United States

Background
Colorectal adenomas are precursors of colorectal cancer (CRC). Pro-inflammatory states associated with colorectal adenomas significantly predispose for colorectal tumorigenesis via upregulation of cyclooxygenase-2 (COX-2) and prostaglandin E2 (PGE2) synthesis. The PGE2/COX-2 axis allows cancer cells to progress and evade attack from immune cells via increasing immunosuppressive cell activity, disrupting antigen-presenting and T-cell functions, and interrupting innate leukocyte responses. The purpose of this study was to demonstrate that PGE2 can upregulate expression of PD-1, an immunosuppressive signaling receptor, on THP-1-derived macrophages and CD8+ T-cells. 

Methods
THP-1 cells were differentiated into macrophages via phorbol 12-myristate-13-acetate (PMA) administration. Following serum starvation and incubation with three different concentrations of PGE2 (0.1μM, 0.3μM, and 1μM), the cells were analyzed with western blot and qRT-PCR. CD8+ T-cells were isolated from mice spleens using biotin-antibody cocktail and anti-biotin microbeads and then stimulated with rIL-2 and CD3/CD28 Dynabeads. The cells were serum starved, incubated with two different concentrations of PGE2 (0.1μM and 0.3μM), and analyzed utilizing flow cytometry and qRT-PCR.

Results
Western blot analysis of THP-1-derived macrophages incubated in 0.1μM, 0.3μM, and 1.0μM PGE2 demonstrated increased PD-1 expression. qRT-PCR analysis of THP-1-derived macrophages incubated in 0.1μM, 0.3μM, and 1.0μM PGE2 demonstrated 3.030-fold, 5.712-fold, and 2.683-fold increased PD-1 mRNA expression, respectively. qRT-PCR of mouse CD8+ T-cells incubated in 0.1μM and 0.3μM PGE2 demonstrated 1.874-fold and 9.594-fold increased PD-1 mRNA expression, respectively. Flow cytometry demonstrated that 9.14% and 9.35% of mouse CD8+ T-cells incubated in 0.1μM and 0.3μM PGE2 expressed PD-1, respectively, while 4.65% of control cells expressed PD-1. 

Conclusion
Our in vitro data shows PGE2 inducing PD-1 expression in THP-1-derived macrophages and mouse CD8+ T-cells. Greater PD-1 expression would allow cancer cells to progress safe from the host immune system. These data demonstrate that PGE2 produced in the colorectal tumor microenvironment promotes immune evasion by suppressing tumor-infiltrating CD8+ T-cell cytotoxicity and macrophage phagocytosis via induction of PD-1. Because of PGE2’s immunosuppressive effects, NSAIDs and COXIBs are effective at preventing and treating CRC by inhibiting PGE2 synthesis via COX-2 blockade.

AMA Research Challenge 2022

PGE2 Promotes Colorectal Tumorigenesis via Increased PD-1 Expression on Macrophages and CD8+ T-Cells

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Charles Lopresto, DO

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Welcome to the largest national, multi-specialty research event featuring research from 750 medical students, residents and fellows, and international medical graduates.


technical paper

Background: Pulmonary Artery Pulsatility index (PAPi) is calculated by pulmonary artery pulse pressure divided by central venous pressure and is directly proportional to right ventricular contractility and stroke volume, and inversely related to pulmonary artery afterload and filling pressures. While PAPi as a prognostic hemodynamic measure has been studied in various adult populations, including to predict right ventricular (RV) failure following left ventricular assist device (LVAD) implantation and to predict renal outcomes following heart transplant (HTx), it has not been evaluated in children post HTx. We hypothesize that PAPi will correlate with known hemodynamic parameters and predict acute kidney injury (AKI) following pediatric HTx.

