ABSTRACT IMPACT: This research takes a transcriptomic approach to parse genes and molecular pathways that underlie the fear memory circuitry and, in doing so, identifies therapeutic targets that can further be developed into treatments for fear disorders, such as post-traumatic stress disorder. OBJECTIVES/GOALS: Normal fear learning produces avoidance behavior that promotes survival, but excessive and persistent fear after trauma can lead to development of phobias and post-traumatic stress disorder (PTSD). Our goal is to understand the mechanism and identify novel genetic targets underlying fear responses. METHODS/STUDY POPULATION: Involvement of the basolateral amygdala (BLA) in fear acquisition is well established and requires activation of N-methyl-D-aspartic acid receptors (NMDARs). At a cellular level, NMDAR activation leads to production of nitric oxide (NO) by a process mediated by interaction between postsynaptic density protein 95 (PSD95) and neuronal nitric oxide synthase (nNOS). To elucidate mechanisms underlying the role of the PSD95-nNOS-NO pathway in conditioned fear, here we use rodent behavioral paradigms, pharmacological treatment with a small molecule PSD95-nNOS inhibitor, RNA-sequencing, and an AAV-mediated knockdown of the nNOS gene in the BLA. RESULTS/ANTICIPATED RESULTS: We show that treatment with ZL006 attenuates rodent cued-fear consolidation. Additionally, with RNA-sequencing, expression of 516 genes was altered in the BLA following fear expression; of these genes, 83 were restored with systemic ZL006 treatment. Network data and gene ontology enrichment analyses further revealed that cGMP effects, insulin-like growth factor binding, and cognition-related pathways were significantly altered. Finally, we show that a BLA-specific knockdown of nNOS attenuates cued fear consolidation, without adverse effects on other memory and motor behaviors. DISCUSSION/SIGNIFICANCE OF FINDINGS: Via a model of NMDAR-mediated fear consolidation, our results reveal novel pathways and genetic targets that underlie plasticity of fear memory circuitry. Importantly, these results will inform future therapeutic strategies for targeting fear related disorders like PTSD.

ABSTRACT IMPACT: This study will provide the essential characterization of intrinsic neural activity in human brain organoids, both at the single cell and network levels, to harness for translational purposes. OBJECTIVES/GOALS: Brain organoids are 3D, stem cell-derived neural tissues that recapitulate neurodevelopment. However, to levy their full translational potential, a deeper understanding of their intrinsic neural activity is essential. Here, we present our preliminary analysis of maturing neural activity in human forebrain organoids. METHODS/STUDY POPULATION: Forebrain organoids were generated from human iPSC lines derived from healthy volunteers. Linear microelectrode probes were employed to record spontaneous electrical activity from day 77, 100, and 130 organoids. Single unit recordings were collected during hour-long recordings, involving baseline recordings followed by glutamatergic blockade. Subsequently, tetrodotoxin, was used to abolish action potential firing. Single units were identified via spike sorting, and the spatiotemporal evolution of baseline neural properties and network dynamics was characterized. RESULTS/ANTICIPATED RESULTS: Nine organoids were recorded successfully (n=3 per timepoint). A significant difference in number of units was seen across age groups (F (2,6) = 6.4178, p = 0.0323). Post hoc comparisons by the Tukey HSD test showed significantly more units in day 130 (51.67 ±14.15) than day 77 (16.33 ±14.98) organoids. Mean firing rates were significantly different in organoids based on age, with drug condition also trending toward significance (F (6,12) = 9.97; p = 0.0028 and p = 0.08 respectively). Post hoc comparisons showed a higher baseline firing rate in day 130 (0.99Hz ±0.30) organoids than their day 77 counterparts at baseline (0.31Hz ±0.066) and glutamate blockade (0.31Hz ±0.045). Preliminary network analysis showed no modularity or small-world features; however, these features are expected to emerge as organoids mature. DISCUSSION/SIGNIFICANCE OF FINDINGS: Initial analysis of brain organoid activity demonstrates changes in single unit properties as they mature. Additional work in this area, as well as further network analyses, will confer better sense of how to rationally utilize brain organoids for translational purposes.

