Jang, Jin Sung

ABSTRACT Objective Fetal surgery (FS) for spina bifida (SB) repair improves infant outcomes; however, spontaneous preterm birth (sPTB) is common. We aimed to evaluate maternal and fetal immune responses following FS for SB repair. Method Pregnant participants undergoing fetoscopic SB repair at 24–26 weeks of gestational age (GA) were prospectively recruited and compared to GA matched low‐risk controls. Blood samples were collected at baseline, 2‐week post‐surgery (POD14), and delivery. Cytokine analysis, flow cytometry, and T cell receptor (TCR) sequencing were performed. Results sPTB occurred in 5/6 of the FS group (0/6 of controls). Maternal blood in the FS group demonstrated increased alpha‐fetoprotein (AFP) and anti‐inflammatory IL‐1RA at baseline and POD14, increased pro‐inflammatory IL‐8 only post‐operatively, and elevated growth factors throughout. T cell phenotypes following FS revealed reduced TH17 frequency but increased activation receptors on cytotoxic T cells. At delivery, the FS cohort had reduced Th2 and Th17 helper T cells and CD4+ memory T cells, but CD8 T cells still showed increased activation. No TCR expansion was detected post‐surgery, suggesting antigen‐independent activation. Cord blood in FS repaired neonates had increased AFP, IL‐8, IL‐1RA. Conclusion Systemic immune changes following FS, particularly T cell maturation and activation, were observed. Further characterization of these mechanisms may help predict and prevent surgery‐induced PTB, improving outcomes.

One of the signicant obstacles to donor heart utilization is that increasingly prolonged cold ischemic times are associated with worsened short- and long-term outcomes. As a result, most transplant centers aim to limit cold ischemic times to less than four hours. The TransMedics Organ Care System (OCS) (Trans Medics, Andover MA) is a portable ex vivo perfusion system designed to store a donor heart in a beating, metabolically active state, until it reaches the recipient, thereby allowing extended time outside the body while minimizing the effects of cold storage. We present a case of a heart transplant recipient supported with the Impella 5.5 as a bridge to transplantation who received a donation after circulatory death (DCD) heart supported by OCS. To our knowledge, this is the rst report of such a case.

ObjectiveTo identify hallmark genes and biomolecular processes in aortitis using high‐throughput gene expression profiling, and to provide a range of potentially new drug targets (genes) and therapeutics from a pharmacogenomic network analysis.MethodsBulk RNA sequencing was performed on surgically resected ascending aortic tissues from inflammatory aneurysms (giant cell arteritis [GCA] with or without polymyalgia rheumatica, n = 8; clinically isolated aortitis [CIA], n = 17) and noninflammatory aneurysms (n = 25) undergoing surgical aortic repair. Differentially expressed genes (DEGs) between the 2 patient groups were identified while controlling for clinical covariates. A protein–protein interaction model, drug–gene target information, and the DEGs were used to construct a pharmacogenomic network for identifying promising drug targets and potentially new treatment strategies in aortitis.ResultsOverall, tissue gene expression patterns were the most associated with disease state than with any other clinical characteristic. We identified 159 and 93 genes that were significantly up‐regulated and down‐regulated, respectively, in inflammatory aortic aneurysms compared to noninflammatory aortic aneurysms. We found that the up‐regulated genes were enriched in immune‐related functions, whereas the down‐regulated genes were enriched in neuronal processes. Notably, gene expression profiles of inflammatory aortic aneurysms from patients with GCA were no different than those from patients with CIA. Finally, our pharmacogenomic network analysis identified genes that could potentially be targeted by immunosuppressive drugs currently approved for other inflammatory diseases.ConclusionWe performed the first global transcriptomics analysis in inflammatory aortic aneurysms from surgically resected aortic tissues. We identified signature genes and biomolecular processes, while finding that CIA may be a limited presentation of GCA. Moreover, our computational network analysis revealed potential novel strategies for pharmacologic interventions and suggests future biomarker discovery directions for the precise diagnosis and treatment of aortitis.

