OBJECTIVES/GOALS: Large-scale clinical proteomic studies of cancer tissues often entail complex workflows and are resource-intensive. In this study we analyzed ovarian tumors using an emerging, high-throughput proteomic technology termed SWATH. We compared SWATH with the more widely used iTRAQ workflow based on robustness, complexity, ability to detect differential protein expression, and the elucidated biological information. METHODS/STUDY POPULATION: Proteomic measurements of 103 clinically-annotated high-grade serous ovarian cancer (HGSOC) tumors previously genomically characterized by The Cancer Genome Atlas were conducted using two orthogonal mass spectrometry-based proteomic methods: iTRAQ and SWATH. The analytical differences between the two methods were compared with respect to relative protein abundances. To assess the ability to classify the tumors into subtypes based on proteomic signatures, an unbiased molecular taxonomy of HGSOC was established using protein abundance data. The 1,599 proteins quantified in both datasets were classified based on z-score-transformed protein abundances, and the emergent protein modules were characterized using weighted gene-correlation network analysis and Reactome pathway enrichment. RESULTS/ANTICIPATED RESULTS: Despite the greater than two-fold difference in the analytical depth of each proteomic method, common differentially expressed proteins in enriched pathways associated with the HGSOC Mesenchymal subtype were identified by both methods. The stability of tumor subtype classification was sensitive to the number of analyzed samples, and the statistically stable subgroups were identified by the data from both methods. Additionally, the homologous recombination deficiency-associated enriched DNA repair and chromosome organization pathways were conserved in both data sets. DISCUSSION/SIGNIFICANCE OF IMPACT: SWATH is a robust proteomic method that can be used to elucidate cancer biology. The lower number of proteins detected by SWATH compared to iTRAQ is mitigated by its streamlined workflow, increased sample throughput, and reduced sample requirement. SWATH therefore presents novel opportunities to enhance the efficiency of clinical proteomic studies.

OBJECTIVES/GOALS:

• Development of gold nanoparticles covalently linked to a photosensitizer for use to enhance radiation therapy. The particles will be thoroughly characterized structurally and mechanistically. The gold particles should enhance radiation activity by closer proximity to the photosensitizer and by increasing particle accumulation in the tumor.

METHODS/STUDY POPULATION:

• Gold nanoparticles were synthesized and coated with amine-terminated poly(ethylene) glycol, then covalently conjugated to chlorin e6, a known FDA-approved photosensitizer. The system was characterized using UV-Vis spectroscopy, transmission electron microscopy, and nanoparticle tracking analysis. The generation of reactive oxygen species was measured after X-irradiation. Enhanced cell killing was evaluated clonogenically in addition to assessment of in vivo efficacy and tumor pathology.

RESULTS/ANTICIPATED RESULTS:

• Conjugation of the particle to the photosensitizer was achieved, and the molecule was detected by UV-Vis spectroscopy. TEM and NTA showed no aggregation of the particles, and an increase in reactive oxygen species generation was observed. The conjugates increased cell killing during radiation treatment, whereas neither the particle alone nor the photosensitizer significantly affected clonogenic survival at the same concentrations. Breast tumors grown in immunocompetent mice showed increased necrotic tissue after a single 20 gy treatment in the presence of the conjugate.

DISCUSSION/SIGNIFICANCE OF IMPACT: Radiation therapy is widely used clinically, but dosage is limited largely to prevent injury to adjacent normal tissue. By increasing the local effect of radiation therapy, our gold conjugate has the potential to augment the effective radiation dose in the tumor, thereby reducing damage to healthy tissue and providing a more effective therapy.

OBJECTIVES/GOALS: To examine the relationship between epigenetic age acceleration (EAA) and depressive symptoms in a cohort of African American women (AAW) with cardiometabolic conditions (CMC) including hypertension, diabetes, obesity; and to explore clinical phenotypes of depressive symptoms in this population. METHODS/STUDY POPULATION: This secondary analysis utilized genomic and longitudinal clinical data from AAW in the InterGEN cohort (n = 250). EWAS data was used to estimate EAA based on the Horvath method, which incorporates the DNA methylation statuses at 353 specific CpG sites and regresses this epigenetic age on chronological age to determine EAA. Pearson’s correlations and linear regression will be used to examine the relationship between EAA and depressive symptoms and a linear mixed model will investigate this relationship over four time points during a two-year period. Clinical phenotyping of depressive symptoms will be explored using a cluster analysis. RESULTS/ANTICIPATED RESULTS: Analysis is underway and will be complete by the time of presentation. We hypothesize that higher EAA will associate with higher depressive symptoms and poorer trajectories over time. We expect that this relationship may be meditated by the presence of CMCs. Exploratory analysis of clinical phenotyping is expected to provide descriptive evidence with respect to specific depressive symptoms or clusters which are most associated with EAA and CMCs. These results will address several gaps. To our knowledge, this is the first study to examine the relationship of EAA and depressive symptoms considering the role of CMC, in a historically understudied population with disproportionate risk. DISCUSSION/SIGNIFICANCE OF IMPACT: Depression limits life quality and quantity and is highly comorbid in CMC. AAW have high risk of comorbidity, and this study furthers knowledge of depression and aging with a clinically accessible marker and aids recognition of a heterogenous phenotype in an undertreated population.

