OBJECTIVES/GOALS: People living with HIV, despite antiretroviral therapy (ART), have increased burden of inflammatory and aging-related comorbidities such as periodontitis. Oral microbiota have been linked to periodontitis, but not in the context of HIV. We aim to compare relationships between the oral microbiome and periodontal disease in HIV+ vs healthy controls. METHODS/STUDY POPULATION: In an ongoing cohort study we have been recruiting pre- and post-menopausal women with HIV+ on ART for ≥6 months and HIV- controls matched by menopausal status (target n = 30 per arm; currently HIV+: n = 30 post- and 9 pre-M; HIV-: n = 15 post- and 6 pre-M). Patients age <18 or on antibiotics within 3 mos., except prophylaxis, are excluded. Patients provide saliva, then subgingival plaque collection during a dental examination through scaling from six index teeth. Standard CDC/AAP classifications of periodontitis are used. We will perform 16S rRNA and ITS sequencing to profile bacterial and fungal communities in saliva and plaque. Linear mixed effect regression and differential abundance analyses will be used to identify microbial and mycobial oral signatures of periodontal disease severity in HIV+ and HIV- populations. RESULTS/ANTICIPATED RESULTS: We found a markedly high prevalence of severe periodontal disease in HIV+ women despite ART (59%, compared to 11% in HIV- controls). In post-menopausal women with HIV, saliva bacterial α- and β-diversity in the saliva differed significantly with periodontal disease severity. Fungal α-diversity was also significantly lower in plaque from teeth with severe loss of tissue attachment (CAL ≥4 mm). We identified bacterial and fungal taxa significantly enriched in post-menopausal HIV+ women with severe compared to no or mild periodontitis. We hypothesize, similarly, associations between the oral microbiome and periodontitis in HIV- controls. However, we expect overall diversity metrics to be significantly altered in HIV+ compared to HIV- patients, indicating long-term dysbiosis despite treatment with ART. DISCUSSION/SIGNIFICANCE OF IMPACT: Contrasting associations between the oral microbiome and periodontal disease with respect to HIV will provide evidence for the role of microbiota in accelerated aging phenotype caused by HIV. Our results would also provide rationale for interventions targeting co-morbidities in people living with HIV to account for both inflammation and dysbiosis.

OBJECTIVES/GOALS: The goal of this study is to compare plasma neurofilament light (NfL) concentrations in asymptomatic children and young adults that carry the gene expansion (GE group) that causes Huntington’s Disease to similar subjects that do not carry this genetic mutation (GNE group). METHODS/STUDY POPULATION: Subjects from the Kids-HD study in the GE group were divided into groups based on their estimated years to motor onset. Each subgroup was compared to the subjects from the GNE group. Additionally, a group of participants with juvenile HD were compared to the GNE group. These comparisons were made by utilizing linear mixed effects regression models that included a random effect per subject and family and also included the covariates of age and parental socioeconomic status. A post-hoc analysis of subjects in the GE group who were within 20 years from their predicted motor onset was conducted to assess the relationship between striatal volume and plasma NfL concentrations. RESULTS/ANTICIPATED RESULTS: GE participants more than 20 years from their predicted motor onset did not have elevated plasma NfL concentrations relative to the GNE group. However, participants who were 15-20 years from their predicted motor onset had a mean NfL concentration of 1.61 pg/uL compared to 1.31 pg/uL in the GNE group (p = 0.036). Participants who were within 15 years from their predicted motor onset had a mean NfL concentration of 2.08 pg/uL, which was also significantly elevated relative to the GNE group (t = 3.03, p = 0.003). Additionally, the participants with juvenile HD had a mean NfL level of 3.22 pg/uL, which was significantly elevated compared to the GNE group (p<0.0001). NfL concentrations were significantly correlated with striatal volume amongst participants who were within 20 years of onset (p = 0.017). DISCUSSION/SIGNIFICANCE OF IMPACT: The huntingtin protein is essential to neurodevelopment but current gene therapies for HD focus on blocking production of this gene. These results will provide guidance on the optimal timing of administration of gene therapies by identifying neurodegeneration decades prior to motor onset of HD.

OBJECTIVES/GOALS: Sudden Unexpected Death in Epilepsy (SUDEP) is a leading cause of death in epilepsy patients. This study aims to determine whether cardiac mechanisms contribute to SUDEP in epilepsy patients with variants in DEPDC5, a gene encoding a member of the mTOR GATOR complex, to identify SUDEP biomarkers. METHODS/STUDY POPULATION: SUDEP has been reported in 10% of epilepsy patients with DEPDC5 loss-of-function variants. We used human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) to measure changes in cellular excitability that are known to be substrates for cardiac arrhythmias. CRISPR-derived isogenic DEPDC5 iPSC-CMs and DEPDC5 patient-derived iPSC-CMs were used in this study. Whole-cell patch-clamp was used to measure voltage-gated sodium current (INa) and calcium current (I>Ca) in single iPSC-CMs in voltage-clamp mode; and to measure action potentials (APs) in 3-dimentional iPSC-CM-derived micro-tissues in current-clamp mode. RESULTS/ANTICIPATED RESULTS: CRISPR generated heterozygous deletion of 1 base-pair in the first coding exon of DEPDC5 gene, resulting in a premature stop codon, simulated the variants identified in DEPDC5 epilepsy patients. In CRISPR generated heterozygousDEPDC5 iPSC-CMs, whole-cell voltage-clamp recordings revealed that INa was increased and ICa was reduced compared with isogenic control iPSC-CMs. Whole-cell current-clamp recordings revealed that AP duration at 80% and 90% of repolarization, APD80 and APD90, respectively, were prolonged compared to isogenic control iPSC-CMs. Similar measurements will be performed for iPSC-CMs derived from DEPDC5 patients. DISCUSSION/SIGNIFICANCE OF IMPACT: This study shows that epilepsy patients with non-ion channel gene variants in DEPDC5 have altered CM excitability, which may serve as a substrate for cardiac arrhythmias in DEPDC5 patients. Importantly, this work may allow us to identify biomarkers for SUDEP risk in these patients in the future. CONFLICT OF INTEREST DESCRIPTION: L.L.I. is the recipient of a collaborative research grant from Stoke Therapeutics.

