Objectives/Goals: The Never-in-mitosis A-related (NEK) kinases are a family of 11 poorly explored kinases that play varying roles in cellular processes. Aberrant NEK activity is implicated in various human diseases, including cancer. We hypothesize that NEK9 drives triple-negative breast cancer (TNBC) motility. Methods/Study Population: Baseline NEK9 expression was determined across five breast cancer and three non-breast cancer cell lines by immunoblot. We expanded the analysis to six additional TNBC cell lines. To determine the impact of NEK9 on TNBC biology, we evaluated the effects of NEK9 pharmacological inhibition with novel inhibitor BA 03-53-11 (BA) at 0µM, 0.5µM, 1µM, and 2µM. We treated TNBC cell lines MDA-MB-231, BT-549, HS578T, and TNBC patient-derived cell line TUBcX-4IC and performed functional assays to assess impacts on cell motility (scratch assay), viability (crystal violet staining), and proliferation (Ki-67 staining). Results/Anticipated Results: NEK9 levels were elevated in the TNBC cell line compared to other breast cancer subtypes, osteosarcoma, and non-cancerous cells. Additionally, NEK9 protein expression was variable across TNBC cell lines. While pharmacological NEK9 inhibition did not significantly alter TNBC cell viability or proliferation, we observed that NEK9 inhibition suppressed cell motility in TNBC cells compared to vehicle control. Taken together, these preliminary results support a role for NEK9 in regulating TNBC progression. Our ongoing studies aim to test NEK9-mediated motility and proliferation in additional TNBC cell lines and evaluate NEK9 as a therapeutic target for this aggressive breast cancer subtype. Discussion/Significance of Impact: TNBC is an aggressive form of breast cancer that lacks targeted therapies. Because aberrant kinase activity underpins oncogenic transformation, kinase inhibitors have emerged as ideal therapeutic agents. Elucidation of NEK9 function in TNBC is essential for the development of novel therapeutics.

Objectives/Goals: To examine gut microbiome–immune interactions that may contribute to respiratory dysfunction in Puerto Rican patients with primary ciliary dyskinesia (PCD), aiming to identify novel biomarkers that inform targeted interventions and address health disparities in rare diseases. Methods/Study Population: This cross-disciplinary pilot study integrates clinical, genomic, and microbiome data from patients with PCD carrying the Puerto Rican RSPH4A founder variant (c.921+3_6delAAGT; n = 8) and healthy controls (n = 8). Fecal samples will undergo 16S rRNA V4 sequencing and bioinformatic profiling using QIIME2, ANCOM, and PICRUSt2. Plasma cytokines (IL-6, IL-8, TNF-α, and IL-10), CRP, and ESR quantify systemic inflammation, while pulmonary tests (FEV₁ and FVC) will assess lung function. Correlation analyses will define microbial taxa linked to inflammatory and respiratory outcomes, elucidating the role of the gut–lung axis in disease progression. Results/Anticipated Results: We anticipate reduced microbial diversity and enrichment of pro-inflammatory taxa (e.g., Bacteroides and Prevotella) in PCD participants. Microbial shifts are expected to correlate with elevated inflammatory cytokines and decreased lung performance. These findings will highlight gut dysbiosis as a modifiable driver of inflammation, offering a foundation for future translational interventions such as diet or probiotic therapies. Discussion/Significance of Impact: This study advances precision medicine for rare lung diseases by elucidating gut–lung axis mechanisms in a genetically defined Puerto Rican PCD cohort, generating foundational data for future microbiome-targeted therapies and strengthening Puerto Rico’s translational research capacity.

