The Mount Sinai Health Hackathon is designed to provide a novel forum to foster experiential team science training. Utilizing a Social Network Analysis survey, we studied the impact of the Mount Sinai Health Hackathon on the nature of collaborative relationships of hackathon participants. After the event, the number of links between participants from different disciplines increased and network density overall increased, suggesting a more interconnected network with greater interdisciplinary communication. This social network approach may be a useful addition to the evaluation strategies for team science education initiatives.

ABSTRACT IMPACT: To track recovery and mitigate additional spinal cord injury (avoiding further paralysis), we are assessing the applicability of an implantable ultrasound device that can monitor the tissue health postoperatively. OBJECTIVES/GOALS: To date, no method has been developed that monitors spinal cord perfusion rate (mL/min/g) or pressure (mmHg) successfully after the surgery. Our goal is to design, construct, and validate (in animal models) a novel sensor that quantifies postoperative tissue perfusion in patients with SCI at the site of and downstream from the injury. METHODS/STUDY POPULATION: A sample size of 10 animals will allow us to test our hypothesis to track tissue perfusion before and after the SCI using ultrasound. After prepping and scrubbing the animal, the skin will be incised with a blade, bony structures will be removed and the spinal cord will be revealed. A 25-g weight will then be dropped from a height of 15 cm, and the animal will be observed for contraction of the lower extremities, a sign that the cord was damaged. Using Doppler ultrasound settings available on commercial transducers, we will investigate the acceptable frequency, as well as proper Doppler mode with and without contrast agents, and with and without elastography (stiffness mapping of the tissue). A range of frequencies will be tested (5 -25 MHz). RESULTS/ANTICIPATED RESULTS: It is expected that at frequencies 12 MHz and above, our radiologist collaborators would be able to easily detect the blood flow. It is also expected that the injury will have a noticeable effect on the changes of this detected blood flow. We aim to present figures demonstrating ultrasound image qualities obtained at various frequencies. We expect three such figures: one for gray scale ultrasound imaging, one for color Doppler and finally, one for spectral Doppler, which is the one mostly used to quantify blood flow. DISCUSSION/SIGNIFICANCE OF FINDINGS: To monitor recovery and mitigate secondary injury in patients with traumatic SCI, there is a need to monitor tissue perfusion intra-operatively. To address this need, we will design, construct, and validate a novel sensor that will postoperatively quantify tissue perfusion for the SCI patients at the site of the injury.

OBJECTIVES/SPECIFIC AIMS: Most incidents of partner violence occur when one or both partners have been drinking, however, the mechanism through which this association exists is unclear. The neural circuits that support self-regulation of emotion and social behavior, as well as autonomic influences on the heart, are co-localized in the brain and represent an integrated bidirectional regulatory system. These physiological regulatory processes are mediated by a neural substrate known as the central autonomic network which includes the peripheral autonomic nervous system. The central autonomic network modulates biobehavioral resources in emotion by flexibly responding to physiological arousal in response to changing situational demands, and serves a fundamental role in emotion regulation and goal-directed motor behavior, and this circuit can be indexed with heart rate variability (HRV). METHODS/STUDY POPULATION: In total, 17 distressed violent (DV) partners (11 females, 6 males) were matched to a sample of distressed nonviolent (DNV) partners (7 female, 6 males) were matched on age, sex, and relationship satisfaction and participated in a placebo-controlled alcohol administration study with an emotion-regulation task during which electroencephalography, HRV, and galvanic skin response (GSR) measures were collected. In the alcohol condition, participants were administered a mixture of 100 proof vodka and cranberry juice calculated to raise their blood alcohol concentration to 0.08%. In the placebo condition, participants consumed a volume of juice equivalent to that consumed in the alcohol condition, but without alcohol. Alcohol and placebo conditions were counter-balanced across participants as were the presentation the blocks of evocative and neutral partner stimuli and emotion-regulation condition (watch vs. do not react). RESULTS/ANTICIPATED RESULTS: Results show that DV partners show greater cortical arousal than DNV partners on measures event-related spectral perturbations, which are mean log event-loced deviations from baseline-mean power at each frequency of the electroencephalography power spectra, when intoxicated and viewing evocative partner stimuli in the “do not react” emotion regulation condition. Results also show a statistically significant 2 (alcohol vs. placebo)×2 (watch vs. do not react)×2 (DV partners vs. DNV partners) interaction of the respiratory sinus arrhythmia measure of HRV when viewing evocative partner behavior (F=7.102, p=0.019, partial η2=0.353). Findings indicate that DV partners have lower HRV than DNV partners across conditions, but particularly when acutely intoxicated and trying not to react to their partners’ evocative behavior. Similarly, results also show a statistically significantly 2 (alcohol vs. placebo)×2 (watch vs. do not react)×2 (DV partners vs. DNV partners) interaction on GSR (F=71.452, p=0.000, partial η2=0.749). GSR findings indicate that DV partners also have lower GSR when acutely intoxicated and trying not to react to their partners’ evocative behavior. DISCUSSION/SIGNIFICANCE OF IMPACT: These results suggest that increases in intimate partner violence under acute alcohol intoxication may be the result of dysfunction of the central autonomic network, especially when DV partners are trying to suppress a behavioral response to their partners’ evocative behavior in conflict. The neurophysiological patterns evidenced by DV partners is consistent with a state of vigilance to threat, and reduced ability inhibit prepotent, but inappropriate responses. They also suggest that HRV may be an important target for intervention with partner with a history of intimate partner violence. One method may be heart rate variability biofeedback which has been shown to increase parasympathetic nervous system functioning, autonomic stability, and emotion regulation.

