OBJECTIVES/SPECIFIC AIMS: Earlier pubertal timing has been associated with risk for depression, particularly in girls (e.g., Keenan etal., 2014). Evidence suggests pubertal timing in girls also relates to alterations in the microstructural properties of brain white matter tracts in late adolescence (Chahal etal., 2018), and structural connectivity of cingulate and frontal regions (Chahal etal., in prep), though differences in pubertal development in both boys and girls have not been examined in the context of brain functional connectivity (FC). Individual differences in the course of puberty may have enduring effects on functional coupling among brain regions that may contribute to the risk for psychopathology. To address this question, we explored the relation between pubertal timing and tempo with depression symptoms (age 16). Then, we examined whether brain network FC (age 16) associates with pubertal indices and predicts concurrent and later depressive symptoms (age 18). METHODS/STUDY POPULATION: Sixty-eight adolescents (37 females) completed the Mini-Mood and Anxiety Symptom Questionnaire (MASQ; Clark & Watson, 1995) at ages 14-18. Gompertz growth curve modelling of pubertal development (age 10-15; Waves 1-6) was used to estimate pubertal timing and tempo per individual, separately for males and females (e.g., Chahal etal., 2018). Resting-state MRI data (age 16) were parcellated into 264 cortical and subcortical regions to create region-to-region FC matrices based on correlations of time-series. Individual matrices were fed to the GraphVar program (Kruschwitz etal., 2015) to assess the interaction of pubertal timing and pubertal tempo with functional network connectivity using Network-based statistic (NBS; Zalesky etal., 2010). Subnetworks showing alterations in relation to pubertal timing and tempo were then examined in association with concurrent (age 16) symptoms and used to predict future depressive symptoms (age 18). RESULTS/ANTICIPATED RESULTS: In all youth, earlier pubertal timing was associated with higher depressive symptoms at age 16 (p<.018). This association was stronger in girls with slower pubertal tempo (p<.039). Interregional connectivity analyses revealed that the interaction of earlier pubertal timing and slower tempo was associated with lower FC between the left cingulate gyrus and right precuneus (p<.0001), regions implicated in emotion processing (i.e., Affective Processing Network) and self-referential thinking (i.e., Default Mode Network). FC of the three other emotion- and self-referential processing network regions (i.g., left insula, superior parietal lobule, and precuneus) was lower in youth with greater age 16 depressive symptoms (p<.0001). Finally, lower FC of of the left and right inferior parietal lobule predicted greater depressive symptoms at age 18 (p<.0001). In summary, FC of overlapping affective and default mode network areas was related to earlier pubertal timing and higher concurrent and future depressive symptoms. DISCUSSION/SIGNIFICANCE OF IMPACT: These findings demonstrate individual differences in pubertal maturation are associated with depressive symptoms and differences in brain connectivity in mid-adolescence. Early pubertal development was associated with greater depression symptoms and lower FC of brain regions involved in emotion regulation and self-referential processing. Further, FC between these regions predicted higher depression symptoms two years later. These neurobiological mechanisms may, in part, underlie the link between off-time pubertal development and the risk for depression. These findings also have important implications for precision psychiatry, as we show that a risk-factor of depression (early pubertal timing) may manifest in developing neurobiology in region-specific ways. Previous network models of depression (e.g., Li etal., 2018) implicated affective network connectivity in sustained negative mood and the default mode/ self-referential network in rumination. Other networks implicated in these past models include the reward network, which may be involved in anhedonia and loss of pleasure. Our study only found associations between affective and self-referential regional connectivity, pubertal maturation, and depression, suggesting that pubertal risk factors may relate more closely with emotion-regulation and self-referential processing deficits.

OBJECTIVES/GOALS: There is an interest in developing a bioprosthetic ovary for ovarian tissue transplantation. The properties of the ovarian extracellular matrix need to be better understood in order to replicate the human ovary. We tested the effects of an encapsulating hydrogel at different rigidities on bovine primordial follicle activation, growth, and survival. METHODS/STUDY POPULATION: Bovine primordial follicles were isolated from ovarian cortex. A mean of 9.9 follicles (range 3-24) were encapsulated per bead in either 1% or 5 % alginate across 4 experiments. The encapsulated follicles were subsequently crosslinked in a calcium sulfate solution. Follicles were then cultured for 8 days with light microscopy imaging taken every other day along with media exchanges. Follicles were then examined using immunofluorescence. Growth and survival curves were constructed and all statistical analyses were performed using Graph Pad Prism 9. RESULTS/ANTICIPATED RESULTS: A total of 372 follicles were encapsulated across 32 beads (16 in 1% alginate and 16 in 5% alginate). There were no differences in initial follicle size between the two conditions (33.53 µm vs. 32.45 µm, p=0.47). At the end of 8 days, there was no difference between follicle size (59.55 vs. 56.06, p=0.48). Additionally, there was no difference in survival between 1% and 5% alginate encapsulation (57.75% vs. 52.43%. p=0.40). Immunofluorescence is being performed on encapsulated follicles to confirm the presence of DDX4, a molecular marker of oocytes, after 8 days in culture. Additional encapsulated follicles have also been submitted for histologic sectioning and hematoxylin and eosin staining to better characterize the viability and health of these follicles after 8 days in culture. DISCUSSION/SIGNIFICANCE: There was no significant difference in growth or survival between primordial follicles cultured in 1% or 5% alginate gels. Immunofluorescent analysis confirmed the presence of viable follicles at the end of 8 days of culture. Future work needs to further explore how factors in the ovarian extracellular matrix impact follicle maintenance and growth.