OBJECTIVES/GOALS: To develop feasible screening methods for activity of the enzyme Glucose-6-phosphate dehydrogenase (G6PD) with point of care applicability. METHODS/STUDY POPULATION: Current knowledge establishes the relevance of G6PD as a critical therapeutic determinant for effective antimalarial therapy due to the occurrence of mutations that lead to post-treatment severe adverse effects. We present our findings on development of cost effective point-of-care screening methodologies to ascertain G6PD deficiency. RESULTS/ANTICIPATED RESULTS: Using Patient Cohort Explorer and data from the Department of Pathology, we established the prevalence of G6PD deficiency at the University of Mississippi Medical Center, Jackson, MS as high as 11.8% (African-American males in all population, n = 2518). Next, for selection of potential target groups, we set up a protocol for recruitment of volunteers based on ethnic background, parental ethnicity, and medical history. G6PD activity was evaluated using point of care methods [Trinity Biotech test or CareSTART Biosensor], and Gold Standard quantitative spectrophotometric assay (LabCorp). Determinations in >20 subjects have showed comparable concordance. If used with a conservative interpretation of the signal, the Trinity Biotech test showed superior potential for use in the field relative to the CareSTART Biosensor. DISCUSSION/SIGNIFICANCE OF IMPACT: We established the prevalence of G6PD deficiency in our medical center. We have also setup tests for point-of-care assessment of G6PD. Pending evaluation of the relative tests performance, we will be in position to screen individuals and select them for a prospective clinical trial to evaluate the safety of antimalarial agents on scope of G6PD deficiency.

OBJECTIVES/GOALS: Atrial fibrillation (AF) not only is the most common sustained cardiac arrhythmia in clinical practice placing patients at increased risk for thromboembolic events. Hispanics despite having a higher risk factor burden for developing AF have a lower overall incidence and prevalence of AF when compared to Non-HispanicWhites (NHW). European ancestry in the African American population was found to be an independent predictor for developing AF. Consequently, we have decided to evaluate if European ancestry is an independent risk factor for Puerto Rican Hispanics(PRH) METHODS/STUDY POPULATION: This project is a secondary analysis of a Puerto Rican population sample (n = 250) fromThePharmacogenetics of Warfarin in Puerto Ricans StudyandAGenomic Approach for Clopidogrel in CaribbeanHispanics; and1000GenomeProject to establish a control group of healthy PRH population. We will evaluate the presence of 111 known single nucleotide polymorphisms(SNP)associated with AF Europeans and determine the frequency in PRH population sample.Using admixture informatic markers (AIM) analysis will determine the percentage of admixture. Statistical analysis will include the use of Pearson Product-MomentCoefficient correlation analysis and multivariate regression. For admixture will use Maximum LikelihoodEstimation Markov Chain Monte Carlo models RESULTS/ANTICIPATED RESULTS: We anticipate that higher-frequency of AF associated European SNPs and overall higher percentage of European admixture will be associated with atrial fibrillation in PRH patients. DISCUSSION/SIGNIFICANCE OF IMPACT: The expected outcomes for this study are to identify the frequency of known genetic loci associated with AF Europeans and validate their use PRH population for machine learning risk factor models.

OBJECTIVES/GOALS: Knocking down the inflammatory response following joint trauma may halt the cytokine cascade and prevent the resulting cyclic degradation of articular cartilage. MK2 inhibiting (MK2i) peptides are an emerging and promising class of pharmaceutical to treat post-traumatic osteoarthritis (PTOA); however, these peptides are susceptible to proteolytic degradation in the extracellular space. Our objective is to encapsulate MK2i in thermoresponsive hollow nanoparticles (hNPs) to knockdown the inflammatory cytokine IL-6 to prevent the cyclic degradation of articular cartilage. METHODS/STUDY POPULATION: NP Synthesis: N-isopropyl acrylamide (NIPAm) cores was initiated by potassium persulfate (KPS) in aqueous solution with sodium dodecylsulfate (SDS) at 70°C under a nitrogen for 2 hours. Then exposed to oxygen for 45 min, followed by a nitrogen purge. NIPAm, 2-acrylamindo-2-methyl-1-propanesulfonic acid (AMPS), N,N’-bis(acryoyl)cystamine (BAC), and Acrylic Acid (AAc), in fluorescent batches rhodamine b isothiocyanate (RBITC), were polymerized around the core to form the shell. NPs were purified using tangential flow filtration. The NPs were dialyzed at 4°C for 14 days to remove the core and form hNPs. Loading & Release: hNPs and MK2i were incubated at 1 mg/ml at 4°C for 24 h. MK2i released into 1x PBS and analyzed on HPLC. IL-6 Expression: Bovine chondrocytes seeded at 10,000 cell/cm2 were stimulated with 20 ng/ml IL-1b daily and treated once with 100 µg/ml MK2i loaded-NP or 100 µg/ml free MK2i treatment on day 2. Analyzed on bovine IL-6 ELISA. In Vivo Intra Articular Injections: 75 µl of 2 mg/ml hNPsRHB or a PBS control was injected into the right knee of 4-month old Fischer 344 (Envigo) rats. Rats were imaged daily for 7 days then euthanized, legs dissected, and imaged. RESULTS/ANTICIPATED RESULTS: Core removal facilitated increased MK2i release from hNPs, Fig 1A, allowing up to 63% after 5 days in PBS. The hNPs generated here offer a continual sustained release of MK2i and hNPs are non-cytotoxic (data not shown) up to 12 mg/ml. MK2i loaded-NPs significantly knocked down IL-6 production after a single treatment after 2 days, Figure 1B, and continued knockdown for up to 4 days. hNPsRBITC was successfully injected into rat joint space and was retained for at least 7-days compared to pre-injection and PBS control, Fig 1 B-C. DISCUSSION/SIGNIFICANCE OF IMPACT: hNPs protect MK2i from ECM degradation and offer continual sustained release into chondrocytes. Core removal allows for MK2i release in vitro with further sustained release compared to previous non-degradable model. The single MK2i treatment lead to a significant IL-6 knockdown bovine chondrocytes for up to 4 days in hNPs. We were able to successfully inject and retain fluorescently labeled hNPs within rat knees for 7 days. Our translational therapeutic shows the promise of delivering a degradable, non-cytotoxic hNP into the joint space to knockdown the inflammatory response to halt the cyclic progression of articular cartilage degradation and progression of PTOA. CONFLICT OF INTEREST DESCRIPTION: The authors declare the following competing financial interest(s): Moerae Matrix, Inc. has a worldwide exclusive license to the CPP (MK2 inhibitor peptide). A. Panitch owns greater than 5% of Moerae Matrix, Inc.

