bluuebunny/crossref_metadata_2025_split huggingface dataset:
Lipid Nanoparticle-Mediated Gene Editing of Human Primary T Cells and Off-Target Analysis of the CRISPR-Cas9 Indels
Authors: Geczy Reka , Thommandru Bernice , Swaminathan Meenakshi , Zhao Rita , Tan Krystal , Low Kevin , Balgi Aruna , Park Stella , Watt Ethan , Thada Vaughn , Rettig Garrett , Zhang Angela , Turk Rolf , Clarke Samuel 
Published: December 2023

Background and Aims: The CRISPR/Cas9 system has emerged as a powerful tool for gene editing of primary cells. In our previous work, we demonstrated a novel lipid nanoparticle (LNP) reagent for the multi-step engineering of gene-edited CAR T cells. We showed high cell viabilities and potent CAR-T mediated killing, even after multiple genetic manipulations. Here, we extend this work by assessing potential off-target editing effects in both LNP-treated T cells and T-cells where the CRISPR reagents were delivered by electroporation. Further, we evaluated multiple Cas9 variants and guide RNA targets.
Methods: LNPs encapsulating wild type or high fidelity S.p. Cas9 mRNA and various TRAC and CD52 targeted guide RNAs (sgRNAs) were produced using our scalable NanoAssemblrTM microfluidics platform. Concurrently, electroporation was performed to deliver equivalent cargoes. Purified primary T cells were cultured, activated, and expanded in serum-free media in plates, flasks, or small bioreactors. The LNPs were added to cells by direct addition for gene editing. Gene expression and cell viability were measured using flow cytometry or colorimetric assays. Multi-target performance of CRISPR-Cas9 editing was evaluated through rhAmpSeq-based targeted next-generation sequencing (NGS) and indels analyzed for with CRISPRAltRations.
Results: TCR or CD52 targeted Cas9 mRNA-LNP addition or electroporation yielded high single and double knockout efficiencies. High-throughput NGS analysis showed strong agreeance to flow cytometry for on-target analysis. We tested a range of sgRNA targets, wild-type and high-fidelity Cas9 mRNAs, and determined off-target editing for all targets and variants investigated. Similar results were obtained when comparing different LNP batch sizes (micro to milligram RNA) and cell culture vessels (0.1 to 45 million cells), demonstrating scalability of both LNP production and cell treatment.
Conclusions: The results from this study further support the utility RNA-LNPs for the genetic engineering of primary T cells. The simple and gentle nature of LNP cell treatment allows for multiple genetic engineering steps for simultaneous expression and deletion of proteins for the next cell therapies. These LNPs can be easily manufactured from small-scale screening of RNA libraries to rapid scale-up for clinical translation.
Prevalence, Epidemiology, Mortality and Morbidity of Thrombotic Events in SLE: A Nationwide Study in the United States
Authors: Singhal Sachi , Jotwani Pooja , Desai Devashish , Styler Michael 
Published: December 2023

CONCLUSION
Thrombotic events (TE) are a major cause of morbidity and mortality in patients with Systemic Lupus Erythematosus (SLE). Data is scant, however, on the prevalence, epidemiological characteristics, in hospital morbidity and mortality of TEs in this population.
To perform a comprehensive epidemiological study on patients with SLE and DVT, PE or atypical thrombosis and their variation by gender, geographic region and hospital status. We also aimed to explore the associated morbidity and mortality for each TE and potential associations with other risk factors such as obesity and smoking.
We conducted a retrospective cohort study, using the 2003-2018 Nationwide Inpatient Sample (NIS) database. All adult patients with a diagnosis of SLE using ICD-9 and 10 codes were included. The ICD 9 and 10 codes were also used to find other TE variables including DVT, PE, Cerebral venous sinus thrombosis (CVST), Splanchnic thrombosis (ST)and Arterial thrombosis (AT).
Of the 513,904 SLE patients included, PE, DVT, ST, CVST and AT were identified in 1.38%, 3.07%, 1.34%, 0.04% and 0.35% patients respectively. Each TE (except CVST) had a statistically significant increase in length of hospital stay, cost of hospitalization and all-cause in-hospital mortality. Higher prevalence of PE, DVT, ST and AT were reported in males with SLE, and higher prevalence of obesity was seen in SLE patients with TEs, except AT, which interestingly was associated with a lower prevalence of obesity. Despite comprising only one third of the cohort, African Americans (AAs) faced the highest burden of PEs and DVTs. No significant differences were seen between geographic regions.
This study is limited by administrative claims data, lack of antiphospholipid antibody status, and only includes inpatient encounters. Additionally, this is an observational retrospective study, and the assessment is limited to association of these thrombotic events in the SLE population.
Arterial, venous and atypical thrombosis exert significant morbidity and mortality in patients with SLE. Racial and gender variations exist in each category of thrombosis, and identifying these differences is crucial while taking management decisions for patients with SLE. Particular attention should be directed to AAs with SLE, who appear to be particularly vulnerable.
Arginine Therapy Supplements Improves the LDH Level in Sickle Cell Patients in Steady States: Preliminary Results from Kinshasa, the Democratic Republic of Congo
Authors: Ngonde Mambakasa Ange Christian 
Published: December 2023