Abstract The human genome encodes large numbers of non-coding RNAs, including divergent antisense transcripts at transcription start sites (TSSs). However, molecular mechanisms by which divergent antisense transcription is regulated have not been detailed. Here, we report a novel ZWC complex composed of ZC3H4, WDR82 and CK2 that suppresses divergent antisense transcription. The ZWC complex preferentially localizes at TSSs of active genes through direct interactions of ZC3H4 and WDR82 subunits with the S5p RNAPII C-terminal domain. ZC3H4 depletion leads to increased divergent antisense transcription, especially at genes that naturally produce divergent antisense transcripts. We further demonstrate that the ZWC complex phosphorylates the previously uncharacterized N-terminal acidic domain of SPT5, a subunit of the transcription-elongation factor DSIF, and that this phosphorylation is responsible for suppressing divergent antisense transcription. Our study provides evidence that the newly identified ZWC-DSIF axis regulates the direction of transcription during the transition from early to productive elongation.

Abstract Introduction: Multiple gene expression profile assays have successfully prognosticated the overall survival (OS) outcomes of patients with Newly Diagnosed Multiple Myeloma (NDMM) and are now available for use in clinical practice. The genes included in these profiles encompass several aspects of cancer biology broadly such as cell-cycle biology, energy metabolism, migration, adhesion, immune evasion etc. Since altered energy metabolism is one of the hallmarks of all cancer cells, our goal was to take a deep-dive into specific genes associated with intracellular central carbon energy metabolism (CCEM) pathways such as oxidative phosphorylation (OxPhos) as well as glycolysis and lactate fermentation (Glyco-Lac) in MM cells to determine their impact on the pathogenesis and prognosis of MM patients. Methods: Gene expression level data via RNA sequencing from the COMMPASS dataset (made available by the Multiple Myeloma Research Foundation (MMRF)) was utilized. The expression levels of all genes associated with the OxPhos (N = 118) and Glyco-Lac (N = 34) pathways were evaluated in CD138+ cells derived from patients with NDMM. Each gene was dichotomized into low and high expression groups -based on their expression level above and below the median fragments per kilobase of transcript per million (FPKM), respectively. The prognostic significance of dichotomizing each of the 152 genes in this NDMM cohort was evaluated by levels of their hazard ratios (HRs) for OS determined by the log-rank method. Genes from the OxPhos and Glyco-Lac pathways were selected based on their significant negative impact on OS, wide representation of their biologic activity throughout the pathway and high independence from each other. This resulted in the selection of 12 distinct genes of interest. A scoring system consisting of assigning a point for every gene where their FPKM was above the median yielded a minimum of 0 and a maximum of 12 for the set of genes in the OxPhos and Glyco-Lac pathways to create an energy metabolism score (EMS). Patients were then categorized into "low" (0-8) or "high" (9-12) EMS groups. In addition, patients were designated as having high-risk cytogenetics if they had any of the following: t(4;14), t(14;16), t(14;20), del17p, and Amp 1q. Survival analysis was performed using the Kaplan-Meier survival analysis and compared via the log-rank method. Finally, the distribution of the EMS score among patients within a spectrum of plasma cell disorders such as, monoclonal gammopathy of undetermined significance (MGUS), smoldering multiple myeloma and MM as well as normal plasma cells was assessed using a previously published transcriptome dataset (GSE6477). Results: A total of 766 patients were included in this study. The selected genes included in this scoring system and their biological relevance in the OxPhos and Glyco-Lac pathways are depicted in Figure A. The distribution of low and high EMS groups across ISS stage and cytogenetic risk are depicted in Figure B and C respectively. The median OS (mOS) and PFS for the high (N = 248) vs. low (N = 518) EMS groups was 55 vs. 94 months (HR: 2.41; (95%CI: 1.85-3.14); P <0.0001) (Figure D) and 24 vs. 42 months (HR: 1.74; (95%CI: 1.42-2.13); P <0.0001). Patients with ISS 2 stage disease categorized as having a high or low EMS score had a similar mOS to patients with ISS 3 and ISS 1 stage disease respectively (Figure E). Similarly, the mOS was similar among patients with standard risk and high-risk cytogenetics when categorized by low and high EMS (Figure F). Finally, the relative expression of most of the 12 select CCEM genes (Figure G) and the probability of a high EMS score increased among patients along the spectrum of plasma cell disorders (Normal plasma cell -> MGUS -> SMM -> MM) (Figure H). Conclusions: The expression levels of specific genes associated with the activity of intracellular CCEM pathways such as glycolysis, lactate fermentation and oxidative phosphorylation in plasma cells provides evidence for the impact of cellular energy metabolism on the pathogenesis and survival outcomes of MM. This provides an opportunity for developing future studies aimed at understanding the energy metabolism pathways of plasma cell malignancies in greater detail so as to exploit them for diagnostic and therapeutic purposes in MM. Figure 1 Figure 1. Disclosures Kumar: Novartis: Research Funding; Amgen: Consultancy, Research Funding; Beigene: Consultancy; Carsgen: Research Funding; Merck: Research Funding; Antengene: Consultancy, Honoraria; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Astra-Zeneca: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Oncopeptides: Consultancy; Roche-Genentech: Consultancy, Research Funding; KITE: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Tenebio: Research Funding; Bluebird Bio: Consultancy; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Consultancy, Research Funding; Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Adaptive: Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Research Funding. Dispenzieri: Alnylam: Research Funding; Sorrento Therapeutics: Consultancy; Oncopeptides: Consultancy; Pfizer: Research Funding; Takeda: Research Funding; Janssen: Consultancy, Research Funding. Kapoor: AbbVie: Research Funding; Takeda: Research Funding; Karyopharm: Consultancy; Cellectar: Consultancy; BeiGene: Consultancy; Pharmacyclics: Consultancy; Sanofi: Consultancy; Amgen: Research Funding; Ichnos Sciences: Research Funding; Regeneron Pharmaceuticals: Research Funding; Glaxo SmithKline: Research Funding; Karyopharm: Research Funding; Sanofi: Research Funding. Dingli: Apellis: Consultancy; Sanofi: Consultancy; Janssen: Consultancy; GSK: Consultancy; Novartis: Research Funding; Alexion: Consultancy. Lin: Gamida Cell: Consultancy; Takeda: Research Funding; Kite, a Gilead Company: Consultancy, Research Funding; Juno: Consultancy; Novartis: Consultancy; Legend: Consultancy; Janssen: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Bluebird Bio: Consultancy, Research Funding; Sorrento: Consultancy; Merck: Research Funding; Vineti: Consultancy. Gertz: Akcea Therapeutics, Ambry Genetics, Amgen Inc, Celgene Corporation, Janssen Biotech Inc, Karyopharm Therapeutics, Pfizer Inc (to Institution), Sanofi Genzyme: Honoraria; Aurora Biopharma: Other: Stock option; AbbVie Inc, Celgene Corporation: Other: Data Safetly & Monitoring; Ionis Pharmaceuticals: Other: Advisory Board; Akcea Therapeutics, Alnylam Pharmaceuticals Inc, Prothena: Consultancy. Fonseca: Aduro: Consultancy; Kite: Consultancy; Juno: Consultancy; Merck: Consultancy; Takeda: Consultancy; Caris Life Sciences: Membership on an entity's Board of Directors or advisory committees; Patent: Prognosticaton of myeloma via FISH: Patents & Royalties; AbbVie: Consultancy; Scientific Advisory Board: Adaptive Biotechnologies: Membership on an entity's Board of Directors or advisory committees; OncoTracker: Consultancy, Membership on an entity's Board of Directors or advisory committees; Mayo Clinic in Arizona: Current Employment; Amgen: Consultancy; BMS: Consultancy; Celgene: Consultancy; Bayer: Consultancy; Janssen: Consultancy; Novartis: Consultancy; Pharmacyclics: Consultancy; Sanofi: Consultancy; GSK: Consultancy.

