ABSTRACT IMPACT: Lipidomics is emerging as a powerful strategy to identify biomarkers for Major Depressive Disorder, as well as therapeutic targets in lipid metabolic pathways. OBJECTIVES/GOALS: Lipidomics is increasingly recognized in precision psychiatry for global lipid perturbations in patients suffering from Major Depressive Disorder (MDD). We will test the hypothesis that lipid metabolism dysregulation is associated with familial risk of depression. METHODS/STUDY POPULATION: Patients with MDD (G1), children (G2), and grandchildren (G3) have been part of a longitudinal study since 1982. If a parent G2 and grandparent G1 have MDD, G3 is considered a high risk of depression. Biospecimens (saliva and serum) were collected for full exome sequencing and RNA analysis. Samples will also be extracted for lipid content and lipids will be identified by mass spectrometry. A panel of nearly 600 lipid species can reliably be identified and quantified using liquid chromatography paired with tandem mass spectrometry (LC-MS/MS). Dysregulated lipids will be correlated with familial risk of depression in samples of G3. RESULTS/ANTICIPATED RESULTS: We hypothesize that dysregulation of lipids and lipid metabolism will be apparent in biospecimens from the high risk compared to the low risk of depression. Also, alterations in RNA transcriptomics of genes involved in lipid metabolic networks are associated with familial risk of depression. Several differential lipid species were previously identified to be associated with MDD. Reduced phosphatidylcholine(PC), phosphatidylethanolamine(PE), phosphatidylinositol(PI), and increased LysoPC, LysoPE, ceramide, triacylglycerol, and diacylglycerol levels have been correlated to MDD. However, these results need to be replicated in independent studies using lipidomics analysis. DISCUSSION/SIGNIFICANCE OF FINDINGS: It is highly likely that completely novel cellular targets will emerge from these studies by uncovering the convergence of lipidomics and genetic variance of lipid metabolic enzymes as biomarkers for predisposition to MDD as well as potential targets for therapeutic development for MDD.

Over the last year, COVID-19 has emerged as a highly transmissible and lethal infection. As we address this global pandemic, its disproportionate impact on Black, Indigenous, and Latinx communities has served to further magnify the health inequities in access and treatment that persist in our communities. These sobering realities should serve as the impetus for reexamination of the root causes of inequities in our health system. An increased commitment to strategic partnerships between academic and nonacademic health systems, industry, local communities, and policy-makers may serve as the foundation. Here, we examine the impact of the recent COVID-19 pandemic on health care inequities and propose a strategic roadmap for integration of clinical and translational research into our understanding of health inequities.

Revisions to the Common Rule and NIH policy require the use of a single Institutional Review Board (sIRB) for the review of most federally funded, multisite research, with the intent of streamlining the review process. However, since initial implementation in 2018, many IRBs and institutions continue to struggle with the logistics of implementing this requirement. In this paper, we report the findings of a workshop held in 2022 to examine why sIRB review remains problematic and propose possible solutions. Workshop participants identified several issues as major barriers, including new responsibilities for study teams, persistent duplicative review processes, the lack of harmonization of policies and practices across institutions, the absence of additional guidance from federal agencies, and the need for greater flexibility in policy requirements. Addressing these problems will require providing additional resources and training to research teams, the commitment of institutional leaders to harmonize practice, and policymakers to critically evaluate the requirement and provide flexibility in applicability.