Methods: Medical records and hemodynamic data were retrospectively reviewed for patients ≤18 years old undergoing HTx at a single center between 1/2007-9/2021. Subgroups by HTx indication were myopathy or congenital heart disease (CHD). PAPi was calculated at routine cardiac catheterizations initially following HTx, and at 6 months, 1 year, and 5 years post HTx, with normal being PAPi ≥2.0. Correlations between PAPi and established hemodynamic measures collected at post HTx catheterizations including right ventricular end diastolic pressure (RVEDp), thermodilution cardiac index (TDCI), and central venous pressure to pulmonary capillary wedge pressure ratio (CVP-to-PCWP ratio) were analyzed using linear regressions. Correlation between PAPi and AKI was analyzed using a Fischer’s Exact test. 

Results: Fifty-three children underwent HTx in myopathy (n=27) or CHD (n=26) subgroups. PAPi correlated with RVEDp (p<0.001), TDCI (p=0.004), and CVP-to-PCWP ratio (p<0.001). At 6 months post HTx, PAPi <2.0 was associated with increased incidence of AKI (n=12/53, p=0.047). In analyzing 5-year trends, the myopathy subgroup had a normal early PAPi with no significant change at any time point. However, the CHD subgroup had a normal early (2.12 ± 0.18) and 6 month (2.45 ± 0.25) PAPi, but had a decline to an abnormal PAPi at 1 year (1.98 ± 0.13) and 5 years post HTx (1.67 ± 0.16). This decline between the 6 month and 5-year catheterizations was statistically significant (p=0.048).

Conclusions: PAPi in children post HTx correlates with established hemodynamic parameters routinely collected post HTx and may serve as a marker predicting renal dysfunction 6 months post HTx. Following HTx, PAPi declined significantly for the CHD subgroup. These findings suggest that incorporating PAPi into existing risk models may substantially improve earlier implementation for advanced therapies, thus improving clinical outcomes. PAPi as a comprehensive prognostic hemodynamic measure post pediatric HTx should be investigated in a larger multi-center study.

Pulmonary Artery Pulsatility Index After Pediatric Heart Transplant

Background
Rett Syndrome (RTT) is a pervasive neurodevelopmental disorder affecting females. RTT results from loss of function mutations in the X-linked gene Methyl CpG Binding Protein 2 (MeCP2*). MeCP2 encodes for the eponymously named protein, a ubiquitously expressed transcription factor critical for neuronal development and mature synapse maintenance. However, this pathogenesis is complicated by unequal mutation distribution; heterozygous mosaicism, one X-chromosome in each cell in lionized, results in a patchwork pattern of protein expression. Differences in the number, type, and location of MeCP2-deficient cells likely underlies the phenotype spectrum among the RTT patients. Whether some cells are more dependent on MeCP2, and therefore contribute more to clinical symptomatology or severity, is unknown. Here we explore that hypothesis in the context of impaired perceptive communication. 

Methods
We used Cre-lox genetic mouse lines to examine the consequences of cell-type restricted Mecp2 mutation to perception of naturally produced ultrasonic vocalizations (USVs). We employed infant pup retrieval behavior as a learned and ecologically relevant readout of auditory perception. Adult rodents naturally learn this behavior in response to pup USVs. We collected electrophysiology recordings and conducted calcium imaging to identify single neuron firing and neuronal population activity patterns in the auditory cortex during retrieval behavior and during passive playback of USVs. 

Results
Cortical cell type identity dictated MeCP2 dependence for supporting auditory processing. Mecp2 mutation restricted to parvalbumin positive (PV) inhibitory interneurons (IN) impaired pup retrieval behavior. However, mutation restricted to somatostatin- or vasoactive-intestinal-peptide- positive IN had no such effect. The inability of PV-Mecp2 mutants to response behaviorally to USVs resulted from degraded stimulus-driven neuronal activity in the auditory cortex, both at the single unit and population levels. That is, PV-restricted Mecp2 mutation impaired inhibitory cortical plasticity.