OBJECTIVES/GOALS: Flavored tobacco sales restrictions (FTSR) may reduce youth tobacco use, but may not be as effective in areas with greater tobacco retailer density (TRD), which is associated with greater tobacco access and more common in low-income areas. We examined the association between FTSRs and e-cigarette use for youth in high and low TRD cities. METHODS/STUDY POPULATION: We analyzed data from the California Healthy Kids Survey using a difference-in-differences (DID) strategy. We compared pre- and post-policy changes in ease of access to e-cigarettes, current use, and ever use one year after implementation among students (9th and 11th graders) attending school in a city with a FTSR (n=20,832) versus without (n=66,126). Separate analyses were conducted for students in cities with low and high TRD, with a median cutoff of 3.3 tobacco retailers per square mile. RESULTS/ANTICIPATED RESULTS: Students attending schools in cities with high TRD (compared with low TRD) had a higher percentage of parents with a high school education or less, and were more likely to identify as Hispanic or Non-Hispanic Asian/Pacific Islander. Among students with low TRD, FTSRs were associated with reduced ease of access to e-cigarettes (DID=0.76, 95% CI: 0.58, 0.99). However, among students with high TRD, FTSRs were associated with increased ease of access (DID: 1.25, 95% CI: 1.02, 1.56) and current use (DID=1.57, 95% CI: 1.31, 1.87). DISCUSSION/SIGNIFICANCE: FTSRs were associated with lower youth e-cigarette access in low, but not high TRD areas. Stronger policies or enforcement may be needed in high TRD areas.

OBJECTIVES/GOALS: Antibiograms are used to guide empiric antibiotic selection. However, it is unclear if antibiotic profiles differ between symptomatic urinary tract infections (UTIs) and asymptomatic bacteriuria (ASB). We aimed to compare antibiotic susceptibility profiles of urinary E. coli isolates from patients with a symptomatic UTI to those with ASB. METHODS/STUDY POPULATION: We conducted a cohort study of 1,140 urinary E. coli isolates from unique patients that received care through Vanderbilt University Medical Center (VUMC) from Nov 2020 – Jun 2021. We included any patient that was seen at VUMC as an inpatient, outpatient or at the emergency department with ≥ 105 colony forming units/mL E. coli detected from a clinical urine specimen. Chart abstractions were performed to capture reported UTI symptoms and demographic information. Descriptive statistics were conducted to compare antibiotic susceptibility profiles (i.e., susceptible, intermediate, resistant) between symptomatic and ASB groups. The risk of detection of a multidrug-resistant organism (MDRO) (intermediate, or resistant to at least one antibiotic in three or more classes) was assessed between groups. RESULTS/ANTICIPATED RESULTS: Among 1,140, 1,018 (89%) and 122 (11%) were symptomatic and ASB, respectively. When comparing symptomatic and ASB, the median ages were 50 and 46. Groups had similar proportions of no indwelling catheter (94% v. 95%) and without diabetes (87% v. 88%). The collection setting between inpatient, emergency department, and outpatient were similar with most being outpatient (79% v. 83%). The proportion of patients who were pregnant, immuno compromised, or had a structural/functional urinary tract abnormality were higher in the symptomatic group. The proportion of isolates resistant and susceptible to tested antibiotics were similar between groups, with only ciprofloxacin showing slightly higher resistance among ASB (16% v. 25%). The risk of MDRO detection was similar between groups (RR: 0.858, 95% CI: 0.64, 1.15). DISCUSSION/SIGNIFICANCE: Antibiotic susceptibility comparison demonstrated similar profiles, which suggests antibiogram use as appropriate to guide ASB treatment. Results offer insight on whether traditional methods for assessing antibiotic susceptibility on population-levels could benefit from further refinement by patient-specific clinical parameters.

OBJECTIVES/GOALS: Pacific Islanders have some of the lowest rates of initiation and duration of exclusive breastfeeding compared to other racial women once they migrate to the United States. The purpose of this study is to identify infant feeding initiation, beliefs, and experiences of Marshallese women living in the United States soon after delivery. METHODS/STUDY POPULATION: This study used an exploratory, descriptive qualitative design with 36 Marshallese women residing in the United States from July 2019 to July 2020. A Community Based Participatory Research Approach was used to design and implement the study. This approach enables the respect and integration of Marshallese cultural values and practices into every aspect of the research All study plans and documents – including recruitment plans and forms, consent plans and forms, retention plans, quantitative surveys, and qualitative interview guides – were developed in partnership with Marshallese research team members. RESULTS/ANTICIPATED RESULTS: Two themes emerged: 1) Infant Feeding Initiation and Practices; and 2) Concerns of Breastfeeding in Public. Within the first theme, two subthemes emerged: 1) Breast and Formula-Feeding; and 2) Return to Work. Within the second theme, two subthemes emerged: 1) Personal Beliefs about Breastfeeding in Public; and 2) Acculturation and Breastfeeding in Public. DISCUSSION/SIGNIFICANCE: This is the first study to document beliefs about exclusive breastfeeding initiation among Marshallese women living in the United States. Findings from this study can be used to develop health education programs to improve exclusive breastfeeding and can be used to stimulate future research in the area of breastfeeding among Pacific Islander women.