OBJECTIVES/GOALS: Treatment options for glioblastoma (GBM) are limited. Prognosis remains dismal, with an 18 month on average survival rate following diagnosis due to treatment resistance and disease recurrence. The goal of this project is to investigate hallmarks of cancer progression that contribute to temozolomide (TMZ) resistance, a first tine treatment for GBM. METHODS/STUDY POPULATION: Two signaling pathways were investigated in TMZ-sensitive and -resistant GBM cell lines and in primary and recurrent patient-derived xenograft (PDX) tumor cells by genetically and pharmacologically inhibiting methionine adenosyltransferase 2A (MAT2A) and adenosylhomocysteinase (AHCY). Cell growth and survival were assessed by measuring protein expression of proliferation, oxidative stress and cell cycle arrest markers. EPIC array analysis and targeted bisulfite sequencing were conducted to identify changes in genome-wide and specific CpG island methylation. The Seahorse XF Analyzer measured mitochondrial respiratory capacity and oxidative metabolism. Induced pluripotent stem cell organoids were co-cultured with PDX tumor cells to determine if treatments mitigate tumor cell invasiveness. RESULTS/ANTICIPATED RESULTS: Compared to parental cells (PC), MAT2A gene expression was increased by 1.7-fold in acquired resistant and de novo resistant GBM cells (RC) [(transcript per million): PC, 7386 ± 0.012; RC, 12925 ± 0.023; n=2; p=2.10e-8]. Compared to TMZ-sensitive cells (TS), TMZ-resistant cells (TR) demonstrated a 56% increase in baseline oxygen consumption rate [(pmol/min): TS, 179 ± 6.7; TR, 279 ± 13; n=18; p=.0012] and 64% increase in maximal respiratory capacity [(pmol/min): TS, 403 ± 29; TR, 659 ± 35; n=6; p DISCUSSION/SIGNIFICANCE: MAT2A and AHCY contribute to TMZ resistance and recurrence by dysregulating methylation programs and upregulating antioxidant programs, respectively. These findings provide a foundation for developing novel combinatory therapeutic strategies and inform clinical studies intended to increase remission and reduce recurrence for GBM patients.

OBJECTIVES/GOALS: Ask the Experts: A Biomedical Innovation Forum, presented by Fast Forward Medical Innovation (FFMI) at the University of Michigan, provided an opportunity to educate biomedical innovators on life science investment trends and technology assessment criteria. METHODS/STUDY POPULATION: FFMI, in partnership with the U.S. Economic Development Administration, recruited an expert group of panelists to be featured at this virtual event. These life science investment experts provided insight on the strategy, timing, and best method for innovators to engaged investors, the specifics of what investors look for in technologies and project teams, and expectations of investors and project teams after the investment is secured. The panel presentation was followed by a poster presentation highlighting projects from the FFMI Hub at the University of Michigan, allowing innovators to have an open and constructive conversation with experts and attendees. RESULTS/ANTICIPATED RESULTS: There was a total of 73 registrants including academic faculty, biomedical innovators, and life science investment professionals from 21 different academic institutions, private companies, and other organizations. 50 attended the panel presentation and poster session. Results (N=5) of an evaluation of the event revealed that 100% of the respondents strongly agreed or agreed that the event met their expectations, while 80% strongly agreed or agreed that they would recommend the event to a colleague. Feedback from poster presenters was also strong with presenters exclaiming they “enjoyed the panel discussion and getting one-on-one time with the panelists,” as well as “a lot of great advice was given by the experts” and “I really liked the poster presentation part in which I got feedback from the investors.” DISCUSSION/SIGNIFICANCE: The data demonstrates how accelerating technology mining activities, proactively seeking and strengthening external partnerships with investors, and scaling commercialization education programs can have a positive impact on the development and launch of biomedical innovations.