OBJECTIVES/SPECIFIC AIMS: Stigma has been recognized as a major impediment to accessing mental health care among Vietnamese and Asian Americans (Leong and Lau, 2001; Sadavoy et al., 2004; Wynaden et al., 2005; Fong and Tsuang, 2007). The underutilization of mental health care, and disparities in both access and outcomes have been attributed to a large extent to stigma and cultural characteristics of this population (Wynaden et al., 2005; Jang et al., 2009; Leung et al., 2010; Spencer et al., 2010; Jimenez et al., 2013; Augsberger et al., 2015). People with neurotic or behavioral disorders may be considered “bad” as many Vietnamese people believe it is a consequence of one’s improper behavior in a previous life, for which the person is now being punished (Nguyen, 2003). Mental disorders can also been seen as a sign of weakness, which contributes to ambivalence and avoidance of help-seeking (Fong and Tsuang, 2007). Equally important is the need to protect family reputation; having emotional problems often implies that the person has “bad blood” or is being punished for the sins of his/her ancestors (Herrick and Brown, 1998; Leong and Lau, 2001), which disgraces the entire family (Wynaden et al., 2005). In these cases, public stigma (as opposed to internal stigma) is the primary reason for delays in seeking help (Leong and Lau, 2001). Other research has also highlighted the influences of culture on how a disorder may be labeled in different settings, although the presentation of symptoms might be identical (see Angel and Thoits, 1987). In Vietnamese culture, mental disorders are often labeled điên (literally translated as “madness”). A điên person and his or her family are often severely disgraced; consequently the individuals and their family become reluctant to disclose and seek help for mental health problems for fear of rejection (Sadavoy et al., 2004). Despite the critical role of stigma in accessing mental health care, there has been little work in trying to understand how stigmatizing attitudes towards mental illness among Vietnamese Americans manifest themselves and the influences of acculturation on these attitudes. Some previous work indicated a significant level of mental illness stigma among Vietnamese Americans, and experiences of living in the United States might interact with the way stigma manifests among this population (Do et al., 2014). Stigma is a complex construct that warrants a deeper and more nuanced understanding (Castro et al., 2005). Much of the development of stigma-related concepts was based on the classic work by Goffman (1963); he defined stigma as a process by which an individual internalizes stigmatizing characteristics and develops fears and anxiety about being treated differently from others. Public stigma (defined by Corrigan, 2004) includes the general public’s negative beliefs about specific groups, in this case individuals and families with mental illness concerns, that contribute to discrimination. Public stigma toward mental illness acts not only as a major barrier to care, but can also exacerbate anxiety, depression, and adherence to treatment (Link et al., 1999; Sirey et al., 2001; Britt et al., 2008; Keyes et al., 2010). Link and Phelan (2001) conceptualized public stigma through four major components. The first component, labeling, occurs when people distinguish and label human differences that are socially relevant, for example, skin color. In the second component, stereotyping, cultural beliefs link the labeled persons to undesirable characteristics either in the mind or the body of such persons, for example people who are mentally ill are violent. The third component is separating “us” (the normal people) from “them” (the mentally ill) by the public. Finally, labeled persons experience status loss and discrimination, where they are devalued, rejected and excluded. Link and Phelan (2001) emphasized that stigmatization also depends on access to social, economic, and political power that allows these components to unfold. This study aims to answer the following research questions: (1) how does public stigma related to mental illness manifest among Vietnamese Americans? and (2) in what ways does acculturation influence stigma among this population? We investigate how the 4 components of stigma according to Link and Phelan (2001) operationalized and how they depend on the level of acculturation to the host society. Vietnamese Americans is the key ethnic minority group for this study for several reasons. Vietnamese immigration, which did not start in large numbers until the 1970s, has features that allow for a natural laboratory for comparisons of degree of acculturation. Previous research has shown significant intergenerational differences in the level of acculturation and mental health outcomes (e.g., Shapiro et al., 1999; Chung et al., 2000; Ying and Han, 2007). In this study, we used age group as a proxy indicator of acculturation, assuming that those who were born and raised in the United States (the 18–35 year olds) would be more Americanized than those who were born in Vietnam but spent a significant part of their younger years in the United States (the 36–55 year olds), and those who were born and grew up in Vietnam (the 56–75 year olds) would be most traditional Vietnamese. The language used in focus group discussions (FGDs) reflected some of the acculturation, where all FGDs with the youngest groups were done in English, and all FGDs with the oldest groups were done in Vietnamese. METHODS/STUDY POPULATION: Data were collected through a set of FGDs and key informant interviews (KIIs) with experts to explore the conceptualization and manifestation of mental illness public stigma among Vietnamese Americans in New Orleans. Six FGDs with a total of 51 participants were conducted. Participants were Vietnamese American men and women ages 18–75. Stratification was used to ensure representation in the following age/immigration pattern categories: (1) individuals age 56–75 who were born and grew up in Vietnam and immigrated to the United States after age 35; (2) individuals age 36–55 who were born in Vietnam but spent a significant part of their youth in the United States; and (3) individuals age 18–35 who were born and grew up in the United States. These groups likely represent different levels of acculturation, assuming that people who migrate at a younger age are more likely to assimilate to the host society than those who do at a later age. Separate FGDs were conducted with men and women. Eleven KIIS were conducted with 6 service providers and 5 community and religious leaders. In this analysis, we focused on mental illness public stigma from the FGD participants’ perspectives. FGDs were conducted in either English or Vietnamese, whichever participants felt more comfortable with, using semistructured interview guides. All interviews were audio recorded, transcribed and translated into English if conducted in Vietnamese. Data coding and analysis was done using NVivo version 11 (QSR International, 2015). The analysis process utilized a Consensual Qualitative Research (CQR) approach, a validated and well-established approach to collecting and analyzing qualitative data. CQR involves gathering textual data through semistructured interviews or focus groups, utilizing a data analysis process that fosters multiple perspectives, a consensus process to arrive at judgments about the meaning of data, an auditor to check the work of the research team, and the development of domains, core-ideas, and cross-analysis (Hill et al., 2005). The study was reviewed and approved by Tulane University’s Internal Review Board. RESULTS/ANTICIPATED RESULTS: Components of public stigma related to mental illness. The 4 components of public stigma manifest to different extents within the Vietnamese Americans in New Orleans. Labeling was among the strongest stigma components, while the evidence of the other components was mixed. Across groups of participants, Vietnamese Americans agreed that it was a common belief that people with mental disorders were “crazy,” “acting crazy,” or “madness.” “Not normal,” “sad,” and “depressed” were among other words used to describe the mentally ill. However, there were clear differences between younger and older Vietnamese on how they viewed these conditions. The youngest groups of participants tended to recognize the “craziness” and “madness” as a health condition that one would need to seek help for, whereas the oldest groups often stated that these conditions were short term and likely caused by family or economic problems, such as a divorce, or a bankruptcy. The middle-aged groups were somewhere in between. The evidence supporting the second component, stereotyping, was not strong among Vietnamese Americans. Most FGD participants agreed that although those with mental disorders may act differently, they were not distinguishable. In a few extreme cases, mentally ill individuals were described as petty thefts or being violent towards their family members. Similarly to the lack of strong evidence of stereotyping, there was also no evidence of the public separating the mentally ill (“them”) from “us”. It was nearly uniformly reported that they felt sympathetic to those with mental disorders and their family, and that they all recognized that they needed help, although the type of help was perceived differently across groups. The older participants often saw that emotional and financial support was needed to help individuals and families to pass through a temporary phase, whereas younger participants often reported that professional help was necessary. The last component, status loss and discrimination, had mixed evidence. While nearly no participants reported any explicit discriminatory behaviors observed and practiced towards individuals with mental disorders and their families, words like “discrimination” and “stigma” were used in all FGDs to describe direct social consequences of having a mental disorder. Social exclusion was common. Our older participants said: “They see less of you, when they see a flaw in you they don’t talk to you or care about you. That’s one thing the Vietnamese people are bad at, spreading false rumors and discrimination” (Older women FGD). One’s loss of status seemed certain if their or their loved one’s mental health status was disclosed. Shame, embarrassment, and being “frowned upon” were direct consequences of one’s mental health status disclosure and subsequently gossiped about. Anyone with mental disorders was certain to experience this, and virtually everyone in the community would reportedly do this to such a family. “You get frowned upon. In the Vietnamese culture, that’s [a family identified as one with mental health problems] the big no-no right there. When everybody frowns upon your family and your family name, that’s when it becomes a problem” (Young men FGD). This is tied directly to what our participants described as Vietnamese culture, where pride and family reputation were such a high priority that those with mental disorders needed to go to a great extent to protect—“We all know what saving face means” as reported by our young participants. Even among young participants, despite their awareness of mental illness and the need for professional help, the desire to avoid embarrassment and save face was so strong that one would think twice about seeking help. “No, you just don’t want to get embarrassed. I don’t want to go to the damn doctor and be like ‘Oh yeah, my brother got an issue. You can help him?’ Why would I do that? That’s embarrassing to myself…” (Young men FGD). Our middle-aged participants also reported: “If I go to that clinic [mental health or counseling clinic], I am hoping and praying that I won’t bump into somebody that I know from the community” (Middle-aged women FGD). Vietnamese people were also described as being very competitive among themselves, which led to the fact that if a family was known for having any problem, gossips would start and spread quickly wherever they go, and pretty soon, the family would be looked down by the entire community. “I think for Vietnamese people, they don’t help those that are in need. They know of your situation and laugh about it, see less of you, and distant themselves from you” (Older women FGD). Culture and mental illness stigma, much of the described stigma and discrimination expressed, and consequently the reluctance to seek help, was attributed to the lack of awareness of mental health and of mental health disorders. Many study participants across groups also emphasized a belief that Vietnamese Americans were often known for their perseverance and resilience, overcoming wars and natural disasters on their own. Mental disorders were reportedly seen as conditions that individuals and families needed to overcome on their own, rather than asking for help from outsiders. This aspect of Vietnamese culture is intertwined with the need to protect one’s family’s reputation, being passed on from one generation to the next, reinforcing the beliefs that help for mental disorders should come from within oneself and one’s family only. Consequently persons with mental health problems would be “Keeping it to themselves. Holding it in and believing in the power of their friends” (Middle-aged FGD) instead of seeking help. Another dimension of culture that was apparent from FGDs (as well as KIIs) was the mistrust in Western medicine. Not understanding how counseling or medicines work made one worry about approaching service providers or staying in treatment. The habit of Vietnamese people to only go see a doctor if they are sick with physical symptoms was also a hindrance to acknowledging mental illness and seeking care for it. Challenges, including the lack of vocabulary to express mental illness and symptoms, in the Vietnamese language, exaggerated the problem, even among those who had some understanding of mental disorders. It was said in the young men FGD that: “when you classify depression as an illness, no one wants to be sick,… if you call it an illness, no one wants to have that sort of illness, and it’s not an illness that you can physically see…” (Young men FGD). Another young man summarized so well the influence of culture on mental illness stigma: “Us Southeast Asian, like, from my parents specifically has Vietnam War refugees. I think the reason why they don’t talk about it is because it’s a barrier that they have to overcome themselves, right? As refugees, as people who have been through the war… [omitted]They don’t want to believe that they need help, and so the trauma that they carry when they give birth to us is carried on us as well. But due to the language barrier and also the, like, they say with the whole health care, in Vietnam I know that they don’t really believe in Western and Eurocentric medicine. So, from their understanding of how, like from their experience with colonization or French people, and how medicine works, they don’t believe in it” (Young men FGD). One characteristic of the Vietnamese culture that was also often mentioned by our FGD participants (as well as KIIs) was the lack of sharing and openness between generations, even within a family. Grandparents, parents, and children do not usually share and discuss each other’s problems. Parents and grandparents do not talk about problems because they need to appear strong and good in front of their children; children do not talk about problems because they are supposed to do well in all aspects, particularly in school. The competitiveness of Vietnamese and high expectations of younger generations again come into play here and create a vicious cycle. Young people are expected to do well in school, which put pressure on them and may result in mental health problems, yet, they cannot talk about it with their parents because they are not supposed to feel bad about school, and sharing is not encouraged. The Asian model minority myth and the expectations of parents that their children would do well in school and become doctors and lawyers were cited by many as a cause of mental health problems among young people. “Our parents are refugees, they had nothing and our parents want us to achieve this American Dream…. [omitted] It set expectations and images for us…. It was expected for all the Asians to be in the top 10, and for, like a little quick minute I thought I wasn’t going to make it, I was crying” (Yong men FGD). As a result, the mental health problems get worse. “If you’re feeling bad about something, you don’t feel like you can talk about it with anyone else, especially your family, because it is not something that is encouraged to be talked about anyway, so if you are feeling poorly and you don’t feel like you could talk to anybody, I think that just perpetuates the bad feelings” (Middle-aged women FGD). Acculturation and mental illness stigma Acculturation, the degree of assimilation to the host society, has changed some of the understanding of mental illness and stigmatizing attitudes. Differences across generations expressed in different FGDs indicated differences in perceptions towards mental illness that could be attributed to acculturation. For example, the young generation understood that mental illness was a health problem that was prevalent but less recognized in the Vietnamese community, whereas a prominent theme among the older participants was that mental illness was a temporary condition due to psychological stress, that it was a condition that only Caucasians had. Some of the components of public stigma related to mental illness seemed to vary between generations, for example the youngest participants were less likely to put a label on a person with mental health problems, or to stereotype them, compared to the oldest and middle-aged participants. This was attributed to their education, exposure to the media and information, and to them “being more Americanized.” However, there was no evidence that acculturation played an important role in changing the other components of public stigma, including stereotyping, separating, and status loss and discrimination. For example, the need to protect the family reputation was so important that our young participants shared: “If you damage their image, they will disown you before you damage that image” (Young men FGD). Young people, more likely to recognize mental health problems, were also more likely to share within the family and to seek help, but no more likely than their older counterparts to share outside of the family—“maybe you would go to counseling or go to therapy, but you wouldn’t tell people you’re doing that” (Young women FGD). The youngest participants in our study were facing a dilemma, in which they recognized mental health problems and the need for care, yet were still reluctant to seek care or talk about it publicly because of fears of damaging the family reputation and not living up to the parents’ expectations. Many young participants reported that it actually made it very difficult for them to navigate mental health issues between the 2 cultures, despite the awareness of the resources available. “I think it actually makes it harder. Only because you know to your parents and the culture, and your own people, it’s taboo, and it’s something that you don’t talk about. Just knowing that you have the resources to go seek it… You want advice from your family also, but you can’t connect the appointment to your family because you’re afraid to express that to your parents, you know? So I think that plays a big part, and knowing that you are up and coming, but you don’t want to do something to disappoint your family because they are so traditional” (Young men FGD). Some participants felt more comfortable talking about mental health problems, like depression, if it was their friend who experienced it and confided in them, but they would not necessarily felt open if it was their problem. Subtle cultural differences like this are likely overlooked by Western service providers. One older participant summarized it well “They [the young generation] are more Americanized. They are more open to other things [but] I think that mental health is still a barrier.” DISCUSSION/SIGNIFICANCE OF IMPACT: This study investigated how different components of public stigma related to mental illness manifest among Vietnamese Americans, a major ethnic group in the United States, and how acculturation may influence such stigma. The findings highlighted important components of public stigma, including labeling and status loss, but did not provide strong evidence of the other components within our study population. Strong cultural beliefs underlined the understanding of mental health and mental illness in general, and how people viewed people with mental illness. Several findings have been highlighted in previous studies with Asian immigrants elsewhere; for example, a study from the perspectives of health care providers in Canada found that the unfamiliarity with Western biomedicine and spiritual beliefs and practices of immigrant women interacted with social stigma in preventing immigrants from accessing care (O’Mahony and Donnelly, 2007). Fancher et al. (2010) reported similar findings regarding stigma, traditional beliefs about medicine, and culture among Vietnamese Americans. Acculturation played a role in changing stigmatizing attitudes as evidenced in intergenerational differences. However, being more Americanized did not equate to being more open, having less stigmatizing attitudes, or being more willing to seek care for mental health issues. Consistent with previous studies (Pedersen and Paves, 2014), we still found some level of stigma among young people aged 18–35, although some components were lessened with an increased level of acculturation. There was also a conflict among the younger generation, in which the need for mental health care was recognized but accessing care was no easier for them than for their parent and grandparent generations. The study’s findings are useful to adapt existing instruments to measure stigma to this population. The findings also have important program implications. One, they can be directly translated into basic supports for local primary and behavioral health care providers. Two, they can also be used to guide and inform the development and evaluation of an intervention and an additional study to validate the findings in other immigrant ethnic groups in the United States. Finally, based on results of the study, we can develop a conceptual framework that describes pathways through which social, cultural, and ecological factors can influence stigma and the ways in which stigma acts as a barrier to accessing mental health care among Vietnamese Americans. The guiding framework then can be validated and applied in future programs aimed to improve mental health care utilization among ethnic minorities.