OBJECTIVES/GOALS: To quantify clinical exam in total knee arthroplasty by answering the following questions: (1) What are the magnitudes of forces applied by surgeons during the varus-valgus exam? (2) Is the choice of tibial insert thickness related to the magnitude of the applied forces? (3) How accurately does a surgeon estimate the gaps in the varus-valgus exam? METHODS/STUDY POPULATION: Three cadaveric knees were implanted with standard TKA trial implants. Four pliable force sensors were wrapped around the foot and ankle of each cadaver to measure the push-pull forces applied during the varus-valgus exam. Six surgeons with varying experience independently conducted a varus-valgus exam in extension and flexion and reported the gaps that they observed. Motion capture was used to measure the gaps between femur and tibia by placing cluster of reflective markers on femur and tibia. Subsequently, each surgeon chose the tibial insert that they thought best fit each knee. The measured peak applied forces were related to the insert thickness and the measured gaps were compared to the observed gaps by surgeons. Since insert thickness was in 1 mm increments, 1 mm gap error was considered a meaningful difference. RESULTS/ANTICIPATED RESULTS: The peak forces varied among surgeons for each cadaver. In cadaver one, the peak forces in varus and valgus in extension were 48±20 and 20±12 N, and in flexion were 27±14 and 8±11 N. Peak forces in cadavers two and three were similar; in varus and valgus in extension, 24±14 and 35±10 N, and in flexion, 23±12 and 20±10 N, respectively. It was observed that the larger the valgus force in extension, the thinner was the inserts choice (β = −0.08 mm/N, p = 0.012). In extension, the difference between estimated gaps and measured gaps was > 1 mm for 36% of all assessments and 91% of gaps were underestimated. Only one measure, however, was underestimated by > 2 mm. In flexion, gap estimates were > 1 mm for 35% of all measurements and 59% of all measurements were overestimated. Four measures were overestimated, and one was underestimated by > 2 mm. DISCUSSION/SIGNIFICANCE OF IMPACT: We found that the applied forces varied among surgeons and a negative association between insert thickness and forces in extension valgus exam. We also found that error in gap estimates among surgeons was > 1 mm a third of the time and that underestimation is more common in full extension, which may lead to using smaller inserts that affect knee stability. CONFLICT OF INTEREST DESCRIPTION: The corresponding author has no COI but my coauthors had the following COI:

1. 1. Royalties from a company or supplier: Zimmer; Stryker; Exactech, Inc; Lima; Mathys Ltd.

2. 2. Speakers bureau/paid presentations for a company or supplier Acelity; Flexion Therapeutics; Smith & Nephew; Exactech, Inc; Mallinckrodt Pharmaceuticals; Stryker.

3. 3B. Paid consultant for a company or supplier Acelity; DePuy Synthes; Exactech, Inc; Flexion Therapeutics; Intellijoint; Smith & Nephew; Zimmer; Stryker

4. 4. Stock or stock options in a company or supplier Imagen; Insight Medical; Intellijoint; Parvizi Surgical Innovation; OrthAlign; Orthobond.

5. 5. Research support from a company or supplier as a Principal Investigator Acelity; Exactech, Inc; Intellijoint; Smith & Nephew; Mallinckrodt Pharmaceuticals; Stryker; Lima.

6. 6. Royalties, financial or material support from publishers (The following conflicts were disclosed) Exactech, Inc.

7. 7. Medical/Orthopaedic publications editorial/governing board Bone and Joint Journal 360; Journal of Orthopaedics and Traumatology; Techniques in Orthopaedics.

8. 8. Board member/committee appointments for a society Knee Society; Eastern Orthopedic Association.