OBJECTIVES/GOALS: The objective of the study is to compare two horizontal bone augmentation techniques (bone expansion and bone splitting) that are currently used for horizontally deficient maxillary ridges. Bone expanded in millimeters (mm), implant stability, and patient satisfaction will be compared with each technique. METHODS/STUDY POPULATION: This pilot (prospective cohort) study will be divided in two sites, a private practice and the Oral and Maxillofacial Surgery (OMS) Clinic at the University of Puerto Rico, School of Dental Medicine. A total of 20 patients will be selected, 10 patients in each site. In both sites, pre-operative and post-operative Cone Beam CT radiographs will be taken to measure bone width. Implant stability will be measured using an Osstell. 2 weeks post-surgery, a patient satisfaction questionnaire will be given to patients. A two-sample T test will be used to compare techniques statistically. RESULTS/ANTICIPATED RESULTS: We anticipate that bone expansion will be as good as (non-inferiority) bone splitting in terms of bone expanded in millimeters to desire width, and implant diameter will not be compromised. We also expect that implants placed with the bone manipulation technique will have a higher implant stability at baseline and less pain, discomfort and swelling in terms of patient satisfaction. DISCUSSION/SIGNIFICANCE OF IMPACT: Our contributions here are expected to illustrate clinical and radiographic bone expansion techniques that will enhance implant placement treatment for implantologists and patient’s experience.

OBJECTIVES/GOALS: The use of P2Y12 receptor inhibitors like Clopidogrel is crucial in the prevention of thrombotic events in patients with coronary artery disease, peripheral arterial disease, and cerebrovascular disease. Variation in the level of platelet inhibition is present in many patients, and it is associated with the occurrence of major adverse cardiovascular events (MACEs). The term High-on treatment platelet reactivity (HTRP) is used to describe impaired antiplatelet inhibition while on Clopidogrel. Multiple factors have been associated with the presence of HTPR in patients with CAD and PAD, including CYP2C19 loss of function polymorphism, drug-drug interactions, and medical comorbidities. Gender differences are another factor that might influence the levels of platelet inhibition while on Clopidogrel and hence, HTPR. Differences by Gender exist in platelet biology, count, and activation. The evidence for the influence of Gender in HTPR is limited, but a possible association has been described. In this study, we described the association of Gender with HTPR and Major Adverse Cardiovascular Events (MACEs) occurrence. The data is from a sample of Hispano-Caribbean patients on Clopidogrel therapy alone or in combination with Aspirin that were retrospectively evaluated from an ongoing trial in Puerto Rico. The result of this study provided evidence of the influence that Gender has on antiplatelet therapy function and MACEs occurrence. METHODS/STUDY POPULATION: The population in the study consisted of Hispano-Caribbean patients using Clopidogrel alone or in combination with Aspirin for coronary artery disease, peripheral arterial disease, or cerebrovascular disease. The sample was obtained from multiple hospital institutions with cardiovascular services in Puerto Rico during the years 2016-2019. Patients were part of the ongoing trial, “Adopting a precision medicine paradigm in Puerto Rico: leveraging ancestral diversity to identify predictors of Clopidogrel response in Caribbean Hispanics.” The sample size consisted of 150 patients. Participants were recruited during routine medical care, pre-admission evaluation for elective cardiac procedures, or during hospitalization in the participating institutions. Platelet reactivity testing was performed with the system Verify Now® to determine PRU values, and High on-treatment platelet reactivity was defined as PRU ≥208. One year after recruitment, the patients were re-evaluated for the occurrence of MACEs. The association of the variables HTPR, occurrence of MACEs, and Gender were assessed using logistic regression in addition to the role of HTPR and Gender for predicting MACE occurrence. The analysis was done using the statistic software Intellectus ©. RESULTS/ANTICIPATED RESULTS: The sample consisted of 67 females and 83 males with and Mean age of 67.87 years and 61.11 years, respectively. The prevalence of HTPR in the sample was 32.67 % (n = 49) with 36% (n = 24) for females, and 30%(n = 25) for males. The mean PRU values were 179.54 for females and 170.81 for males. The percentage of MACEs one year after recruitment was 29.33 % (n = 44) with 43% on females (n = 19), and 57% on males (n = 25). Logistic regression for Gender predicting HTPR was non-significant with a χ2(2) = 0.55, p = .758, and McFadden R2 = 0.00. Also, logistic regression for the effects of Gender and HTPR on the Odds of MACEs occurrence was not significant based on a model with an alpha of 0.05, χ2(2) = 1.99, p = .370, and McFadden R2 = 0.01. DISCUSSION/SIGNIFICANCE OF IMPACT: The sample consisted of 67 females and 83 males with and Mean age of 67.87 years and 61.11 years, respectively. The prevalence of HTPR in the sample was 32.67 % (n = 49) with 36% (n = 24) for females, and 30%(n = 25) for males. The mean PRU values were 179.54 (±70.42) for females and 170.81(±64.89) for males. The percentage of MACEs one year after recruitment was 29.33 % (n = 44) with 43% on females (n = 19), and 57% on males (n = 25). Logistic regression for Gender predicting HTPR was non-significant with a χ2(2) = 0.55, p = .758, and McFadden R2 = 0.00. Also, logistic regression for the effects of Gender and HTPR on the odds of MACEs occurrence was not significant based on a model with an alpha of 0.05, χ2(2) = 1.99, p = .370, and McFadden R2 = 0.01.

