol drug use, cost and age or gender was evaluated by Fisher’s exact test. RESULTS/ANTICIPATED RESULTS: Out of the 613 PD subjects identified, 421 received no cholesterol management, 15 received non-statins, 153 received a statin and 24 received a statin-based combo therapy. While the medical expenses in the general population receiving a statin are roughly three times higher than non-statin users, no significant cost difference was noticed between PD subjects receiving or not statins. However, after adjusting for age and gender, receiving statin vs. non-statin vs combo vs none was significantly associated with total expense (p=0.017) suggesting that cholesterol management decision may play a significant role. DISCUSSION/SIGNIFICANCE OF FINDINGS: Selection of specific cholesterol treatment may play a considerable role in the overall PD expenditure. Duration of statin treatment and type of statin are expected to play a role.

OBJECTIVES/SPECIFIC AIMS: Peripheral neuropathy is the dose limiting toxicity of paclitaxel treatment. Paclitaxel pharmacokinetics (PK), specifically the Cmax and amount of time the concentration remains above 0.05 µM (Tc>0.05), have been associated with occurrence of severe, clinician-documented neuropathy. The objective of this study was to confirm that paclitaxel PK predicts progression of patient-reported neuropathy. METHODS/STUDY POPULATION: This observational trial enrolled breast cancer patients receiving weekly 1-hour paclitaxel infusions (80 mg/m2×12 cycles) at the University of Michigan Comprehensive Cancer Center. Paclitaxel concentration was measured via LC/MS in plasma samples collected at the end of (Cmax) and 16–24 hours after (Tc>0.05) first infusion. Patient-reported neuropathy was collected (EORTC CIPN20) at baseline and each cycle. The rate of neuropathy severity increase per treatment cycle is being modeled for each patient. Cmax and Tc>0.05 values will be introduced into the model to confirm that PK independently contributes to neuropathy progression. RESULTS/ANTICIPATED RESULTS: PK and neuropathy data have been collected from 60 patients for ongoing analysis. Our initial model will characterize the expected severity of neuropathy after each cycle of paclitaxel treatment. The PK-neuropathy model will include either PK parameter to validate their contribution to the progression of neuropathy severity during treatment. We anticipate, based on our preliminary analysis of the first 16 patients, that both PK parameters will significantly contribute to the model but Tc>0.05 will be more strongly associated with neuropathy progression. DISCUSSION/SIGNIFICANCE OF IMPACT: This project will generate a model that can be used to predict a patient’s neuropathy severity throughout treatment using a single, conveniently collected and easily measured PK sample during their first cycle. The next steps of this project include identifying genetic and metabolomic biomarkers that predict which patients experienced more severe neuropathy than would be anticipated based on their paclitaxel PK, and a planned interventional trial of personalized paclitaxel dosing to enhance efficacy and/or prevent neuropathy.