Objectives/Goals: Major depression is a common, debilitating illness. Rumination is common in depression and associated with worse outcomes. Hyperconnectivity within the Default Mode Network, particularly from ventromedial frontal cortex, underlies rumination. This study uses TMS to target this network and symptom. Methods/Study Population: This is a single-site open-label pilot K12 study that aims to recruit n = 20 adult patients with major depressive disorder. The study intervention is 20 once-daily sessions consisting of 1800 pulses of image-guided continuous theta burst stimulation (cTBS) to the ventromedial prefrontal cortex within the default mode network. Participants will undergo three MRI scans: prior to first TMS, early in the treatment course, and post-TMS. The first MRI includes a high-resolution structural scan used for TMS targeting. All scans include resting state and task-based fMRI. Outcomes include rumination (Ruminative Response Scale) and depression (Hamilton Depression Rating Scale) scores, as well as connectivity in functional MRI. Results/Anticipated Results: At the time of this writing, four participants have completed the study; two additional participants dropped out (one due to family circumstances and one due to MRI claustrophobia). The intervention and other study procedures have been well tolerated, and no serious adverse events have occurred. All participants showed decreased depression and rumination scores at the end of the intervention compared to pretreatment baseline; repeated measures general linear models indicate statistically significant effects for both overall depression and rumination. We anticipate updated symptom-based and preliminary fMRI results at the conference. Discussion/Significance of Impact: This image-guided novel TMS protocol appears to be well tolerated in our target population and feasible at our institution. Preliminary results for depression and rumination symptoms, although open label with a very small sample, are promising. We aim to complete the study by July 2026.

Objectives/Goals: The impact of genetic alterations on survival in patients with neuroendocrine prostate cancer (NEPC) is unclear. This study assessed how genetic alterations and tumor mutational burden (TMB) affect overall survival (OS) in patients with NEPC treated at Mayo Clinic. Methods/Study Population: Demographic, clinicopathologic, and genomic data from patients with NEPC treated at Mayo Clinic (2015–2025) were abstracted from electronic medical records. Patients had consented to research and received at least one cycle of therapy. Descriptive statistics summarized patient characteristics, and survival was estimated with Kaplan–Meier analysis. OS was compared by disease type (de novo vs treatment-emergent NEPC), TMB (high vs low), and RB1, TP53, and PTEN alteration status. Results/Anticipated Results: Among 66 patients in the study, the average age at NEPC diagnosis was 67.5 years. There were no significant differences between the age at diagnosis of de novo and treatment emergent NEPC (t-NEPC). t-NEPC made up 77.3% of the cohort. Median follow-up was 46.5 months (IQR 28–120). The most common alteration was TP53. RB1 alterations were associated with significantly shorter OS compared to patients without RB1 alterations (mOS Not Reached (NR) vs 13.8 months; p = 0.0056). Similarly, patients with high TMB had worse overall survival (mOS NR vs 14.9 months; p = 0.077). We did not observe worse OS in patients with alterations in TP53 or PTEN. RB1 alteration and high TMB were the strongest predictors of worse survival. Discussion/Significance of Impact: RB1 alterations and high TMB predict poor survival in patients with NEPC, highlighting potential biomarkers for risk stratification. TP53 and PTEN did not significantly affect OS. Early genomic profiling and targeted management of actionable alterations such as BRCA and ATM may improve survival and inform future translational studies.

Objectives/Goals: The Clinical and Translational Research Funding Program (CTRFP) awards 12-month community-engaged pilot projects. Following cycles of proposals with underdeveloped community engagement plans, the study section recommended earlier community feedback. The CTRFP early review process integrates lived experiences in proposal development. Methods/Study Population: Each CTRFP competitive letter of intent (LOI) submission includes a partnership description with roles, project engagement plans, and aims. The early review process includes 1) match community health consultants with specific lived experiences to LOIs; 2) separately, match academics with specific community-engaged research experiences to LOIs; 3) meet with each reviewer to collect verbal feedback; 4) provide written feedback from both community and academic perspectives to applicants one month prior to proposal due date; and 5) consult with applicants as requested leading up to proposal submission. Applicants must state how they incorporated feedback into their proposal. This process is managed through the Institute of Clinical and Translational Sciences’ community engagement function. Results/Anticipated Results: Three cycles of early review have been integrated into the CTRFP, involving 11 community and 5 academic reviewers. * 29% of LOIs reviewed did not submit. Two realized their partnerships were not ready and within 4 months applied for partnership development funding. * An early reviewer said, “It’s one thing to have a bird’s eye view of an issue, but another to talk to people that grew up in it and know it in their day to day. My voice is totally necessary. I break down ‘food insecurity’ from this surface concept into this issue impacting real people like me and my family.” * Applicants said feedback led to better research design and engagement activities, community benefit, and bigger partnerships. * 22% of proposals scored in the fundable range in 2021. In 2024, 66% of proposals scored in the fundable range. Discussion/Significance of Impact: Early review of CTRFP applications proposing community-engaged research demonstrates accountability to valuing lived experience. Staggering funding with programs that support community engagement, such as partnership development funding and community studios, can efficiently improve proposals not adequately prepared for submission.