ABSTRACT IMPACT: The potential to use vaginal pH as a low cost, non-invasive diagnostic test at the point of CIN2 diagnosis to predict worsening of cervical disease. OBJECTIVES/GOALS: We previously reported that persistence/progression of cervical intraepithelial neoplasia-2 (CIN2) was uncommon in women living with HIV (WLH) from the Women’s Interagency HIV Study (WIHS, now MWCCS). Here we examined additional factors that may influence CIN2 natural history. METHODS/STUDY POPULATION: A total of 337 samples from 94 WLH with a confirmed CIN2 diagnosis were obtained from the MWCCS. 42 cervicovaginal HPV types and 34 cervicovaginal cytokines/chemokines were measured at CIN2 diagnosis (94 samples) and 6-12 months prior to CIN2 diagnosis (79 samples). Covariates, including CD4 count and vaginal pH, were abstracted from core MWCCS visits. Logistic regression models were used to explore CIN2 regression (CIN1, normal) vs. persistence/progression (CIN2, CIN3). Log rank tests, Kaplan Meier method, and Cox regression modeling were used to determine CIN2 regression rates. RESULTS/ANTICIPATED RESULTS: The most prevalent HPV types were HPV54 (21.6%) and 53 (21.3%). 33 women (35.1%) had a subsequent CIN2/CIN3 diagnosis (median 12.5 years follow-up). Each additional hr-HPV type detected at the pre-CIN2 visit associated with increased odds of CIN2 persistence/progression (OR 2.27, 95% CI 1.15, 4.50). Higher vaginal pH (aOR 2.27, 95% CI 1.15, 4.50) and bacterial vaginosis (aOR 5.08, 95% CI 1.30, 19.94) at the CIN2 diagnosis visit associated with higher odds of CIN2 persistence/progression. Vaginal pH >4.5 at CIN2 diagnosis also associated with unadjusted time to CIN2 persistence/progression (log rank p=0.002) and a higher rate of CIN2 persistence/progression (adjusted hazard ratio [aHR] 3.37, 95% CI 1.26, 8.99). Cervicovaginal cytokine/chemokine levels were not associated with CIN2 persistence/progression. DISCUSSION/SIGNIFICANCE OF FINDINGS: We found relatively low prevalence of HPV16/18 in this cohort. Elevated vaginal pH at the time of CIN2 diagnosis may be a useful indicator of CIN2 persistence/progression and the rate of persistence/progression.

ABSTRACT IMPACT: This work investigates C. difficile strains in soil as a potential exposure for gut colonization and community-acquired infection of C. difficile. OBJECTIVES/GOALS: Identifying environmental sources of C. difficile can inform how non-hospital reservoirs can potentially contribute to C. difficile exposure and subsequent gastrointestinal colonization. The objective of the study was to identify C. difficile and toxin genes across various soil sources. METHODS/STUDY POPULATION: This was a cross-sectional study utilizing soil samples obtained throughout Texas, USA. All samples were collected between August and November of 2019 and 2020. Samples were taken from human and animal high contact areas, such as recreational parks. Samples were stored at -80oC until processing. DNA extractions were performed using the DNeasy Powersoil Pro Kit (Qiagen) per manufacturer’s instructions. Real-time PCR was also performed on extracted DNA using the Microbial DNA qPCR Multi-Assay Kit for Clostridium difficile Pathogenicity (Qiagen) for the identification of C. difficile, toxin A (TcdA), and toxin B (TcdB) genes. RESULTS/ANTICIPATED RESULTS: A total of 137 soil samples including dry dirt, sand, and wet soil near water sources were collected and processed for the presence of C. difficile. These included samples from parks and trails (42.3%), water sources (36.5%), and other public spaces (21.2%). C. difficile was identified in 59 (43.1%) soil samples: 6 (4.4%) with Toxin A and 2 (1.5%) with toxin B production. C. difficile was most prevalent among samples taken from parks and trails (50.0%), followed by other public spaces (48.3%), and water sources (32.0%). The median (IQR) Cq value for the C. difficile gene was 39.24 (33.45-40.47) among samples that tested positive. DISCUSSION/SIGNIFICANCE OF FINDINGS: We identified a high prevalence of Clostridioides difficile in soil samples, though toxin gene detection prevalence was low. Future studies will analyze other sources, including water and varying surface samples to obtain a comprehensive view of C. difficile in the environment.