OBJECTIVES/SPECIFIC AIMS: The goals of this retrospective cohort study is threefold: 1) to assess how many pregnant women at Boston Medical Center from 2012 to 2017 have had their vitamin D status checked prior to and during pregnancy, 2) determine associations between vitamin D levels, birth outcomes and demographics and 3) assess how many of those found to have lower than satisfactory vitamin D levels (<30ng/mL) received interventions, including receiving vitamin D supplementation and/or being referred to an appropriate specialist such as an endocrinologist or a nutritionist. METHODS/STUDY POPULATION: Our study population is mothers over age 18 who received care at Boston Medical Center during their pregnancy from 2012 to 2017. Our primary outcomes are vitamin D utilization rates and associations between vitamin D levels with clinical outcomes during pregnancy and at birth. Secondary outcomes are demographic predictors of mothers who receive vitamin D testing and those who have complications associated with low vitamin D. We will conduct multiple linear regressions to check for associations between vitamin D levels, birth outcomes and demographic variables. We will adjust vitamin D levels with maternal BMI. De-identified clinical data was gathered from Boston University Medical Center’s (BUMC) Clinical Data Warehouse. This retrospective study was approved with a HIPAA waiver by the BUMC Institutional Data Warehouse. All statistical analysis was completed using SAS version 9.4 and was primarily done by the student PI and reviewed by Dr. Hossein, the co-investigator who is trained as a statistician and geneticist. The team also utilized Boston University’s Biostatistics, Epidemiology & Research Design (BERD) team to check the feasibility of the statistical methods. RESULTS/ANTICIPATED RESULTS: We anticipate that our descriptive demographic data will reflect the medical center’s predominantly black/Hispanic and low-income profile. Based on previous literature, we expect low vitamin D levels to have positive associations with gestational diabetes, pre-eclampsia, and preterm birth. Analyses are currently actively in progress and we expect to have results before the ACTS conference date in March, 2019. DISCUSSION/SIGNIFICANCE OF IMPACT: Vitamin D is an essential part of the human body system. It is well documented in current literature that vitamin D is correlated with bone health, mental health and maternal health. Moreover, there is evidence that maternal vitamin D supplementation prevents vitamin D deficiency in newborns. Previous literature suggests that low vitamin D may be associated with gestational diabetes, pre-eclampsia, and pre-term births. Boston Medical Center is Massachusetts’ largest urban medical center and acts as its only safety-net hospital, serving predominantly low-income and socially marginalized patient populations. There is limited existing research on assessment of maternal vitamin D in urban hospital settings. Pregnant women rarely receive vitamin D screenings as part of their prenatal checkups as current national and regional guidelines do not require pregnant women to be screened for vitamin D deficiency or insufficiency. The results will demonstrate the potential effects vitamin D supplementation, or lack thereof, in expectant mothers living in urban, safety net communities. We hope to inform prenatal care practices and attitudes of vitamin D supplementation in maternal health with the results of our study.

OBJECTIVES/GOALS: A clinical research coordinator (CRC) digital badge offers a novel approach to addressing the ongoing shortage of CRCs at academic medical centers. We sought to pilot this low-cost, short-term training digital badge to train entry level CRCs and to attract a diverse pool of individuals. METHODS/STUDY POPULATION: Experienced clinical research professionals outlined competencies needed by study coordinators working in an academic center using a Delphi approach. The competencies aligned with 6 of the 8 domains established by the Joint Taskforce for Clinical Trial Competencies. Five course modules were created in a standardized format consisting of instructional videos, handouts, and a 25-question quiz (passing grade: 90%) which was hosted in the university learning management system. Beta testers were recruited to complete surveys at the conclusion of each module addressing both module performance and content accuracy. An item analysis was performed on the 25-question quiz. RESULTS/ANTICIPATED RESULTS: Nine volunteers were recruited for the pilot. Seven completed the badge. All participants, with years of experience from 0 to over 10 in clinical trials, found the course to be informative. The majority (84%) of responses on course logistics were positive or neutral. The quiz item analysis identified most questions (86%) as either Easy or Neutral, which is ideal for a Level 1 Badge addressing foundational skills. Forty percent of the quiz questions scored high on the Discrimination Index, indicating that the quiz could identify those who performed well and those challenged by the quizzes. One negative comment received stated that course was too intense to be completed in 4 weeks. DISCUSSION/SIGNIFICANCE: The results of this pilot study demonstrated that this badging course offers a low cost, easily accessible option for recruiting and training entry level clinical research professionals. The proof-of-concept study encouraged our initiation of the program and attempt to broaden our reach.