Effective stakeholder engagement increases research relevance and utility. Though published principles of community-based participatory research and patient-centered outcomes research offer guidance, few resources offer effective techniques to engage stakeholders and translate their engagement into improvements in research process and outcomes. The Indiana Clinical and Translational Sciences Institute (Indiana CTSI) is home to Research Jam (RJ), an interdisciplinary team of researchers, project management professionals, and design experts, that employs human-centered design (HCD) to engage stakeholders in the research process. Establishing HCD services at the Indiana CTSI has allowed for accessible and innovative stakeholder-engaged research. RJ offers services for stakeholder-informed study design, measurement, implementation, and dissemination. RJ’s services are in demand to address research barriers pertaining to a diverse array of health topics and stakeholder groups. As a result, the RJ team has grown significantly with both institutional and extramural support. Researchers involved in RJ projects report that working with RJ helped them learn how to better engage with stakeholders in research and changed the way they approach working with stakeholders. RJ can serve as a potential model for effectively engaging stakeholders through HCD to improve translational research.

Understanding the clinical risk factors for COVID-19 disease severity and outcomes requires a combination of data from electronic health records and patient reports. To facilitate the collection of patient-reported data, as well as accelerate and standardize the collection of data about host factors, we have constructed a COVID-19 survey. This survey is freely available to the scientific community to send electronically for patients to complete online. This patient survey is designed to be comprehensive, yet not overly burdensome, to gather data useful for a range of clinical investigations, and to accommodate a wide variety of implementation settings including at a COVID-19 testing site, at home during infection or after recovery, and/or for individuals while they are hospitalized. A widely adopted standardized survey that can be implemented online with minimal resources can serve as a critical tool for combining and comparing data across studies to improve our understanding of COVID-19 disease.