OBJECTIVES/GOALS: My research aims to discover African American breast cancer genetic risk factors. Interested in genetic predisposition, I search for inherited variants that could explain why African American women are disparately diagnosed at younger ages and with aggressive subtypes compared to other ethnicities. METHODS/STUDY POPULATION: Our study cohort, the Alabama Hereditary Cancer Cohort (AHCC), consists of African Americans that have had a breast cancer diagnosis indicative of hereditary breast cancer. Whole genome sequencing is conducted for AHCC breast cancer cases. Hypothesizing that African American-specific protein-truncating variants explain inherited risk, our control cohort consists of whole exome sequencing data of (~2500) African Americans from the Type 2 Diabetes Exome Sequencing Project on dbGAP. Single variant and gene-based association tests are being conducted to identify risk variants/genes. Prime editing is conducted to introduce risk variants into cancer cell lines for functional analyses. RESULTS/ANTICIPATED RESULTS: Preliminary studies, involving 60 breast cancer cases, have already revealed multiple African American-specific genetic variants in the nuclear and mitochondrial genome that are statistically linked to breast cancer risk. We are in the process of increasing our breast cancer sample size, aiming for significantly higher confidence. Prime editing for selected novel variants has begun in breast cancer cell lines. Functional assays will then be carried out to observe differences in cell proliferation, cell migration, and spheroid formation in the genetically edited compared to unedited cell lines. DISCUSSION/SIGNIFICANCE: African Americans are underrepresented in breast cancer research. This study reduces research participation gaps and identifies genetic risk variants, leading to better risk assessments and screening methods. Such discoveries can also lead to new therapeutic targets, reducing breast cancer deaths.

Objectives/Goals: The goal of this proposal is to better understand how informing African Americans of their genetic risk affects their behavior as part of a cardiovascular disease (CVD) risk reduction intervention. Aim 1: To determine the effect of genetic risk knowledge on CVD health behavior. Aim 2: To determine the effect of genetic risk knowledge on secondary variables. Methods/Study Population: Method: Fifty participants from the Baton Rouge metropolitan area will be recruited. Participants must be African American adults over the age of 18. Potential participants will be recruited using community-based efforts that have been successful in recruiting this population specifically. Participants will be randomized into one of two groups. Genetically Unblinded Group (GU) will be “genetically unblinded” after baseline orientation. Genetically Blinded Group (GB) will be “genetically blinded” until the end of the study. This study design ensures that we can measure the impact of knowledge of genetic risk on participant behavior. Results/Anticipated Results: Baseline participants’ characteristics (body mass index, blood glucose, and cholesterol) will be summarized by intervention group, with counts and percentages for categorical variables and means and 95% confidence intervals for continuous variables. Primary Outcomes: Attendance in intervention sessions will be counted across groups. Effect on genetic risk knowledge will be determined via comparing the difference between the increased healthy lifestyle behaviors at endpoint between Genetically Unblinded (Cases) and Genetically Blinded Groups (Control). Secondary and Tertiary Outcomes: Mean change in secondary outcomes in the GU group will be compared against the mean change in the GB group. Participant’s survey responses and changes in physical measurement from baseline to endpoint will be observed. Discussion/Significance of Impact: This study empowers African Americans in Baton Rouge by providing genetic risk knowledge for cardiovascular disease. By addressing social determinants of health, it promotes behavior change, improves health outcomes, and fosters trust, potentially reducing health disparities and advancing health equity.

ABSTRACT IMPACT: Baseline presentation in AT (higher upper trapezius PPT with no difference at calf or tendon) may suggest a mechanism for persistent symptoms: with more advantageous central pain processing and no tradeoff peripherally, they may choose to continue their usual activities without regard for further damage to the affected tendon. OBJECTIVES/GOALS: Exercise-induced hypoalgesia, a reduction in pain with exercise, is often observed in healthy populations but is not well established in Achilles tendinopathy (AT). The aim was to compare pressure-pain threshold (PPT) at baseline and after fatiguing isometric exercise in AT and healthy controls. METHODS/STUDY POPULATION: 21 participants were recruited for the study: 7 AT (26.5 ±8.8 yrs), 14 control (22.1 ±3.2 yrs). After a familiarization session, participants completed an experimental session that involved performance of intermittent maximal voluntary isometric contractions (MVICs) (2s:2s duty cycle) in a Biodex3 dynamometer (Biodex Medical, Shirley, NY) for 4 minutes. PPT was measured at the medial gastrocnemius (calf), Achilles tendon, and upper trapezius at baseline and immediately following the fatiguing isometric task using a Somedic Algometer (Somedic AB, Sweden). Data are expressed as Mean(SD). Change in PPT is expressed as a percentage of baseline PPT. Units for PPT are kPa. A priori alpha was set to 0.05. RESULTS/ANTICIPATED RESULTS: There was no change in tendon or calf PPT following isometric exercise in AT (tendon: p=0.78; calf: p=0.76), while both increased (i.e., exercise-induced hypoalgesia) in controls (tendon: 9.5(17.8), p=0.03; calf: 21.3(22.7), p<0.01). Neither group experienced a post-exercise change in upper trapezius PPT (AT: p=0.35; control: p=0.37). There was no between-group difference in baseline calf (p=0.14) or tendon (p=0.19) PPT. However, baseline and post-exercise upper trapezius PPT were significantly higher in AT (baseline: 335.6(194.8); post-exercise: 321.2(170.1)) than in controls (baseline: 193.7(75.1), p<0.01; post-exercise: 198.1(79.1), p<0.01). DISCUSSION/SIGNIFICANCE OF FINDINGS: These findings suggest: (1) in persons with AT, central pain processing is altered at baseline, but unaffected in response to isometric fatiguing exercise; and (2) in persons with AT, peripheral pain processing is unaffected at baseline, but is altered in response to this mode and dosage of fatiguing isometric exercise.

