bluuebunny/crossref_metadata_2025_split huggingface dataset:
Manufacture and In Vivo Support of Gene-Edited, Regulatory-like, T-Cells (edTreg) Using a Synthetic IL2 Receptor
Authors: Uenishi Gene I. , Mallari Cornell , Gamboa Luis , West Samuel , Jacobs Chester , Dahl Noelle , Boucher Greg , Chin Samantha , Aeran Rangoli , Wodziak Dariusz , Gebremeskel Simon , Vo Karen , Ito Cade E. , Smith Laura , Patel Chandra , Abe Keith , Stankovich Basha , Sommer Karen , Torgerson Troy R. , Rawlings David J. , Scharenberg Andrew M. , Cost Greg J. 
Published: November 2019

Regulatory T-cells (Treg) depend on signals from IL2 and their endogenous T-cell receptors to survive, proliferate, and maintain suppressive activity. We have developed a strategy for engineering effector CD4 T-cells into edited, regulatory-like T-cells (edTregs) for treatment of autoimmune disease. edTregs contain a rapamycin-activated synthetic IL2 receptor (designated a chemical-induced signaling complex or CISC) for use in selective cell manufacturing and in vivo trophic support. Our strategy uses gene editing to integrate an MND/ CISC expression cassette into the native FOXP3 locus, resulting in the CISC system cis-linked to expression of the native FOXP3 gene. The resulting high-level and stable expression of FOXP3 converts peripheral T-cells to edTregs with immunosuppressive activity. Cis-linked expression of the CISC system allows for selective cell expansion and in vivo support using rapamycin. Using an optimized protocol, we obtained efficient HDR rates across cells from multiple healthy donors. Edited cells were enriched to >90% purity and expanded 20-50-fold over a 10 day period of culture in rapamycin. Importantly, we demonstrated sustained in vivo suppressive activity of edTregs in a xeno-GvHD mouse model, and successful trophic support of these edTregs by rapamycin (via the CISC) in the absence of effector CD4 T-cells. Along with preliminary data showing successful editing of CD4-positive T-cells from auto-immune disease patients, our data provide pre-clinical proof-of-concept data supporting clinical use of CISC-edTreg in conjunction with rapamycin support for therapy of autoimmune disease.

Disclosures
Uenishi: Casebia Therapeutics: Employment. Mallari:Casebia Therapeutics: Employment. Gamboa:Casebia Therapeutics: Employment. Boucher:Casebia Therapeutics LLc: Employment. Chin:Casebia Therapeutics LLc: Employment. Aeran:Casebia Therapeutics: Employment. Wodziak:Casebia Therapeutics: Employment. Gebremeskel:Casebia Therapeutics: Employment. Vo:Casebia Therapeutics: Employment. Ito:Casebia Therapeutics: Employment. Patel:Casebia Therapeutics: Employment. Abe:Casebia Therapeutics: Employment. Stankovich:Casebia Therapeutics LLc: Employment. Torgerson:Shire: Consultancy; CSL Behring: Consultancy; ADMA Biosciences: Consultancy; UCB: Consultancy. Scharenberg:Generation Bio: Equity Ownership; Casebia Therapeutics LLc: Employment, Equity Ownership; Alpine Biosciences: Consultancy, Equity Ownership. Cost:Casebia Therapeutics: Employment.
Characteristics of Sickle Cell Clinic Visits and Hospital Admissions in a Large Urban Academic Hospital
Authors: Odikadze Levani , McLemore Morgan L. , Stoyanova Neli , El Rassi Fuad A 
Published: November 2019