Aim: SCD is a congenital hemolytic anemia of autosomal recessive inheritance. The disease has the characteristics of a chronic inflammatory disease. Due to chronic sickling process by intracellular polymerization of red blood cell and precipitation of mutated sickle hemoglobin. It was recently well-established that arginase released from lysed RBCs consumes the eNOS substrate arginine, with a depletion of plasma arginine, the essential substrate for NO production by NO synthase, leading to acute and chronic complications. L-arginine was shown to significantly increase NO levels and decrease oxidative stress and lipid peroxidation, in recent studies. Despite high incidence of the disease in sub-Saharan Africa, there is still very little information on the benefits of arginine dietary supplement, in sickle cell patients.
Methods: We carried out a retrospective study to assess the benefit of arginine supplement on the level of LDH. The study group consisted of 31 patients who received 500 mg of Arginine per day for SCDs from 1 to 14 years old and 1000 mg per day from 15 years old. Three follow-up periods were defined: (i) before starting treatment with hydroxyurea (HU); (ii) a period of taking HU, greater than 3 months; (iii) a period of taking HU+ arginine supplement, greater than 3 months.
Results: The median age of this group was 12 years (range; 2 years-43 years). During the three periods of the study, the LDH value was above the norms, respectively 649,7 ± 364,2 U/L in period 1, 661,6 ± 367 U/L in period 2, and 529,9± 346,3 U/L in period 3. By comparing the different periods of the study, There is a significant increase (p=0.4589; Kruskal Wallis test) in the mean LDH value in the period 2 compared to the period 1. There is a significant decrease in the mean LDH value in period 3 compared to period 1, and period 2 ( p= 0,4589, Kruskal Wallis test).
Conclusion: This study found that the sickle cell patients who received arginine supplements with HU decreased significantly the level of LDH. However, the present study had some limitations. These include its hospital-based retrospective design limiting the potential to make conclusions. On the other hand, further investigations including a large sample in a prospective study with a control group and other biological parameters data will be interesting in our midst.
Comparison of Time to Relapse between Low-Fixed Dose Versus Standard-Dose Rituximab in the Treatment of ITP, TTP, and AIHA
Authors: Powell Frankie , Hasan Yousef , Shi Runhua , Ramadas Poornima 
Published: December 2023