Conclusion
Our findings demonstrate a disproportionate dependence among cortical cell populations on MeCP2. This argues for cell-type specific pharmacological or genetic therapies, and a personalized medicine approach to the treatment of mosaic genetic disorders, including RTT. Moreover, these findings expand our understanding of neurotypical processes of audition and the contributions of various cortical subtype populations to those processes.

Rett Syndrome Mutation in Select Cortical Cell Populations Disrupts Auditory Processing

Background
Disease modifying antirheumatic drugs (DMARDs) are used to treat Rheumatoid Arthritis (RA).
The rates of total shoulder arthroplasty (TSA) in RA patients since the implementation of
DMARDs therapy is unknown. Furthermore, the utilization rates of TSA among various age
cohorts as well as by procedure type (anatomic (ATSA) and reverse (RTSA) total shoulder
arthroplasty) are not well defined in this patient population. The primary aim of this study was to
assess TSA trends from 2011-2019 for RA patients with and without filled DMARD
prescriptions.
Methods
A retrospective trends analysis utilizing a national claims database was performed investigating
the trends of TSA in RA patients with and without DMARD from 2011-2019. Patients were
identified using International Classification of Diseases codes and database-specific drug codes
were used to identify DMARD prescriptions. Patients were divided into two cohorts: RA patients
who underwent a TSA with or without DMARD prescription claims. Patients were also stratified
by age groups and procedure type for analysis. Linear regression and compound annual growth
rate (CAGR) were performed to compare trends of surgical utilization. Significant changes
between trends were determined using an interaction term between the trend and calendar year
within the regression. Statistical analysis were performed with Stata version 17.0.
 2018 American Medical Association. All rights reserved.
2
Results
From 2011-2019, incidence of TSA in RA patients without filled DMARD prescriptions
increased significantly more than RA patients with filled prescriptions (CAGR = 9.43% vs -
0.54%; linear regression P<0.001; quadratic P<0.001). Among the age groups, incidence of TSA
in RA patients with and without filled DMARD prescriptions significantly increased among all
age groups (P<0.001). The rate of change of TSA incidence in RA patients was significantly
higher increased in patients without filled DMARD prescription for all age groups compared to
RA patients with filled prescriptions (P<0.001). By procedure type, the incidence of RTSA were
significantly increasing while incidences of ATSA were significantly decreasing in both groups
(P<0.001).
Conclusion
Our findings show a greater increase in TSA surgeries among patients younger than 59 without
prescription claims suggesting that DMARD therapy may play a beneficial role in preventing or
delaying the need for TSA in RA patients.

Shoulder Arthroplasty Trends in Patients with Rheumatoid Arthritis Receiving DMARD Therapy

Background and Purpose: Deep burns and wounds remain difficult to treat effectively. Recent advances in tissue engineering and biotechnology have led to the advent of several skin substitutes (SS) including xenografts, bioscaffolds, and tissue-engineered dermal templates, though no product has been proven superior to current standards of care (SOC) for burns or wounds. We aim provide a systematic review and meta-analysis on clinical trials (CT) and randomized controlled trials (RCT) investigating the available skin substitute options for burn and wound treatment.

Methods: A systematic review of the PubMed, Scopus, and CINAHL databases was performed using PRISMA guidelines for CTs and RCTs published between 2017 and 2022. Articles were screened for relevance using Covidence®. Studies were then grouped by indication for use in burn or wound treatment. Meta-analysis was performed using RevMan v.5® on burn studies that assessed mean time to complete re-epithelialization and wound studies that assessed the proportion of wounds completely closed at 12 months. 

Results: 48 studies (39 RCT; 9 CT) were included and stratified by indication: burns (20 studies) or wounds (28 studies). For burns, 4 studies compared SS to SOC silver sulfadiazine cream as assessed by mean time to complete re-epithelialization (190 burns) and were included in meta-analyses. Fish skin xenografts (FSX) significantly outperformed SOC (p=0.0003) with burn depths ranging from superficial partial thickness to deep 2nd degree. For wounds, 13 studies compared SS to SOC alginate dressings assessed by the proportion of wounds completely healed at 12 weeks (2529 wounds) and were included in meta-analysis. Dehydrated human amnion and chorion allografts (dHACA; p=0.01) and human reticular acellular dermal matrices (HR-ADM, p<0.00001) all significantly outperformed SOC.