Introduction: Although high rates of emergency department (ED) visits by sickle cell disease patient population is a significant health-care system burden, there are limited studies available that explore outpatient/ED visit patterns. The purpose of this study was to evaluate the pattern of medical care utilization including clinic visits and hospital admission in adults with sickle cell disease in a large urban academic center with a dedicated 24 hour/7 day a week sickle cell emergency room.
Methods: Patients seen in the Acute Care at Grady Memorial Hospital Comprehensive Sickle Cell Center (GMHSCC) over a three years from January 2015 to December 2017 where analyzed. The Acute Care center at GRMHSCC is a unique 24 hour 7 day a week dedicated emergency room for sickle cell patients. Patient demographics, specialized sickle cell clinic visits and hospital admissions were analyzed descriptively. Independent samples t-test was used for comparison of hospitalization and acute care visits between males/females and phenotype groups.
Results: The total of 1,277 patients were included in analysis representing 12,129 acute care visits. 683 (53.5%) females and 593 males (46.5%). Mean age was 32.9 years. Genotypes where similar to known expected distributions, 878 patients had Hb SS (68.8%), 301 had SC (23.6%), 23 had SB0TH (1.8%) and 74 had SB+TH (5.8%). The mean number of acute care visits over 3 years for SS phenotype was 9.71, SC phenotype 9.74, SB+Th phenotype 5.73 and SB0TH phenotype 10.48. Overall admission rate was 16%. There was a significant statistical difference between the mean number of admissions among SS and SC phenotype patients (p=0.009). Rate of hospitalization per clinic visit for SS phenotype was 18.19% , for SC phenotype 10.78% , SB+TH phenotype was 15.58% and SB0TH phenotype 9.55%;
Conclusions and future objectives: We have demonstrated that a dedicated Sickle Cell emergency room is able to achieve a significantly lower admission rate than standard Emergency rooms. Preliminary analysis shows that the distribution of acute care visits is distributed across the genotypes at the known distribution. In addition there is no difference in mean number of sickle cell acute visits between patients with SS and SC phenotype. This is interesting as Hgb SC is considered a less severe form of the disease. There was a statistically significant higher admission rate for SS patients compared to SC patients.
Future analyses will look at additional parameters associated with acute care visits and admission.

Disclosures
No relevant conflicts of interest to declare.
Statins Enhance Killing of Multiple Myeloma Cells By the BCL-2 Inhibitor Venetoclax and the MCL-1 Inhibitor S63845
Authors: Fruman David , Juarez Dennis , Diep Grace , Nguyen Falisha , Gupta Vikas A. , Boise Lawrence H. 
Published: November 2019

The BCL-2 inhibitor venetoclax has begun to change the landscape of therapy for diverse blood cancers. Combining venetoclax with standard of care agents has led to impressive response rates, and increased depth and durability of response compared to monotherapy. However, there is a continuing need to develop safe and effective combinations in multiple myeloma (MM), where the BELLINI triple combination trial (venetoclax-dexamethasone-bortezomib) was closed to accrual due to adverse events. We reported that HMG-CoA-reductase inhibitors (statins), a safe and widely prescribed class of drugs, enhance the efficacy of venetoclax in human leukemia and lymphoma cell lines and in a retrospective study of patients with chronic lymphocytic leukemia (JS Lee et al., Sci. Transl. Med.2018, 10(445)). In MM, the subset with t11;14 translocation is BCL-2-dependent and sensitive to venetoclax, whereas other MM subtypes are more dependent on MCL-1. Here we show that statins sensitize MM cell lines and primary cells to killing by either venetoclax or the MCL-1 inhibitor S63845. The compound pitavastatin is particularly potent, active in vitro at a concentration (100 - 300 nM) that is clinically achievable. In OPM2 cells, pitavastatin synergizes with venetoclax and increases apoptosis in triple combination with dexamethasone. Pitavastatin also potentiates venetoclax killing in dexamethasone-resistant MM1R cells. In MCL-1-dependent MM cells, statins increase sensitivity to S63845. Notably, statins do not increase killing of normal human lymphocytes by either venetoclax or S63845. Statin responsiveness in MM cell lines correlates with p53-independent upregulation of PUMA, BAX and BAK, and gene editing experiments suggest that all three contribute to the apoptotic mechanism. Statin-induced apoptosis and PUMA expression are rescued by addition of geranylgeranyl-pyrophosphate, and recapitulated by an inhibitor of geranylgeranyl-transferase I. Together these findings establish a mechanistic framework to explain how statins promote apoptosis by BH3 mimetic drugs like venetoclax and S63845. Further, these results highlight the potential of statins to enhance therapeutic efficacy of BCL-2 and MCL-1 inhibitor regimens in MM, with possibly lesser toxicity than other combination strategies.

Disclosures
Boise: Genentech Inc.: Membership on an entity's Board of Directors or advisory committees; AstraZeneca: Honoraria, Research Funding.