Background:
The optimal Rituximab dosing regimen for the treatment of immune thrombocytopenia purpura (ITP), autoimmune hemolytic anemia (AIHA), or thrombotic thrombocytopenia purpura (TTP) has not been well established. To our knowledge, there has not been a randomized trial examining the time to relapse with the different Rituximab dosing regimens (low-fixed dose of 100 mg x 4 weekly doses versus standard dose of 375 mg/m2 x 4 weekly doses) in the treatment of ITP, TTP, and AIHA. Previous comparative studies have shown that the weekly low-fixed Rituximab dosing regimen has been shown to be just as effective in treating these diseases as demonstrated by similar response and duration of response. Therefore, we conducted a comparative analysis to evaluate the time to first recurrence after being treated with Rituximab.
Methods:
We collected clinical data from a single center of patients with diagnosis of ITP, TTP, or AIHA and were treated with 4 weekly low-fixed or standard dose Rituximab. Time to first recurrence was measured to compare if there was a significant difference between the two Rituximab dosing regimens. Kaplan-Meier time to recurrence estimates were used to generate time to recurrence curves.
Results:
Of the 30 patients analyzed, 10 patients were diagnosed with AIHA, 12 patients with ITP, and 8 with TTP. All patients had a complete response (CR) per the specific disease CR definition prior to first disease recurrence. 17 patients were treated with standard dose Rituximab and 13 patients were treated with low-fixed dose Rituximab. 17 patients did not have a recurrence in the selected time frame of the study. Recurrence was observed in 40% of the AIHA patients, 50% of the TTP patients, and 37.5% of TTP (P=0.8297). 47% females had a recurrence compared to 36% males (P=0.7084). Recurrence was observed in 61% patients who received the low-fixed dose Rituximab and 29% of patients who received the standard dose Rituximab (P=0.1376). The median time of recurrence was 16 and 27 months among patients who received low-fixed dose vs standard dose Rituximab, respectively (P=0.1914).
Conclusions:
The use of low-fixed dose Rituximab versus standard dose Rituximab for treatment of ITP, TTP, and AIHA appears to have a higher recurrence rate but did not meet statistical significance. The median time difference in recurrence was 9 months. This could potentially be significant in a larger study population. Further investigation through large, randomized controlled trials is needed to assess if there is a significant difference in recurrence rate and time to recurrence between the two Rituximab dosing regimens.
BRG1 Regulates Adult B-Cell Acute Lymphoblastic Leukemia Proliferation and Apoptosis By Interacting with c-MYC and Activating PI3K/AKT Pathway
Authors: Kang Qian , Zhao Peng , Ni Ming , Fang Qin , Wang Jishi 
Published: December 2023

Background and objective：Adult B-cell acute lymphoblastic leukemia（B-ALL）has historically had a dismal prognosis, with limited treatment options and cure rates less than 40%. Only 25% of patients older than 50 years old were alive 5 years after diagnosis, highlighting the need for further improvements in treatment for older adult patients (≥40 years). Recently years, genome profiling revealed that cancer cells frequently depend on chromatin regulatory activities to maintain a malignant phenotype and proliferation. Brahma-related gene 1（BRG1）, a core catalytic ATPase subunits of the SWI/SNF chromatin remodelling complex, has been reported to support oncogenic transcriptional program and sustain leukaemogenic transformation in leukemia. However, the specific role and mechanisms of BRG1 in B-ALL development and progression remains largely unknown. Therefore, elucidating the underlying maintenance mechanisms is essential to develop novel effective treatment strategies.
Methods：For clinical sample, we analyzed the mRNA expression level of BRG1 in newly diagnosed B-ALL patients(n=576) and healthy donors(n=74) in the GSE13159 subset from GEO database. Then, quantitative reverse transcription-PCR (RT-PCR) and Western blot were used to detect the expression of BRG1 in 38 newly diagnosed B-ALL patients and 25 healthy donors in our department. For further exploring the effects of BRG1 on B-ALL , lentiviral transfection were used to knock down BRG1 in SUP-B15 and Nalm-6 cells and overexpress BRG1 in RS4:11 cell, and CCK-8, Edu, clone formation assay, cell cycle, apoptosis, IF, RT-PCR and Western Blot were used to detect the effects. In addition, TMT proteomics was used to analyze the differential proteins between sh-ctrl and sh-BRG1 of Nalm-6 cell. Western Bolt was used to detect the changes of key proteins in related pathways.
Results: Our investigation of SMARCA4 revealed that BRG1 is over-expressed in the 576 newly diagnosed B-ALL patients from GSE13159 dataset, as well as in 38 of our department. Downregulated BRG1 inhibited growth and increased the apoptosis of ALL, while ectopic expression of BRG1 showed the opposite effects. Mechanistically, TMT proteomics analysis and vitro experiments further revealed that BRG1 exerted these functions through its interaction with c-MYC and regulated the expression of CDK4/6, BCL-2 and BCL-xl.
Conclusion: Our study demonstrates that BRG1 upregulation is closely associated with proliferation and anti-apoptosis in adult B-ALL.
IDH1/2-Mutated B-ALL Presents As Burkitt's Leukemia/Lymphoma-like ALL
Authors: Qian Yu , Jin Jie 
Published: December 2023