OBJECTIVES/SPECIFIC AIMS: Recent evidence from resting-state fMRI studies have shown that brain network connectivity is altered in patients with neurodegenerative disorders. However, few studies have examined the complete connectivity patterns of these well-reported RSNs using a whole brain approach and how they compare between dementias. Here, we used advanced connectomic approaches to examine the connectivity of RSNs in Alzheimer disease (AD), Frontotemporal dementia (FTD), and age-matched control participants. METHODS/STUDY POPULATION: In total, 44 participants [27 controls (66.4±7.6 years), 13 AD (68.5.63±13.9 years), 4 FTD (59.575±12.2 years)] from an ongoing study at Indiana University School of Medicine were used. Resting-state fMRI data was processed using an in-house pipeline modeled after Power et al. (2014). Images were parcellated into 278 regions of interest (ROI) based on Shen et al. (2013). Connectivity between each ROI pair was described by Pearson correlation coefficient. Brain regions were grouped into 7 canonical RSNs as described by Yeo et al. (2015). Pearson correlation values were then averaged across pairs of ROIs in each network and averaged across individuals in each group. These values were used to determine relative expression of FC in each RSN (intranetwork) and create RSN profiles for each group. RESULTS/ANTICIPATED RESULTS: Our findings support previous literature which shows that limbic networks are disrupted in FTLD participants compared with AD and age-matched controls. In addition, interactions between different RSNs was also examined and a significant difference between controls and AD subjects was found between FP and DMN RSNs. Similarly, previous literature has reported a disruption between executive (frontoparietal) network and default mode network in AD compared with controls. DISCUSSION/SIGNIFICANCE OF IMPACT: Our approach allows us to create profiles that could help compare intranetwork FC in different neurodegenerative diseases. Future work with expanded samples will help us to draw more substantial conclusions regarding differences, if any, in the connectivity patterns between RSNs in various neurodegenerative diseases.

OBJECTIVES/GOALS: Aging is the biggest risk factor for cancer, yet little is known about cancer immunotherapy effects. Here we investigate melanoma response to αPD-1, αPD-L1 and αPD-L2 in young vs. aged hosts. We look at different immune outcomes as possible mechanism. METHODS/STUDY POPULATION: We tested αPD-1 (100 μg/mouse), αPD-L1 (100 μg/mouse) or αPD-L2 (200 μg/mouse) in aged (18-24 months) and young (3-8 months) mice challenged orthotopically with B16. Tumors and draining lymph nodes (TDLN) were analyzed by flow. Bone marrow-derived DC were generated with GM-CSF. RESULTS/ANTICIPATED RESULTS: We reported that αPD-1 treats young and aged with B16 and αPD-L1 only treats young. aPD-L2 treated B16 in aged but, remarkably, not young, the first anti-cancer single agent immunotherapy exhibiting this property. Efficacy in young (aPD-1, aPD-L1) and aged (aPD-1, aPD-L2) correlated with increased T cell stem cells (TCSC) and total tumor-infiltrating lymphocytes (TIL), but TCSC differed by age and treatment (e.g., distinct CCR2, CXCR5, CXCR3, PD-1 and TIM- expression). Aged expressed significantly more T-cell PD-1 and up to 40-fold more PD-L2 versus young in myeloid and NK cells, and TCSC. Bone marrow-derived DC experiments suggest aged DC are destined for high PD-L2 versus young. DISCUSSION/SIGNIFICANCE OF IMPACT: Treatment differences in aged vs. young could depend on immune checkpoint or TCSC differences. We are now identifying mechanisms for increased PD-L2 and contributions to aPD-L2 efficacy in aged, and testing TCSC effects. Our work can improve cancer immunotherapy in aged hosts and further provide important insights even in young hosts.