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/SPECIFIC AIMS: Qualitative approaches help explore poorly understood phenomenon, and are highly engaging, enabling both sides of an encounter greater connection. Historically, Deaf communities have been marginalized and oppressed, with their linguistic needs unrecognized and ignored. As a result, Deaf participants are rarely involved in clinical research. Like other marginalized communities, the Deaf community experiences health disparity compared with others, especially in low- and middle-income settings. The purpose of this project was to assess the feasibility of conducting qualitative research with Deaf Dominicans. METHODS/STUDY POPULATION: We implemented a partnered research process with 59 Deaf community members in the Dominican Republic, conducting preliminary thematic analysis through reviews of interviews and on-site debriefings. RESULTS/ANTICIPATED RESULTS: Participants were highly engaged with the Deaf-Deaf research encounters, indicating satisfaction with both the process and with the opportunity to communicate their needs and interests. Preliminary findings indicated Deaf Dominicans were highly engaged, confirming their interest, and often stated that they felt they were being listened for the first time. Indeed, some participants claimed that this was the first time they communicated their experiences as Deaf Dominicans and appreciated the opportunity to relate this experience to Deaf interviewers. DISCUSSION/SIGNIFICANCE OF IMPACT: This experience confirms that the Deaf Dominican community can be mobilized and will participate in Deaf-Deaf research.

OBJECTIVES/GOALS: The Virtual CTSA Visiting Scholars Program allows KL2 scholars to serve as visiting professors at host CTSA consortium institutions. This program facilitates connections with faculty outside scholars home institutions, as well as fostering collaboration among different CTSA hubs. Stanford aimed to incorporate participants into its KL2 program. METHODS/STUDY POPULATION: Stanford is one of 57 host institutions for the Virtual CTSA Visiting Scholars Program. The program includes a virtual meeting between each participant and a chosen faculty at the host institution. Program participants also give a virtual Grand Rounds lecture, open to the entire CTSA consortium. As well as encouraging Spectrum KL2 Scholars to participate in the virtual exchange, the Spectrum KL2 program incorporated the visiting scholars into weekly activities over the course of 3 months. Visiting scholars were invited to participate virtually in KL2 educational sessions, which provide career development training and mentoring. As meetings transitioned to in-person, KL2 scholars were allowed to attend virtually when needed. Hybrid in-person sessions were also conducive to participation by visiting scholars. RESULTS/ANTICIPATED RESULTS: Stanfords Spectrum KL2 program was virtual (Zoom) for academic year 2020-2021 and included 1:1 mentoring sessions, weekly career development seminars, and 1:1 peer virtual lunches to integrate the 9 junior faculty scholars. Visiting scholars joined the weekly Zoom meetings when their schedules allowed. KL2 faculty mentored visiting scholars for 3 months, exchanging ideas and forming collaborations. Each visiting participant was paired with a KL2 Scholar, who provided 1:1 peer mentorship. In addition, visiting scholars presented a work-in-progress seminar, to obtain feedback before the more formal Grand Rounds lecture. For academic year 2021-2022, Spectrum KL2 meetings include virtual and hybrid in-person sessions, allowing visiting scholars to join by Zoom during the 3-month virtual exchange program. DISCUSSION/SIGNIFICANCE: What set Stanfords Virtual CTSA Visiting Scholars Program apart was faculty engagement and mentorship provided to visiting participants. By incorporating visiting scholars virtually into the ongoing KL2 education program, participants could engage fully with mentors and scholars, even surpassing opportunities of pre-pandemic on-site visits.

OBJECTIVES/GOALS: Sleep is critical for healing, however pediatric intensive care unit (PICU) sound is above recommended levels (i.e., 45 A-weighted decibels [dBA]). This observational study identifies sources of PICU sound and compares sources between times of high (i.e., dBA≥45) and low (i.e., dBA < 45) levels. METHODS/STUDY POPULATION: The sound environment of 10 critically ill children 1 to 4 years of age was monitored via a bedside dosimeter and video camera for 48 hours, or until PICU discharge. Dosimeter and video data were uploaded to Noldus Observer XT and time synchronized. A reliable, previously published coding scheme developed to identify sound sources in the adult ICU was modified for the pediatric population. Sound sources (e.g., clinician/family/child [verbal vs. non-verbal] vocalization, patient care, medical equipment) were identified via instantaneous sampling of video data at each minute of recording. The proportion of sampling points with each sound source are compared between times of high and low sound levels, and between day (7:00-18:59) and night (19:00-6:59) shift. RESULTS/ANTICIPATED RESULTS: Video coding is ongoing, with high inter-rater reliability (κ̅=0.99, SD DISCUSSION/SIGNIFICANCE: Medical equipment sound is ubiquitous in the PICU. Clinicians should optimize the PICU sound environment for sleep, including minimizing equipment alarms, conversation, general activity, and screen media during child rest. Large-scale studies are needed to confirm findings from this small cohort.