North Carolina Central University (NCCU) and Duke Cancer Institute implemented an NCI-funded Translational Cancer Disparities Research Partnership to enhance translational cancer research, increase the pool of underrepresented racial and ethnic group (UREG) researchers in the translational and clinical research workforce, and equip UREG trainees with skills to increase diversity in clinical trials. The Cancer Research Education Program (C-REP) provided training for UREG graduate students and postdoctoral fellows at Duke and NCCU. An innovative component of C-REP is the Translational Immersion Experience (TIE), which enabled Scholars to gain knowledge across eight domains of clinical and translational research (clinical trials operations, data monitoring, regulatory affairs, UREG accrual, biobanking, community engagement, community outreach, and high-throughput drug screening). Program-specific evaluative metrics were created for three broad domains (clinical operations, basic science/lab research, and population-based science) and eight TIE domains. Two cohorts (n = 13) completed pre- and post-surveys to determine program impact and identify recommendations for program improvement. Scholars reported statistically significant gains in knowledge across three broad domains of biomedical research and seven distinct areas within TIE. Training in translational research incorporating immersions in clinical trials operation, biobanking, drug development, and community engagement adds value to career development of UREG researchers.

ABSTRACT IMPACT: By demonstrating the feasibility of a multimodal, interdisciplinary intervention for prediabetes, the current project aims to provide a template for the prevention of diabetes and associate comorbid conditions. OBJECTIVES/GOALS: To determine if a multi-modal, interdisciplinary intervention delivered to a group of prediabetic patients will result in reduced rates of diabetes progression. This project is a retrospective evaluation that will exam the feasibility and possibly efficacy of this intervention. METHODS/STUDY POPULATION: We will enroll 50 participants for the clinic, aged 21-60 inclusive. Patients will have a Body Mass Index >27kg/m2 with a diagnosis of prediabetes. Patients must be non-pregnant, using approved contraception, and agree to not become pregnant for 1 year after enrollment. After enrollment, the initial treatment period is for 1 year and includes a 12 week low calorie diet plan, a 6-month intensive behavioral and lifestyle modification plan followed by a 6 month behavior reinforcement extension. Weight management medications may be used if appropriate for the patient from a clinical perspective during the 6-month intensive behavioral/lifestyle modification. RESULTS/ANTICIPATED RESULTS: It is anticipated that there will be decreased weight with a mean weight loss goal of approximately >10%. Furthermore, it is expect that there will be improvement of other markers of metabolic disease. These include improvement of lipid values (LDL-C, HDL-C, Triglycerides, Total Cholesterol) as well as blood pressure with expected blood pressures of below 130/80 in greater than 50% of participants. Finally, It is expected that 50% or greater participants will have improvement of glycemic control. It is anticipated that greater than 50% of participants will have improvement of glycemic control and achieve normoglycemia. These values will be determined based upon fasting glucose or A1c. DISCUSSION/SIGNIFICANCE OF FINDINGS: The significance of this intervention is enormous. By demonstrating feasibility in this trial, we can work toward both assessing efficacy and possibly dissemination of this model program. If these interventions provide durable changes at scale, this could help slow the epidemic of obesity and obesity related comorbid conditions.