Objectives/Goals: Youth with painful disorders of gut–brain interaction (DGBIs) report school-related challenges: stigma from school staff and poorer performance. The present study explored how stigma from school staff correlates with poorer school functioning through internal locus of control and pain interference. Methods/Study Population: Participants included 133 youth aged 8–18 years (Mage= 13.8). Youth completed a series of validated self-report questionnaires assessing pain-related stigma from school personnel, pain interference (i.e., the degree to which pain prevents engagement in daily activities), health-related locus of control (LOC – the belief that one’s health is determined by their own actions), and school functioning. A sequential mediation analysis was conducted using 95% bootstrapped confidence intervals to ascertain the indirect effect of school-based stigma on school functioning through internal LOC and pain interference. An alpha level of 0.05 was determined a priori. Results/Anticipated Results: School-based stigma was significantly correlated with poorer school functioning, even after controlling for internal LOC and pain interference, β = -0.22, p = .003. Evidence suggests a partial mediation model, such that school-based stigma is indirectly associated with poorer school functioning via internal LOC and pain interference (β = -0.028, 95% CI [-0.069, -0.002]). More specifically, greater school-based stigma correlates with a greater internal LOC (β = 0.19, p= .03). In turn, a greater internal LOC correlates with greater pain interference (β = 0.32, p<.001). Finally, greater pain interference correlates with poorer school functioning (β = -0.46, p< .001). Discussion/Significance of Impact: Stigma from teachers and nurses may cause youth with DGBIs to feel more responsible for their pain. Consequently, as they internalize this stigma, their pain may present a greater barrier to engagement in daily activities, including participation in school activities.

Objectives/Goals: Interdisciplinary teams are central to modern science, but measuring their success remains difficult. Traditional metrics like publications or citations often miss key contributions, such as community engagement, mentorship, and lasting collaboration. New approaches are needed to capture these goals. Methods/Study Population: We introduce a framework aligning success metrics with a team’s organizing priorities, illustrated through two contrasting models: Team A, a time-bound, community-focused group emphasizing rapid deliverables and public health partnerships, and Team B, a mentorship-driven, development-focused group prioritizing capacity building and long-term growth. These teams share a commitment to advancing translational research but differ in how they pursue these goals. Team A emphasized centralized coordination, shared ownership of goals, yielding timely scholarly outputs, and setting the stage for future collaborations. Team B prioritizes long-term development, sustained collaboration, and the mentorship of early-career faculty, even if traditional academic productivity metrics accrue more gradually. Results/Anticipated Results: These contrasting approaches raise an important question: how should success be defined and measured for teams with very different goals, structures, and outputs? From these cases, we derive four Collaborative Guideposts: Time Perspective, Unit of Assessment, Desired Outcomes, and Sustainability, which provide a systematic way to identify and select metrics that meaningfully capture a team’s contributions and impact. By offering a structured yet adaptable framework, these guideposts enable teams to track progress, monitor outcomes, and ensure alignment with their unique goals, operating environments, and stages of development. This approach contributes to advancing validated models of team science by providing a practical method for tailoring evaluation criteria to diverse collaborative contexts. Discussion/Significance of Impact: By grounding evaluation in team-specific goals, this framework supports more accurate assessments of collaborative impact. Guideposts provide a flexible structure to align metrics with context, guide discussions of outcomes, and evaluate teams on productivity, collaboration, mentorship, and long-term scientific progress.