OBJECTIVES/SPECIFIC AIMS: To develop a social network model of collaborations within and external to the University of Rochester Medical Center (URMC) CTSI using data from the annual Research Performance Progress Report (RPPR) as well as other sources, to provide longitudinal evaluation of the CTSI’s engagement with key stakeholder groups. METHODS/STUDY POPULATION: The annually submitted RPPR follows a specific format with well-defined sections. The Highlights, Milestones and Challenges Report includes areas in which CTSI function leaders provide details about program integration and innovation, including collaborations with other functions or external groups. The Highlights, Milestones and Challenges Report was qualitatively coded to identify function-collaborator dyads. Each entity in the dyad became a node in the network. Nodes were connected by edges named by the dyads. The network included two types of nodes. The first were CTSI internal functions/programs, i.e. the entities that submitted RPPR sections and formed an interconnected sub-network. The second type of nodes were entities external to the CTSI (collaborators, internal or external to the CTSI site). These entities were named by functions submitting RPPR narratives. External nodes with similar meanings were consolidated. Duplicate edges were removed. CTSI-external nodes were grouped into five stakeholder categories: URMC, University of Rochester (UR), community, other CTSA institutions, CTSA consortium. Thus, these nodes were connected to the CTSI internal nodes, but not to each other. A second source of collaboration data was function-reported internal metrics. As part of the internal metric data collection, functions list partners who play a role in improving metric data or who are responsible for providing data. Partners identified in the internal metrics data, but not specified in the RPPR, were added to the network. RESULTS/ANTICIPATED RESULTS: Twenty-three internal CTSI functions submitted an RPPR and represent the CTSI internal nodes. Internal CTSI functions identified 235 collaborations (edges): 125 collaborations with other CTSI internal functions, 57 collaborations with URMC entities, 14 with UR entities, 15 with the external community, 15 with other institutions (CTSA hubs and other universities), and 9 with CTSA consortium entities. Thirty-eight of the collaborations were identified in the internal metrics partners section. In total, the network comprised 104 nodes. Graph density was.022 for full network and.21 for the CTSI internal sub-network. The global clustering coefficient, a measure of connectivity, for the CTSI internal sub-network was.252. DISCUSSION/SIGNIFICANCE OF IMPACT: The RPPR provides an underutilized source of data for annually repeated analyses of internal and external CTSI collaborations and is a way to enhance use of this routinely collected information. Analyses of the network yield metrics for measuring CTSI reach and impact on stakeholder groups over time. For example, measures such as number of nodes representing entities external to CTSI and average vertex degree of the CTSI Internal nodes track aspects of CTSI collaborations. Visualizations using different layouts or highlighting different sub-networks provide a representation of CTSI engagement with the communities of stakeholders as well as insights to relationships between functions, regions of collaboration, and areas of gaps. These data also provide an important new mechanism to engage the CTSI leadership and function leads in understanding how their work contributes to the overall network and synergies they have with each other.

ABSTRACT IMPACT: This work will develop a novel drug delivery system that has improved biocompatibility and controlled release than current systems and allow for customizable loading and drug delivery to unique patient and treatment requirements. OBJECTIVES/GOALS: The goal of my project is a novel hybrid core/shell nanoparticle system for controlling in vivo chemotherapeutic concentration. The current goal is to confirm core and shell polymeric nanoparticle formation via emulsion technique and validate predictive model developed to optimize shell formation efficiency and control shell thickness. METHODS/STUDY POPULATION: Though early results are promising, they are not proof that the desired core/shell structure is being formed via my novel process. I constructed a theoretical model to use to optimize and control the process for precise shell thicknesses. Therefore, the current experimental plan focus is to not only visually confirm the predicted formation of my core/shell design but use these experiments to validate the model.