OBJECTIVES/GOALS: Acetaminophen (Tylenol, APAP) toxicity has been well documented and well explored over the last 50 years. However, there has been no investigation into identification of specific metabolites that can predict which patients will have adverse reactions to therapeutic doses of APAP. METHODS/STUDY POPULATION: 205 subjects recruited from the Denver, CO community received the highest recommended daily dosing of APAP, 4 grams, for 16 days. Subjects were grouped by 1) alanine aminotransferase (ALT) at any monitored time point above 60units/L (n = 20) vs 2) no increase in ALT at any time point (n = 185). Blood was collected at days 0, 4, 7, 16, and 31. Samples were run on ultra-high performance liquid chromatography mass spectrometry with 27 heavy-labeled standards for metabolites documented to be associated with APAP metabolism. Data will be analyzed to look for significant changes in metabolite and demographic variable expressions using t-tests, chi square and logistic regression, as appropriate. RESULTS/ANTICIPATED RESULTS: It is expected that there will be greater elevations of conjugated non-toxic APAP metabolites (APAP-glucuronide, APAP-sulfate) in subjects whose ALT did not elevate because of successful hepatoprotection. Conjugated APAP metabolites are expected to only be present in samples taken after APAP therapy initiation confirming exposure as compared to being predictive of toxic response. Increases in lactate and cysteine in pre-exposure samples would allow for prediction of APAP toxicity as they are expected to have increased expression in subjects whose ALT became elevated which is indicative of increased hepatic damage due to oxidative damage. DISCUSSION/SIGNIFICANCE OF IMPACT: Identification of metabolites and/or demographic factors associated with toxic response to APAP prior to administration could advise APAP recommendations. Quantification of post-APAP administration metabolites would identify extent of successful hepatoprotective mechanisms.

OBJECTIVES/GOALS: The detection of liver fibrotic changes at an early and reversible stage is essential to prevent its progression to end-stage cirrhosis and hepatocellular carcinoma. Liver biopsy, which is the current gold standard for fibrosis assessment, is accompanied by several complications due to its invasive nature in addition to sampling errors and reader variability. In this study, we evaluate the use of quantitative parameters extracted from hybrid ultrasound and photoacoustic imaging to detect and monitor fibrotic changes in a DEN rat model. METHODS/STUDY POPULATION: Liver fibrotic changes were induced in 34 Wistar male rats by oral administration of Diethylnitrosamine (DEN) for 12 weeks. 22 rats were imaged with B-mode ultrasound at 3 different time points (baseline, 10 weeks and 13 weeks) for monitoring liver texture changes. Texture features studied included tissue echointensity (liver brightness normalized to kidney brightness) and tissue heterogeneity. 12 rats were imaged with photoacoustic imaging at 4 time points (baseline, 5 wks, 10 wks, and 13 wks) to look at changes in tissue oxygenation. Hemoglobin oxygen saturation (sO2A) and hemoglobin concentration (HbT) in the right and left lobes of the liver were measured. 8 rats were used as controls. Liver tissue samples were obtained following 13 weeks from DEN start time for METAVIR histopathology staging of fibrosis. RESULTS/ANTICIPATED RESULTS: Texture features studied showed an increase with time in DEN rats. Normalized echointensity increased from 0.28 ± 0.06 at baseline to 0.46 ± 0.10 at 10 weeks (p < 0.0005) and 0.53 ± 0.15 at 13 weeks in DEN rats (p < 0.0005). In the control rats, echointensity remained at an average of 0.25 ± 0.05 (p = 0.31). Tissue heterogeneity increased over time in the DEN-exposed rats from a baseline of 208.7 ± 58.3 to 344.6 ± 52.9 at 10 weeks (p < 0.0005) and 376.8 ± 54.9 at 13 weeks (p = 0.06) however it stayed constant at 225.7 ± 37.6 in control rats (p = 0.58). The quantitative analyses of the photoacoustic signals showed that blood oxygen saturation significantly increased with time. At 5 weeks sO2AvT increased by 53.83 % (± 0.25), and HbT by 35.31 % (± 0.07). Following 10 weeks of DEN; sO2AvT by 92.04 % (± 0.29), and HbT by 55.24 % (± 0.1). All increases were significant p < 0.05. In the 13th week, however, the values of all of these parameters were lower than those in the 10th week, however, the decrease was statistically insignificant. DISCUSSION/SIGNIFICANCE OF IMPACT: Quantitative features from B-mode ultrasound and photoacoustic imaging consistently increased over time corresponding to hepatic damage, inflammation and fibrosis progressed. The use of this hybrid imaging method in clinical practice can help meet the significant need for noninvasive assessment of liver fibrosis.