OBJECTIVES/GOALS: The aim of this study was to quantify balance impairments in stance in individuals with degenerative cervical myelopathy (IwDCM) in response to external perturbations. IwDCM have damage to their spinal cord due to degeneration of the cervical vertebral column, but little is known about balance. METHODS/STUDY POPULATION: Recovery time following a perturbation may be an important measure of balance. Changes in recovery time were measured in 7 IwDCM (2m, 58.59±15.00y) and 6 controls without DCM (2m, 56.91±11.04y) as they stood on an instrumented treadmill and received cued (predictable) and uncued (unpredictable) lateral pulls to the waist at 12% (high) and 6% (low) pull magnitudes. Individuals stood with feet together, shoulder width, and wide. Recovery time was defined as the time following pull onset when the absolute value of the center of pressure velocity returned to <1x baseline standard deviation. Repeated measures ANOVA was performed on recovery time. RESULTS/ANTICIPATED RESULTS: We anticipate that feet together standing, unpredictable, higher magnitude perturbations will be most challenging evidenced by longer recovery times. For waist pull recovery time, there was a trend for a Group x Predictability x Magnitude x Stance Width interaction (p = 0.1) which we anticipate being greater with additional participants. There were significant Group x Predictability x Stance Width (p = 0.01) and Group x Magnitude x Predictability (p = 0.01) interactions. IwDCM had exaggerated recovery times in narrow and wide stances with unpredictable pulls. IwDCM recovered more slowly in response to unpredictable higher magnitude pulls. DISCUSSION/SIGNIFICANCE OF IMPACT: Balance responses in IwDCM are most impaired in narrow stances and when perturbations are unpredictable. Rehabilitation should focus on shortening latency of response timing and increasing power utilization during balance response to promote quicker recovery.

OBJECTIVES/GOALS: Mammals require iron for hemoglobin, respiration, immunity and as cofactor in enzymes. But free iron is toxic from the production of reactive oxygen species. Ferroportin is the sole exporter of cellular iron and it crucially determines cellular and systemic iron levels. Labile iron must be tightly regulated. This requires structural understanding. METHODS/STUDY POPULATION: We built structure of human ferroportin (FPN1) using the ab ignition prediction approaches of Rosetta/Robetta and by comparative modeling with distance restraints in MODELLER. Templates selected were from solute carrier protein families of distantly related orthologs and homologs including a proton coupled peptide transporter (PDB ID: 4IKV) and the bacterial iron transporter in outward-open and inward-open states, (PDB ID: 5AYM, 5AYO). Each model was validated by experimental mass spectrometry data. The energy minimized structural model was inserted into a lipid bilayer, placed in a rectangular simulation box, covered with TIP3P water solvent balanced with counterions and conditioned. Finally, we carried out 350 nanoseconds molecular dynamics simulations. RESULTS/ANTICIPATED RESULTS: Our first model of FPN1 (571aa), using Rosetta/Robetta ab initio approach, resembles the structure of the proton-dependent transporter, POT and consists of 12 transmembrane helices. The membrane spanning helices veer away from the orientation in the structure of 4IKV. The alternate model using MODELLER and the method of satisfaction of constraints, returned one template, the structure of Bdellovibrio bacteriovorus iron (Fe2+) transporter homolog (5AYN, 440aa) with sequence identity of 19%. Aligning FPN1 on the template sequence incorporating structural information revealed better conservation (29%). This model also comprises 12 transmembrane helices in two bundles separated by a large intracellular loop. The iron binding site predicted in both models match the structures of distant bacterial homologs. DISCUSSION/SIGNIFICANCE OF IMPACT: We are using these experimentally verified structures and functional data to answer questions about the mechanism of ferroportin iron transport, structural dynamics and the significance of mutations in ferroportin seen in different populations, especially the Q248H mutation found in Africans and black Americans with moderate to high prevalence.

OBJECTIVES/GOALS: Our lab previously identified a population of vascular smooth muscle (SMC)-derived progenitor cells (AdvSca1-SM) which expand robustly in response to disease and can differentiate into multiple cell types. We now aim to define the role of these AdvSca1-SM cells in atherosclerotic plaque progression. METHODS/STUDY POPULATION: Goal one uses SMC lineage tracing mice and a model of atherosclerosis to track reprogramming of SMCs to AdvSca1-SM cells in the setting of disease. Arteries are analyzed using flow cytometry and immunofluorescence to quantify changes in number of mature SMCs and AdvSca1-SM cells. Goal two uses AdvSca1-SM lineage tracing mice with high cholesterol-induced atherosclerosis and plaque neovascularization. Arteries are analyzed to quantify expansion of AdvSca1-SM cells, subsequent re-differentiation into mature SMC, endothelial cells, or macrophages, and contribution to plaque neovascularization. Mechanistic findings from both goals are being investigated in diseased human coronary arteries. RESULTS/ANTICIPATED RESULTS: Flow cytometry from SMC lineage tracing mice revealed a 7- to 13-fold expansion of AdvSca1-SM cells in carotid arteries (p<0.001) and aortas (p = 0.03) after 6 weeks of western diet; no differences in macrophage numbers were observed. Additional SMC and AdvSca1-SM cell lineage tracing mice are on atherogenic diets to assess early and advanced atherosclerosis. We predict that AdvSca1-SM cells will contribute to macrophage accumulation as well as plaque neovascularization in the setting of severe atherosclerosis. Translational relevance of mechanisms driving SMC reprogramming and AdvSca1-SM cell contribution to plaque progression are being applied to studies of diseased human coronary arteries. DISCUSSION/SIGNIFICANCE OF IMPACT: Our data suggest a role for AdvSca1-SM cells in atherosclerosis. Ongoing work will clarify the mechanisms driving plaque-associated AdvSca1-SM expansion and define the ultimate fates of these cells. In vivo modulation of this process could provide the basis for future anti-atherosclerotic therapies. CONFLICT OF INTEREST DESCRIPTION: AD - CCTSI TOTTS TL1TR002533; SL - 18POST34030397 from the American Heart Association; AJ – no conflicts; KS - 1F31HL147393 from the National Heart, Lung, and Blood Institute, NIH; MM – no conflicts; RT – no conflicts; KSM – no conflicts; RAN - R01CA236222 from the National Cancer Institute, NIH, and 2018-03 from the Lungevity Foundation; and MCMW-E - R01 HL121877 from the National Heart, Lung, and Blood Institute, NIH, and 25A8679 from the Chernowitz Foundation.