OffLabel Disclosure:
Venetoclax is a BCL-2 inhibitor that is approved for some indications. I will discuss the use of venetoclax for multiple myeloma, a disease for which venetoclax is not approved.
A Meta-Analysis of Genetic Abnormalities and Next Generation Sequencing of Primary Cases of Castleman Disease
Authors: Butzmann Alexandra , Przybyl Joanna , Sridhar Kaushik , Kumar Jyoti , Brar Nivaz , Zehnder James L , Warnke Roger A , Ohgami Robert S. 
Published: November 2019

Castleman disease is a rare lymphoproliferative disorder known to represent at least four distinct clinico-pathologic sub-types. Large advancements in our clinical and histopathologic description of these diverse diseases have been made, resulting in subtyping based on degree of nodal involvement (unicentric versus multicentric) and according to viral infection (human herpes virus 8 - HHV8 and Human Immunodeficiency Virus - HIV). However, we continue to lack a foundational understanding of the biologic mechanisms driving at least a subset of patients with these diseases. In recent years, significant molecular and genetic abnormalities that may be associated with this disease have been described; yet we are still unclear as to a unified vision of what loci and genes are involved in at least a subset of cases of Castleman disease.
In our study we performed a meta-analysis of all cases (unicentric and multicentric) of Castleman disease described in literature to date, and correlate cytogenetic, molecular and genetic abnormalities with disease subtypes, phenotypes and clinical outcomes. We further performed targeted DNA deep sequencing and RNA-sequencing on a cohort of Castleman Disease cases.
Our results demonstrate that genetic abnormalities in the MAPK-pathways, PI3K-pathway and Interleukin signaling are frequent in unicentric Castleman disease, whereas abnormalities in DNA methylation are seen more commonly in idiopathic multicentric Castleman disease, along with abnormalities in MAPK-pathways.
We conclude that UCD and iMCD may share similar pathways of MAPK pathogenesis but can also be distinguished by cytokine and other DNA modification pathways that differ.

Disclosures
Ohgami: Agilent technologies: Other: received support/funding.
Mitigation of Venous Thromboembolism Risk through Favorable Lifestyle: The ARIC Study
Authors: Evans Christina , Hong Ching-Png , Folsom Aaron R , Heckbert Susan , Smith Nicholas , Wiggins Kerri , Cushman Mary 
Published: November 2019

Background: Venous thromboembolism (VTE) is a common disease with a strong genetic basis. Unhealthy lifestyle factors contribute to risk, but it is unknown whether healthier lifestyle can mitigate the risk for VTE in those at high genetic risk. We studied whether greater adherence to the American Heart Association's (AHA's) cardiovascular health metric called Life's Simple 7 (LS7) is associated with a lower rate of VTE in individuals with high genetic risk score (GRS) for VTE.
Methods: We followed 9,026 middle-aged white participants from the Atherosclerosis Risk in Communities (ARIC) Study, a prospective cohort of 15,792 individuals enrolled in 1987-89. A validated GRS was used, comprising 5 well known genetic conditions associated with VTE (factor V Leiden, prothrombin 20210A, non-O blood group, factor XI rs4241824, and fibrinogen gamma FGG rs2066865). Only white participants were included, as the GRS did not predict VTE in others. AHA's LS7 categories of inadequate, average, and optimal health were determined based on smoking status, body mass index, physical activity, diet, total cholesterol, blood pressure, and fasting glucose. VTE events were adjudicated by expert medical record review. We calculated hazard ratios (HRs) and 95% confidence intervals (CI) of incident VTE by LS7 categories, stratified by GRS (low, intermediate, high), adjusting for age, sex, and education. HRs were also calculated for individual LS7 components stratified by GRS.
Results: There were 466 incident VTE over 22.8 years of follow-up. Compared to those with optimal health, those with inadequate LS7 score had higher rates of VTE (5.7% vs. 3.9%). In Figure 1, compared to the high GRS / inadequate LS7 group, the HR of VTE in the low GRS group with optimal health was lowest at 0.39 (95% CI 0.25-0.61), but moreover, the HR in the high GRS group with optimal health was also attenuated to 0.65 (95% CI 0.48-0.89). The pattern of association was similar for provoked and unprovoked VTE. Of the LS7 components, obesity was most strongly related to VTE. In Figure 2, compared to obese / high GRS participants, the HR of VTE with normal weight / low GRS was 0.36 (95% CI 0.23-0.57), while the HR in high GRS / normal weight participants was reduced by 45%, at 0.55 (95% CI 0.4-0.76).
Conclusion: Among all participants, even those at high genetic risk, healthier lifestyle factors, particularly obesity, were associated with decreased incidence of VTE. Further studies should determine the impact of lifestyle change among patients at high genetic risk of VTE, such as in thrombophilic families.