Objectives/Goals: The objective of this study is to quantify the physical properties of the human vitreous noninvasively in vivo, by applying Brillouin Spectroscopy. Our goal is to measure the Brillouin shift and linewidth of the human vitreous in normal aging, diabetes, and myopia. Methods/Study Population: Our study population will include three groups: patients with emmetropia and no diabetes, patients with diabetes without proliferative retinopathy, and patients with axial myopia. We will exclude patients with any prior vitreoretinal surgery or other systemic or ocular conditions that may affect the vitreous such as rhegmatogenous retinal detachment or systemic collagen disorders. We will measure baseline characteristics including age, axial eye length, diabetes duration, and A1c level. We will perform a complete dilated eye exam and obtain imaging including optical coherence tomography in addition to Brillouin microscopy of the anterior and mid vitreous. Results/Anticipated Results: We anticipate the following findings based on evidence from ex vivo studies which show a decrease in vitreous viscosity in the anteroposterior axis physiologically, in addition to a normal decrease in vitreous viscosity with age. In diabetes, we expect to find an increase in the viscosity of the vitreous proportional to the duration and severity of diabetes due to glycation of the vitreous. In myopia, we expect to find accelerated age-related decline of the viscosity of the vitreous, proportional to the degree of myopia present. Discussion/Significance of Impact: Aging, diabetes, and myopia are risk factors for the development of blinding conditions like diabetic retinopathy and rhegmatogenous retinal detachment. By quantifying the mechanical changes of the vitreous related to these risk factors, we hope to identify patients at high risk for vision loss for preventive treatment and monitoring.

Objectives/Goals: There remains an urgent need for the development of safe and effective therapeutics for the long-term management of pain. We evaluated our novel analgesic in multiple pre-clinical models to assess its abuse liability, analgesic efficacy in models of chronic pain, and its mechanism of action in ex vivo spinal cord slices. Methods/Study Population: We evaluated SSA3’s abuse liability by utilizing an IV self-administration paradigm. In this experiment, rats are placed in a chamber and have the option to press two different levers. Abuse liability was determined based on extent of lever pressing on the drug-infusion lever. To evaluate chronic pain efficacy, mice underwent a spared nerve injury (SNI) which induces sustained hypersensitivity in their hind paw. Mice were then treated with SSA3, and their tactile hypersensitivity was assessed. Finally, the mechanism of action was investigated through patch clamp electrophysiology recordings taken from excised spinal cord slices in mice. We recorded currents from neurons involved in the pain signaling pathway before and after application of our compound to observe drug efficacy at the cellular level. Results/Anticipated Results: Our results demonstrated that in a pre-clinical IV self-administration paradigm, SSA3 exhibited no abuse liability, indicated by a lack of drug intake or escalation of drug intake. Results from our SNI model of chronic pain reveal that when SSA3 is administered directly onto the spinal cord via intrathecal injection, it is efficacious in reversing the pain hypersensitivity established by the model, suggesting a spinally mediated mechanism of action. Finally, we observed that SSA3 effectively reduced the amplitude of NMDA receptor-mediated excitatory postsynaptic currents in a concentration-dependent manner. This reduction in amplitude suggests that our compounds analgesic efficacy comes from the inhibition of spinally mediated NMDA receptor signaling. Discussion/Significance of Impact: This investigation into the therapeutic potential of substituted agmatine for the management of chronic pain revealed several key insights. Our drug is efficacious in models of chronic pain and, importantly, lacks the abuse liability commonly seen with analgesic drugs. Our drug’s mechanism of action involves inhibition of spinal NMDA receptors.

Objectives/Goals: Autism spectrum disorder (ASD) is a complex neurodevelopmental condition with unclear biological causes, potentially influenced by the gut-brain axis. This study sought to explore the infant gut microbiome to identify whether specific changes in the microbiota during early infancy are associated with later diagnosis of ASD. Methods/Study Population: Study participants included a subset of children from MARBLES (Markers of Autism Risk in Babies – Learning Early Signs), an ASD enriched-risk cohort study. Infant stool samples were collected from these children between 0 and 7 months of age. At 36 months of age, children were clinically classified as having ASD (n=42), or non-typically developing without ASD (Non-TD, n=16), or typically developing (TD, n=79). The fecal microbiome was investigated using 16S rRNA gene sequencing to evaluate whether the gut microbial composition during early infancy was associated with later neurodevelopmental diagnoses. Results/Anticipated Results: Overall, no significant differences in alpha diversity or beta diversity, or bacterial abundance, were found between groups of infants who developed ASD or Non-TD compared to those who went on to have TD. However, our findings highlight some early alterations in gut microbial composition during infancy that may relate to later neurodevelopmental outcomes. For example, infants who developed ASD showed slightly lower Veillonella and Flavonifractor levels, suggesting minor early microbial distinctions potentially linked to neurodevelopment. These differences are less pronounced than the effects of delivery mode or diet on the infant gut microbiome. Discussion/Significance of Impact: Although we found no significant differences in bacterial abundance in early infancy linked to later neurodevelopmental outcomes, our results highlight minor microbial shifts that align with findings in older children with ASD, which may indicate early gut microbiome dysbiosis, but further research is needed.