OBJECTIVES/GOALS: Human placentation requires complex coordination between maternal and fetal cell types but remains incompletely understood. We hypothesize that uterine natural killer (uNK) cells, an immune cell type that increases in abundance during the implantation window, is essential for appropriate implantation and placentation. METHODS/STUDY POPULATION: We plan to examine stromal cell (SC) decidualization, spiral artery remodeling, and EVT invasion, processes vital for early pregnancy establishment, in the presence or absence of secretory phase uNK cells. Fetal extravillous trophoblasts (EVTs) will be isolated from first trimester pregnancy tissue; maternal SCs, endothelial cells (ECs) and uNK cells will be obtained from secretory phase uterine tissue. SCs will be placed in monoculture and coculture with uNK cells and prolactin will be measured to evaluate decidualization. To study EVT invasion, we will utilize our novel “implantation-on-a-chip” device to determine how addition of uNK cells affects EVT migration through a collagen-matrigel matrix. In this system, we will also examine spiral artery remodeling with or without uNK cells via TUNEL staining. RESULTS/ANTICIPATED RESULTS: We anticipate that uNK cell addition to SCs will lead to a significant increase in SC prolactin levels, suggesting a role of uNK cells in endometrial decidualization. In vitro, we expect the addition of uNK cells will increase EC apoptosis and promote EVT invasion. DISCUSSION/SIGNIFICANCE OF IMPACT: Although decidual NK cells are known to participate in placentation, the role of pre-pregnancy uNK cells is unknown. uNK cell involvement in processes important for the earliest stages of pregnancy would provide a potential marker for abnormal placentation and offer avenues for intervention to decrease placentation associated perinatal morbidity.

OBJECTIVES/GOALS: Non-alcoholic steatohepatitis (NASH) is a leading cause of cirrhosis in the world for which no anti-fibrotic therapies exist. We hypothesized that BMS-22 and maraviroc (MVC), chemokine receptor 2 (CCR2) and 5 (CCR5) antagonists, respectively, would diminish the fibrogenic activity of "fat-exposed" murine pHSCs. METHODS/STUDY POPULATION: pHSCs were isolated from livers of 6 week old male mice following 4 weeks on a NASH-inducing choline-deficient high fat diet (CDAHFD, “fat-exposed”) or standard diet (SD) and passaged in vitro. Early passage (6-12) pHSCs were plate-adhered and TGF-b-treated (10ng/mL) to maximally activate their pro-fibrogenic genes, collagen 1α1 (Col1A1), tissue inhibitor of metalloproteinase 1 (TIMP1), or α-smooth muscle actin (ACTA2). CDAHFD and SD pHSCs were then treated for 48 hours with increasing doses of BMS-22 or MVC (range: 0.3-120ng/mL) to determine (1) the degree of attenuation of the pro-fibrogenic response as measured by qPCR of fibrogenic genes (Col1A1, TIMP1,ACTA2); (2) enhancement of a fibrolytic response as measured by qPCR of matrix metalloproteinases (MMP) 2, 9 and 13 genes; and (3) pHSC migration using the scratch assay. Cell viability and CCR2 and CCR5 gene expression in response to escalating doses of antagonists were also measured. RESULTS/ANTICIPATED RESULTS: Plate- and TGF-b activated CDAHFD pHSCs had a 2-fold greater, dose-dependent attenuation of their pro-fibrogenic activity in response to BMS-CCR2-22 and MVC, when compared with plate- and TGF-b activated SD pHSCs, as measured by reductions in collagen 1α1 (Col1A1) and α-smooth muscle actin (ACTA2) gene expression. TIMP1 gene expression was unaffected by drug treatment for 48 hours. Cell viability was not affected up to doses of 30ng/mL of each drug. pHSCs also demonstrated a dose-dependent increase in CCR2, CCR5 and MMP-9 gene expression in response to surface receptor antagonism. Migration assays comparing CDAHFD and SD pHSCs in response to escalating doses of MVC and BMS-22 are ongoing and expected to demonstrate a significantly decreased migratory capacity of CDAHFD pHSCs than SD pHSCs in response to therapy, reflecting the increased susceptibility of the “fat-exposed” pHSCs to anti-fibrotic therapy than normal pHSCs. DISCUSSION/SIGNIFICANCE OF IMPACT: Anti-fibrotic drugs that dampen pro-fibrogenic activities of “fat-exposed” pHSCs are urgently needed. CCR2 and CCR5 antagonists, BMS-22 and MVC, respectively, can selectively dampen the pro-fibrogenic response of fat-exposed pHSCs, and must be considered for future trials in human NASH. CONFLICT OF INTEREST DESCRIPTION: Dr. Jill Smith has a patent licensing agreement with Immune Therapeutics, Inc.