Background: Burkitt leukemia/lymphoma (BL) is a highly aggressive hematological malignancy, which is characterized by the FAB L3 morphology of monotonous medium-sized cells containing abundant basophilic cytoplasm and lipid vacuoles, with mature B cell immunophenotype. Interestingly, some B-cell acute lymphoblastic leukemia (B-ALL) could resemble BL in morphology, but with immature B-cell immunophenotype. In this study, we aimed to explore the molecular and clinical characteristics of these “BL-like B-ALL” patients.
Methods: We analyzed the molecular and clinical characteristics of B-ALL patients in the First Affiliated Hospital of Zhejiang University. RNA-seq analysis of BL-like B-ALL was performed to study the mechanisms of L3 morphology.
Results: We analyzed the population of patients with B-ALL of FAB L3 morphology, among 68 patients, 17 cases with IDH1 mutation and 1 case with IDH2 mutation. In subgroups of 260 B-ALL patients, OS and EFS were significant shorter in BL-like patients compared with others (median 567 days vs not reached, median 238 days vs 566 days; p=0.0024, 0.0122, respectively). Gene set enrichment analysis (GSEA) analysis of RNA-seq with 27 cases of BL-like B-ALL patients revealed significant enrichment in fatty acid metabolism pathway, compared with 203 B-ALL patients in TARGET database. Therefore, we measured levels of serum triglyceride (TG), total cholesterol (TC), low-density lipoprotein-cholesterol (LDL-C), high-density lipoprotein-cholesterol (HDL-C) in 251 B-ALL patients at diagnosis, among which TG and TC levels of 62 BL-like B-ALL patients were significant higher (p=0.0006, p=0.0040), while LDL-C and HDL-C levels showed no significant differences between two groups. Furthermore, we tracked a IDH1-mutated BL-like patient and found that IDH1 mutation and L3 morphology were not detected in remission, but both reappeared with disease relapse. Meanwhile, IDH1/2-mutated B-ALL patients in our center had significant shorter EFS compared with other B-ALL patients (p=0.0087).
Conclusion: IDH1/2-mutated B-ALL presents as Burkitt's leukemia/lymphoma-like ALL. BL-like ALL patients have higher levels of lipid metabolism with worse prognosis.
Pre-treatment 18f-FDG PET/CT for Assessing and Predicting Bone Marrow Involvement in Patients with Newly Diagnosed Peripheral T-Cell Lymphoma
Authors: Chen Jing , Cai Zhen , Zhao Yi 
Published: December 2023

Objective: To explore the value of 18F-fluorodeoxyglucose (FDG)-positron emission tomography (PET)/computed tomography (CT) in assessing and predicting bone marrow involvement (BMI) in patients with newly diagnosed peripheral T-cell lymphoma (PTCL) prior to treatment.
Methods: This retrospective study included 201 eligible patients of PTCL who received bone marrow biopsy (BMB) and PET/CT between January 2017 and December 2022. Extra subgroup analysis was designed considering the variation of avidity of FDG in different distinct entities. The status of bone marrow (BM) by PET was assessed using a visual examination and a quantitative index (the highest SUV of BM divided by the mean SUV of the liver [M/L]). Prognostic value of BM findings was also evaluated regarding progression free survival (PFS) and overall survival (OS).
Results: The sensitivity and specificity of PET/CT for patients with confirmed PTCL by BMB were 43.2% and 90.2%, respectively. As for subgroup analysis, the kappa value reflecting the agreement between BMB and visual analysis in angioimmunoblastic T-cell lymphoma (AITL), peripheral T-cell lymphoma, not otherwise specified (PTCL-NOS), anaplastic large cell lymphoma (ALCL) as well as nodal T-follicular helper (TFH) cell lymphoma (PTCL-TFH) was 0.315, 0.307, 0.426 and 0.581, respectively. (Give the number of cases in each lymphoma group) PET score was an independent factor for PFS (P =0.011) but not for OS (P =0.840). In addition, 25 patients with stage I to II disease had no evidence of BMI by either PET/CT or BMB. Image-guided biopsy was also recommended when PET/CT showed a focal FDG uptake outside the iliac crest. Quantitative analysis revealed the differences in the two-level of M/L; however, they were not statistically significant for both PFS and OS (P =0.246 and P =0.227).
Conclusion: PET/CT has a complementary role in assessing BMI and prognosis in PTCL patients.
Long COVID-19 Infections and Organizing Pneumonia in a Series of Patients with Indolent Lymphoma Treated with Rituximab
Authors: Savage David , Xavier Marin , Mason James , Chang Jacqueline , Escobar Steven 
Published: December 2023