Objectives/Goals: This research develops a Ruthenium (III)–Fruquintinib complex for lung cancer treatment. Goals include structural characterization using MALDI-TOF and UV–Vis spectroscopy, synthesis optimization, and antitumor activity assessment comparing efficacy against Fruquintinib alone through cell studies. Methods/Study Population: The Ruthenium(III)–Fruquintinib complex is synthesized through an optimized coordination chemistry protocol ensuring reproducibility. Small-scale reactions defined baseline conditions, monitored by UV–Vis spectroscopy for ligand–metal charge transfer. After successful crystallization, the method was scaled up consistently. Structural characterization using MALDI-TOF mass spectrometry will be performed in collaboration with the MSRC, University of Puerto Rico, with infrared spectroscopy as a complementary validation. Biological evaluation will use NSCLC cell lines to compare cytotoxic efficacy against Fruquintinib alone, employing MTT assays to determine IC50 and potential synergistic effects. Results/Anticipated Results: Lung cancer remains the leading cause of cancer-related mortality, with 125,070 projected deaths in 2024. Current treatments for metastatic disease include surgery, chemotherapy, radiation, and targeted therapy. Fruquintinib (FQ), a VEGFR inhibitor, showed promise in early NSCLC trials but limited efficacy in advanced stages. Ruthenium (III) complexes, based on the hard–soft acid–base principle, offer potential anticancer properties. Combining FQ with Ruthenium (III) may enhance therapeutic outcomes. Preliminary synthesis yielded reproducible crystals confirmed by UV–Vis signals. Future MALDI-TOF characterization and biological assays will assess structural validity and antitumor potential. Discussion/Significance of Impact: The Ruthenium (III)–Fruquintinib complex could enhance Fruquintinib efficacy in advanced NSCLC. This approach demonstrates the potential of metal-based drug design to improve cancer therapies, offering insights into combinatorial strategies and advancing coordination chemistry in medicinal research.

Objectives/Goals: Despite the growing importance of environmental and occupational health (EOH) in public health, little is known about how DrPH graduates with EOH-focus perceive training adequacy, navigate careers, and leadership roles. This study explores the alignment between dissertation, career, and leadership outcomes among EOH-focused DrPH graduates. Methods/Study Population: This cross-sectional study analyzed 60 DrPH dissertations completed between 2000 and 2024 with EOH foci from U.S-based public health schools and programs. Secondary data were drawn from dissertations in ProQuest dissertation database, institutional records, Council of Education on Public Health (CEPH) records, and LinkedIn profiles. Variables examined include program characteristics, dissertation design, dissertation-career alignment, post-DrPH leadership role, and perceived preparedness for career and leadership roles. Data were summarized by descriptive statistics. Results/Anticipated Results: Majority (57%) of graduates were female; 81.7% trained at CEPH-accredited programs. Prior bachelor’s degrees were mostly in biology (20%) and master’s in public health (33.3%). Dissertation themes were environmental (38.3%), occupational (33.3%), or both (28.4%). Methods were largely quantitative (41%), cross-sectional (80%), with primary data (51.7%), though 36.7% lacked a theoretical framework. Post-DrPH employment was mainly in academia (35%), private sector (28.5%), and government (15%), evenly split between EOH and non-EOH sectors. Graduates held leadership roles globally (e.g., WHO, FDA, NASA, CDC, and NGOs) as professors, EOH specialists, directors, consultants, senior researchers, CEOs, and VPs. Discussion/Significance of Impact: DrPH graduates with EOH focus demonstrated strong academic preparedness and diverse leadership capabilities across academia, government, and global agencies. Versatility is reflected in equal EOH/non-EOH employment. Limited methodological diversity and weak theoretical use suggest areas for program curriculum improvement.