OBJECTIVES/GOALS: Oligodendrocytes (OL) are glial cells of the central nervous system (CNS) responsible for the energy demanding task of generating myelin sheaths during development and remyelination after demyelinating injury. One metabolite shown to significantly increase ATP production in OL is the nitrogenous organic acid, creatine. Creatine plays an essential role in ATP buffering within tissues with highly fluctuating energy demands such as brain and muscle. Interestingly, mature OL, which are the cells capable of myelin production, are the main cells in the CNS expressing the rate-limiting enzyme for creatine synthesis, guanidinoacetate methyltransferase (Gamt). Patients with mutations in Gamt display intellectual disabilities, impaired myelination and seizures. Therefore, we hypothesize that creatine may be essential for developmental myelination and improve remyelination. METHODS/STUDY POPULATION: To investigate these hypotheses, we developed a new transgenic mouse model with LoxP sites flanking exons 2-6 of the Gamt gene where excision leads to expression of a green fluorescent tag allowing us to track the cells normally expressing Gamt. RESULTS/ANTICIPATED RESULTS: In this mouse model, we show a 95% (±0.47%, n = 3) co-localization of Gamt within mature OL during postnatal (P) day P14. Next, we show that knocking out Gamt leads to a significant reduction in OL in the major CNS white matter tract, the corpus callosum, at P14 and P21 (P14: 0.007, n = 3; P21: 0.04, n = 3). Here, we also investigate whether dietary creatine can enhance remyelination in the cuprizone model of toxic demyelination. DISCUSSION/SIGNIFICANCE OF IMPACT: These studies highlight the important role creatine plays in developmental myelination and investigate whether creatine can provide a therapeutic value during a CNS demyelinating insult.

OBJECTIVES/GOALS: This study’s goal is to determine whether intraoperative treatment of facial nerves with polyethylene glycol (PEG) fusion technology improves facial paralysis outcomes. Improved facial nerve regeneration in facial paralysis patients would lead to improved recovery time and effectiveness. METHODS/STUDY POPULATION: 30 rats were utilized; 15 underwent facial nerve regeneration without PEG fusion, and 15 with PEG fusion. Facial paralysis was initiated on the left by transection of the buccal and marginal mandibular branches of facial nerve. The buccal branch was repaired though microsuture technique. Neurorrhaphy sites of rats in the PEG group were exposed to calcium free saline, methylene blue, and polyethylene glycol. Nerve continuity was assessed post-operative in 5 animals in each group through electron microscopy. Functionality was assessed in the other 10 per group by EMG and whisker analysis after surgery, and weekly for 8 weeks. At 8 weeks, nerves and distal muscles were histologically analyzed. RESULTS/ANTICIPATED RESULTS: PEG fusion technology immediately restored axonal continuity following surgery, demonstrated by electron microscopy. Electrophysiology was also similarly restored across the site immediately, determined through intraoperative nerve stimulation, in the PEG fusion group. The nonintervention group showed dramatically reduced functional recovery than the PEG fusion group following surgery, shown by lower whisking activity and poor electrophysiology outcomes. Furthermore, the PEG fusion group showed statistically significant higher fascicle counts, myelination diameter, axonal diameter, and distal muscle fibers histologically. DISCUSSION/SIGNIFICANCE OF IMPACT: This study demonstrates that polyethylene fusion technology may improve facial reanimation outcomes. PEG is already a FDA-approved drug, and thus the pathway to translational clinical application of this work may thus be streamlined, bringing new options to patients with facial paralysis.

OBJECTIVES/GOALS: The goal of this study is to evaluate and optimize the characterization of cystoid macular edema (CME) using an investigational swept source (SS)-OCT system. Our knowledge of CME in preterm infants is limited; optimizing its characterization is a critical step in understanding its impact on vision. METHODS/STUDY POPULATION: In this IRB-approved protocol, 118 preterm infants were imaged in the Duke intensive care nursery (ICN) with a novel lightweight, hand-held, high-speed, SS-OCT system following routine clinical eye exams. SS-OCT images were deidentified, automatically segmented using custom software (DOCTRAP), measured for several retinal layer thicknesses, and reviewed by masked expert graders for the presence and severity of CME. Reliability of SS-OCT measures will be assessed, and the association between CME status and retinal layer thicknesses will be calculated using logistic regression modeling. RESULTS/ANTICIPATED RESULTS: The prevalence of CME overall and by severity will be calculated. The distribution of several retinal layer thicknesses will be reported and compared by infant CME status and, when edema is present, by CME severity. Reproducibility and repeatability will be reported for objective variables, and intra-grader and inter-grader agreement will be reported for subjective variables. Multivariate logistic regression coefficients and odds ratios will be calculated for each retinal layer thickness variable. DISCUSSION/SIGNIFICANCE OF IMPACT: This study will use a novel SS-OCT system to identify retinal thickness measures that may be objective markers of CME status. This will refine the characterization of CME and provide a framework for correlating CME with functional outcomes like visual acuity. CONFLICT OF INTEREST DESCRIPTION: SC and CT have unlicensed patents on relevant technologies. CT receives royalties from Alcon and Hemosonics and consultation fees from EMMES.