OBJECTIVES/SPECIFIC AIMS: The objective of this study is to assess the efficacy of combined AF and NP repairs to prevent degenerative changes and restore native disc morphology in an in vivo large animal model. We hypothesize that combined repairs will prevent disc degeneration following injury to a greater extent than the individual repairs after 6 weeks in vivo, as demonstrated through disc height measurements and disc morphology. METHODS/STUDY POPULATION: A total of 8 skeletally mature female Finn sheep were used in this study. Following a previously described method, IVDs from L1 to L6 of the lumbar spine were exposed using a lateral access, extraperitoneal approach5. IVDs were randomized into 5 treatment groups: 1) intact discs, 2) discs injured via a 3 cm x 1 cm box annulotomy and partial nucleotomy, 3) injury followed by a high density collagen (HDC) AF patch, 4) injury followed by injection of a modified hyaluronic acid (HA) into the NP, and 5) injury followed by both the HDC AF patch and HA NP injection. The HDC treatment was 15 mg/mL type-I collagen mixed with 0.06mM riboflavin, injected at the defect site and crosslinked in situ with blue light. The NP injection was HA modified with C16 side chains to increase the viscosity of the hydrogel (HYADD 4®)6. At 6 weeks post-operatively, sheep were sacrificed and had 3T magnetic resonance images (MRI) taken of their lumbar spine. Disc height analysis and Pfirrmann grading were performed on each segment using MR images. Additionally, quantitative MRI analyses were performed using a custom MATLAB algorithm that segments NP from the surrounding tissue and directly measures the NP volume. ANOVA with Tukey’s HSD was used to determine statistical significance between groups for disc height and quantitative MRI analyses, and the Kruskal-Wallis test with Mann-Whitney tests was used to statistically analyze Pfirrmann Grades. All animal use followed approved IACUC protocol. RESULTS/ANTICIPATED RESULTS: As shown in axial MR images (Figure 1A), intact discs had hyperintense NP with a distinct border to the AF. The discs receiving injury with no treatment had hypointense NP with no distinct border between the AF and NP. Individual and combined treatment with the HA NP injection and HDC AF patch appeared to preserve the hyperintense NP signal and AF/NP border. Intact control discs were not degenerated and had an average Pfirrmann grade of 1 (Figure 1B), while injured, untreated discs had significant degeneration with an average Pfirrmann grade of 3. Discs receiving the HA NP injection and collagen AF patch individually showed fewer signs of degeneration than the injured alone, and the combined treatment resulted in the least amount of degeneration with Pfirrmann grades not significantly different than the intact controls. Disc height index confirmed the trends seen in the Pfirrmann grades (Figure 1C), where injured discs lost 20% of the intact disc height, the individual NP and AF repairs restored 5-10% of intact disc height, and the combined repairs preserved 90% of the intact disc height. The NP voxel count of all treatment groups were similar to the intact controls (Figure 1D). DISCUSSION/SIGNIFICANCE OF IMPACT: The objective of this study was to determine how combined AF and NP can prevent degenerative changes to the disc in a large animal in vivo model. Pfirrmann grading and disc height index results show that the greatest preservation of disc morphology was seen with combined AF and NP repairs, while the individual strategies prevented degenerative changes better than injury with no treatment. It appears the HA NP injection restores lost water content to the disc following injury, and the AF collagen patch plays a role in maintaining the NP repair within the disc. This is the first study to our knowledge to attempt combined AF and NP repairs in an in vivo large animal model. Combining NP and AF repairs leads to significantly improved outcomes following disc injury, which warrants the translation of combined repairs into the clinic to improve patient outcomes with degenerative disc disease involving NP and AF.

OBJECTIVES/SPECIFIC AIMS: Neurologic disorders are among the most significant health challenges facing society today. Although different neurologic disorders are often thought to be distinct from one another, evidence suggests similar processes may contribute to pathology in different diseases. Previous studies suggest that common disease mechanisms contribute to the development of epilepsy and tauopathy. The purpose of this study is to better characterize this relationship and explore potential therapeutic avenues to slow disease progress. METHODS/STUDY POPULATION: This study uses the pilocarpine-induced status epilepticus model of temporal lobe epilepsy to explore the effect of severe seizures on tau pathology. Brains were collected from mice at 6 or 24 hours after induced status epilepticus. Homogenates were analyzed via Western blot to look for changes in tau phosphorylation or activity of two major regulators of tau phosphorylation, GSK3β and PP2A. These data show that changes in tau phosphorylation dynamics occur at a much earlier time point after status epilepticus than has previously been described. RESULTS/ANTICIPATED RESULTS: GSK3β activity increased within 6 hours and remained elevated by 24 hours. PP2A activity initially decreased but returned to normal by 24 hours. These data show that changes in tau phosphorylation dynamics occur at a much earlier time point after status epilepticus than has previously been described. DISCUSSION/SIGNIFICANCE OF IMPACT: The current project supports previous observations that seizures promote tau phosphorylation in vivo, but suggests that changes begin much earlier than previously thought. Further work is needed to understand how post-seizure changes in tau phosphorylation develop over longer periods of time. Additionally, future work will characterize the effect of tauopathy on electrical activity in vivo and in vivo.