OBJECTIVES/GOALS: Same-day discharge (SDS) after mastectomy without reconstruction (MwoR) is cost-effective with equivalent complications rates when compared to patients discharged the day after surgery. Despite this, facilities SDS rates after MwoR vary from <10%-80% (20% overall). We aimed to explore barriers to and strategies for implementing SDS after MwoR. METHODS/STUDY POPULATION: We conducted semi-structured interviews with surgeons currently performing MwoR and practicing in the state of Michigan. Recruitment was done through purposeful and snowball sampling methods. Interviews ranged from 40-60 minutes in length. The interview guide was based on the Tailored Implementation of Chronic Disease framework. Interviews were transcribed then analyzed by 3 independent reviewers. A framework matrix was created to identify common themes. RESULTS/ANTICIPATED RESULTS: Participants (n=15) included general surgeons, breast surgeons, surgical oncologists, and both university and non-university affiliated practices were represented. Surgeons either discharged patients the same-day or the day after MwoR. All surgeons who didnt routinely discharge patients the day of surgery (n=9) believed their facility had the resources to implement this practice and it would be safe for most patients. Identified barriers to same-day discharges included provider preoperative expectation setting, patient anxiety about managing their wound drain, and practice standards at their facility. Potential strategies for implementation included incorporating drain teachings with patients before surgery and explaining the benefits of same-day discharge with other providers at your facility. DISCUSSION/SIGNIFICANCE: We identified novel barriers to same-day discharge after MwoR. These barriers are potentially targetable with interventions addressing patient anxiety surrounding drain management, and education of providers across the care continuum.

OBJECTIVES/GOALS: To tackle population-level health disparities, quality dashboards can leverage individual socioeconomic status (SES) measures, which are not always readily accessible. This study aimed to assess the feasibility of a population health management strategy for colorectal cancer (CRC) screening rates using the HOUSES index and heatmap analysis. METHODS/STUDY POPULATION: We applied the 2019 Minnesota Community Measurement data for optimal CRC screening to eligible Mayo Clinic Midwest panel patients. SES was defined by HOUSES index, a validated SES measure based on publicly available property data for the U.S. population. We first assessed the association of suboptimal CRC screening rate with HOUSES index adjusting for age, sex, race/ethnicity, comorbidity, and Zip-code level deprivation by using a mixed effects logistic regression model. We then assessed changes in ranking for performance of individual clinics (i.e., % of patients with optimal CRC screening rate) before and after adjusting for HOUSES index. Geographical hotspots of high proportions of low SES AND high proportions of suboptimal CRC screening were superimposed to identify target population for outreach. RESULTS/ANTICIPATED RESULTS: A total of 58,382 adults from 41 clinics were eligible for CRC screening assessment in 2019 (53% Female). Patients with lower SES defined by HOUSES quartile 1-3 have significantly lower CRC screening compared to those with highest SES (HOUSES quartile 4) (adj. OR [95% CI]: 0.52 [0.50-0.56] for Q1, 0.66 [0.62-0.70] for Q2, and 0.81 [0.76-0.85]) for Q3). Ranking of 26 out of 41 (63%) clinics went down after adjusting for HOUSES index suggesting disproportionately higher proportion of underserved patients with suboptimal CRC screening. We were able to successfully identify hotspots of suboptimal CRC (area with greater than 130% of expected value) and overlay with higher proportion of underserved population (HOUSES Q1), which can be used for data-driven targeted interventions such as mobile health clinics. DISCUSSION/SIGNIFICANCE: HOUSES index and associated heatmap analysis can contribute to advancing health equity. This approach can aid health care organizations in meeting the newly established standards by The Joint Commission, which have elevated health equity to a national safety priority.