ABSTRACT IMPACT: We developed a novel semi-automated computational platform to quantify spatial characteristics of preserved peripheral visual field loci in retinitis pigmentosa. This work may inform future research on therapies to prevent visual field loss in retinitis pigmentosa and related diseases. OBJECTIVES/GOALS: Retinitis pigmentosa (RP) causes progressive and severe peripheral visual field (pVF) loss in some but not all patients. The characteristics of pVF in RP are incompletely understood. We developed a novel semi-automated computational platform to quantify the spatial characteristics of pVF. METHODS/STUDY POPULATION: We analyzed preserved pVF size and location in both eyes of RP patients using the Goldmann V4e isopter. We developed a custom algorithm in MATLAB to align and average the binary V4e isopter segmentations, and generated a two-dimensional probability map of the spatial distribution of preserved pVF loci along the radial and circumferential dimensions. To adjust for disease duration, cases were categorized by the time from self-reported symptom onset. Probability maps of pVF preservation were generated for categories of disease duration using unsupervised K-means clustering. Analyzing cases with longitudinal data, we identified loci of stable pVF over time. RESULTS/ANTICIPATED RESULTS: A total of 152 patients were included (N=304 eyes). The mean age was 46.7 years and 49.3% were male. Disease duration was categorized as <20 years (N=72, 47.4%), 20-40 years (N=60, 39.5%), or >40 years (N=20, 13.2%). Longitudinal data (3.2 -5.7 years of follow-up) were available in 65 patients (42.8%). Probability plots of preserved pVF loci in the cross-sectional dataset showed that the median percentage of preserved pVF loci were located between 50 ºand 80 ºeccentricity and between the 30 ºto 50 ºmeridians, with highly concordant inter-ocular symmetry. Probability plots in the longitudinal dataset showed that inferotemporal pVF loci were most likely to be preserved over time. DISCUSSION/SIGNIFICANCE OF FINDINGS: Semi-automated quantification of pVF loci is a useful platform to analyze spatial characteristics of the visual field in RP. Certain portions of the pVF may be relatively resistant to functional decline. Understanding the molecular basis of pVF resilience will inform further research on RP therapy.

CTSI Career Development Award (KL2) programs provide junior faculty with protected time and multidisciplinary, mentored research training in clinical and translational science research. The KL2 Visiting Scholars Program was developed to promote collaborative cross-CTSA training, leverage academic strengths at host CTSAs, and support the career development of participating scholars through experiential training and the development of new partnerships. This manuscript provides a detailed programmatic description and reports outcomes from post-visit and outcomes surveys. Since 2016, 12 scholars have completed the program, with 6 scheduled to complete it in 2021. Post-visit surveys (n = 12) indicate all scholars reported the program valuable to career development, 11 reported benefit for research development, and 11 expansion of collaborative networks. Outcomes surveys (n = 11) revealed subsequent scholar interaction with host institution faculty for 10 scholars, 2 collaborative grant submissions (1 funded), 2 planned grant submissions, 1 published collaborative manuscript, and 3 planned manuscript/abstract submissions. The Visiting Scholars Program is a cost- and time-efficient program that leverages the academic strengths of CTSAs. The program enhanced KL2 scholar training by expanding their professional portfolio, promoting research development, and expanding collaborative networks. Resources to support the program are shared in this report to expedite the development of similar programs at regional and national levels.

ABSTRACT IMPACT: This study provides public health and K-12 school districts with a pragmatic, flexible, adaptable model showing COVID-19 transmission dynamics, using local data and program elements that are modifiable and with an online model for easy use, to enable safe and equitable re-opening and maintenance of in-person learning. OBJECTIVES/GOALS: School closures resulting from the COVID-19 pandemic disrupt student education and health and exacerbate inequities. Public health agencies and school districts currently lack pragmatic models to assess the effects of potential strategies for resuming and maintaining in-person learning on outcomes such as transmission and attendance. METHODS/STUDY POPULATION: This study explored how various combinations of transmission-mitigating interventions affect health and learning outcomes in a range of underlying epidemiological conditions. The CTSA science team developed a conceptual framework and an agent-based simulation model with parameters including prevalence, transmission, testing, preventive and responsive actions, infection control, population behavior and awareness, and the potential impact of vaccine adoption and exemption policies. The team partnered with a large school district to ensure relevance of the program components to decision-making. RESULTS/ANTICIPATED RESULTS: The model shows that no single program element or condition ensures safety. Combining interventions can result in synergy in the mitigation efforts. Even without testing, an efficient health screening process with forthcoming risk reporting, combined with on-campus infection control, can reduce on-campus transmission. The resulting model is accessible online to enable exploration of likely scenarios. It is adaptable as COVID-19 science evolves, including for testing and vaccines. DISCUSSION/SIGNIFICANCE OF FINDINGS: This research provides public health agencies and school districts with a model that couples local conditions with programmatic elements to help inform the local COVID-19 response, recognizing that decisions about the school community are often complex politically, technically, and operationally when it comes to addressing a health crisis.