OBJECTIVES/SPECIFIC AIMS: Prostate cancer is the second most common cancer in the world for men. For reasons still unclear, aggressive PCa disproportionately affects males of African ancestry (MoAA). Incidence and mortality rates are highest in MoAA as they have consistently shown a 2.3–3.0-fold higher risk of mortality compared with Caucasian men. This aggressiveness of PCa may be due to specific biological factors. Studies have established that microRNAs (miRNAs), essential in post-transcriptional gene regulation, are dysregulated in PCa. miR-1207-3p is encoded at the PVT1 gene locus, which is located downstream of MYC on the 8q24 PCa susceptibility locus. The chromosomal region 8q24 is associated with aggressive PCa. Yet miR-1207-3p in PCa in MoAA has never been investigated. Moreover, studies have shown that PVT1/MYC co-operation is a fundamental feature in all cancers with 8q24 amplification and 98% of the 8q24 amplicons contained concurrent amplification of the MYC and PVT1 loci. Moreover, MYC has been linked to PCa aggressiveness, and has been reported to be downstream of androgen receptor (AR) in some PCa. However, the mechanisms regulating c-MYC have never been studied in MoAA. We have recently demonstrated that miR-1207-3p has prognostic value in PCa, and directly binds to the 3′ UTR of Fibronectin type III domain containing 1 (FNDC1) to regulate a novel FNDC1/fibronectin (FN1)/AR pathway upregulated in metastatic PCa. However, the relevance of this novel and clinically significant miR-1207-3p molecular pathway in PCa in MoAA is unknown. Therefore, we hypothesized that miRNA 1207-3p, encoded at the 8q24 PCa susceptibility locus, is a PCa biomarker with clinical applications in MoAA. Our specific aim was to assess the clinical relevance of miR-1207-3p, FNDC1, FN1, AR, and MYC expression in aggressive PCa in a cohort of West African Black males. METHODS/STUDY POPULATION: Consequently, we aimed to determine the expression profile of miRNA-1207-3p, FNDC1, FN1, AR, and MYC in histologically confirmed normal prostate (n=24), benign prostate (n=44) and malignant prostate tissue (n=29) from prostatectomy or transrectal ultrasound-guided biopsies in patients recruited at the University College Hospital, Ibadan, Nigeria, a sub-Saharan Black African population. In total, 17 patients had tumor tissues with Gleason score ≥8. Tissues were collected in compliance with Institutional Ethics Board approved protocols. RNA extraction, cDNA synthesis, and quantitative-PCR were performed to analyze mRNA expression of miRNA-1207-3p, FNDC1, FN1, AR, and MYC. Statistical analysis were performed using SPSS software. All data were analyzed by the 1-way ANOVA test. Tukey post-hoc test was performed to determine the differences in mean expression between normal and tumor prostate tissues. p<0.05 were considered significant. RESULTS/ANTICIPATED RESULTS: We discovered that miR-1207-3p is significantly underexpressed in prostate tumor tissues [0.09±0.02, 95% CI (0.04, 0.136), p=0.000] in comparison with normal prostate tissue in MoAA [0.92±0.15, 95% CI (0.60, 1.244), p=0.000]. This is the first description of miR-1207-3p differential expression in human clinical PCa in MoAA. In contrast, FNDC1 was significantly overexpressed in prostate tumor tissues [21.93±8.21, 95% CI (4.97, 38.89), p=0.003] in comparison with normal prostate tissues in MoAA [1.57±0.45, 95% CI (0.625, 2.51), p=0.003]. The same positive correlation with advanced disease held true for FN1 [tumor: 13.66±3.53, 95% CI (6.38, 20.93), p=0.000; normal: 1.07±0.235, 95% CI (0.58, 1.56), p=0.000], AR [tumor: 20.49±6.74, 95% CI (6.50, 34.48), p=0.000; normal: 0.94±0.20, 95% CI (0.52, 1.36), p=0.000], and c-MYC [tumor: 33.93±8.43, 95% CI (16.53, 51.33), p=0.000; normal: 1.94±0.36, 95% CI (1.18, 2.70)]. The significantly increased mean expression for FNDC1, FN1, AR, and c-MYC in prostate tumor tissues in comparison with normal prostate tissues indicate that their overexpression is associated with increased risk of cancer progression in MoAA. DISCUSSION/SIGNIFICANCE OF IMPACT: These results show that the underexpression of miR-1207-3p and the overexpression of FNDC1, FN1, AR and MYC is associated with aggressive PCa in MoAA. miR-1207-3p, and it molecular target FNDC1, may be useful biomarkers for prognostication of PCa in MoAA.

OBJECTIVES/SPECIFIC AIMS: The aims of this study are (1) to develop and characterize a novel nonhuman primate model of pneumococcal pneumonia that mimics human disease; and (2) determine whether Streptococcus pneumoniae can: (a) translocate to the heart, (b) cause adverse cardiac events, (c) induce cardiomyocyte death, and (d) lead to scar formation during severe pneumonia in baboons. METHODS/STUDY POPULATION: Six adult baboons (Papio cynocephalus) were surgically tethered to a monitoring system to continuously assess their heart rate, temperature, and electrocardiogram (ECG). A baseline transthoracic echocardiogram, 12-lead ECG, serum troponin-I levels, brain natriuretic peptide, and heart-type fatty acid binding protein (HFABP) levels were obtained before infection and at the end of the experiment to determine cardiovascular damage during pneumococcal pneumonia. Animals were challenged with 108 colony-forming units of S. pneumoniae in the right middle lobe using flexible bronchoscopy. Three baboons were rescued with ampicillin therapy (80 mg/kg/d) after the development of pneumonia. Cardiac damage was confirmed by examination of tissue sections using immunohistochemistry as well as electron and fluorescence microscopy. Western-blots and tissue staining were used to determine the presence of necroptosis (RIP3 and pMLKL) and apoptosis (Caspase-3) in the cardiac tissue. Cytokine and chemokine levels in the heart tissue were determined using Luminex technology. RESULTS/ANTICIPATED RESULTS: Four males (57%) and three (43%) females were challenged. The median age of all baboons was 11 (IQR, 10-19) years old, which corresponds to a middle-aged human. Infected baboons consistently developed severe pneumonia. All animals developed systemic inflammatory response syndrome with tachycardia, tachypnea, fever, and leukocytosis. Infection was characterized by initial leukocytosis followed by severe leukopenia on day 3 postinoculation. Non-specific ischemic alterations by ECG (ST segment and T-wave flattering) and in the premortem echocardiogram were observed. The median (IQR) levels of troponin I and HFABP at the end of the experiment were 3550 ng/mL (1717–5383) and 916.9 ng/mL (520.8–1323), respectively. Severe cardiomyopathy was observed using TEM and H&E stains in animals with severe pneumonia. Necroptosis was detected in cardiomyocytes of infected animals by the presence of pMLKL and RIP3 in cardiac tissues. Signs of cardiac remodeling indicated by disorganized collagen deposition was present in rescued animals but not in the other animals. DISCUSSION/SIGNIFICANCE OF IMPACT: We confirmed that baboons experience cardiac injury during severe pneumococcal pneumonia that is characterized by myocardial invasion, activation of necroptosis, and tissue remodeling in animals rescued by antimicrobial therapy. Cardiac damage by invading pneumococci may explain why adverse cardiac events that occur during and after pneumococcal pneumonia in adult human patients.