OBJECTIVES/SPECIFIC AIMS: This study’s aims are to optimize the isolation and growth of chondrocytes from pig auricular cartilage; to identify the ideal seeding conditions onto 3D printed auricular bioscaffolds to maximize chondrocyte growth; and to investigate what quantity and types of host tissue can grow on the bioscaffold. Primary outcomes will include comparisons between different seeding conditions in various objective measures of bioscaffold growth and survival as listed in the methods section. Secondary outcomes will include continued optimization of bioscaffolds to minimize extrusion rates and maximize morphologic and histologic similarity to human auricular cartilage. METHODS/STUDY POPULATION: For chondrocyte-seeded scaffolds, cartilage will be collected from freshly harvested porcine auricular tissue and digested in type II collagenase. Chondrocytes derived from the harvest will be seeded into auricular PCL scaffolds using a type I collagen/hyaluronic acid composite gel, which has been previously shown to support chondrogenesis. For scaffolds containing cartilage, punch biopsies will be collected and embedded in specific areas of the scaffold previously shown to experience excessive stress/strain compared to the rest of the construct. From there, five of each chondrocyte-seeded bioscaffolds, chondrocyte-unseeded bioscaffolds, and cartilage-containing bioscaffolds will be implanted into athymic rats. Total follow up will be for six months, with outcomes as measured by clinical assessments, morphologic measurements, radiological imaging, histological analysis, biomechanical evaluation, and photodocumentation. Once these measures are obtained, we will work closely with Dr. Myra Kim, an adjunct professor with the Biostatistics Department, to appropriately analyze differences between the models. RESULTS/ANTICIPATED RESULTS: We believe that while all scaffolds (chondrocyte-seeded, chondrocyte-unseeded, and cartilage-containing) will be structurally sound, the chondrocyte-seeded scaffolds and cartilage-containing scaffolds will exhibit improved soft tissue coverage and have lower exposure and fracture rates. Additionally, between the two, we posit that there will not be appreciable differences histologically, radiologically, or morphologically. DISCUSSION/SIGNIFICANCE OF IMPACT: Auricular reconstruction is a geometrically complex and technically challenging problem. Reconstruction hinges on the physical characteristics of the deformity, patient preferences, and reconstructive materials available. The current gold standard for auricular reconstruction uses autologous rib cartilage as foundational support for overlying soft tissue and these techniques involve freehand carving of the cartilage, requiring high levels of technical skill. Harvesting the materials for this procedure is invasive, and the outcomes of the surgery are largely variable and sometimes undesirable. As alternatives, implantable scaffolds including those made from high density porous polyethylene (commercially referred to as MedPor) have been investigated. However, many of these have proven inadequate due to factors including infection, extrusion, and morphologic and biomechanical dissimilarity from native tissue. 3D printing represents an exciting new avenue through which to address many of these difficulties. Our group has previously demonstrated the successful design, production, and implantation of 3D-printed models: in auricular reconstruction, we have demonstrated the successful creation and implementation of a 3D printed ear scaffold into an athymic rodent model. We now turn our attention to optimization of seeding of our ear scaffold with chondrocytes derived from porcine auricular cartilage or with cartilage punch biopsies, all while maintaining emphasis on regulatory feasibility. With success in this arena, we will be able to provide a much less invasive and technically challenging alternative to the current gold standard, create patient-specific bioscaffolds which are more form fitting and individualized, and provide children with ear malformations better alternatives and treatments for their conditions.

OBJECTIVES/SPECIFIC AIMS: The promise and potential of connected personal health records (PHRs) has not come to fruition. This may be, in part, due to the lack of user-centered design and of a patient-centric approach to curating personal health data for use by patients. Co-design with end-users could help mitigate these issues by ensuring the software meets user’s needs, and also engages patients in informatics research. Our team partnered with patients with multiple chronic conditions to co-design a patient-centric PHR. This abstract will describe our experience with the co-design process, highlight functionalities desired by patients, and showcase the final prototype. METHODS/STUDY POPULATION: We conducted 3 design sessions (90 min per session) with patients as co-designers and employed an iterative process for software development. Patients were recruited from Chapel Hill and surrounding areas. The initial design session laid the foundation for future sessions, and began with brainstorming about what patients thought their ideal version of an engaging connected PHR would look like in terms of features and functionalities. After each software iteration, our entire design team, including our patient co-designers, was shown the prototype during a subsequent design session. Once the final prototype was developed, usability testing was conducted with patient participants. Our team then conducted a final design session to debrief about the final prototype. RESULTS/ANTICIPATED RESULTS: We started with an initial group of 12 patients (6 males) who all had diabetes and an additional comorbidity such as hypertension and hyperlipidemia. Age of participants ranged from 30 to 77 years with an average age of 56. The majority of participants were Caucasian with 1 Asian and 2 African Americans. Hemoglobin A1c values ranged from 6.0% to 9.2% with approximately half having A1c values less than the goal of 7.0%. Half the patients were aware of PHRs, majority had smartphones, and all participants had access to the Internet and used email. Two of the patients were retired engineers who had prior experience with software design. The other sessions had between 7 and 8 participants at each session, and 7 patients completed the 90-minute usability testing session. There was a core group of 7 patients who were engaged in the design and testing sessions throughout the entire 9-month study. Key features of the PHR that emerged from design sessions included the following: (1) allow for annotation of data by patients (particularly important for lab values like glucose or for physical activity); (2) calendars, to do list, and reminder functions should be linked so that an entry in one of these allows for auto-population of this data within the other sections; (3) notifications whenever new data from the electronic health record or other sources are pushed to the PHR account; (4) allow for drag and drop of photos of pills/medications taken via smartphone or from other sources so that medication list has photo of actual pills or pill bottle; (5) allow for patients to customize the order of sections in the PHR dashboard so that the sections most important to the individual patient can be displayed more prominently; (6) allow for notifications from pharmacies to be pushed to the PHR (eg, confirmation of receipt of prescription requests or alert that prescription is ready to pick up); and (7) graphical display of trends over time (patients would like to select the measures and time frames to plot for display). Patients cited the importance of data provenance so that patient-entered data Versus provider or electronic health record data could be easily differentiated. Patients also highlighted the importance of having this PHR be a “one-stop shop for all their health data” and to have meaningful data dashboards for the different types of information needed to comprehensively manage their health. Patients wished for a single PHR that could easily bring together data from multiple patient portal accounts to avoid having to manage multiple accounts and passwords. They felt that heat map displays such as those used on popular fitness tracking websites were not intuitive and that the color-coding made interpretation challenging. Participants noted that engagement in the design process made them feel that they contributed towards developing software that could not only positively impact them individually but others as well. Every patient indicated the desire to participate on future design projects. Of the 19 tasks evaluated during usability testing, only 5 tasks could not be completed (eg, adding exercise to the calendar, opening the heat map, etc.). Patients felt that the overall PHR design was clean and aesthetically pleasing. Most patients felt that the site was “pretty easy to use” (6 out of 7). The majority of participants would like to use this PHR in the future (5) and would recommend this PHR to their friends/family to use (6). DISCUSSION/SIGNIFICANCE OF IMPACT: Involving patients directly in the design process for creating a patient-centric connected PHR was essential to sustaining engagement throughout the software life cycle and to informing the design of features and functionalities desired by patients with chronic conditions.