OBJECTIVES/GOALS: The purpose of this study is to understand how hemodynamics during dialysis in End-Stage Renal Disease (ESRD) patients on hemodialysis (HD) affect white matter health and how those effects cause cognitive impairments. METHODS/STUDY POPULATION: We collected demographic data, comorbidities, intradialytic measurements of blood pressure and cerebral oximetry, cognitive measures in several domains using NIH Toolbox Cognition Battery, diffusion-weighted and anatomical MRIs for 20 participants on HD. Specific tracts were identified using tractography and were used to calculate the average DTI measurements in each tract. Regression analysis was used to examine the relationship between mean DTI measurements of white matter integrity and cognitive performance scores. In addition, we compared diffusion MRI and T1 anatomical images of 16 healthy age-matched controls from a previous study. RESULTS/ANTICIPATED RESULTS: In our cohort 18 participants had imaging data that could be used in the analysis. We found widespread decreases in DTI white matter integrity compared to healthy age-matched controls, mean whole-brain fractional anisotropy was .3218 in the HD cohort and .3472 in controls p = .0018. Decreased integrity was found in most of the tracts identified but more decreased in tracts implicated in cognition. Partial regression analysis identified significant relationships between the white matter integrity of the left superior longitudinal fasciculus and overall fluid cognitive performance, R = .5525, p = .0174 before multiple comparisons correction when controlling for differences due to age. DISCUSSION/SIGNIFICANCE OF IMPACT: We found a widespread decrease in white matter integrity and significant correlations between cognitive performance and specific tract integrity in our HD cohort using regions identified by tractography imaging analysis. This analysis shows that HD patients have decreased white matter health and identifies several tracts that are important for cognitive performance in HD patients.

OBJECTIVES/GOALS: The purpose of this secondary data analysis was to identify latent subgroups of seriously ill adults based on multiple chronic conditions and mortality risk using the CCI. This study was conducted by performing a secondary analysis of data from a randomized controlled trial of seriously ill patients receiving palliative care. METHODS/STUDY POPULATION: A cross-sectional analysis of baseline CCI data was conducted. 381 seriously ill adults receiving palliative care were in the original study. Latent subgroups were identified based on the CCI by conducting a latent class analysis in MPlus. The LCA was modeled on each of the 19 disease items as binary latent predictor variables, an additional binary variable representing presence of any disease not accounted for by the CCI, and a final categorical variable representing the total CCI score divided based on clinically significant cutoffs including zero, low (> = 1-<2), moderate (> = 2-<5), and high CCI (> = 5). RESULTS/ANTICIPATED RESULTS: Three distinct latent subgroups were identified based on the CCI. Latent subgroup 1 included those with a low-moderate CCI consisting of MCC and non-Metastatic Cancers (n = 178), with 45% of this group having chronic obstructive pulmonary disease. The second two subgroups included individuals with a high CCI or a score greater than or equal to 5. Latent subgroup 2 (n = 64) was comprised of individuals with MCC and non-metastatic cancer. Latent subgroup 3 (n = 139) included individuals with metastatic cancer. DISCUSSION/SIGNIFICANCE OF IMPACT: In a sample of seriously ill adults with MCC, latent subgroups were identified consisting of individuals with low, moderate, or high CCI. The low to moderate CCI group consists of individuals with chronic conditions including COPD, congestive heart failure, myocardial infarction, cardiovascular disease. There were two subgroups with high CCI scores and the differentiating factor between the two subgroups was the presence of metastatic cancer in latent subgroup 3. The identification of latent subgroups sets the groundwork for further analyses to compare differences in symptom burden, quality of life, and functional status between groups. The findings have the potential to inform future studies seeking to better characterize seriously adults with MCC based on their disease burden and mortality risk.

OBJECTIVES/GOALS: Precision care may engage smokers and providers in treatment but is understudied in the community. We piloted guideline-based care (GBC) alone or with Respiragene, a lung cancer polygenic risk score (PRS, 1-10), or metabolism-informed choice of medication using the nicotine metabolite ratio (NMR). METHODS/STUDY POPULATION: Daily smokers (n = 58) with stored biospecimens in the Southern Community Cohort Study were randomized 1:1:1 to GBC, PRS, or NMR, counseled to quit smoking, and co-selected FDA-approved cessation medication (nicotine replacement, varenicline) with a tobacco counselor. In PRS, precision motivational counseling was guided by PRS (i.e., lung cancer risk 10-40-fold that of never-smokers). In NMR, precision medication recommendations consisted of varenicline for faster metabolizers (NMR≥0.31) and nicotine replacement for slow metabolizers (NMR<0.31). Feasibility was defined as achieving at least 50% provider engagement (med prescription) and at least 50% patient engagement (self-reported med use). RESULTS/ANTICIPATED RESULTS: Participants were median age 59, 72% female, 81% Black, 60% with incomes <$15,000; median cigarettes/day was 15 (IQR 8-20) and 52% reported time-to-first cigarette <5 minutes, illustrating moderate nicotine dependence. Providers confirmed medication prescriptions for 40% of patients (32% GBC, 50% PRS, 37% NMR) and 83% of patients reported using medication (prescribed or unprescribed) during the study (90% GBC, 80% PRS, 79% NMR). At 6-month follow-up, 27% (n = 15) reported cessation (39% GBC, 16% PRS, 26% NMR). Among persistent smokers, 46% reported smoking at least 50% fewer cigarettes/day compared to baseline (45% GBC, 38% PRS, 57% NMR). Small sample size precluded statistical comparisons. DISCUSSION/SIGNIFICANCE OF IMPACT: Precision interventions to quit smoking are feasible for community smokers, who engaged at high rates. However, only 40% of providers supported patients’ quit attempts with medication prescriptions. Future research should test strategies to raise provider engagement in precision smoking treatment. CONFLICT OF INTEREST DESCRIPTION: R.F.T. has consulted for Quinn Emmanual and Apotex on unrelated topics. H.A.T. reported providing input on design for a phase 3 trial of cytisine proposed by Achieve Life Sciences and being a principal investigator of National Institutes of Health–sponsored studies for smoking cessation that include medications donated by the manufacturers. Other authors declare no potential conflicts of interest.