Objectives/Goals: While there are many efforts to teach artificial intelligence and data science (AI & DS) methods, the importance of problem formulation in translational research (TR) has received limited attention. The goal of this presentation is to revisit this challenging pedagogical topic and formalize approaches in inculcating its design principles. Methods/Study Population: AI & DS didactic courses cover problem formulation implicitly in their various course modules. Also, problem formulation is a prominent education theme of quantitative fields such as mathematics and physics. Also, data scientists working with TR usually spend considerable time in the data design aspects to quantitively solve the health problem at hand. We used three approaches to building a pedagogical framework for DS problem formulation in TR: (1) review existing literature on problem formulation in TR, applied mathematics and physics; (2) obtain structured responses around teaching problem formulation from AI & DS instructors; and (3) add exemplar DS in TR studies we actively supported. Results/Anticipated Results: Conceptualizing complex problems in a quantifiable manner can often be challenging. The typical processes involved in formulation included starting with a real-world and relatable context, iteratively defining the problem and chunking it into portions related to individual entities, abstracting each chunk to a quantifiable and working statement, identifying appropriate data resources and their constraints, and transforming data into reasonable representations. Strategies to teach problem formulation we identified through our analysis include active learning (problem-based learning and question formulation techniques), stimulating metacognition (problem defining, contextualizing, experimentation, and creativity), and group learning with think aloud and constructive feedback. Discussion/Significance of Impact: Instilling problem formulation skills, acquired over years of experience, in condensed timelines, requires formal pedagogical approaches. Ability to formulate a problem in multiple ways increases the likelihood of finding a tractable solution and can be covered in TR and DS courses and enable increased use of real-world data resources in research.

Objectives/Goals: Nearly 1 in 5 children with a suicide-related emergency department (ED) visit will have a suicide attempt within 18 months. Our objective was to determine demographic and clinical characteristics associated with time to ED revisit with self-injurious thoughts and behaviors (SITB) within 6 months. Methods/Study Population: We performed a retrospective cohort study of children aged 6 to 17 years with a primary mental health ED visit at 38 US children’s hospitals from January 2021 to December 2023. Data were obtained from the Pediatric Health Information System administrative database. We included the first index mental health ED visit for each child. Primary outcome was time to first ED revisit with a SITB diagnosis within 6 months of a patient’s index mental health ED visit, defined by ICD-10-CM diagnosis codes. We evaluated demographic and clinical characteristics associated with time to SITB revisit using a multivariable Cox Proportional Hazards Regression model. Results/Anticipated Results: We included 201,214 unique children, 8.7% had an ED revisit with SITB within 6 months. Among children with an SITB revisit, 74.6% had SITB diagnosed at the index visit. Multiple demographic characteristics were associated with longer time to revisit including male sex (HR 0.76, 95% CI 0.74-0.79) compared to female, private insurance (HR 0.83, 95% CI 0.80-0.86) compared to public, and Hispanic race and ethnicity (HR 0.81, 95% CI 0.77-0.85) compared to non-Hispanic White. Clinical characteristics associated with a shorter time to SITB revisit included index visit diagnoses for intentional self-harm (HR 1.32, 95% CI 1.27-1.38), suicidal ideation (Hazard Ratio (HR) 1.65, 95% CI 1.59-1.71), and mood disorder (HR 1.38, 95% CI 1.33-1.44). Discussion/Significance of Impact: Three in four children with a SITB ED revisit had SITB diagnosed at the index visit. Children with index visit suicidal ideation were at highest risk of SITB revisit. These findings highlight missed opportunities to identify and address pediatric SITB at the index ED visit and to prevent ED revisits.

