Citations

Abstract While chimeric antigen receptor T-cell therapy targeting CD19 (CART19) has shown remarkable success in the treatment of hematological malignancies, the durable response rates remain approximately 40% and there are limited solutions for CART cell therapy in the treatment of solid tumors. To further understand mechanisms of resistance, including CART cell exhaustion, we employed three independent approaches: 1) RNA and ATAC sequencing on unstimulated vs. exhausted healthy donor CART19 cells by utilizing an in vitro model for exhaustion, 2) RNA and ATAC sequencing on pre-infusion CART19 cell products from responders and non-responders in the Zuma-1 clinical trial that led to the FDA approval of axi-cel CART19 therapy, and 3) a genome-wide CRISPR knockout screen in healthy donor CART19 cells using our in vitro model for exhaustion. In each of these approaches, IL-4 was identified as a regulator of CART cell dysfunction. In approach 1, ingenuity pathway analysis of genes that were both differentially accessible and expressed in exhausted compared with unstimulated CART19 cells revealed IL-4 as a top upstream regulator (p = 5E-6). In approach 2, IL-4 was one of two genes that were both upregulated and more accessible in CART19 cell products from non-responders (p < 5E-2). Finally, in approach 3, gene ontology enrichment analysis of genes that were positively selected during the genome-wide CRISPR knockout screen, revealed regulation of the IL-4 pathway as one of the top affected pathways (p = 1E-4). Together, our data indicates a role for IL-4 in CART cell dysfunction caused by exhaustion. Investigating this mechanism further, we saw an increase in the production of IL-4 as CART cells became exhausted (p = 4E-3). Treatment of CART19 cells with human recombinant IL-4 (hrIL-4) resulted in dysfunction as evident by a decrease in antigen specific cytotoxicity (p = 4E-3) and proliferative ability (p= 6.5E-2), as well as exhaustion-specific signs of dysfunction such as an increase in the expression of the inhibitory receptor, TIM-3 (p = 3E-3) and an increase in the transcription of the exhaustion-related transcription factor EOMES (p = 1E-2). Finally, we tested whether IL-4 neutralization enhances CART19 cell functions. Using a CD19+ JeKo-1 xenograft mouse model, we compared the combination treatment of CART19 cells and an IL-4 neutralizing monoclonal antibody (10 mg/Kg, clone # MP4-25D2) to CART19 cells and an IgG control. IL-4 neutralization in combination with CART19 cells resulted in reduced tumor burden (p = 4.6E-2), increased CART cell proliferation (p = 8E-3), and prolonged overall survival (p= 5E-2). In summary, our data indicates that 1) IL-4 induces CART cell dysfunction through a state of exhaustion and 2) IL-4 neutralization with a monoclonal antibody enhances CART cell therapy. As such, this novel combination therapy holds the potential to be translated to the clinic to improve durable responses from CART cell therapy. Citation Format: Carli Stewart, Michelle J. Cox, Reona Sakemura, Ekene J. Ogbodo, Ismail Can, Claudia Manriquez Roman, Kun Yun, Olivia Sirpilla, James H. Girsch, Truc Huynh, Elizabeth L. Siegler, Jenny J. Kim, Mike Mattie, Nathalie Scholler, Simone Filosto, Saad S. Kenderian. IL-4 depletion leads to the improvement of CART cell therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1153.