OBJECTIVES/GOALS: One in six children in the U.S. has a Neurodevelopmental Disability (NDD). Prechtl’s General Movement Assessment (GMA) is a qualitative predictor of early motor dysfunction. However, no quantitative biomechanical assessment exists to more accurately identify all patients with NDD. METHODS/STUDY POPULATION: With UAMS IRB approval, as part of a larger study, healthy infants were filmed while lying supine on a force plate for 2 minutes. We studied 12 healthy full-term infants (gestational age: 38.9±1.5 weeks, age: 2.1-7.0 months; 7M, 5F; length: 64.0±5.2 cm; weight: 7.2±1.3 kg). Within our data set there were 3 infants transitioning to fidgety period (≤3 months), 4 in the fidgety period, (3-5 months), and 5 that matured beyond fidgety period (>5 months). Center of pressure (COP) path-lengths were gathered from the force plate at 1000 Hz. We grouped our data with K-means clustering and performed statistical analysis with ANOVA. RESULTS/ANTICIPATED RESULTS: We divided our data into 3 distinct clusters. The first group contained infants with moderate variability of movements which included 2 infants between 3 and 5 months and 2 infants slightly outside of this range. The second group, with mild variability in movements, included 4 infants between 2 and 3 months as well as 2 infants just older than 5 months. The third group, with little variability in movements, included 2 infants older than 5 months. A GMA reader (TJ) qualitatively confirmed these findings with video footage. Using a threshold of p<0.05, data sets within the clusters were similar and significantly different from other clusters. DISCUSSION/SIGNIFICANCE OF IMPACT: Fidgety infants have greater variability in COP patterns than their mature counterparts. We anticipate additional COP measurements will correspond with qualitative GMA analyses. Artificial Intelligence-based quantification of the GMA may be useful in earlier detection or prediction of NDD outcomes.

OBJECTIVES/GOALS: Virtual surgical planning and 3D printing enable streamlined surgeries and increased complexity. These technologies, however, require CT scans and radiation exposure. This project’s goal is to optimize and demonstrate the accuracy of Black Bone MRI for surgical planning in reconstructive surgery. METHODS/STUDY POPULATION: Four common craniofacial surgeries were planned and performed on cadaver specimens (maxillary advancement, orbital floor reconstruction with patient-specific implants, cranial vault reconstruction, and fibular free flap reconstruction of the mandible). For each surgical procedure, ten cadaver heads were used. Five of each surgery were planned and 3D printed guides were created utilizing Black Bone MRI versus five with CT scans. Following mock surgeries, all specimens underwent a post-operative CT scan. 3d reconstruction was performed and surgical accuracy compared to the plan was assessed using GeoMagic Wrap, assessing average post-operative deviation from plan. RESULTS/ANTICIPATED RESULTS: In all surgeries, guides created from Black Bone MRI demonstrated high accuracy to surgical plan. Average osteotomy (cut) deviation from plan was not statistically significantly different when Black Bone MRI was used compared to CT scans for planning and guide creation in the wide variety of craniofacial surgeries performed. The average deviation of post-operative anatomy from pre-operative plan was also not statistically significant when Black Bone MRI versus CT scans were utilized in the surgeries. These results then enabled the translational application of this technology clinically, and we demonstrate a clinical reconstructive craniofacial case planned utilizing Black Bone MRI. DISCUSSION/SIGNIFICANCE OF IMPACT: This study demonstrates that virtual surgical planning and 3d surgical guide creation can be performed using Black Bone MRI with comparable accuracy to CT scans in a wide variety of craniofacial procedures. This could dramatically reduce radiation exposure for patients. The successful segmentation, virtual planning, and 3d printing of accurate guides from Black Bone MRI demonstrate potential to change the pre-operative planning standard of care. This project, overall, also demonstrates the development of new solutions to advance clinical care, thus serving as an example of moving translational science from a concept to the operating room.