Background: Patients with hematologic malignancies have an increased risk of complications and mortality from COVID-19 (SARS-CoV-2). These patients are often more susceptible to protracted COVID-19 infections (e.g., long COVID-19). In addition, patients treated with the CD20-targeted immunotherapy rituximab in B-cell lymphomas are at greater risk for a protracted clinical course with COVID-19, including care in the ICU for respiratory failure. Hypogammaglobulinemia is an independent predictor of increased risk for long-COVID19. Immune depletion both leads to a persistent detection of SARS-CoV-2 and an inability to mount a sustained immune response. Other authors have reported on the appearance of a delayed organizing pneumonia (OP) after acute COVID-19 infection, but these patients were not immunocompromised by rituximab. In this case series, we report on three patients with indolent lymphoma who had the delayed presentation of long COVID-19 after an acute infection. All three were previously treated with rituximab, and each case was notable for prolonged OP that was refractory to steroid therapy.
Methods: Summative data for the three patient cases was aggregated from the medical record including available physician notes, diagnostic body imaging, and laboratory data.
Results: In total, we found three cases of long COVID-19 in patients with indolent lymphoma from our institution (Table 1). Patient 1 received just two doses of rituximab, Patient 2 received three doses of rituximab, and Patient 3 was treated with rituximab intermittently for nearly three years. All three patients experienced their first COVID-19 infections in August 2022 and had low IgG. This was followed by complications including acute hypoxemic respiratory failure, pulmonary embolism, and organizing pneumonia in these patients two to three months thereafter. These patients continue to have ongoing chronic hypoxemic respiratory failure requiring immune suppression and continuous oxygen support.
Conclusions: Immune depletion with rituximab in patients with indolent lymphomas can put them at significant risk of long COVID-19, OP, and chronic hypoxemic respiratory failure after contracting SARS-Cov-2. Such patients may benefit from prolonged immune suppression, but the exact duration remains unclear. The mechanism of long COVID-19 and refractory OP is unclear and complex. Possible mechanisms may include inadequate clearance of virus or a dysfunctional immune response.
Table 1: Lymphoma patients who received rituximab and then developed long COVID-19
CD56briCD27+CD11b+/- NK Cells Mediated Remission of Agvhd Via Immunoregulation
Authors: Ni Ming , Zhao Peng , Kang Qian , Pan Chengyun , Hu Xiuying , Hu Tianzhen , Fang Qin , Wang Jishi 
Published: December 2023

Background: In our previous study, CD56briCD27+CD11b+/- NK cells show the potential of the remission of aGvHD, while the mechanism behind it is still unknown. Thus, in the present study, samples from patient after allo-HSCT within 100 days has been investigated deeply.
Method: 90 samples of patients after allo-HSCT were collected in the affiliated hospital of Guizhou medical university. Immunomonitoring with respect to the phenotype and function of CD56briCD27+CD11b+/- NK cells was performed on rested peripheral blood mononuclear cells (PBMCs) using multiparametric flow cytometry. Furthermore, CD56briCD27+CD11b+/- NK cells have been co-cultured with T cells and ELISPot as well as CFSE staining has been performed to determine the affect of NK cells on T cells. Sorted CD56briCD27+CD11b+/- NK cells have been analyzed by bulk RNA sequencing to elucidate the profile of gene expression. On the other hand, CD56briCD27+CD11b+/- NK cells have been administrated into xgeneic mice agvhd model to observe whether CD56briCD27+CD11b+/- NK cells influence on the onset of aGvHD.
Result: (1) Our result shows a positive correlation between CD56briCD27+CD11b+/- NK cells and Th22, Tfh as well as Th1 like Tfh cells. Interestingly, a dramatic higher expression of homing marker CD62L has been observed on CD56briCD27+CD11b+/- NK cells. Thus we hypothesis that the CD56briCD27+CD11b+/- NK cells may migrate into the lymphoid tissues and regulate the differentiation of naïve T cells. (2) On the other hand, an elevated percentage of NKG2D has been found on CD56briCD27+CD11b+/- NK cells in patients with aGvHD. Meanwhile a high expression of Fas and exhaustion markers have been observed on CD4+ and CD8+ T cells. These results suggest that T cells may be eliminated by CD56briCD27+CD11b+/- NK cells. (3) Both the cytokine secretion and proliferation of T cells has been inhibited by CD56briCD27+CD11b+/- NK cells. (4) CD56briCD27+CD11b+/- NK cells could delay the onset of aGvHD in mice model.
Conclusion: According to these findings, the CD56briCD27+CD11b+/- NK cells may result in the remission of aGvHD via immunoregulation by regulation of T cells.