Abstract BackgroundOverexpression of c-Myc is required for the progression of pre-malignant plasma cells in monoclonal gammopathy of undetermined significance (MGUS) to malignant plasma cells in multiple myeloma (MM). c-Myc also increases glutamine anaplerosis into the tricarboxylic acid (TCA) cycle within cancer cells. Whether increased glutamine anaplerosis is associated with the progression of pre-malignant to malignant plasma cells is unknown.MethodsHuman volunteers (N = 7) and patients with MGUS (N = 11) and MM (N = 12) were prospectively recruited to undergo an intravenous infusion of 13C-labelled glutamine followed by a bone marrow aspiration to obtain bone marrow cells and plasma.ResultsDespite notable heterogeneity, stable isotope resolved metabolomics (SIRM) revealed that the mean 13C-labelled glutamine anaplerosis into the TCA cycle was higher in malignant compared to pre-malignant bone marrow plasma cells relative to the remainder of their paired bone marrow mononuclear cells. RNA sequencing demonstrated a higher relative mRNA expression of c-Myc and glutamine transporters such as ASCT2 and SN2 in malignant compared to pre-malignant bone marrow plasma cells. Finally, higher quantitative levels of TCA cycle intermediates in the bone marrow plasma differentiated MM from MGUS patients.ConclusionMeasurement of the in vivo activity of glutamine anaplerosis into the TCA cycle provides novel insight into the metabolic changes associated with the transformation of pre-malignant plasma cells in MGUS to malignant plasma cells in MM.