Conclusions: FSXs were superior to SOC in the treatment of burns and both dHACAs and HR-ADMs were superior to SOC for wound treatment.

State-of-the-art Skin Substitute and Scaffold Designs for Burns and Wounds: A Systematic Review and Meta-Analysis

Background: Despite the significant advances in detecting pulmonary nodules, the management plan of small tumors with malignant potential remains controversial. The objective of this study was to evaluate the overall survival of patients with ≤8 mm non-small cell lung cancer (NSCLC) who undergo stereotactic body radiation therapy (SBRT) versus wedge resection.

Methods: Kaplan-Meier analysis, multivariable Cox proportional hazards modeling, and propensity score-matched analysis were performed to evaluate the overall survival of patients with ≤8 mm NSCLC in the National Cancer Database (NCDB) from 2004-2017 who underwent SBRT versus patients who underwent wedge resection. The above-mentioned matched analyses were repeated for patients with no comorbidities. Patients who were coded in the NCDB as having undergone radiation because surgery was contraindicated due to patient risk factors (e.g., comorbid conditions, advance age, etc.) and those with a history of prior malignancy were excluded from analysis.

Results: Of the 1,505 patients who had NSCLC ≤8 mm during the study period, 1,339 (89%) patients underwent wedge resection, and 166 (11%) patients underwent SBRT. In both unadjusted analysis and in multivariable Cox modeling and propensity score-matched analysis, wedge resection was associated with improved survival when compared to SBRT. These results were consistent in a sensitivity analysis limited to patients with no comorbidities.

Conclusion: In this national analysis, patients with NSCLC ≤8 mm undergoing wedge resection experienced improved survival when compared to SBRT. This finding can be used to help inform the treatment decision-making process for tumors that are being discovered incidentally or through lung cancer screening. 





Stereotactic Body Radiation Therapy Versus Wedge Resection for Early-Stage Node-Negative Non-Small Cell Lung Cancer Tumors ≤8 mm: A National Analysis

Hannah Kelly, Grant Fabrizio, Lauren Yap, Isela Melendez-Carpio, Susan M. Cheng, H. Carrie Chen

Diversity in medical education is imperative for future physicians as they prepare to treat heterogeneous patient populations. For aspiring physicians, mentorship plays an important role in navigating the pre-medical journey. Currently, gaps in medical mentorship are monopolized by costly, third-party “admission consulting” agencies. Inaccessibility to these services disproportionately impacts underrepresented minority and first-generation students. We addressed this gap by implementing and evaluating an extension of Georgetown’s Students Helping Aspiring Physicians Excel (SHAPE) initiative, housed within the Office of Diversity, Equity and Inclusion, called SHAPE Summer Mentors program. This free, 7-week mentorship experience includes an individualized mentor/mentee matching process, an interactive weekly curriculum addressing common secondary application essay themes, and consistent quality time and support from trained first-year medical student mentors.

Program participants from the 2021-2022 AMCAS application cycle answered pre- and post- program surveys about their background and experiences regarding stress, confidence, and support.  Sixteen questions used a 5-point likert scale. Data analysis included descriptive statistics and two-tailed t-test for independent means.