Abstract Thyroid cancer is the most common endocrine cancer in the US, and its incidence is rising. Most thyroid cancer deaths are attributed to treatment-refractory, metastatic tumors. Thyroid stimulating hormone receptor (TSRH) expression is largely limited to the thyroid gland and is abundantly expressed on thyroid tumor cells, making TSRH a compelling target for advanced thyroid cancer diagnostics and therapeutics. Therefore, we developed a novel TSHR-targeted chimeric antigen receptor (CAR) T cell therapy to treat aggressive thyroid cancers. TSHR-CAR constructs were cloned into a lentiviral CAR construct containing 4-1BB and CD3ζ. First, we demonstrated potent TSHR-CART antigen-specific anti-tumor activity in vitro. Then, NOD-SCID-γ-/- (NSG) mice were inoculated subcutaneously with TSHR+ tumor cells and randomized by tumor volume to treatment with TSHR-CART cells or control Untransduced T cells (UTD). Treatment with TSHR-CART cells resulted in dose-dependent antitumor activity and prolonged survival. De-differentiated anaplastic thyroid cancers (ATC) downregulate TSHR. Our TSHR immunohistochemistry results corroborated these findings and displayed minimal TSHR protein expression, precluding successful TSHR-CART treatment. We therefore sought to sensitize these tumors with MAPK inhibitors, as a strategy to upregulate TSHR expression in patients with metastatic thyroid cancer. TSHR expression was upregulated in patient-derived xenograft (PDX) ATC models after one week of daily administration of the MAPK inhibitors (p=0.0024). After confirming that MAPK inhibition does not dampen TSHR-CART effector functions, we tested sequential and combination therapy of TSHR-CART with MEK and BRAF inhibition in vivo. NSG mice were engrafted with ATC BRAF-mutant PDX tumors and randomized by tumor volume to daily oral treatment with placebo or trametinib (MEK inhibitor) plus dabrafenib (BRAF inhibitor). One week later, mice received either UTD or TSHR-CART. Mice conditioned with trametinib plus dabrafenib (p=0.0018) and subsequently treated with TSHR-CART showed superior antitumor activity. However, the improved antitumor activity in this setting was transient. We therefore tested the durability of TSHR upregulation following MEK/BRAF inhibition and demonstrated that TSHR upregulation lasts less than 48-72 hours after discontinuation. Finally, we tested the combination of TSHR CART cells with MEK/BRAF inhibitors in ATC BRAF-mutant PDX tumors. Here, combining TSHR-CART cells with MEK/BRAF inhibitors result in durable control of the tumors. Collectively, our findings indicate that MEK/BRAF inhibition of de-differentiated thyroid cancers upregulated TSHR expression and enhanced TSHR-CART antitumor activity. This work represents a viable strategy to improve outcomes of patients with aggressive, metastatic thyroid cancers. Citation Format: Claudia Manriquez Roman, Kendall J. Schick, Justyna J. Gleba, Truc N. Huynh, Elizabeth L. Siegler, James L. Miller, Aylin Alasonyalilar Demirer, Matthew L. Pawlush, Ahmet Biligili, Long K. Mai, Erin Tapper, Leo R. Sakemura, Michelle J. Cox, Carli M. Stewart, Ismail Can, Ekene J. Ogbodo, Gaofeng Cui, Georges Mer, Gloria R. Olivier, Yushi Qiu, Robert C. Smallridge, Zubair C. Abba, Han W. Tun, John A. Copland, Saad S. Kenderian. Addition of MAPK inhibitors to prime and sensitize poorly differentiated thyroid cancers as a strategy to improve TSHR-CART cell therapy antitumor activity. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5074.

Background. Normothermic regional perfusion (NRP) has gained widespread adoption in multiple European countries. The aim of this study was to examine the influence of thoracoabdominal-NRP (TA-NRP) on the utilization and outcomes of liver, kidney, and pancreas transplantation in the United States. Methods. Using the US national registry data between 2020 and 2021, donation after circulatory death (DCD) donors were separated into 2 groups: DCD with TA-NRP and without TA-NRP. There were 5234 DCD donors; among them 34 donors were with TA-NRP. After 1:4 propensity score matching, the utilization rates were compared between DCD with and without TA-NRP. Results. Although the utilization rates of kidney and pancreas were comparable ( P = 0.71 and P = 0.06, 94.1% versus 95.6% and 8.8% versus 2.2%, respectively), that of liver in DCD with TA-NRP was significantly higher ( P < 0.001; 70.6% versus 39.0%). Among 24 liver transplantations, 62 kidney transplantations, and 3 pancreas transplantations from DCD with TA-NRP, there were 2 liver grafts and 1 kidney graft that failed within 1 y after transplantation. Conclusions. TA-NRP in the United States significantly increased the utilization rate of abdominal organs from DCD donors with comparable outcomes after transplantation. Increasing use of NRP may expand the donor pool without compromising transplant outcomes.

