Universal CAR cell therapy: Challenges and expanding applications.

Chimeric Antigen Receptor (CAR) T cell therapy has gained success in adoptive cell therapy for hematological malignancies. Although most CAR cell therapies in clinical trials or markets remain autologous, their acceptance has been limited due to issues like lengthy manufacturing, poor cell quality, and demanding cost. Consequently, "Off-the-shelf", universal CAR (UCAR) cell therapy has emerged. Current concerns with UCAR therapies revolve around side effects such as graft versus host disease (GVHD) and host versus graft response (HVGR). Preclinical research on UCAR cell therapies aims to enhance efficacy and minimize these side effects. Common approaches involve gene editing techniques to knock out T cell receptor (TCR), human leukocyte antigen (HLA), and CD52 expression to mitigate GVHD and HVGR risks. However, these methods carry drawbacks including potential genotoxicity of the edited cells. Most recently, novel editing techniques, such as epigenetic editing and RNA writer systems, have been developed to reduce the risk of GVHD and HVGR, allowing for multiplex editing at different sites. Additionally, incorporating more cell types into UCAR cell therapies, like T-cell subtypes (DNT, γδT, virus-specific T cells) and NK cells, can efficiently target tumors without triggering side effects. In addition, the limited efficacy of T cells and NK cells against solid tumors is being addressed through CAR-Macrophages. In summary, CAR cell therapy has evolved to accommodate multiple cell types while expanding applications to various diseases, including hematologic malignancies and solid tumors, which holds tremendous growth potential and is promised to improve the lives of more patients in the future.

Bioinformatics analysis of immune infiltration in human diabetic retinopathy and identification of immune-related hub genes and their ceRNA networks.

Diabetic retinopathy (DR) is the most common microvascular complication in diabetic patients, and recent studies have shown that immune regulatory mechanisms are closely associated with retinal damage in DR. Therefore, this study focused on exploring immune cells and immune-related genes (IRGs) in DR and gaining insight into the ceRNA mechanisms by which IRGs regulate DR progression. Four datasets from human DR model retinal tissues were obtained from the Gene Expression Omnibus (GEO) database. R software was first used to identify differentially expressed mRNAs (DE-mRNAs) in the dataset GSE160306-mRNAs, then the distribution of immune cells in the gene matrix was analyzed by xCell and ImmuCellAI, ImmPort and InnateDB database were used to obtain immune-related hub genes (IRHGs) in the DR, and finally the STRING online tool and Cytoscape to construct the immune-related ceRNA network. The datasets GSE102485, GSE160308 and GSE160306-lncRNAs were used to validate the results of the ceRNA network further. The results of immune cell infiltration analysis showed that macrophages are important immune cells in DR; immune-related gene screening showed that FCGR2B is an IRHG in DR, and 2 immune-related ceRNA networks of IRHG were obtained: DDN-AS1/miR-10a-5p/FCGR2B and LINC01515/miR-10a-5p/FCGR2B. Our study suggests that infiltration of immune cells, especially the immune role of macrophages, is an important component of DR progression; the immune-related hub gene FCGR2B and its ceRNA network may be a key regulatory network for DR progression. The discovery of key immune cells, IRHG and ceRNA networks in this study may provide new prospects for early intervention and targeted treatment of DR.

Multi-omics integration reveals the oncogenic role of eccDNAs in diffuse large B-cell lymphoma through STING signalling.

BACKGROUND: Extrachromosomal circular DNAs (eccDNAs), a type of double-stranded DNAs (dsDNAs) that facilitate the activation of the DNA sensing machinery, have been implicated in the progression and prognosis of various diseases. While the roles of eccDNAs remain contentious, their significance in diffuse large B-cell lymphoma (DLBCL) has not been reported. METHODS: Circular DNA sequencing (circle-seq) was used to demonstrate the expression profile of eccDNAs in DLBCL, and atomic force microscopy to validate the presence of eccDNAs. CCK-8 and scRNA-seq techniques were employed to uncover the activation of eccDNA in the STING pathway, leading to enhanced cell proliferation. Chemotherapeutic drugs were used to test the hypothesis that DNA damage induces the production of eccDNA, thereby activating the STING pathway independent of cGAS. GEO databases were used for verification of the prognosis of the eccDNA-related genes, and animal models were used to investigate the synergistic effects of DNA damage therapy in combination with STING inhibitors on anti-tumour responses. RESULTS: EccDNAs were widely expressed in DLBCL and associated with the prognosis of patients. Elevated abundance of eccDNAs promoted the progression of DLBCL. Chemotherapeutic drugs-induced DNA damage triggered the generation of eccDNAs, resulting in the activation of the STING signalling in a cGAS-independent manner. Moreover, inhibition of STING exerted a synergistic anti-tumour effect with cisplatin. CONCLUSIONS: EccDNAs induced by DNA damage exert an oncogenic role in DLBCL via activating the STING signalling independently of cGAS. This finding offers a rational therapeutic strategy combining chemotherapy with targeting STING. HIGHLIGHTS: EccDNAs induced by DNA damage exert an oncogenic role in DLBCL via activating the STING signalling independently of cGAS. The combined treatment of chemotherapeutic drugs with STING inhibitor significantly delayed the tumor progression, providing new insights into the therapeutic strategy for patients with DLBCL, particularly the relapsed and/or refractory (R/R) ones.