One hundred twenty-one pre-medical mentees participated in the program; 79 (65.3%) completed the pre-program survey and 76 (62.8%) completed the post-program survey. Of those participating, 30 (24.8%) identified as underrepresented in medicine, 62 (51.2%) as first- or second-generation immigrants, and 38 (31.4%) as first-generation college students in the United States. Responses from the pre- to post-program surveys indicated that after going through the SHAPE Summer Mentors program, participants felt more supported in their medical school application process (4.08 to 4.33, p=0.0383) and less stressed about applying (4.46 to 4.08, p=0.0098). Participants also reported improved confidence in their ability to write secondary essays (3.28 to 3.92, p<0.0001) and improved comfort with interviews (3.23 to 4.04, p=0.0027). Additionally, there was a notable trend in which participants had increased confidence that they will be accepted to medical school (2.99 to 3.29, p=0.0639). Upon program completion, 100% of participants felt more prepared for the application cycle.

Preliminary evaluation results of the SHAPE Summer Mentors program indicate that the program is successful in supporting participants and improving their confidence through the medical school application process. The SHAPE Summer Mentors program’s student-led curriculum offers an innovative solution to reduce barriers and increase access for pre-medical students from a variety of backgrounds, including underrepresented minority and first-generation students in medicine.

Students Helping Aspiring Physicians Excel (SHAPE) Summer Mentors: A Novel Peer-to-Peer Mentorship Program for Medical School Admissions

Background
Patients diagnosed with breast cancer at our safety-net hospital (SNH) are significantly more likely to be diagnosed at later stages, and this late-stage-at-diagnosis is most highly associated with lack of screening mammogram. We showed that a two-part intervention - phone call and assistance with scheduling - significantly increased mammography uptake by 12% among our safety-net population. Implementing this finding system-wide proved challenging and resource heavy. Prior studies show success with text message reminders in low-income populations, but none evaluate the utility of scheduling by text. Here, we seek to determine whether a text-based intervention with reminder and scheduling could increase mammography uptake.
Methods
A randomized controlled study included 843 women aged 50-65 who had not had a screening or diagnostic mammogram in the past two years, but had established care at a primary care clinic (PCC) within our SNH. One-third of the participants were assigned to each of the following groups: Intervention 1 (text reminder with specific scheduling options), Intervention 2 (text reminder with open-ended scheduling options) and usual-care control. Participants in both intervention groups could engage in two-way texting, and up to three texts were sent to each patient. Differences in percent mammography uptake at 3 months were compared between intervention and control groups using a two-tailed chi-square.
Results
Patients receiving a text-based reminder and scheduling opportunity were significantly more likely to get mammograms within 3 months than those in the usual care control group (10.2% and 6.2%, respectively; chi2 = 5.6279, p < 0.017). 14.8% of participants responded to text message. 9% of participants scheduled an appointment for mammogram via text, of which 63% received mammogram. Mammography compliance did not differ by type of scheduling offered (specific vs. general) or by PCC.
Conclusion
Reminders and scheduling through two-way text-messaging is effective in increasing mammography uptake among primary care clinic patients in an urban safety-net setting. Scheduling component was a key feature in the success of the intervention. The safety-net is a diverse and hard-to-reach population. Future studies should combine different types of interventions to design a multi-pronged approach that is tailored to this population with unique needs.

Text-based Intervention Increases Mammography Uptake among Overdue Patients at an Urban, Safety-Net Hospital: A Randomized Controlled Study

Background
Neuroblastoma is the most common extracranial childhood tumor. Despite intensive multi-modality therapies, approximately 50% of high-risk patients die from progressive disease. Although retinoic acid-based therapies provide clinical benefit, patients eventually relapse due to resistance, underscoring the critical need for novel treatments. Amplification of MYCN, an oncogene present in 40-50% of high-risk neuroblastoma cases, is associated with poor patient prognosis and poor response to retinoid therapies. However, MycN is “undruggable,” prompting efforts to indirectly impair its expression or function. The aryl hydrocarbon receptor (AhR) is a transcription factor whose role in neuroblastoma remains incompletely characterized. Here, we investigate AhR as a novel tumor promoter that regulates MycN and retinoic acid pathways in neuroblastoma. 