OBJECTIVES/GOALS: Our research has three main aims: 1. Measure attitudes toward a SARS-CoV-2 vaccine among BIPOC. 2. Assess the effectiveness of race-conscious public health messages in changing attitudes toward a SARS-CoV-2 vaccine. 3. Test the efficacy of financial incentives to increase uptake of a SARS-CoV-2 vaccine. METHODS/STUDY POPULATION: We surveyed 784 unvaccinated residents of L.A. County. To recruit participants, we collaborated with Qualtrics. The survey randomized participants to one of three public health messages, as well as one of two financial compensation schemes. Twenty-five participants completed semi-structured interviews via Zoom or telephone. Interviews were audio recorded, translated into English if needed, and transcribed. The inductive, semi-structured interview guide focused on three domains: i) concerns and distrust toward a COVID vaccine, ii) policy interventions and/or government action related to a COVID vaccine, iii) non-pharmacological policy interventions related to the COVID-19 pandemic. Major emergent themes were identified and analyzed using Watkins (2012) team analysis of qualitative data steps. RESULTS/ANTICIPATED RESULTS: Many BIPOC remain vaccine hesitant: 2/3 of the survey respondents stated that they did not intend to or were not sure if they planned to get vaccinated. Follow-up interviews show that fear of vaccine side effects, bodily autonomy in choosing to get vaccinated are major concerns. However, public health measures like masking and physical distancing remain preferred safety methods for BIPOC residents. Misinformation and overcommunication in public health messaging concerning vaccine eligibility may be a major barrier to vaccine uptake among BIPOC. DISCUSSION/SIGNIFICANCE: Real world financial compensation mechanisms need to provide large enough compensation to avoid a crowding out of altruistic vaccination motivations and to effectively incentivize increased vaccine uptake. Additionally, short race-conscious public health messages were ineffective at improving vaccine attitudes.

OBJECTIVES/GOALS: Breast cancer has an increased requirement for lipids. The sigma-2 receptor plays a critical role in the effective uptake of lipoproteins by forming a complex with the LDL Receptor. We investigate the role of the sigma02 receptor in modulating lipid uptake pathways in breast cancers, and how this can be leveraged as a viable therapeutic strategy. METHODS/STUDY POPULATION: CRISPR/Cas9 will be used to ablate TMEM97 in the MDAMB231 and MCF7 cell lines. This study seeks to identify pathways that are dysregulated upon TMEM97 knockout (KO) by characterizing RNASeq data to identify differentially expressed genes and perform pathway analysis. RESULTS/ANTICIPATED RESULTS: Knockout of TMEM97 in breast cancer cells is is expected to decrease lipid uptake. Treatment with statins in these knockout cells is expected to result in decreased cell viability and result in a quiescent cell population. DISCUSSION/SIGNIFICANCE: This is an important mechanistic study to understand the importance of lipid homeostasis in cancer cell proliferation and how it can be targeted to improve therapeutic anti-tumor strategies. Understanding the pathways that TMEM97 modulates is vital for therapeutic strategies to curb the proliferation of breast cancer cells.

OBJECTIVES/GOALS: Analyze how the Endowment HIREC ‘s Mentoring and Career Coach Model A productive mentoring relationship is essential to advance researchers into being independent and bring extramural funds. METHODS/STUDY POPULATION: Provide Hispanic researchers mentoring and career coaching to strengthen their pathway as researcher. The HiREC’s Career Coach and Mentoring Component (CCMC) is an innovated approach to support long-lasting research mentoring relationships in our institution. This approach was developed to advance research to eliminate health disparities, promote multidisciplinary translational research in a Minority Institution and sustain research infrastructure and services, career, and workforce development initiatives. Promising Faculty are target and early and mid-career investigators interested in pursuing a research career. To implement the CCMC with the Visiting Endowed Chair a HiREC Advisory Leadership Group in Mentoring will be established, with researchers from Puerto Rico, and US mainland. RESULTS/ANTICIPATED RESULTS: Three Hispanic mid-career women from the School of Medicine and one from the School of Health Professions from the University of Puerto Rico received a HiREC Advanced Research Award of $50,000. The awardees achieved their goals; completed their research plan, research infrastructure needs, peer-reviewed publications, and submission of a competitive grant. They also provided successful perspectives on mentoring relationships in a Minority institution. Each one showed the mentor’s and mentee’s experiences as fundamental for their research advancements, productivity, leadership, and successful results. HiIREC’s mentoring component with the Visiting Endowed Chairs improves a healthy work environment and expands the research agenda for each awardee sustaining the institutional research culture. DISCUSSION/SIGNIFICANCE: A productive mentoring relationship is essential to advance researchers into being independent and bring extramural funds. Four mentees received formal, long-term guidance and endowment funds for their research infrastructure requirements with successful outcomes. HiREC contributes to building up an institutional mentoring program.

OBJECTIVES/GOALS: Health insurance status is associated with differences in access to healthcare and health outcomes. The objective of this study was to test the hypothesis that among infants born in the United States, maternal private insurance compared with public Medicaid insurance would be associated with a lower infant mortality rate (IMR). METHODS/STUDY POPULATION: This ecological study used data from the Center for Disease Control and Prevention (CDC) WONDER expanded linked birth and infant death records database 2017-2018. We included hospital-born infants from 20 to 42 weeks of gestational age (wga) if the mother had either private or Medicaid insurance. We excluded infants with congenital anomalies and infants who died due to congenital anomalies. We used negative-binomial regression adjusted for race, sex, multiple birth, and any maternal pregnancy risk factors (as defined by the CDC) to determine the difference in IMR between private and Medicaid insurance. Chi-square or Fishers exact test was used to compare differences in categorical variables between groups. RESULTS/ANTICIPATED RESULTS: We included 6,901,328 infants; 53.6% had private insurance and 46.4% were insured by Medicaid. Privately insured infants had a lower IMR compared with Medicaid insured infants (2.84/1000 vs. 5.32/1000; adjusted relative risk (aRR) 0.71; 95% confidence intervals (CI) 0.62 to 0.81; p<.0001). The privately insured had higher rates of 1st trimester prenatal care compared to those with Medicaid (85.6% vs. 66.6%; p<.00001). Rates of infant morbidity and maternal morbidity (per CDC definitions) were lower among the privately insured compared to those with Medicaid (both p<.00001). The privately insured had lower rates of preterm (9.1% vs. 11.0%), extremely preterm (0.5% vs. 0.7%), low birth weight (7.1% vs. 9.6%), and extremely low birth weight (0.5% vs. 0.7%) births compared to those with Medicaid (all p<0.001). DISCUSSION/SIGNIFICANCE: Private insurance is associated with a lower IMR compared to Medicaid insurance. Privately insured pregnancies also have higher rates of early prenatal care, less morbidity, and less preterm and low birth weight births. There may be opportunities to improve access to care and pregnancy outcomes among Medicaid insured pregnancies in the United States.