Disclosures
Heckbert: National Institutes of Health: Other: Grants.
Tiered Oral Therapy Protocol for Sickle Cell Vaso-Occlusive Crisis
Authors: Guy Margaret , Magee Lauren , Keiser Jessica , Carter Nicole , Vaughan DaleMarie , McHenry Chantal , Sop Daniel , Hartigan Sarah , Smith Wally R 
Published: November 2019

Background: Adults with sickle cell disease (SCD) often present with extremely painful acute vaso-occlusive crises (VOC). VOC guidelines quote studies supporting early initiation of opioids, but are lacking in a standardized approach to achieve adequate analgesia and subsequent de-escalation. Various SCD studies comparing continuous basal versus demand bolus patient-controlled analgesia (PCA) show conflicting outcomes. Thus the objective of our pilot was to develop a standardized approach to pain management utilizing PCA and a Tiered Oral Therapy Protocol (TOTP) in an effort to affect healthcare utilization by decreasing length of stay without increasing readmission rates.
Methods: We studied admissions of 31 SCD (any genotype) patients (8 males, 23 females) ≥ 18 years old, all African-American, admitted for VOC, and treated by hospitalist providers using TOTP on one medical unit at Virginia Commonwealth University Health System from July 2018 to June 2019. Inpatient admissions for one calendar year prior to the initial TOTP admission served as the patient's historical control. TOTP is guided by the patient's individualized treatment plan and starts with continuation of their home basal dosing, demand dose only PCA, and immediate-release oral opioids for additional breakthrough pain. TOTP goals are aggressive up-titration of opioids until adequate analgesia. After a period of stability the next phase encourages timely, aggressive down-titration of opioids to speed discharge. We primarily compared length of stay (LOS), hospital charges, and 30-day readmission rates for TOTP vs historical controls. Secondarily, we compared adverse safety events: opioid-related rapid responses, naloxone administration, or falls.
Results: Utilization of TOTP resulted in reductions in LOS (21%, 4.7 days controls vs. 3.7 days TOTP, p&lt;0.014), and hospital charges ($1,627,497.73 controls vs. $944,528.96 TOTP, p=0.1403), and an insignificant increase in the 30-day readmission rate (35.6% controls vs. 42.7% TOTP, p=0.5027). Hospital charge data is difficult to interpret as a full year of post TOTP encounters are not available for all patients. There were no TOTP vs. historical control differences in opioid-related safety events.
Conclusion: Standardization of therapy using the TOTP improved length of stay and reduced charges for patients with sickle cell disease admitted for acute VOC without increasing adverse safety events or significantly impacting the 30 day readmission rate. We have enlarged this pilot to include other VCU medical units, to test whether we can reproduce our initial one-unit success. Further, we recommend TOTP implementation and testing in other hospitals to exclude secondary trends, since TOTP was but one among several interventions in a VCU Adult SCD Medical Home.
Figure

Disclosures
Smith: Novartis: Consultancy, Honoraria.
Mutation of Murine Rpl5 reveals a New Model for Diamond Blackfan Anemia Characterized By Defective Erythropoiesis
Authors: Lemay Philippe , Jones Morgan A , Mohamed Fatma , Guyot Marie-Claude , Ludlow Alexander , Li Qing , Engel James Douglas , Franceschi Renny , Rothstein Thomas L. , Kaartinen Vesa , Kibar Zoha , Singh Sharon A 
Published: November 2019