OBJECTIVES/GOALS: Primary graft dysfunction (PGD) is acute lung injury in the first three days after lung transplant. Patients that experience PGD have increased mortality and an increased risk of chronic lung allograft dysfunction. The pathogenesis is thought to be an ischemia-reperfusion injury but is incompletely understood and there are no specific therapies. We investigated the role of the microbiome in PGD and associations with inflammation and markers of aspiration. METHODS/STUDY POPULATION: We collected airway lavage samples from lung transplant donors before procurement and recipients after reperfusion. We extracted DNA, amplified the bacterial 16S rRNA gene, and sequenced on the Illumina MiSeq platform. QIIME2 and Deblur were used for bioinformatic analysis. R packages were used for downstream analysis and visualizations. The host response was quantified using the Milipore 41-plex Luminex and an ELISA for pepsin. Clinical data was collected by the Penn Lung Transplant Outcomes Group. PGD was assessed by degree of hypoxemia and chest X-ray findings in the 72 hours after transplant. RESULTS/ANTICIPATED RESULTS: There was no significant difference in alpha diversity (Shannon index, p = 0.51), biomass (via comparison of 16S amplicon PicoGreen, p = 0.6), or beta diversity (Weighted UniFrac, p = 0.472, PERMANOVA) between subjects with PGD grade 3 (n = 36) and those that did not (n = 96). On taxonomic analysis, we found an enrichment of Prevotella in donor and recipient lungs that went on to develop PGD (p = 0.05). To follow up this finding we measured immune response and pepsin concentrations in recipient lungs. We found elevated levels in 35/41 cytokines measured in subjects that developed PGD as well as an elevation in pepsin and a correlation between pepsin concentration and Prevotella relative abundance (Figure 1). Additionally, Prevotella relative abundance had statistically significant positive correlations with multiple cytokines such as IL-6 (Pearson’s = 0.26, p = 0.009) and eotaxin (Pearson’s = 0.24, p = 0.016). DISCUSSION/SIGNIFICANCE OF IMPACT: There is an enrichment of oral anerobes in lung allografts that eventually develop PGD. This is associated with elevated levels of pepsin and markers of inflammation. These lines of evidence suggest aspiration contributes to priming the allograft for PGD.

OBJECTIVES/GOALS: This project seeks to understand how personalized medicine can optimize care for patients with colorectal cancer. It identifies opportunities for personalized medicine to improve clinical outcomes, and uses cost-effectiveness analysis to assess the clinical and financial impact of this approach. METHODS/STUDY POPULATION: This project uses two methods to understand the impact of personalized medicine. First, this project has used SEER-Medicare data in conjunction with Clinical Pharmacogenetics Implementation Consortium guidelines to identify medications used by patients with colorectal cancer that can be impacted by genetic variants. This data will then be combined with population genetic variant rates to understand the likely impact screening for a given variant will have on medication response and adverse events. Medication use frequencies and genetic variant rates are then used to populate cost-effectiveness models that simulate the clinical and financial outcomes, identifying optimal genes to screen. RESULTS/ANTICIPATED RESULTS: The first result will be a comprehensive overview of treatment patterns for patients with colorectal cancer in the United States, as well as the treatments used for disease-induced comorbidities. The second result will be the identification of genetic variants based on population rates and medication utilization that should be screened in this patient population. The final result will be a breakdown of the clinical and financial outcomes associated with implementing screening for the identified genes. Preliminary results from a two-gene cost-effectiveness analysis demonstrates that screening for variants in those genes improves both clinical and financial outcomes. DISCUSSION/SIGNIFICANCE OF IMPACT: This project demonstrates how current treatment approaches can be optimized via personalized medicine. It uses epidemiological methods to identify opportunities to integrate genetic findings from other diseases, and uses cost-effectiveness analysis to understand the impact of transforming care. CONFLICT OF INTEREST DESCRIPTION: Stocks-Aurinia, Syndax, Adaptimmune, Rigel pharma