OBJECTIVES/GOALS: Older adults are included in clinical research infrequently compared to their burden of chronic illness. The goal for this study is to learn from older adults about their lived experiences with research and use this knowledge to develop tools and solutions aimed at increasing their inclusion. METHODS/STUDY POPULATION: This study utilized the 5T Model (developed by Duke CTSA) and Community Engagement Studio (CES) (developed by Vanderbilt CTSA) to connect and engage with community experts (older adults and those who work with older adults) in Oregon. Two CES were completed with 14 community experts and 4 investigators interested in including older adults in their studies. Participants took part in a 2-hour facilitated discussion to gain insight from their perspectives on research. The 5T Model was shared with participants and used to guide the discussion and elicit feedback on the model and identify gaps in resources and training needed for investigators to enhance inclusion of older adults in research. RESULTS/ANTICIPATED RESULTS: Trust, relationships, education, and diversity were themes identified across all of the 5Ts. Participants discussed the need for inclusion and diversity within research, with an emphasis on those at the oldest ages, rural populations, and lower socioeconomic status. Participants acknowledged both investigators and participants require more education, with a great need for improving health literacy for research participants. Participants saw trust and relationships as an integral part of older adult inclusion in research, with the relationship being not only that between investigator and participant, but between them and the communities that support older adults, including family members. DISCUSSION/SIGNIFICANCE: This study highlighted the voices of older adult research participants, allowing for participant-informed findings and solution development. Future directions will focus on developing and refining tools and resources for investigators and expanding to other underrepresented populations.

Objectives/Goals: To create, train, and evaluate the FAST-PACE (Promoting Academic and Community Engagement) Toolkit that catalyzes academic-community translation science teams during a public health emergency. The toolkit is a road map based on the Research Readiness and Partnership Protocol (R2P2), which was developed from the Flint Water Crisis. Methods/Study Population: A literature review was conducted by the Michigan Institute for Clinical & Health Research Community Engagement (MICHR CE) program and the Community-Based Organization Partners (CBOP), to identify important and common elements in disaster response protocols with a set of key interviews (n = 31) to glean perspectives from community leaders. Key findings were extracted and reviewed to generate guidelines and recommendations for the R2P2 protocol. The co-developed FAST-PACE Toolkit launched its expansion statewide to address emergencies and health disparities of communities in crisis. The iterative process consisted of community report-outs, gathering input from stakeholders, via discussion, and evaluation surveys. The feedback was used to develop, enhance, and tailor the toolkit and training content. Results/Anticipated Results: Data from training (n = 8) of the critical elements of the FAST-PACE Toolkit, which provides guidance for academic and community team members that includes 1) assessing community assets and needs; 2) engaging in clear and bidirectional communication; 3) facilitating transparency and equitable partnering; 4) identifying health equity and justice issues; and 5) conducting the evaluation of research. The training will be disseminated in-person and virtually across the state of Michigan resulting in participants sharing community-identified health issues and social determinants of health to assist MICHR CE to suggest resources to address health impacts. Discussion/Significance of Impact: The FAST-PACE Toolkit borne from the flint water crisis and confounded by other crises used CEnR principles to create a translation science roadmap. It equips communities and collaborating academic institutions across the state to respond to public health crises and fosters equitable translation science partnerships built on respect and trust.

OBJECTIVES/GOALS: Infectious keratitis is a potentially sight-threatening disease. Its epidemiology has been considered on various national and regional levels, which demonstrates unique patterns in patient risk factors, causative organisms, antimicrobial resistance and clinical outcomes. This study will examine the patterns specific to the Rochester, NY area. METHODS/STUDY POPULATION: This project will be a ten-year retrospective study, examining all patients who were diagnosed with infectious keratitis at the University of Rochester Medical Center between 2011 and 2021. The study population is selected from a compiled list of those patients with an ICD diagnosis code including ’keratitis’ in the eRecord that falls within our chosen date range. Participants were excluded if the patient’s physician documented that the keratitis is most likely not infectious in etiology. If eligible for the study, there is documentation of patient demographics, clinical risk factors, clinical course, culture data from corneal scraping and antimicrobial resistance patterns, if available. Once data is collected, it will be analyzed and compared to pre-existing regional and national data. RESULTS/ANTICIPATED RESULTS: Between 2011 and 2021, there were 1652 patients with ICD diagnoses of keratitis at the University of Rochester Medical Center. Of these patients, we anticipate approximately 1,200 to meet our inclusion criteria. Some of the major risk factors for developing infectious keratitis in this population include contact lens use, immunocompromised state (elderly, diseased or iatrogenic) and corneal trauma. Clinical complications include progression to endophthalmitis, need for interventional surgery, or failure to improve on clinical exam after 2 weeks of antimicrobials. Causative organisms are most often bacterial. Certain organisms are associated with specific risk factors while other organisms predict greater risk of complications. We also anticipate patterns of antimicrobial resistance to emerge. DISCUSSION/SIGNIFICANCE: This study will provide insights into the risk factors and clinical course of infectious keratitis according to causative organisms in the Rochester, NY area. This can help us mitigate risk factors and help prognosticate for patients, once diagnosed. Also, patterns of antimicrobial resistance can inform empiric antimicrobial recommendations.