Methods
Human neuroblastoma cells were modified to over- or under-express AhR, or were treated pharmacologically with the AhR antagonist, clofazimine (CLF). To assess tumorigenicity, invasion, clonogenicity, and tumor growth were measured. For MycN regulation studies, AhR and MycN protein expression were detected by immunoblotting. Cells were treated with retinoic acid after AhR depletion or inhibition and examined by microscopy. 

Results
We found AhR depletion decreases clonogenicity and tumor growth in vivo. Notably, treatment with clofazimine (CLF), an FDA-approved anti-leprosy drug we previously identified to be a novel AhR antagonist, has the same effect. CLF is orally bioavailable, safe, cost-effective, and approved for use in children, representing a promising new therapy for neuroblastoma patients. Our work further reveals that AhR depletion and inhibition reduces MycN expression and induces differentiation. Finally, we found AhR depletion and inhibition potentiates retinoid-induced neuroblastoma differentiation. 

Conclusion
Our work uncovers a novel role for AhR as a tumor promoter in neuroblastoma and could lead to an already FDA-approved drug, CLF, as a promising new therapy for neuroblastoma patients that improves efficacy of retinoic acid-based treatments. 


The Aryl Hydrocarbon Receptor (AhR) as a Novel Therapeutic Target in Neuroblastoma

Immigrant Haitian women in New York City (NYC) have the highest rate of Severe Maternal Morbidity of 494.0 per 10,000 deliveries, compared to 229.8 for U.S.-born women. There is little study into the needs and barriers of the Haitian immigrant community, which is one of the largest growing immigrant populations in the United States today. We sought to understand pregnant and postpartum Haitian women’s barriers to healthcare services in NYC and explore ways to improve their overall health outcomes.

The Barriers to Healthcare Pregnant and Postpartum Haitian Women Experience in New York City

Background 
Patients with severe aplastic anemia (SAA) develop high-risk infections. Prior data suggest efficacy and safety of granulocyte transfusion (GT) as an infection management tool in this population. A multicenter randomized control trial of GT in patients with severe neutropenia of varying etiologies did not reveal a significant difference in overall survival (OS) or microbial response but was limited by low enrollment and variable granulocyte dose. In this study of SAA patients who received GTs at our institution, we characterize patients’ clinical and survival outcomes and the role of GTs as a bridge to curative therapy. 

Methods 
SAA patients treated with immunosuppressive therapy who received GTs between 2011 and 2020 are included in our analysis. The primary outcomes are survival from first GT, including OS, and number of patients surviving to discharge and HSCT in patients awaiting HSCT. Secondary outcomes include evaluation of response at 30 days after initiation of GTs based on clinical, microbiological, and radiographical criteria. Cox proportional-hazards model was utilized to determine impact of neutrophil response and infection type on survival. Kaplan-Meier was used to assess OS. 

Results 
Twenty-eight SAA patients of median age 20 years (range 6-65 years) underwent 30 GT courses of median 8 products per course (range 1-39 products), over median 23.5 days (range 3-103 days), with a per-dose median 1.28 x 10^9 granulocyte cells/kg (range 0.45-4.52 x 10^9). Infection type included bacterial (n=15), fungal (n=12), and mixed (n=3). OS from initial GT was 50% with a median length of follow up at 550.5 days (range 2-4213 days). 18 of 28 (64.3%) patients survived to hospital discharge. Of the 21 patients awaiting HSCT during granulocyte therapy, 18 (85.7%) received HSCT. Patients achieving complete, partial or stable response at 30 days had significantly improved OS compared to non-responders (p=0.00004). Sex, type of infection, or % of days with ANC >200 cells/mcL during the granulocyte course, were not predictive of survival (p=0.52, p=0.7, p=0.28 respectively). 

Conclusion 
GTs of adequate dose and course duration are a valuable adjunctive therapy to manage severe infections in SAA patients and may be particularly valuable as a bridge to undergoing curative treatment with HSCT.

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PGE2 Promotes Colorectal Tumorigenesis via Increased PD-1 Expression on Macrophages and CD8+ T-Cells

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