Ferroptosis and Lipid Metabolism in Acute Myocardial Infarction.

Acute myocardial infarction (AMI) is triggered by the blockage of coronary arteries, leading to restricted blood flow to the myocardium, which results in damage and cell death. While the traditional understanding of cell death primarily revolves around apoptosis, a new player in the game has emerged: ferroptosis. This novel form of cell death relies on iron and is propelled by reactive oxygen species (ROS). Lipid metabolism, an indispensable physiological process, plays a vital role in preserving cellular homeostasis. However, when this metabolic pathway is disrupted, the accumulation of excess waste increases, specifically lipid peroxides, which are strongly linked to the occurrence and progression of AMI. As a result, comprehending this complex interaction between ferroptosis and lipid metabolism could pave the way for new therapeutic approaches in tackling AMI.

Integrated analysis and validation of ferroptosis-related genes and immune infiltration in acute myocardial infarction.

BACKGROUND: Acute myocardial infarction (AMI) is indeed a significant cause of mortality and morbidity in individuals with coronary heart disease. Ferroptosis, an iron-dependent cell death, is characterized by the accumulation of intracellular lipid peroxides, which is implicated in cardiomyocyte injury. This study aims to identify biomarkers that are indicative of ferroptosis in the context of AMI, and to examine their potential roles in immune infiltration. METHODS: Firstly, the GSE59867 dataset was used to identify differentially expressed ferroptosis-related genes (DE-FRGs) in AMI. We then performed gene ontology (GO) and functional enrichment analysis on these DE-FRGs. Secondly, we analyzed the GSE76591 dataset and used bioinformatic methods to build ceRNA networks. Thirdly, we identified hub genes in protein-protein interaction (PPI) network. After obtaining the key DE-FRGs through the junction of hub genes with ceRNA and least absolute shrinkage and selection operator (LASSO). ImmucellAI was applied to estimate the immune cell infiltration in each sample and examine the relationship between key DE-FRGs and 24 immunocyte subsets. The diagnostic performance of these genes was further evaluated using the receiver operating characteristic (ROC) curve analysis. Ultimately, we identified an immune-related ceRNA regulatory axis linked to ferroptosis in AMI. RESULTS: Among 56 DE-FRGs identified in AMI, 41 of them were integrated into the construction of competitive endogenous RNA (ceRNA) networks. TLR4 and PIK3CA were identified as key DE-FRGs and PIK3CA was confirmed as a diagnostic biomarker for AMI. Moreover, CD4_native cells, nTreg cells, Th2 cells, Th17 cells, central-memory cells, effector-memory cells, and CD8_T cells had higher infiltrates in AMI samples compared to control samples. In contrast, exhausted cells, iTreg cells, and Tfh cells had lower infiltrates in AMI samples. Spearman analysis confirmed the correlation between 24 immune cells and PIK3CA/TLR4. Ultimately, we constructed an immune-related regulatory axis involving XIST and OIP5-AS1/miR-216a/PIK3CA. CONCLUSION: Our comprehensive analysis has identified PIK3CA as a robust and promising biomarker for this condition. Moreover, we have also identified an immune-related regulatory axis involving XIST and OIP5-AS1/miR-216a/PIK3CA, which may play a key role in regulating ferroptosis during AMI progression.

RBM25 binds to and regulates alternative splicing levels of Slc38a9, Csf1, and Coro6 to affect immune and inflammatory processes in H9c2 cells.