OBJECTIVES/GOALS: Our overall objective is to develop a directly observable and reproducible method of enhanced blood flow through tumor vessels (i.e. dynamic control) at the time of systemic treatment delivery. Our central hypothesis is that the dynamic control of tumor vessels will improve (1) systemic drug delivery and (2) effector cell trafficking to target tumor. METHODS/STUDY POPULATION: B16 melanoma cells were inoculated into C57BL/6 (B6) mice (male and female) in both regional (hind leg) and systemic (flank) models. Dynamic control consisted of an IV saline bolus (500 ul) and phenylephrine (10 ug). Tumor vessel response was observed in real-time through window chambers using intravital microscopy (IVM). Dynamic control was combined with melphalan (20 mg/ml) either regionally (isolated limb perfusion) or systemically. Outcomes included tumor growth, survival, IHC, and toxicity. Dynamic control will be combined with adoptive transfer of effector T cells. B6 mice will be inoculated with B16/OVA (flank with window chamber) and treated with fluorescently labeled (calcein), OVA-specific CD8+ T cells from OT-1 transgenic mice. IVM, IHC, and flow cytometry will be used to measure T cell trafficking. RESULTS/ANTICIPATED RESULTS: Dynamic control (1) restored blood flow in non-functional tumor vessels and (2) increased and then transiently reversed blood flow in functional vessels. Vessel diameters did not change, suggesting that shunting of systemic blood to the tumor vasculature accounted for the observed changes. Dynamic control augmented tumor responses in our regional therapy model of melanoma. Increases in DNA adduct formation (melphalan mechanism of action) detected by IHC, decreased tumor growth, and increased survival were observed with dynamic control. There was no increased limb toxicity. Similarly, dynamic control augmented responses in our systemic therapy model (decreased tumor growth and improved survival). We anticipate that dynamic control will improve trafficking of effector T cells in the next set of experiments. DISCUSSION/SIGNIFICANCE OF IMPACT: Heterogeneous responses to systemic therapies represent a major gap in current cancer treatment. An essential requirement for any effective therapy is its ability to reach tumor via the tumor-associated vasculature. We have therefore developed an approach to enhance drug delivery (dynamic control), which we also plan to test in clinical trials.

OBJECTIVES/SPECIFIC AIMS: Specialized pro-resolving lipid mediators (SPM) actively counter proinflammatory cascades. A deficit of SPMs is one possible mechanism through which inflammation leads to the development of atherosclerotic disease. The purpose of this study is to characterize the profiles of intermediates of SPM synthesis pathways and end-product SPMs in the plasma of patients with peripheral artery disease (PAD). METHODS/STUDY POPULATION: A cross-sectional sample of 52 patients with PAD was recruited at the San Francisco Veterans Affairs Medical Center. PAD was defined as the presence of claudication symptoms and an ankle-brachial index <0.9, or a history of revascularization for claudication. Patients were excluded if they were taking immunosuppressive medications, had a severe acute illness (infection, surgery, illness, critical limb ischemia) within the last 30 days, or had severe hepatic, renal, or nonvascular inflammatory disease. Intermediates of SPM synthesis pathways and end-product SPMs were measured in plasma samples of patients by liquid chromatography-tandem mass spectrometry. RESULTS/ANTICIPATED RESULTS: The average age of the cohort was 69±6.3 and patient comorbidities reflected common comorbidities associated with PAD (hypertension 96%, hyperlipidemia 87%, diabetes mellitus 42%, coronary artery disease 34%). Rutherford categories, measurements of PAD symptom severity, ranged from 0 to III (0 10%, I 40%, II 27%, III 23%). Three EPA products were measured: 18-hydroxyeicosapentaenoic acid (18-HEPE), resolvin E1 (RvE1), and resolvin E2 (RvE2). 18-HEPE, an intermediate of SPM synthesis, was detectable in the plasma of every patient (median: 105 pg/mL, IQR: 54.9–195), whereas the SPM end-products, RvE1 and RvE2, were only detectable in 6 and 10 patients, respectively. In total, 7 DHA products were measured: 14-hydroxydocosahexaenoic acid (14-HDHA), 17-HDHA, resolvin D1 (RvD1), resolvin D2 (RvD2), protectin D1, protectin DX, and maresin 1. The intermediates 14-HDHA (median: 6546 pg/mL, IQR: 3329–12061) and 17-HDHA (median: 644 pg/mL, IQR: 340–1056) were detectable in the plasma of every patient. However, the end-products RvD1, RvD2, protectin D1, protecin DX, and maresin 1 were identified in less than half of the cohort. DISCUSSION/SIGNIFICANCE OF IMPACT: We report the presence of several intermediates of SPM synthesis pathways (18-HEPE, 14-HDHA, and 17-HDHA) in every patient but the presence of SPM end-products in only a limited portion of the cohort. These results suggest that some patients with PAD may have a deficit of SPMs. Further investigation is required to better understand the role of SPMs and mediators of resolution of inflammation in PAD.