Introduction: Medical education is trending towards an outcome-based curriculum that prepares medical graduates to excel in a rapidly evolving, team-centred healthcare delivery system. The Mayo Medical School (MMS) has recently redesigned its curriculum to introduce early clinical relevance, optimise course integration, provide flexibility and promote active learning. This study aims to evaluate the role that curricular flexibility plays in students’ perceived learning, satisfaction and performance. Materials and Methods: First-year students completed a 5-point Likert scale survey regarding satisfaction with and perceived learning from various components of the flexible curriculum during 2 different academic blocks. Students’ academic performance was assessed by a National Board of Medical Examiners (NBME) Subject Examination after each block. Results: In comparing student-rated satisfaction and perceived learning from didactics versus other curricular components for Block 3 (Pathology and Immunology) and Block 5 (Gross Anatomy and Radiology), students rated didactics higher in all cases in which there was a statistically significant difference in the ratings. There was a statistically significant positive correlation between the amount learned and satisfaction for all curricular components with the exception of Block 5 independent study. During Block 3, only interest in the subject matter correlated positively with the NBME score, while during Block 5, only time spent in class correlated negatively with the NBME score. Conclusions: Although various components of the flexible curriculum do not appear to affect satisfaction and perceived learning, their potential influence on graduate outcomes, in terms of delivering healthcare providers who are patient-centred, creative thinkers and compassionate leaders should not be discounted. Key words: Curriculum reform, Medical curriculum, Outcomes-based education, Student satisfaction

Abstract Postmortem computed tomography (PMCT) has been integrated into the practice of many forensic pathologists. To evaluate the utility of PMCT in supplementing and/or supplanting medicolegal autopsy, we conducted a prospective double‐blind comparison of abnormal findings reported by the autopsy pathologist with those reported by a radiologist reviewing the PMCT. We reviewed 890 cases: 167 with blunt force injury (BFI), 63 with pediatric trauma (under 5 years), 203 firearm injuries, and 457 drug poisoning deaths. Autopsy and radiology reports were coded using the Abbreviated Injury Scale and abnormal findings and cause of death (COD) were compared for congruence in consensus conferences with novel pathologists and radiologists. Overall sensitivity for recognizing abnormal findings was 71% for PMCT and 74.6% for autopsy. Sensitivities for PMCT/autopsy were 74%/73.1% for BFI, 61.5%/71.4% for pediatric trauma, 84.9%/83.7% for firearm injuries, and 56.5%/66.4% for drug poisoning deaths. COD assigned by reviewing PMCT/autopsy was correct in 88%/95.8% of BFI cases, 99%/99.5% of firearm fatalities, 82.5%/98.5% of pediatric trauma deaths, and 84%/100% of drug poisoning deaths of individuals younger than 50. Both autopsy and PMCT were imperfect in recognizing injuries. However, both methods identified the most important findings and are sufficient to establish COD in cases of BFI, pediatric trauma, firearm injuries and drug poisoning in individuals younger than 50. Ideally, all forensic pathologists would have access to a CT scanner and a consulting radiologist. This would allow a flexible approach that meets the diagnostic needs of each case and best serves decedents' families and other stakeholders.

Background: The cortical silent period (CSP) and long-interval intracortical inhibition (LICI) are putative markers of γ-aminobutyric acid receptor type B (GABAB)-mediated inhibitory neurotransmission. We aimed to assess the association between LICI and CSP in youths. Methods: We analyzed data from three previous studies of youth who underwent CSP and LICI measurements with transcranial magnetic stimulation and electromyography. We assessed CSP and LICI association using Spearman rank correlation tests and multiple linear regression analyses adjusted for demographic and clinical covariates. Results: The sample included 16 healthy participants and 45 participants with depression. The general mean (SD) age was 15.5 (1.7), 14.3 (1.7) for healthy participants, and 15.9 (1.6) years for participants with depression. Measures were nonnormally distributed (Shapiro–Wilk, p < 0.001). CSP and LICI were not correlated at 100-millisecond (ρ = −0.2421, p = 0.06), 150-millisecond (ρ = −0.1612, p = 0.21), or 200-millisecond (ρ = −0.0507, p = 0.70) interstimulus intervals using Spearman rank correlation test. No correlations were found in the multiple regression analysis (p = 0.35). Conclusions: Although previous studies suggest that cortical silent period and long-interval intracortical inhibition measure GABAB receptor-mediated activity, these biomarkers were not associated in our sample of youths. Future studies should focus on the specific physiologic and pharmacodynamic properties assessed by CSP and LICI in younger populations.