1. 1. Gel-Suspended SEM: nanoparticles suspended in gel matrix, bisected to reveal inner structure

2. 2. Fluorescent Conjugation Microscopy: visually-distinct dyes used to show polymer distribution and validated against the theoretical model predictions.

3. 3. Modified Hydrophobic Dye Release: different mixtures of polymers with release showing if previous promising results due to core/shell structure RESULTS/ANTICIPATED RESULTS: As stated, the experiments will confirm the core/shell nanoparticle structure, validate the developed theoretical model, or provide direct evidence against any formation. This core/shell structure is key to the current design for controlling payload release rate and thus in vivo drug concentration. For the gel-suspension experiment, the interior core will be labeled with ultrasmall SPIONs and thus any layers within the particles will be distinct. While this result is qualitative, high magnification fluorescent microscope images will be analyzed using image processing software to determine core/shell formation efficiency and compared to estimated efficiencies from the model. Finally, the mixed release will clarify previous experiments’ release mechanism and either support or disprove shell influence. DISCUSSION/SIGNIFICANCE OF FINDINGS: The significance of this work is twofold: core/shell particles have been proven to provide variable control of release on the micron scale but not yet at the nanoscale, allowing for a circulating, targeted system that can finely control release. The process is also novel for producing this type of structure, at highly consistent quality and size.

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 streamline the standards and procedures for operating a research-specific, clinical data warehouse, acheived by defining roles, introducing a common language, and categorizing dataset types to provide transparency regarding data security risks inherent in the use of patient data. METHODS/STUDY POPULATION: We established a Bioethics committee responsible for ensuring clinical data is securely procured, maintained, and extracted in a manner that adheres to all federal, state, and local laws. We created an operational framework in the form of an umbrella IRB protocol and shared it with the bioethics committee for feedback and approval. The protocol was approved first by the bioethics committee and subsequently by the IRB. It was then disseminated across the institution and published online for continuous reference and use by committee members, researchers, and the data warehouse service team. RESULTS/ANTICIPATED RESULTS: The resulting framework defined the roles of researchers, data warehouse service team members, and honest brokers; explains the procedures for accessing and securely delivering data; and lists six categories of datasets according to type and implicit risks: datasets that are preparatory for research/aggregate counts, anonymized datasets, coded datasets, limited datasets, identified datasets for recruitment purposes, and defined identified cohort datasets. The protocol is approved and in use enterprise-wide, has reduced the number of questions from stakeholders, and has given researchers, IRB members, and informatics staff confidence in the use of the clinical research data warehouse. DISCUSSION/SIGNIFICANCE: We offer our framework to CTSAs interested in streamlining their data warehouse operations. We believe the adoption of this framework will establish strong procedures for ensuring compliance with IRB requirements, data privacy, and data security while reducing barriers to clinical research.