OBJECTIVES/GOALS: There is an imperative need to initiate translational genetic studies of hidradenitis suppurativa (HS). Such work requires large cohorts and no HS registries exist. Precision medicine initiatives provide new resources and methods for efficiently constructing cohorts, but empirically informed best practice guidelines are needed. METHODS/STUDY POPULATION: Traditional methods for building cohorts rely on clinical encounters to identify patients and collect phenotype data. Precision medicine initiatives aim to decrease the time and cost of data collection by using alternative sources, including electronic health records (EHR) and remote collection of patient-reported data. The public’s use of the Internet to obtain and exchange health-related information coupled with the success of direct-to-consumer genetic companies suggests that it is feasible to remotely ascertain research participants for genetic studies. Importantly, Internet cohorts provide an opportunity to include research participants who are disconnected from healthcare, and thus remain hidden from research that relies on EHR or clinical services. RESULTS/ANTICIPATED RESULTS: First, to conduct studies in EHR we are developing an analytic pipeline for the automated extraction of an accurate HS diagnosis using natural language processing of clinical notes. In our preliminary work we are also using ICD codes to build cohorts in two EHR systems with and without linked genetic data. Second, we have developed Internet advertising campaigns for symptom-based recruitment. Informed consent and patient-reported data is collected on-line through a series of short surveys. Patients who complete the surveys and express interest in participating in genetic studies are sent saliva collection kits and return mailing material. Finally, we have established an HS biobank that has DNA from 300 participants identified through clinical services. Enrollment is on going. DISCUSSION/SIGNIFICANCE OF IMPACT: Our goal is to assemble an HS cohort that is large enough to power genetic discoveries. Our work is generating empirical evidence for precision medicine guidelines and will improve our knowledge about HS. The methods we are developing can be applied to efficiently create new cohorts for genetic studies of other diseases across different clinical areas.

OBJECTIVES/GOALS: There is no objective, biologic tool to detect concussion or guide clinical management. We previously showed that saliva microRNA (miRNA) levels differ in children with concussion and may predict symptom duration. The purpose of this study was to validate the utility of saliva miRNA and define longitudinal trends during the recovery period. METHODS/STUDY POPULATION: We collected concussion symptom burden (SCAT-5), cognitive performance (DANA), balance measures (ClearEdge), and saliva from 150 children (7-21 years) with concussion over 5 time-points: 0-2, 3-6, 7-14, 15-29, and 30-60 days post-injury. Saliva miRNA levels within the 443 concussion samples were quantified with RNA sequencing and compared to 218 samples from age- and sex-matched controls (healthy and post-exercise participants). Non-parametric ANOVA assessed RNA levels across time-points, and between concussions/controls. Machine learning was used to build logistic regression algorithms differentiating concussions/controls, and symptomatic/recovered concussion participants. Relationships between miRNAs and concussion phenotypes were explored with Spearman’s Rank correlations. RESULTS/ANTICIPATED RESULTS: Fifteen miRNAs differed across control and concussion participants (FDR < 0.05). Within concussion participants, all 15 miRNAs trended back toward control levels by 30-60 days post injury. A regression algorithm employing 6 of the 15 miRNAs differentiated control and concussion participants with an area under the curve (AUC) of 0.78 in a training set (n = 244) and 0.84 in a naïve test set (n = 24). Similarly, 6 miRNAs were able to differentiate symptomatic (SCAT-5 symptom score > 7) and asymptomatic concussion participants with an AUC of 0.73 in a training set (n = 219) and 0.76 in a naïve test set (n = 44). Furthermore, 5 miRNAs showed significant (R > 0.3; FDR < 0.05) associations with subjective and/or objective measures of concussion-related symptoms. DISCUSSION/SIGNIFICANCE OF IMPACT: Saliva miRNAs levels are altered in children with concussion, and display predictable longitudinal trends following injury. Saliva miRNA measurement represents a non-invasive, objective tool that could be rapidly assessed to provide biologic evidence for clinicians managing pediatric concussion. CONFLICT OF INTEREST DESCRIPTION: I serve as a paid consultant and scientific advisory board member for Quadrant Biosciences, who has funded a portion of this work and licensed the findings from the Penn State College of Medicine.

OBJECTIVES/GOALS: It has been previously shown that pediatric high-grade glioma (pHGG) survival is different between sexes. We set out to find out whether there are sex-specific differences in the genomic landscapes of pHGG that may underlie this sex disparity. METHODS/STUDY POPULATION: We downloaded Illumina 450k DNAm data from ArrayExpress and GeneExpressionOmnibus. The minfi package was used to process raw DNAm data. Sex chromosomes and CpGs that are common SNPs were removed. Surrogate variables (SVs) were estimated via the sva Bioconductor package. Differentially methylated CpGs were identified by fitting a multiple linear regression model for the DNAm level at each CpG, with independent variables being sex (a binary variable) and the estimated SVs. RNAseq data was downloaded from Cavatica, and differential gene expression analysis was carried out via the DESeq2 package. RESULTS/ANTICIPATED RESULTS: In the pediatric glioblastoma (GBM) DNAm data [58 female & 91 male IDH wt samples; ages 0.1–21 yrs;], we found 7,371 differentially methylated cytosines (DMCs) at FDR≤0.05. Of the DMCs, 289 had DNAm differences between male and female samples ≥10%. The majority of probes (68%) were in CpG islands, shelves, or shores. We also found 4 differentially methylated regions (DMRs) between sexes (FWER≤0.1). In the adult GBM DNAm samples [32 F & 32 M IDH wt samples; ages 22–75 yrs], we found only 117 DMCs at FDR≤0.05, and no DMRs. In the RNAseq dataset [68 F & 54 M pHGG samples, ages 0.08–30.6 yrs], we found 383 differentially expressed genes (at FDR≤0.05), and 16 of them (4%) overlapped a DMC. DISCUSSION/SIGNIFICANCE OF IMPACT: Our findings demonstrate that pHGG exhibits sex-specific methylome differences. Interestingly, this difference is greater in the pediatric population as compared to adults. The pHGG transcriptome also differs by sex, which may be related to differential DNAm in a minority of cases.