Abstract Problem Prenatal exposure to metabolic dysregulation arising from maternal obesity can have negative health consequences in post‐natal life. To date, the specific effects of maternal obesity on fetal immunity at a cellular level have not been well characterized. Method of Study Using cord blood mononuclear cells (CBMCs) and cord plasma (n = 9/group) isolated from infants born to women with a high body mass index (BMI>25kg/m 2 ) compared to women with a normal BMI (18‐25kg/m 2 ), we evaluated differences in immune cell populations using single‐cell mass cytometry (CyTOF). CBMCs were matched according to potentially confounding variables, such as maternal and gestational age, ethnicity, smoking status, and gravidity. Statistical results were adjusted for fetal sex. Data were analyzed by viSNE and FlowSOM softwares in Cytobank ™ . Results In newborn CBMCs from women with high BMI, we observed changes in frequency and phenotype of immune cell populations, including significant increases in CD4 + T cells and decreases in myeloid cell populations. IL‐12p40 and MDC concentrations were significantly elevated in the high BMI group compared to control. Conclusion This study demonstrates an association between maternal obesity and fetal immunity. Our results warrant following long‐term immunologic outcomes and associated clinical risks in children born to women with a high pre‐pregnancy BMI.

e19013 Background: Mutation profiling to assess for potentially druggable mutations in NSCLC is being offered at an increasing number of cancer centers throughout North America and internationally. Although data continue to accumulate for the potential value of mutation testing in designing chemotherapeutic regimens, the treatment impact of obtaining information beyond assessment of EGFR and ALK status remains unclear. How best to obtain and clinically utilize these data, including information from exome and whole genome sequencing, also remains unclear. Methods: Patients were reviewed electronically in a multidisciplinary conference regarding indications for testing and results of mutation profiling from various methods, including the mass-spec based LungCarta test, targeted NexGen sequencing, exome, and whole genome sequencing. Outcomes of the multidisciplinary review were communicated back to treating physicians. Results: Mutation testing was performed on 200 patients using a variety of approaches. The majority (>150) were surgically resected stage I and II tumors. Mutations in at least 1 major cancer driver gene, including EGFR, KRAS, MET, BRAF and PIK3CA, were found in 47% of all patients tested. EGFR mutations were present in 14.8% of patients tested, KRAS 21.3%, BRAF 2.6%, PIK3CA 3.2%, and MET 4.5%. A total of 8 patients underwent either exome or whole genome sequencing. A limited number of patients (<10) had mutation results that impacted treatment decisions from this cohort. Conclusions: Mutation profiling can influence treatment decisions in NSCLC, but at a low frequency. The role of exome and or whole genome sequencing for patients with NSCLC is evolving and remains undefined.

Abstract Multiple myeloma (MM) is a malignant plasma cell disorder with well-defined primary genetic/cytogenetic abnormalities. Investigating genetic heterogeneity in the MM cells, may have an impact on therapeutic approaches and assessment of response to treatment. Single cell genomic profiling technology provides an opportunity for us to identify genetic heterogeneity during the pathogenesis of MM progression and select the most appropriate clinical interventions. We performed single cell whole transcriptome analysis in 597 CD138 positive cells derived from 15 patients at different stages of MM progression. Using Coefficient of Variation (CV) approach, we identified 790 genes and defined four main groups reflective of increasing levels of aggressiveness at the single cell level. Differential gene expression and gene set enrichment analyses showed that protein homeostasis pathways, unfolded protein response and mTORC1 related gene sets were significantly enriched at the most aggressive level consistent with the MM cell progression and predicts the overall survival. Our results provide novel insights into the cellular heterogeneity within each MM patient and a foundation for therapeutic opportunities including proteasome inhibitors. Citation Format: Jin Sung Jang. Molecular signature of multiple myeloma progression through single-cell RNA-seq [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4354.