Objectives/Goals: Cross-institutional collaboration describes the development of cross-institutional partnership between two CTSI’s to overcome common challenges faced by both entities. Community engagement work discusses ways that Strategic Doing fostered sustainable initiatives to improve wellness in rural communities. Methods/Study Population: Through collaborative efforts, the Indiana and Penn State University Clinical & Translational Science Institutes were able to plan and implement community engagement initiatives to advance health outcomes among under-resourced populations in each of their respective communities. The institutions shared and utilized key principles of an educational series entitled “Strategic Doing (SD)” that provided instruction on developing and facilitating complex collaborations. Leveraging SD practices, both CTSI’s community engagement teams guided groups of local community leaders and stakeholders in meaningful, action-oriented partnerships to promote better health and wellness. Results/Anticipated Results: At Indiana CTSI, six staff completed Strategic Doing Institute Agile Leadership training. As a result, community partners in 11 counties implemented evidence-based strategies, including a community garden with cooking classes, a mobile kitchen for nutrition education, and wellness passports at community events with food samples, screenings, and educational programs. At Penn State, twelve staff (9 CTSI, 3 community partners) completed the same training. Of 8 post training evaluation respondents, 75% were extremely likely and 25% somewhat likely to apply the training to their work. Trainees are leading task forces that received CTSI funding focused on substance use, healthy aging, and public health infrastructure and health literacy. Discussion/Significance of Impact: SD trainees have reported transformative changes when facilitating community-academic partnerships. By using a scientifically based framework, task forces have experienced enhanced synergy, resulting in increased engagement and accountability that led to improved wellness in the community.

Objectives/Goals: The University of Michigan (U-M) is a large research enterprise with a broad infrastructure. This poses challenges for clinical research professionals (CRPs) to create and utilize professional networks. To address this issue, we created a CoP to provide networking opportunities and support to CRPs at schools and colleges across the university. Methods/Study Population: We began by creating a document that detailed the needs of CRPs, defined CoP goals, and described the initial cohort, establishing a shared vision and structured guidance for recruitment. The CoP was designed to meet monthly for 6 months with the ability to continue participation as new cohorts are added. Facilitators identified 30 CRPs for cohort 1 representing a variety of locations and research types across the university. CRPs had at least 2 years of experience and demonstration of leadership responsibilities. CRPs were invited to submit a brief interest form committing to support the CoP. Communication tools were also provided so that participants could connect between meetings. Results/Anticipated Results: The program began in June 2025 with 15 participants accepting the initial invitation. Twelve CRPs attended the kick-off event, which included establishing group norms, goals, and a speed networking activity to promote cross-department networking. Participants ranged in experience from less than 3 years to over 10 years and represented 11 different departments. Of the 15 participants, 10 had 8 or more years of experience. Members selected topics including Breaking Out of Your Silo, Mentoring Up, and Time Management, and a mixed in-person and virtual schedule was chosen by the initial cohort members. Program evaluation is ongoing, including session attendance, participant willingness to continue in the CoP, sense of community, value provided by CoP elements, and feasibility of continued programming. Discussion/Significance of Impact: CoPs can be created at large institutions with careful planning and intentionality. We established a strong initial cohort that can maintain continuity and mentor future members. Communication tools for continued engagement between meetings help cross-department networking, which is challenging to maintain in a busy research environment.

Objectives/Goals: Lowering of FUS with antisense oligonucleotide (ASO) has shown promise in a subtype of amyotrophic lateral sclerosis (FUS-ALS). As inflammation is associated with ALS progression, we characterized immune trajectories in FUS-ALS patients treated with FUS ASO to understand the relationship inflammation may have with treatment response. Methods/Study Population: Twelve participants (10 with a diagnosis of ALS, 1 symptomatic carrier, and 1 asymptomatic carrier) received monthly intrathecal administrations of FUS ASO (i.e., jacifusen). Motor function was measured using the ALS Functional Rating Scale-Revised. Axonal injury was measured by cerebrospinal fluid (CSF) neurofilament light chain (NfL) concentration. 10x Genomics single-cell RNA sequencing (scRNA-seq) with T- cell receptor (TCR) sequencing was performed on paired peripheral blood mononuclear cells (PBMCs) and CSF cells. Proteomic analysis was conducted using the Olink Explore HT platform. Complementary analyses included spectral flow cytometry validation and immunohistochemical staining of post-mortem lumbar spinal cord. Results/Anticipated Results: We observed a proportional shift starting at 6 months, whereby T-cell proportions decreased, and myeloid cell proportions increased over treatment. Decreases in T-cell proportions over treatment were not accompanied by increased CD8+ T-cell infiltration in the ventral horn of the lumbar spinal cord. Differential expression revealed long-term, chronic activation and lysosomal dysregulation in myeloid cells. While CSF monocyte proportions were positively associated with CSF NfL concentration, positive associations with ALSFRS-R score were observed only in PBMC T-cell proportions, particularly cytotoxic subclusters. Proteomic analysis also highlighted a shift toward long-term innate immune cell activation associated with reduced ALSFRS-R score, accompanied by decreases in CSF lysosomal proteases. Discussion/Significance of Impact: These findings provide insight into the immune dynamics of FUS-ALS in the context of CNS FUS knockdown, as well as their potential relationship with clinical outcomes. Further, the long-term shift toward monocyte activation and persistent lysosomal dysregulation signature outside the CNS highlights potential avenues for combination therapy.