Background: Alternative splicing (AS) is a biological process that allows genes to be translated into diverse proteins. However, aberrant AS can predispose cells to aberrations in biological mechanisms. RNA binding proteins (RBPs), closely affiliated with AS, have gained increased attention in recent years. Among these RBPs, RBM25 has been reported to participate in the cardiac pathological mechanism through regulating AS; however, the involvement of RBM25 as a splicing factor in heart failure remains unclarified. Methods: RBM25 was overexpressed in H9c2 cells to explore the target genes bound and regulated by RBM25 during heart failure. RNA sequencing (RNA-seq) was used to scrutinize the comprehensive transcriptional level before identifying AS events influenced by RBM25. Further, improved RNA immunoprecipitation sequencing (iRIP-seq) was employed to pinpoint RBM25-binding sites, and RT-qPCR was used to validate specific genes modulated by RBM25. Results: RBM25 was found to upregulate the expression of genes pertinent to the inflammatory response and viral processes, as well as to mediate the AS of genes associated with cellular apoptosis and inflammation. Overlap analysis between RNA-seq and iRIP-seq suggested that RBM25 bound to and manipulated the AS of genes associated with inflammation in H9c2 cells. Moreover, qRT-PCR confirmed Slc38a9, Csf1, and Coro6 as the binding and AS regulatory targets of RBM25. Conclusion: Our research implies that RBM25 plays a contributory role in cardiac inflammatory responses via its ability to bind to and regulate the AS of related genes. This study offers preliminary evidence of the influence of RBM25 on inflammation in H9c2 cells.

Promising roles of non-exosomal and exosomal non-coding RNAs in the regulatory mechanism and as diagnostic biomarkers in myocardial infarction. / 非外泌体/å¤–æ³Œä½“éžç¼–ç RNAsåœ¨å‚ä¸Žå¿ƒè‚Œæ¢—æ­»è°ƒèŠ‚åŠä½œä¸ºè¯Šæ–­æ€§ç”Ÿç‰©æ ‡å¿—ç‰©æ–¹é¢çš„ä½œç”¨.

Non-exosomal non-coding RNAs (non-exo-ncRNAs) and exosomal ncRNAs (exo-ncRNAs) have been associated with the pathological development of myocardial infarction (MI). Accordingly, this analytical review provides an overview of current MI studies on the role of plasma non-exo/exo-ncRNAs. We summarize the features and crucial roles of ncRNAs and reveal their novel biological correlations via bioinformatics analysis. The following contributions are made: (1) we comprehensively describe the expression profile, competing endogenous RNA (ceRNA) network, and "pre-necrotic" biomarkers of non-exo/exo-ncRNAs for MI; (2) functional enrichment analysis indicates that the target genes of ncRNAs are enriched in the regulation of apoptotic signaling pathway and cellular response to chemical stress, etc.; (3) we propose an updated and comprehensive view on the mechanisms, pathophysiology, and biomarker roles of non-exo/exo-ncRNAs in MI, thereby providing a theoretical basis for the clinical management of MI.

Role of LncRNAs in the Pathogenesis of Coronary Artery Disease.

Coronary artery disease (CAD), caused by coronary artery occlusion, is a common cardiovascular disease worldwide. Long non-coding RNAs (lncRNAs) are implicated in the regulation of endothelial cell injury, angiogenesis, plaque formation, and other pathological mechanisms in CAD by acting on different cell types. Some lncRNAs are significantly upregulated in CAD patients; however, other lncRNAs are significantly downregulated. Differential expression of lncRNAs in CAD patients enables them to be exploited as potential biomarkers to evaluate disease progression and diagnosis/prognosis in CAD patients. In this study, we reviewed the role of lncRNAs in the development of different clinical subtypes of CAD.

Longitudinal Associations of Work Stress with Changes in Quality of Life among Patients after Acute Coronary Syndrome: A Hospital-Based Study.

(1) Background: Targeting a sample of Chinese employees in this study, the correlation of work stress with changes in quality of life (QoL) was explored subsequent to acute coronary syndrome (ACS). (2) Methods: Patients suffering from the first ACS episode, with regular paid work before ACS, were eligible for this one-year longitudinal study. Effort-reward imbalance (ERI), together with job strain (JS) models, were employed to evaluate work stress before discharge, and QoL prior to discharge (baseline), as well as at 1, 6, and 12 months following discharge, were measured using the 8-Items Short Form (SF-8), in addition to the Seattle Angina Questionnaire (SAQ). Moreover, generalized estimating equations were used to determine the relationship of work stress to longitudinal QoL variations. (3) Results: After adjusting for covariates, high work stress at the baseline measured by JS was associated with the slow recovery of both mental health (p < 0.01) and physical health (p < 0.05) in SF-8, while ERI-measured work stress was related to slower improvement in SF-8 physical health (p < 0.001), SAQ-angina stability (AS) (p < 0.05), SF-8 mental health (p < 0.001), and SAQ-angina frequency (AF) (p < 0.05). After mutual adjustment for JS and ERI, high work stress as assessed by JS displayed no correlation with any QoL alteration (all p > 0.05), whereas ERI-determined work stress at a high level still presented a relationship to slow improvement in SF-8 physical health, SAQ-AS, SF-8 mental health, and SAQ-AF (all p < 0.05). (4) Conclusion: Work stress was associated with slow recovery of QoL in patients with ACS across one year. For ACS patients, ERI was a stronger predictor of QoL variations than JS.