ABSTRACT IMPACT: This change will improve primary care physicians and pediatrics ability to identify, intervene and prevent obesity related renal damage in the vulnerable population of young adults OBJECTIVES/GOALS: Obesity related glomerulopathy has a reversible stage manifested as hyperfiltration. Early intervention depends on the ability to identify hyperfiltration. Hyperfiltration prevalence is underestimated using the currently recommended formula We investigated whether calculating BSA-adjusted GFR will more readily identify hyperfiltration. METHODS/STUDY POPULATION: We extracted data from a large urban, multi-institutional Electronic Health Records (EHR) clinical data research network to construct an EHR data base of 60,549 women and girls ages 12-21 years from the New York metropolitan area. EGFR was calculated in two ways, 1) according to age appropriate formula, and 2) according to age appropriate formula and adjusted to body surface area (BSA). BMI-for-age values were classified according to the World Health Organization schema and grouped according to the CDC definitions. BSA was calculated according to the Du-Bois formula. Hyperfiltration was defined by a threshold of 135ml/min. The Bland Altman method assessed the agreement between formulas across the different BMI groups. RESULTS/ANTICIPATED RESULTS: Serum creatinine values were similar across different BMI groups. Comparing eGFR values, hyperfiltration rates were similar across BMI groups, ranging between 4%-6.6%. BSA-adjusted GFR was different across BMI groups: hyperfiltration rates were 0.81% for the underweight group, 2.56% for the normal weight, 12.18% for the overweight and 39% in the obese group. This trend of hyperfiltration paralleled the the rise in urine creatinine across BMI groups. DISCUSSION/SIGNIFICANCE OF FINDINGS: BSA-adjusted GFR more sensitively detects hyperfiltration due to obesity than does eGFR. Calculating BSA-adjusted GFR will improve primary care and pediatric physicians’ ability to identify, intervene and prevent early ORG. Changes in body composition may account for the increasing discordance between BSA-adjusted and eGFR as BMI rises.

ABSTRACT IMPACT: Given the association between lower time to treatment and better clinical outcomes in stroke patients, identifying factors correlated with reduced proximity and thus greater time to stroke care can aid efforts to reduce disparities in stroke outcomes. OBJECTIVES/GOALS: The objective of this study is to quantify the relationship between distance to the nearest certified stroke hospital and census-derived demographics of age, race/ethnicity, income, and insurance status. METHODS/STUDY POPULATION: This is a cross-sectional study. Population data for all census tracts in the contiguous United States were obtained from the US Census Bureau’s 2014-2018 American Community Survey. Stroke hospitals were identified from national or state level certification databases and were required to offer at least IV tPA. The main outcome is driving distance in kilometers from each census tract to the nearest certified stroke center, which was calculated using OSMnx, a Python package to retrieve, model and analyze real-world street networks. Quantile regression analysis was used to compare relationships between distances and tract-level demographics of age, race/ethnicity, income, and insurance status. RESULTS/ANTICIPATED RESULTS: 2,423 stroke centers and 71,929 census tracts containing 316,995,649 individuals were included. 49,918 (69%) tracts were urban. Demographic disparities in proximity to certified stroke care were greater in non-urban areas than in urban areas. Higher representation of individuals with age ≤65 years were associated with increased median distance to a certified stroke center in non-urban areas, but not urban areas. Median distance was greater with greater representation of American Indian or uninsured populations in urban and non-urban census tracts. Higher median income was associated with decreased median distance in non-urban census tracts and greater median distance in urban census tracts. DISCUSSION/SIGNIFICANCE OF FINDINGS: Reduced proximity to stroke care exists in areas with greater representation of elderly, American Indian, or uninsured persons; and low median income. These disparities are magnified in non-urban settings. Such knowledge can aid efforts to address and reduce disparities in stroke outcomes.