Background
Ribosome dysfunction is implicated in several abnormal developmental conditions and diseases in humans. Heterozygous germline ribosomal protein (RP) variants are found in the majority of individuals with Diamond Blackfan anemia (DBA) with RPL5 (uL18) being the second most frequently mutated gene. Patients with RPL5 mutations are for unknown reasons at increased risk of congenital defects when compared to other genotypes. There is currently no available Rpl5 mouse model with anemia and therefore hematopoiesis in this genotype has not been well studied.
Methods
We characterized a novel mutant mouse with a growth and kinky tail defect, and increased mortality. The mouse was generated in a chromosome 11 balancer ENU mutagenesis screen. Following genetic mapping and whole exome sequencing, we identified a heterozygous intronic Rpl5 missense variant, which segregated with all affected mice. We first performed molecular analyses to validate the impact of this variant on RPL5 and ribosome levels, and then examined the development and physiology of multiple tissues by histology, micro-CT, echocardiogram and measurement of erythroid differentiation to determine the etiology of the observed increased mortality in newborn Rpl5 mutant mice.
Results
This variant led to decreased Rpl5 mRNA and protein, and consequential polysome defects (decreased 60S and 80S ribosome subunits) consistent with RPL5 haploinsufficiency. All Rpl5+/- neonates exhibited deficient terminal erythroid differentiation. A fraction of the mutants progressed to severe anemia leading to increased mortality while others appeared to undergo spontaneous remission. Surviving adult/aged mice exhibited no anemia although preliminary studies indicate reduced numbers of hematopoietic progenitors. Most newborn mutant mice had large ventricular septal defects (VSD) while surviving adult mice had small VSDs on echocardiogram. Rpl5+/- mice were significantly smaller without craniofacial abnormalities as assessed by micro-CT or histology. Bone and cartilage staining of newborn mice revealed that the kinky tail defect resulted from delayed endochondral ossification.
Conclusion
This model recapitulates the major developmental and erythroid differentiation defects of DBA in an animal model. We propose that this novel Rpl5 mutant provides an opportunity to study the mechanism of the variable penetrance of anemia and the congenital defects normally encountered in DBA.

Disclosures
Singh: Emmaus Medical, Inc: Consultancy.
Platelet Phenotype Prediction from Whole Genome Sequencing in 621 Sickle Cell Disease Patients
Authors: Montemayor Garcia Celina , Weisman Julie K. , Wang Xunde , Gandla Divya , Long James Owen , Dalgard Clifton L. , Pirooznia Mehdi , Klein Harvey , Thein Swee Lay 
Published: November 2019

Introduction: Patients with sickle cell disease (SCD) are at increased risk of alloimmunization. Platelet refractoriness is a serious known complication and often seen in SCD patients who are heavily transfused and/or in the bone marrow transplantation (BMT) setting. Next generation sequencing (NGS) is an emerging and promising genotyping strategy in the context of blood typing, due its high throughput and its ability to detect both known and novel variants in the patient and donor population. Here we describe an algorithm to predict common and rare human platelet antigens (HPA) from NGS data, and its validation through Sanger sequencing.
Design/Methods: Whole genome sequencing (WGS) was performed on stored blood samples from 621 SCD patients enrolled in 2 IRB-approved clinical studies. Our open source software RyLAN (Red Cell and Lymphocyte Antigen prediction from NGS) was utilized to translate WGS data into predicted RBC and platelet phenotypes. The 29 genomic variants interpreted by RyLAN in 6 HPA genes were correlated with Sanger sequencing.
Results: Our study cohort consisted of 621 SCD patients (485 HbSS, 21 HbSβ0, 29 HbSβ+, 84 HbSC, 1HbS O Arab, and 1 HbSD). The mean age was 34.3 years, and 46% were male. Previous red cell transfusions were recorded in 62% of patients, and 3% were documented as never transfused. RyLAN software was executed as a singularity container in multithreaded mode, completing analysis of all 621 .bam WGS files in 18 hours. RyLAN predicted 237 unique extended platelet phenotype combinations in this cohort, with an average read depth of 33 in genomic areas of interest. Predictions for 10 platelet antigens in 26 participants, including rare phenotypes like HPA-25bw+ and HPA-13bw+, were confirmed by bidirectional Sanger.
Conclusions: We describe an efficient, open-source algorithm used to interpret 6 HPA genes from WGS in a large SCD cohort. WGS, in conjunction with the RyLAN algorithm, demonstrated 100% accuracy in predicting common and rare HPA genomic variants. Future studies are needed to refine WGS algorithms in SCD, and to examine the possible value of this technology in HPA alloantibody identification workups, optimal platelet product allocation, and donor recruitment.

Disclosures
No relevant conflicts of interest to declare.