ABSTRACT IMPACT: This work will help to understand a novel therapeutic approach to a common type of acute myeloid leukemia. OBJECTIVES/GOALS: FMS-like tyrosine kinase 3 (or FLT3) mutations occur in ˜30% of acute myeloid leukemia (AML) cases. FLT3 tyrosine kinase domain (TKD) mutations are particularly important in relapsed/refractory FLT3 mutant AML, which portends poor prognosis. This study describes a therapeutic approach to overcoming resistance conferred by FLT3-TKD mutations. METHODS/STUDY POPULATION: To understand the efficacy of a novel type 1 FLT3 inhibitor (NCGC1481), as a monotherapy and combination therapy, several assays were utilized to interrogate functionality of these therapies. Cell lines and patient samples containing aspartate 835 to tyrosine mutations (D835Y, the most common TKD alteration) and phenylalanine 691 to leucine (F691L) were utilized to examine the effects of NCGC1481 with and without other targeted therapies like MEK inhibitors. Specifically, assays measuring viability, cell death using flow cytometry, in vitro clonogenicity, cellular signaling, and xenograft survival were examined in these FLT3-TKD AML models. Synergy was also measured using well described methods, which also allowed for appropriate dose finding for drug combination experiments. RESULTS/ANTICIPATED RESULTS: Our novel type 1 FLT3 inhibitor (NCGC1481) was particularly effective in the most common FLT3 TKD mutant, D835Y. NCGC1481 reduced viability and cell signaling, while also inducing cell death and prolonging xenograft survival in the FLT3-D835Y model system. In contrast, clinically approved FLT3 inhibitors were less effective at suppressing AML cells expressing FLT3-D835Y. In the case of FLT3-F691L, most of the FLT3 inhibitors tested, including NCGC1481, suppressed canonical FLT3 signaling, but did not significantly reduce viability or leukemic clonogenicity. However, when NCGC1481 was combined with other targeted agents like MEK inhibitors, at synergistic doses, eradication of the FLT3-F691L AML clone was substantially increased. DISCUSSION/SIGNIFICANCE OF FINDINGS: In AML, response to FLT3 inhibitor therapy is often short-lived, with resistance sometimes occurring via FLT3-TKD mutations. Given the dismal prognosis of relapsed FLT3 mutant AML, novel therapies are necessary. This study describes efficacy of a novel FLT3 inhibitor, along with its synergistic activity when combined with other targeted agents.

OBJECTIVES/GOALS: Alzheimer’s Disease (AD) displays numerous pathological features, including amyloid-beta deposition, extensive neuroinflammation, and vascular fibrosis. However, putative therapeutic options for alleviating these features remain limited, emphasizing the need to develop comprehensive treatments for patients with AD. METHODS/STUDY POPULATION: CSF from human AD patients treated with nilotinib (n=12), a tyrosine kinase inhibitor, or placebo (n=11) was collected and sequenced, and significantly altered miRNAs were identified and analyzed for alterations to disease-associated genes via gene ontology analysis. TgAPP mice were injected intraperitoneally with one of two novel tyrosine kinase inhibitors, BK40143 or BK40197, or DMSO (n=12 per group) daily for six weeks, during which memory deficits between groups were measured, before brains were harvested for analysis of amyloid-beta load via ELISA, microglial activation via Sholl analysis, and vascular collagen levels via immunohistochemistry. RESULTS/ANTICIPATED RESULTS: CSF obtained from AD subjects treated with nilotinib revealed significantly increased (p<0.05) levels of miRNAs regulating autophagy, neuroinflammation, and collagen production compared to placebo. These results were validated in vivoin TgAPP mice, who displayed improved recall on the novel object recognition test and Morris water maze following treatment with our drugs, correlating with decreased levels of brain amyloid-beta (30% decrease, p=0.002), decreased microglial reactivity and activation (40% decrease, p=0.01), and decreased vascular fibrosis (50% decrease, p=0.005) along small brain blood vessels compared to controls. DISCUSSION/SIGNIFICANCE: These data identify tyrosine kinase inhibition as a valid therapeutic strategy for alleviating various pathological features associated with AD and warrant further investigation as a treatment option for human patients as a means of slowing cognitive decline.