OBJECTIVES/GOALS: We use a tissue engineered, biomimetic, 3D model to study the pathogenesis of breast implant-associated anaplastic large cell lymphoma (BIA-ALCL) by comparing the effect of silicone implant shell on proliferation of patient-derived BIA-ALCL to its precursor T cells within the breast microenvironment. METHODS/STUDY POPULATION: Patient-derived breast tissue was processed for component adipocytes, ductal organoids, and stromal vascular fraction. These were suspended within 50 µl of 0.3% type I collagen matrix to which was added 200,000 cells/mL of either patient-derived BIA-ALCL cells or T progenitor cells. These were then plated into 6mm wells. As a control, both BIA-ALCL cells and T progenitor cells were suspended within type I collagen alone at the same seeding density without breast components. Before plating, wells were lined circumferentially with either textured, smooth, or no implant shell. These were 1cm by 2cm pieces dissected from the whole implant. Wells were imaged using confocal microscopy over 8 days. RESULTS/ANTICIPATED RESULTS: Unstimulated T progenitor cell count showed no significant increase in any of the conditions tested. The change in cell count over 8 days was 3.85% in each condition (p = 0.3352). A Tukey’s multiple comparison test comparing each condition revealed no significant increase in cell count over 8 days for all six conditions. Notably, our previous studies have shown proliferation of BIA-ALCL cells to be significantly more robust in the biomimetic platform compared to collagen-only groups, regardless of implant shell type (p < 0.01). BIA-ALCL cells grew nearly 30% faster in textured and smooth shell biomimetic groups compared to biomimetic wells lacking implant shell. DISCUSSION/SIGNIFICANCE OF IMPACT: Towards elucidating BIA-ALCL’s etiopathology, we show that silicone implant shell has a significant effect on proliferation of BIA-ALCL cells, but not their precursor T cells. If breast implant silicone shell is not a sufficient stimulus for T cell proliferation, co-stimulatory factors are required.

OBJECTIVES/GOALS: Traumatic interpersonal experiences are associated with higher rates of chronic pain, increased pain severity and poorer functioning. The objective of this ongoing project is to obtain prevalence rates for various forms of interpersonal trauma among individuals with chronic pain, and to explore the potential mediating effect of heightened sensory and social sensitivity on the experience of pain. METHODS/STUDY POPULATION: Patients at Michigan Medicine between the ages of 18 and 65 complete an online survey. Patients are being recruited through a tertiary-care, outpatient pain clinic, as well as through an online health research portal. We aim to recruit 700 participants; we currently have 59.6% of our goal (n = 417). Participants also have the option to be included in a registry from which we can recruit for future studies. Approximately 85% of our participants have agreed to be in the registry. RESULTS/ANTICIPATED RESULTS: Preliminary data show that, of the 263 (63.4%) participants for whom data on chronic pain is available, 167 (63.5%) report chronic or persistent pain over the previous 3 months. Of these, 54% reported some form of childhood abuse or neglect. Approximately 41% reported four or more adverse childhood experiences. Additionally, of the 122 participants (73%) who were in a current romantic relationship, 20% reported some form of physical violence victimization from their romantic partner. We anticipate that interpersonal trauma will be associated with poorer perceptions of social relationships, higher sensory sensitivity, and higher perceived stress. DISCUSSION/SIGNIFICANCE OF IMPACT: The PASE Project parent study will be used to better understand prevalence rates for various forms of interpersonal trauma in our chronic pain population. Future analyses and studies will explore alternative pathways linking interpersonal trauma to the experience of pain through sensory and social sensitivity, which will inform interventions aimed at reducing pain among patients with a history of trauma.