ABSTRACT IMPACT: Melatonin use could alleviate virus-induced respiratory illnesses. OBJECTIVES/GOALS: Melatonin was identified as a potential repurposable drug in the fight against SARS Cov-2. Its ability to attenuate some virus inoculation effects raises the question whether melatonin use could alleviate virus-induced respiratory illness. Here we evaluated the occurrence of respiratory conditions in melatonin users and non-users surveyed. METHODS/STUDY POPULATION: Records from the Medical Panels Expenditure Survey (MEPS) database made available by the Agency for Healthcare Research and Quality were used to evaluate whether melatonin may be associated with reduced viral respiratory disease burden. First, all subjects reporting melatonin use (1996-2017) were collected along with records for all subjects reporting respiratory diseases as identified by consolidated ICD-9/10 codes. Second, all diagnosis codes were retrieved for all individuals identified in the first step. In total there were 201,490 occurrences of the specified conditions among 180,468 unique individuals. The relative risk of specific respiratory disease occurrence was computed for melatonin users and non-users. Population estimates for melatonin use were also determined. RESULTS/ANTICIPATED RESULTS: Among 221 melatonin users, 132 had at least one respiratory illness. Among the 180,468 total subjects reporting at least one respiratory condition, melatonin use was associated with a lower rate of the common cold, pharyngitis, strep throat, scarlet fever, and sinusitis. Furthermore, melatonin was associated with a significantly reduced risk of common cold (RR 0.760, CI 0.587-0.985) and sinusitis (RR 0.407, CI 0.186-0.890). Due to low subject counts, the reduced risk observed for scarlet fever and strep throat was not considered significant. Melatonin users had a higher relative risk of allergic rhinitis (RR 1.393, CI 1.043-1.862) and asthma (RR 2.166, CI 1.672-2.806), probably due to melatonin active prescribing in these patients as sleep aid. DISCUSSION/SIGNIFICANCE OF FINDINGS: Although melatonin showed a lower relative risk of certain viral respiratory conditions, the low melatonin user numbers and their heterogeneous distribution over the time interval led to highly variable population estimates. Yet, our data suggests that melatonin may alleviate viral respiratory illness and deserves further investigation.

OBJECTIVES/SPECIFIC AIMS: The objective of this study was to determine if trimethylamine N-oxide (TMAO) alone could acutely alter cardiac contractile function on a beat-to-beat basis. METHODS/STUDY POPULATION: CD1 adult mouse hearts were extracted, attached to a force transducer, oxygenated, and paced within an organ bath. Changes in contractility were measured after pipetting or reverse perfusing TMAO through the aorta via a modified Langendorff apparatus to facilitate TMAO delivery into the myocardium. To determine if our findings translated to the human heart, we performed contractility experiments using human right atrial appendage biopsy tissue retrieved during cardiopulmonary bypass procedures. To investigate whether TMAO alters contractile rate, in a separate series of experiments, the atria and sinoatrial node of isolated hearts were kept intact to allow for spontaneous beating without artificial pacing and were treated with TMAO or vehicle. In addition, intracellular calcium measurements were performed on spontaneously beating embryonic rat cardiomyocytes after TMAO or vehicle treatment. RESULTS/ANTICIPATED RESULTS: We found acute exposure to TMAO, diluted into the organ bath, increased average contraction amplitude 20% and 41% at 300 µM and 3000 µM, respectively (p<0.05, n=6). Langendorff reverse perfusion of mouse hearts ex vivo with 300 µM TMAO generated an even greater response than nonperfusion peripheral exposure and increased isometric force 32% (p<0.05, n=3). Consistent with what we observed in mouse hearts, incubation of human atrial muscle tissue with TMAO at 3000 µM increased isometric tension 31% compared with vehicle (p<0.05, n=4). TMAO treatment (3000 µM) also increased average beating frequency of ex vivo mouse hearts by 27% compared with vehicle (p<0.05, n=3) and increased the spontaneous beating frequency of primary rat cardiomyocytes by 13% compared with vehicle treatment (p<0.05, n=4). DISCUSSION/SIGNIFICANCE OF IMPACT: TMAO, at pathological concentrations, directly increases the force and rate of cardiac contractility. Initially, these inotropic and chronotropic effects may be adaptive during CKD; however, chronic increases in isometric tension and beating frequency can promote cardiac remodeling and heart failure. Further translational studies are needed to understand the intricate relationship between the microbiome, kidneys, and heart and to examine if TMAO represents a therapeutic target for reducing cardiovascular mortality in CKD patients.