OBJECTIVES/GOALS: Patients frequently need or desire fat grafting to improve common issues such as implant visibility and contour deformity, often done as a second, staged procedure following immediate reconstruction. This study aimed to identify which patient factors and reconstructive techniques predict the need for revision with AFG after IBBR METHODS/STUDY POPULATION: Patients who underwent IBBR with either tissue expanders or implants following mastectomy from 2017 to 2021 were identified. Demographics, comorbidities, and the postoperative course were reviewed. The primary outcome variable was AFG after the initial reconstruction. Univariate and regression analyses were performed to identify factors predictive of AFG. RESULTS/ANTICIPATED RESULTS: Five-hundred twenty-nine patients were included in our analysis, with 43% having AFG. Univariate regression displayed single-stage reconstruction (OR=0.53, 95% 0.37-0.75) and previous radiation (OR 0.59, 95% 0.35-0.99) negatively predicted the need for AFG, while bilateral breast reconstruction (BBR) was a predictor (OR 2.32, 95% 1.58-3.4). On multivariate analysis, decreasing age and BBR remained predictive of AFG. The odds of AFG decreased by 3% for every one-unit increase in age (95% CI [0.96, 0.99]). Interestingly, neither pre-pectoral breast reconstruction nor specimen weight:implant ratio was associated with increased need for AFG on univariate/multivariate analysis. DISCUSSION/SIGNIFICANCE: Patients requiring AFG were likely younger and had undergone BBR with tissue expanders. Plane of implant did not appear to affect need for AFG. Knowledge of these predictive factors may help plastic surgeons in preoperative counseling before implant-based breast reconstruction.

OBJECTIVES/SPECIFIC AIMS: To develop an algorithm that identifies post-concussion syndrome (PCS) cases and controls from among patients with mild traumatic brain injury (mTBI) in a large academic biobank. METHODS/STUDY POPULATION: The Vanderbilt University Medical Center’s (VUMC) electronic medical record (EMR) research database includes longitudinal medical record data on 2.5 million people. DNA and genotype data were also available for >225,000 of these individuals. Our algorithm used a combination of billing codes and natural language processing to apply inclusion and exclusion criteria. We defined PCS cases as those with a PCS billing code (ICD-9 310.2 or ICD-10 F07.81) and/or symptoms of PCS within 1–6 months of a qualifying mTBI. We will compare the positive predictive value of our algorithm to that of 2 simpler case selection schemes: (1) 1 instance of the PCS billing code anywhere in the medical record; and (2) 2 or more instances of the PCS billing code anywhere in the medical record. RESULTS/ANTICIPATED RESULTS: An mTBI was diagnosed in 28,720 patients regularly attending VUMC, and 528 of these patients were classified as PCS cases by our algorithm. The characteristics of our EMR sample reflected known risk factors for PCS. Our cases were more likely than controls to be female (49.4% vs. 38.4%), to have sustained a previous TBI (31.0% vs. 12.0%) and to have comorbid mood disorders. Our PCS cases were also more likely than controls to be <18 years of age (42.4% vs. 33.6%) and to have a sports-related keyword associated with the mTBI (44.1% vs. 25.2%), emphasizing the relevance of PCS to young athletes. Nonetheless, the number of PCS cases identified by our algorithm was small, and within the VUMC EMR, there were 5039 patients with 1 PCS billing code, and 2457 patients with 2 or more PCS billing codes anywhere in their EMR. Our next step is to calculate the positive predictive values of each selection scheme by manually reviewing the EMR of a selection of cases. Ultimately, we will implement the selection scheme that maximizes both positive predictive value and sample size, and in future work, we will genotype the selected patients to better understand the genetic architecture of PCS. DISCUSSION/SIGNIFICANCE OF IMPACT: EMR and biobanks are the future of human health research, and we asked whether complex algorithms or simple billing codes were best for studying the genetics of recovery after mTBI within the VUMC EMR. Our results are relevant to other studies of brain injury phenotypes within biobanks, including recovery from moderate or severe TBI, recovery from stroke, or the occurrence of delirium after routine surgery, and will help transform biobanks into fruitful research tools.

This abstract is based on unpublished data. OBJECTIVES/GOALS: The estimates of unbiased first-degree relatives (FDRs) risk of cancers would enhance genetic counseling of at-risk FDRs in families where the pancreatic cancer (PC) proband carrying a germline variant. This study aims at quantifying gene-specific risks of six cancers among FDRs of PC patients with germline variants in cancer-associated genes. METHODS/STUDY POPULATION: In the prospective, clinic-based Mayo Clinic Biospecimen Resource for Pancreas Research registry, 4,562 PC patients had previously undergone germline genetic testing for pancreatic cancer-associated genes through either research studies or clinical testing. Of these, 234 PC probands were found to carry germline pathogenic/likely pathogenic variants (PLPV) among 9 genes of interest and had provided detailed demographic and cancer data on their FDRs by questionnaire. We focused on six cancer types (ovary, breast, uterus, pancreas, colon, and malignant melanoma) in FDRs as reported by the probands. Standardized incidence ratios were calculated to estimate risk of six cancers among FDRs of PC patients carrying PLPV by gene. RESULTS/ANTICIPATED RESULTS: 1,670 FDRs (mean age 58.1+17.8SD; 48.9% female) were included in the study. We found significantly increased risk of ovarian cancer in female FDRs of PC probands who carry PLPV in BRCA1 (SIR 9.49, 95%CI:3.06-22.14) or BRCA2 (3.72, 95%CI:1.36-8.11), and breast cancer risks were higher with BRCA2 (2.62, 95%CI:1.89-3.54). Uterine cancer risk was increased in FDRs of PC probands who carry PLPV for Lynch Syndrome mismatch repair (MMR) (6.53, 95%CI:2.81-12.86). PC risk was also increased (ATM 4.53, 95% CI:2.69-7.16; BRCA2 3.45, 95%CI:1.72-6.17; CDKN2A 7.38, 95%CI:3.18-14.54; PALB2 5.39, 95%CI:1.45-13.79). Increased colon cancer risk was observed in FDRs of probands who carried MMR PLPV (5.83, 95%CI:3.70-8.75), while melanoma risk was elevated for FDRs of probands with CDKN2A PLPV (7.47, 95%CI:3.97-12.77). DISCUSSION/SIGNIFICANCE: PLPV in nine syndrome-associated genes in PC probands are associated with increased risk of six cancers in FDRs. The findings underscore the importance of risk estimation of various other cancers in PC families for screening, early detection, intervention, and cascade genetic testing.