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: Supported by the State of Alabama, the Alabama Genomic Health Initiative (AGHI) is aimed at preventing and treating common conditions with a genetic basis. This joint UAB Medicine-HudsonAlpha Institute for Biotechnology effort provides genomic testing, interpretation, and counseling free of charge to residents in each of Alabama’s 67 counties. METHODS/STUDY POPULATION: Launched in 2017, as a state-wide population cohort, AGHI (1.0) enrolled 6,331 Alabamians and returned individual risk of disease(s) related to the ACMG SF v2.0 medically actionable genes. In 2021, the cohort was expanded to include a primary care cohort. AGHI (2.0) has enrolled 750 primary care patients, returning individual risk of disease(s) related to the ACMG SF v3.1 gene list and pre-emptive pharmacogenetics (PGx) to guide medication therapy. Genotyping is done on the Illumina Global Diversity Array with Sanger sequencing to confirm likely pathogenic / pathogenic variants in medically actionable genes and CYP2D6 copy number variants using Taqman assays, resulting in a CLIA-grade report. Disease risk results are returned by genetic counselors and Pharmacogenetics results are returned by Pharmacists. RESULTS/ANTICIPATED RESULTS: We have engaged a statewide community (>7000 participants), returning 94 disease risk genetic reports and 500 PGx reports. Disease risk reports include increased predisposition to cancers (n=38), cardiac diseases (n=33), metabolic (n=12), other (n=11). 100% of participants harbor an actionable PGx variant, 70% are on medication with PGx guidance, 48% harbor PGx variants and are taking medications affected. In 10% of participants, pharmacists sent an active alert to the provider to consider/ recommend alternative medication. Most commonly impacted medications included antidepressants, NSAIDS, proton-pump inhibitors and tramadol. To enable the EMR integration of genomic information, we have developed an automated transfer of reports into the EMR with Genetics Reports and PGx reports viewable in Cerner. DISCUSSION/SIGNIFICANCE: We share our experience on pre-emptive implementation of genetic risk and pharmacogenetic actionability at a population and clinic level. Both patients and providers are actively engaged, providing feedback to refine the return of results. Real time alerts with guidance at the time of prescription are needed to ensure future actionability and value.

OBJECTIVES/GOALS: Research Education Towards Opportunities (RETO) Mentoring Offering Training Opportunities for Research (MOTOR) 1 – 2 and the elective courses (INTD 5998/ MDCL 101) in CTR of Title V Coop were designed to provide the participants from higher education institution (HEI) in Puerto Rico (PR), interdisciplinary – interprofesional knowledge in CTR. METHODS/STUDY POPULATION: Since April 2017, Research Education Towards Opportunities (RETO) and Mentoring Offering Training Opportunities for Research (MOTOR) 1 - 2 trainings were offered as part of the Title V Coop. In addition, since January 2020, as part of the institutionalization of the trainings in CTR, two elective courses (INTD 5998 and MDCL 101) were created-offered. The trainings/courses present the main concepts underlying CTR performance through lectures, workshops and presentations, in hybrid modalities, as well as the services-resources of the Center Research Education and Science Commnunication Opportunities (CRESCO). These programs have given students (undergraduate and graduate) and faculty the opportunity to get started in CTR and to integrate in Clinical and Translational Mentoring Teams (CTMT). RESULTS/ANTICIPATED RESULTS: Eight (8) cycles of RETO-MOTOR 1 and seven (7) cycles of RETO-MOTOR 2; two sessions of INTD 5998 and one session of MDCL 101 were offered. The RETO-MOTOR 1 training was completed by 219 participants and RETO-MOTOR 2 by 130 participants. The INTD 5998 course was completed by 22 students and the MDCL 101 course by 18 students. A total of 389 participants have been initiated in the CTR. Of the trainings, 90% indicated that the knowledge acquired in CTR was invaluable, 85% understand that the most significant achievement, as students, was present at a scientific conference, and 100% indicated interest in continuing to do CTR. Of the courses, 100% indicated that they were a good learning experience, helped them increase their knowledge in CTR, met their expectations and would recommend other students to take the course. DISCUSSION/SIGNIFICANCE: The RETO-MOTOR 1, RETO-MOTOR 2 trainings and CTR courses provide a based of research knowledge and valuable interprofessional experience for those who whish to start in the clinical and translational research. The Title V Cooperative Project provides this opportunity to undergraduate and graduate students such as faculty of HEI in PR.