OBJECTIVES/GOALS: Cell-cell (CC) and cell-matrix interactions (CM) are known to affect drug sensitivity of cancer cells, but are not effectively recapitulated using 2D platforms. This research aims to determine how cell and matrix interactions confer drug resistivity in 3 distinct culturing models: 2D (no CM/limited CC), 3D spheroids (CC) and 3D fibronectin (both). METHODS/STUDY POPULATION: We examined four breast cancer cell types. The cells were derived from a nonmetastatic primary tumor (HMLE-E2) or overt bone-metastasis (BM). Transglutaminase 2 (TGM2), a matrix crosslinking protein, is overexpressed in metastatic bone tumors and may play a key role in matrix-conferred drug resistivity. In a gain-of-function model, TGM2 was upregulated in HMLE-E2 cells and compared to shTGM2 knockdown BM cells. Growth rates were analyzed using metabolic activity over 8 days, and drug sensitivity to Neratinib (0-1000 nM) was analyzed via cell titer. To account for the different transport properties of the 3 distinct culture environments, we developed a mathematical model for each condition, allowing us to normalize the drug sensitivity results across models to effectively compare true biological resistivity. RESULTS/ANTICIPATED RESULTS: We observed that increased cellular levels of TGM2 significantly increase the growth rate and drug resistivity of cells on fibronectin matrices. Interestingly, in 2D cultures, TGM2 expression was correlated with higher Neratinib resistivity but did not affect growth rates. In spheroid models without a significant matrix component, that rely solely on cell-cell junctions, high levels of TGM2 were correlated with lower survival rates. Lower levels of TGM2 are correlated with a more epithelial phenotype, and using our mathematical model we have identified significant transport differences between high and low TGM2 spheroids. We theorize that the low TGM2 spheroids have denser packing, which lowers the rate of diffusion and, thus reduces the effective concentration of the drug to the majority of the cells. DISCUSSION/SIGNIFICANCE OF IMPACT: Our studies indicate that the cellular response to drugs can be altered by changes in both transport properties of the tissue and the CM interactions. By systematically investigating the effects of CC interactions and CM interactions, we can use mathematical models to delineate physical means of drug resistivity from a biologically driven resistance.

OBJECTIVES/GOALS: Real-world studies have been gaining momentum in providing evidence of treatment effectiveness and hold great potential for facilitating the drug regulatory process. The U.S. Food and Drug Administration (FDA) has recognized this by providing a framework for using real-world data (RWD) to generate real-world evidence (RWE). The objective of this study is to assess the current level of RWE implementation in clinical studies. METHODS/STUDY POPULATION: Using keywords relevant to RWE, we reviewed studies on drugs, biologics, and medical devices published on PubMed in 2018. Information regarding the therapeutic area of focus, intervention type, study design, primary outcome, and data source was recorded. Further analyses of the three main therapeutic areas of study (oncology, cardiology, and infectious diseases) were performed to determine how RWE was being utilized. In addition, a broad “real-world” search was performed on Clinicaltrials.gov, from which we extracted relevant observational and Phase I, II, II/III, III, III/IV, and IV studies. A supplemental PubMed search was used to evaluate published studies in order to identify which field these trials were concentrated in and the outcome of interest.” RESULTS/ANTICIPATED RESULTS: After application of “real-world” search terms to PubMed, 995 hits were generated and of these, 311 studies were excluded. More than half of the studies were observational and retrospective in nature (64%) with 70% examining drug/biologic outcomes. RWE data sources were largely dominated by medical records and claims data. The primary uses of RWE across oncology, cardiology, and infectious diseases included supporting drug product effectiveness, assessing safety, and evaluating treatment patterns. Of the 207 RWE studies identified on ClinicalTrials.gov, 66 were cancer randomized controlled trials (RCTs), a majority of which were used for post-marketing safety evaluations. Further research will be conducted to determine the precise role of RWE in all studies (e.g. historical comparator, label expansion). DISCUSSION/SIGNIFICANCE OF IMPACT: By examining the use of RWE in regulatory decision making, we can inform stakeholders of the extent to which robust RWE studies complement evidence generated by RCTs. Thought to reflect a product’s performance in a broader and more diverse population, RWE can provide greater insight to clinical trial conduct and ultimately transform patient outcomes.

OBJECTIVES/GOALS: The goal was to understand the effectiveness of a novel clinical trial educational module and a corresponding initiative designed and disseminated by the Southern California Clinical and Translational Science Institute (SC-CTSI) to increase the quality of clinical trials conducted in academia. METHODS/STUDY POPULATION: The CRCs (Clinical Research Coordinators) for the initiative are asked to complete the online training. Possible study protocols are picked to be monitored by the CRCs. The monitor is instructed to study the protocol extensively and prepare for their monitoring visit. The trained monitor from the initiative then reaches out to the CRC of the study that is to be monitored and carries out the monitoring visit. Afterwards, the monitor sends initiative personnel the monitoring report, which is evaluated to see if the monitor checked everything they should have during the visit. The PI of the study is contacted with highlights from the monitoring report and improvements that they can make. RESULTS/ANTICIPATED RESULTS: The first study monitored was a site of a large NIH-sponsored study where the consent forms were signed electronically. It was found that the monitor could not access the consent forms. Therefore, the monitor could not do source data verification. The PI of the study said that they would be raising this issue with the NIH. During the monitoring visit of the second study chosen for the initiative, patient binders were specifically examined for informed consent and source documentation completeness. The charts of patients were also reviewed. The only deviation found was a missing signature in the Investigator Site File. For the last two studies, data will be reported. DISCUSSION/SIGNIFICANCE OF IMPACT: Monitors were not only able to monitor efficiently, but also able to point out deficiencies in the monitoring practices of large studies. This model could be expanded to other academic institutions to establish quality management systems to ensure data integrity and subject protection.