OBJECTIVES/GOALS: To compare the opioid drug requirements amongst those individuals with high levels of catecholamines in blood and acute post-procedural pain, by ICD9/10 codes (experimental) to those with normal levels of catecholamines and acute post-procedural pain (AP-PP) only (controls) METHODS/STUDY POPULATION: In collaboration with both the Informatics and the Biostatistics Departments at CTSI and under the auspices of the IRB at the University of Rochester, we completed the collection of ~8,000 electronic health records(EHRs) of adults 18 years and older with surgical appointments at Strong Memorial Hospital (SMH), who met inclusion criteria, from January 2006 to September 2019 and received Fentanyl therapy for AP-PP management. Subjects were categorized in a two-arm-matched case-control fashion. A ratio of 1(Experimental):1(Control) was utilized. Analytic comparisons were completed using normal distribution statistical methods with p >0.1 for significance. RESULTS/ANTICIPATED RESULTS: After removal of duplicates and exclusion of EHRs, a total of 17 subjects met inclusion criteria for the experimental group. We matched controls (n = 17) with experimental subjects for age, gender and surgical procedure for accurately compare opioid requirements in the postoperative recovery. Mean age of subjects was 69(+/-10.1235) years old. Most of subjects were females (70%). Mean Fentanyl requirement was significantly different in the experimental group 466.17(625.621)mcg compared to 215.58(353.323)mcg in the controls (p value 0.07832) DISCUSSION/SIGNIFICANCE OF IMPACT: It is suggested that healthy individuals with genetic variations in pain pathways including; the COMT and MAOA rendered individuals with higher levels of catecholamines in the body driving abnormal responses to pain sensitivity. We emulated this genetic variation for clinical purposes using ICD10/9 codes of those with conditions related to higher catecholamine levels in the body. Based on our preliminary results, we suggest that COMT and MAOA genetic variations could impact opioid drug use and the current opioid dependency and epidemics in the U.S. This study will address remarkable questions and identify strategies about this topic.

OBJECTIVES/GOALS: The Monte Carlo dose calculation method is often considered the “gold standard” for patient dose calculations and can be as radiation dose measurements. Our study aims to develop a true Monte Carlo model that can be implemented in our clinic as part of our routine patient-specific quality assurance. METHODS/STUDY POPULATION: We have configured and validated a model of one of our linear accelerators used for radiation therapy treatments using the EGSnrc Monte Carlo simulation software. Measured dosimetric data was obtained from the linear accelerator and was used as the standard to compare the doses calculated with our model in EGSnrc. We will compare dose calculations between commercial treatment planning systems, the EGSnrc Monte Carlo model, and patient-specific measurements. We will implement the Monte Carlo model in our clinic for routine second-checks of patient plans, and to recalculate plans delivered to patients using machine log files. RESULTS/ANTICIPATED RESULTS: Our Monte Carlo model is within 1% agreement with our measured dosimetric data, and is an accurate representation of our linear accelerators used for patient treatments. With this high level of accuracy, we have begun simulating more complex patient treatment geometries, and expect the level of accuracy to be within 1% of measured data. We believe the Monte Carlo calculation based on machine log files will correlate with patient-specific QA analysis and results. The Monte Carlo model will be a useful tool in improving our patient-specific quality assurance protocol and can be utilized in further research. DISCUSSION/SIGNIFICANCE OF IMPACT: This work can be implemented directly in clinical practice to ensure patient doses are calculated as accurately as possible. These methods can be used by clinics who do not have access to more advanced dose calculation software, ensuring accuracy for all patients undergoing radiotherapy treatments.

OBJECTIVES/GOALS: Our goal is to determine if a precision medicine approach to guide anti-platelet therapy for patients with ACS, post PCI, is feasible for a diverse urban population. Also, we will evaluate if guided therapy reduces major adverse cardiovascular events (MACE) while remaining economically sustainable. METHODS/STUDY POPULATION: This prospective, pragmatic study will enroll two-hundred patients with ACS undergoing PCI, receiving DAPT. Patients will receive point-of-care CYP2C19 genotyping. Patients with at least one loss-of-function (LoF) allele will be recommended prasugrel. Those without LoF alleles, will be recommended to take prasugrel for 7 days then clopidogrel for 7 days, followed with platelet reactivity phenotyping. Patients with HPR >208 P2Y12 reaction units will take prasugrel; the remainder will take clopidogrel. We will review electronic health records and contact patients at baseline, then at 1, 3, 6, and 12 months to collect data for cardiovascular and health-related quality of life (HRQoL) outcomes. RESULTS/ANTICIPATED RESULTS: Feasibility and clinical utility will be measured by the proportion of patients with a genotype or phenotype leading to a clinical recommendation of alternative therapy and whether or not recommendations were accepted by clinicians. Effectiveness will be measured by combined MACE (composite of cardiovascular death, non-fatal myocardial infarction, and non-fatal stroke), stent thrombosis, and major and minor bleeding over the study period. Cost of testing, 30-day hospital readmission, and HRQoL questionnaires will be included for pharmacoeconomic analysis from an institutional perspective. DISCUSSION/SIGNIFICANCE OF IMPACT: There are no studies investigating the clinical utility of implementing guided anti-platelet selection, combining CYP2C19 genotyping and HPR phenotyping. We anticipate incorporating this precision medicine approach to guide P2Y12 inhibitor selection will be feasible while improving patient outcomes.