The goal of this study was to assess the utility of participatory needs assessment processes for continuous improvement of developing clinical and translational research (CTR) networks. Our approach expanded on evaluation strategies for CTR networks, centers, and institutes, which often survey stakeholders to identify infrastructure or resource needs, using the case example of the Great Plains IDeA-CTR Network. Our 4-stage approach (i.e., pre-assessment, data collection, implementation of needs assessment derived actions, monitoring of action plan) included a member survey (n = 357) and five subsequent small group sessions (n = 75 participants) to better characterize needs identified in the survey and to provide actionable recommendations. This participatory, mixed-methods needs assessment and strategic action planning process yielded 11 inter-related recommendations. These recommendations were presented to the CTR steering committee as inputs to develop detailed, prioritized action plans. Preliminary evaluation shows progress towards improved program capacity and effectiveness of the network to respond to member needs. The participatory, mixed-methods needs assessment and strategic planning process allowed a wide range of stakeholders to contribute to the development of actionable recommendations for network improvement, in line with the principles of team science.

ABSTRACT IMPACT: Our work helps show universities that embedding dedicated sexual health clinics within university health and wellness clinics may expand the amount of students they see for sexual health screenings during a time of increased sexual behavior and exploration. OBJECTIVES/GOALS: The National College Health Association reports that college students have frequent, condomless sex. Student health and wellness clinics (SHWC) offer sexual health services, but few have dedicated sexual health clinics (SHC). We evaluated sexual health service use at a university SHWC after implementation of a dedicated SHC two half-days per week. METHODS/STUDY POPULATION: This was a retrospective analysis of data collected from patients receiving sexual health screening at the University of Alabama at Birmingham (UAB) SHWC between January 2015 and June 2019. Demographic variables, sexual behaviors, reason for testing, and rates of STIs were extracted from the electronic medical record and were compared by clinic (SHC vs. SHWC). Data on screening visits of patients over 18 were included in the final analysis. Variables were summarized with frequencies and percentages. Univariate models were fit, and multi-variable models will be fit, selecting variables with p values of 0.1 or less. Odds ratios with corresponding 95% confidence intervals for univariate analysis are presented. The study was approved by the UAB Institutional Review Board. RESULTS/ANTICIPATED RESULTS: A total of 5025 STI screenings were performed. Males (OR 4.13; 3.61-4.72), undergraduates (OR 1.33; 1.15-1.54), and persons reporting sex with the same sex (OR 1.88; 1.56-2.28), were significantly more likely to seek care at the SHC. Students with symptoms were more likely to seek care at the SHWC (OR 0.53; 0.47-0.61), while persons who reported contact with STIs were more likely to seek care at the SHC (OR 2.88; 2.22-3.74). The overall percentage of positive screenings was 9.3% for chlamydia (CT), 3.0% for gonorrhea (GC), 0.8% for trichomoniasis (TV), 0.7% for syphilis, and 0.3% for HIV with higher percentages of positive for CT (OR 1.60; 1.30-1.96) and GC (OR 2.02; 1.44-2.85) in the SHC. A greater percentage of positives for TV (OR 0.37; 0.14-0.96) was found in the SHWC. DISCUSSION/SIGNIFICANCE OF FINDINGS: Based on demographics of persons utilizing services, embedding a dedicated SHC within a university SHWC may expand populations reached for STI screening. With higher percentages of patients testing positive for CT and GC, a SHC may allow for greater diagnosis and treatment of STIs in general screening and persons presenting as contacts.