ABSTRACT IMPACT: Our work might lead to a new treatment for patients with acute myeloid leukemia OBJECTIVES/GOALS: Acute myeloid leukemia (AML) is a devastating hematologic malignancy, with dismal 5-year survival. Chimeric antigen receptor (CAR) T cells have been approved for B cell malignancies but not for AML. The goal of this study is to explore the safety and efficacy of CAR T cells targeting CD105 (endoglin) to treat AML. METHODS/STUDY POPULATION: We have constructed human and murine CAR T cells targeting CD105. The CARs were created by sequencing the V(D)J regions of hybridomas and designing single chain variable fragments that target CD105 which were subsequently introduced in a CAR backbone via Gibson assembly. The CAR T cells were produced via transduction using retrovirus or lentivirus. Leukemia cell lines were assessed for CD105 expression with flow cytometry. Killing assays were performed via measurement of luminescence of target cells after co-culture with CAR T cells. Activation assays were performed with co-culture of CAR T cells and target cells and measurement of activation markers with flow cytometry. To assess in vivo efficacy and safety, murine CAR T cells were infused into C57BL/6J mice carrying B16 melanoma after lymphodepletion. RESULTS/ANTICIPATED RESULTS: All human leukemia cell lines assessed (Nalm6, MOLM-14, MV4-11, Kasumi-1, THP-1) expressed some degree of endoglin apart from the T cell leukemia Jurkat. Human CD105 CAR T cells were activated by co-culture with leukemia cell lines and effectively killed leukemia cells in vitro in a CD105-specific manner. Murine CAR T cells killed efficiently both murine solid tumors (B16 melanoma) and murine leukemias (C1498) in vitro. Murine CAR T cells did not exhibit any toxicity when infused after low-dose lymphodepletion (cyclophosphamide 100mg/kg) but caused significant morbidity after higher doses (cyclophosphamide 200mg/kg). Murine CAR T cells delayed the growth of B16 melanoma in immunocompetent mice. DISCUSSION/SIGNIFICANCE OF FINDINGS: We have constructed human and murine CD105 CAR T cells with excellent activity in vitro. The activity of human CD105 CAR T cells in xenografts and the biologic relevance of the toxicity of murine CD105 CAR T cells in humans needs to be further investigated. CD105 CAR T cells might prove an important therapeutic option for patients with AML.

OBJECTIVES/SPECIFIC AIMS: This study aims to estimate patient attitudes and receptiveness towards stem cell-based cell replacement devices to management Type 1 Diabetes. The primary outcomes of this study are mean response values to questions interrogating patient attitudes, knowledge, and receptiveness. METHODS/STUDY POPULATION: A RedCap survey was generated for this study. 100 participants will be drawn from Mayo Clinic Rochester, MN patients living with Type 1 Diabetes. RESULTS/ANTICIPATED RESULTS: Response values will be used to estimate broader patient attitudes. DISCUSSION/SIGNIFICANCE OF IMPACT: The response values of this survey will help inform future directions of cell replacement device development. Additionally, understanding patient attitudes may be useful in crafting more effective education strategies.

ABSTRACT IMPACT: This study aims to provide insight into naturally acquired immunity against severe malaria, thereby laying the foundation for the design of novel vaccine candidates to prevent severe disease as well as monoclonal antibody therapies to treat severe malaria. OBJECTIVES/GOALS: Severe malaria is caused by parasite surface antigens that contain high sequence diversity. Nevertheless, P. falciparum-exposed individuals develop antibody responses against these antigens. Our goal is to isolate antibodies with broad reactivity to understand how disease protection is acquired. METHODS/STUDY POPULATION: Our study cohort consists of Ugandan adults living in a malaria-endemic region with high transmission intensity, who are protected against severe malaria. Using fluorescently labeled probes of parasite surface antigens, we have isolated antigen-specific B cells from these donors. We then expressed the corresponding monoclonal antibodies in vitro. These antibodies were screened against a library of variant surface antigens to determine antibody breadth and potential to inhibit interaction of the parasite surface antigen with host receptors, a critical step in pathogenesis. Additionally, using a panel of variant surface antigen mutants, we have predicted the epitopes targeted by the broadest monoclonal antibodies. RESULTS/ANTICIPATED RESULTS: We have identified three monoclonal antibodies with exceptionally broad reactivity and inhibitory activity against our panel of severe disease-inducing variant surface antigens. We have identified two major sites targeted by these broadly reactive antibodies. The first site was associated with the largest breadth, but limited inhibitory potential, while the second site showed high-affinity antibody binding and inhibition of receptor binding. Interestingly, two of these three antibodies were very similar in structure, even though they were isolated from different donors. Isolation of antigen-specific B cells from additional donors will enable us to identify how common such broadly reactive antibodies are and allow the identification of additional epitopes DISCUSSION/SIGNIFICANCE OF FINDINGS: This study is the first to isolate broadly reactive antibodies that are likely to protect against severe malaria in naturally immune individuals. Further characterization of antibody-antigen interactions will inform the development of this surface antigen as a vaccine candidate for malaria.

OBJECTIVES/GOALS: Chronic or new symptoms after infection with severe-acute-respiratory-coronavirus-2 (SARS-CoV-2) has been termed post-acute sequelae of Covid-19 (PASC) or Long Covid. Our objective is to present results from COVID-OUT, a phase 3 double-blind, randomized controlled trial of early outpatient treatment of Covid-19 with repurposed medications. METHODS/STUDY POPULATION: COVID-OUT enrolled adults age 30 to 85 with overweight or obesity who had proof of SARS-CoV-2 infection and fewer than 7 days of symptoms. In this 2 by 3 factorial design trial of metformin, ivermectin, fluvoxamine, or exact-matching placebo of each medication, participants were randomized 1:1:1:1:1:1 to the 6 treatment allocations. This abstract focuses on whether early treatment with metformin prevented Long Covid. Immediate release metformin was titrated to 1500mg daily over the first 6 days. We assessed the incidence of clinician-diagnosed Long Covid with follow up through 10 months after enrollment. We also assessed where participants were diagnosed with Long Covid, and where they received Long Covid treatment. RESULTS/ANTICIPATED RESULTS: Of 1124 participants, 98 (8.7%) report having a healthcare provider make a diagnosis of long covid. By arm, 6.9% (39/564) of metformin participants report having a diagnosis for long covid as compared with 10.5% (59/560) of matched placebo controls. The absolute reduction attributable to metformin was 3.6% (95%CI, 0.3% to 7.0%; P=0.031) with a relative risk reduction of 34% (95%CI, 3% to 55%). The metformin cost per long covid case averted was $28 (95%CI, $15 to $306). 10-month follow-up data will be available at the time of presentation as well as an analysis of baseline factors associated with the development of Long-Covid, independent of treatment allocation in the trial. DISCUSSION/SIGNIFICANCE: Metformin reduced the incidence of clinician-diagnosed long covid by 34% in a double-blind randomized placebo-controlled trial, and previous research published in-vitro activity by metformin against SARS-CoV-2 and other RNA viruses. Further investigation of metformin as early treatment for SARS-CoV-2 is warranted.