Objectives/Goals: Colorectal cancer (CRC) disproportionately affects the U.S. Deep South. Socioeconomics, genetics, and healthcare access contribute to increased incidence and mortality. A next-generation endoscope with improved contrast and automatic detection may boost early detection and narrow expertise gaps between top centers and rural clinics. Methods/Study Population: We implemented excitation-scanning hyperspectral imaging (Ex-HSI) to discriminate normal and cancerous mouse colorectal tissues by detecting changes in autofluorescence. Ex-HSI permits detection of all emitted light above a cutoff wavelength. Ex-HSI reduces acquisition time and improves signal-to-noise ratio compared to traditional emission scanning. We used azoxymethane/dextran sodium sulfate (AOM/DSS) to induce colitis and nodule formation in the mouse CRC model. Ex-HSI data was complemented with transmitted light and confocal images. Histology sectioning with H&E staining was obtained for “gold standard” validation. Results/Anticipated Results: Inflammation and bleeding were observed, consistent with AOM/DSS treatment. Nodules were visible using 4x and 20x objectives. Background-subtracted Ex-HSI data was corrected to a flat spectral response. Excitation spectra were extracted from select regions within each field of view. Excitation spectra displayed multiple peak excitation wavelengths, suggesting multiple autofluorescent molecules present. Further investigation will utilize spectral unmixing, principal component analysis, and neural networks to interpret image data. Discussion/Significance of Impact: A hyperspectral endoscope that detects early-stage cancers and precancers would enhance CRC screening in the Deep South. This research is supported by NIH P01HL066299, NIH UL1TR00141, S10RR027535, S10OD02860, NSF MRI1725937, CCTS T32TR004767, and the Center for Lung Biology, University of South Alabama.

Objectives/Goals: Our objectives were to determine enteric microbiome characteristics in fluoroquinolone-resistant Enterobacterales (FQRE)-colonized and non-FQRE-colonized hematopoietic cell transplant (HCT recipients, and to understand the enteric microbiome-specific factors associated with the development of FQRE BSI in FQRE-colonized HCT recipients. Methods/Study Population: This was a prospective cohort study of patients undergoing HCT at Weill Cornell Medicine from November 2016 to August 2019. All participants received levofloxacin prophylaxis during post-HCT neutropenia. Stool samples were collected prior to levofloxacin prophylaxis initiation and during the week after HCT. Our overall study population consisted of 26 FQRE-colonized HCT recipients and 69 non-FQRE-colonized HCT recipients. Samples underwent selective quantitative culture for FQRE, as well as 16S rRNA sequencing. Raw sequence data were processed and aligned to a custom SILVA database. Analysis of 16S rRNA sequencing was conducted in R using phyloseq. Results/Anticipated Results: We saw significant differences in microbiome composition in comparing pre-HCT samples from FQRE-colonized HCT recipients to non-FQRE-colonized HCT recipients, including significantly higher relative abundances of Bacteroidales and Enterobacterales in colonized recipients. MaAslin2 demonstrated significantly differential abundance of Bacteroides and Escherichia-Shigella ASVs. We observed a significant decrease in Enterobacterales relative abundance in non-FQRE-colonized recipients after HCT, but no such change in FQRE-colonized recipients. FQRE-colonized recipients who developed FQRE BSI had a lower relative abundance of Bacteroidales as compared to those who did not. No significant difference in FQRE colonization density as measured by quantitative FQRE stool culture was observed. Discussion/Significance of Impact: FQRE-colonized HCT recipients represent a unique subgroup of HCT recipients. Colonized HCT recipients with FQRE BSI had lower relative abundance of Bacteroidales compared to those without BSI. Further work is needed to determine the mechanism behind this association and to further characterize the microbiome in this subgroup of HCT recipients.