New developments in non-exosomal and exosomal ncRNAs in coronary artery disease.

Aim & methods: Non-exosomal and exosomal ncRNAs have been reported to be involved in the regulation of coronary artery disease (CAD). Therefore, to explore the biological effects of non-exosomal/exosomal ncRNAs in CAD, the authors searched for studies published in the last 3 years on these ncRNAs in CAD and summarized their functions and mechanisms. Results: The authors summarized 120 non-exosomal ncRNAs capable of regulating CAD progression. In clinical studies, 47 non-exosomal and nine exosomal ncRNAs were able to serve as biomarkers for the diagnosis of CAD. Conclusion: Non-exosomal/exosomal ncRNAs are not only able to serve as biomarkers for CAD diagnosis but can also regulate CAD progression through ceRNA mechanisms and are a potential target for early clinical intervention in CAD.

ncRNAs are increasingly found to play regulatory roles in coronary artery disease (CAD), and transcriptome studies offer greater advantages for controlling CAD at its source. Therefore, the authors conducted an accurate search and summary of studies on CAD and ncRNAs published in the past 3 years to analyze the main pathological mechanisms in CAD progression, aiming to provide a research basis for clinical treatment of CAD.

Identification of immune-related hub genes and miRNA-mRNA pairs involved in immune infiltration in human septic cardiomyopathy by bioinformatics analysis.

Abstract: Septic cardiomyopathy (SCM) is a serious complication caused by sepsis that will further exacerbate the patient's prognosis. However, immune-related genes (IRGs) and their molecular mechanism during septic cardiomyopathy are largely unknown. Therefore, our study aims to explore the immune-related hub genes (IRHGs) and immune-related miRNA-mRNA pairs with potential biological regulation in SCM by means of bioinformatics analysis and experimental validation. Method: Firstly, screen differentially expressed mRNAs (DE-mRNAs) from the dataset GSE79962, and construct a PPI network of DE-mRNAs. Secondly, the hub genes of SCM were identified from the PPI network and the hub genes were overlapped with immune cell marker genes (ICMGs) to further obtain IRHGs in SCM. In addition, receiver operating characteristic (ROC) curve analysis was also performed in this process to determine the disease diagnostic capability of IRHGs. Finally, the crucial miRNA-IRHG regulatory network of IRHGs was predicted and constructed by bioinformatic methods. Real-time quantitative reverse transcription-PCR (qRT-PCR) and dataset GSE72380 were used to validate the expression of the key miRNA-IRHG axis. Result: The results of immune infiltration showed that neutrophils, Th17 cells, Tfh cells, and central memory cells in SCM had more infiltration than the control group; A total of 2 IRHGs were obtained by crossing the hub gene with the ICMGs, and the IRHGs were validated by dataset and qRT-PCR. Ultimately, we obtained the IRHG in SCM: THBS1. The ROC curve results of THBS1 showed that the area under the curve (AUC) was 0.909. Finally, the miR-222-3p/THBS1 axis regulatory network was constructed. Conclusion: In summary, we propose that THBS1 may be a key IRHG, and can serve as a biomarker for the diagnosis of SCM; in addition, the immune-related regulatory network miR-222-3p/THBS1 may be involved in the regulation of the pathogenesis of SCM and may serve as a promising candidate for SCM therapy.

MicroRNA-582-5p targeting Creb1 modulates apoptosis in cardiomyocytes hypoxia/reperfusion-induced injury.

BACKGROUND: Myocardial ischemia-reperfusion injury (MIRI) caused by the reperfusion therapy of myocardial ischemic diseases is a kind of major disease that threatens human health and lives severely. There are lacking of effective therapeutic measures for MIRI. MicroRNAs (miRNAs) are abundant in mammalian species and play a critical role in the initiation, promotion, and progression of MIRI. However, the biological role and molecular mechanism of miRNAs in MIRI are not entirely clear. METHODS: We used bioinformatics analysis to uncover the significantly different miRNA by analyzing transcriptome sequencing data from myocardial tissue in the mouse MIRI model. Multiple miRNA-related databases, including miRdb, PicTar, and TargetScan were used to forecast the downstream target genes of the differentially expressed miRNA. Then, the experimental models, including male C57BL/6J mice and HL-1 cell line, were used for subsequent experiments including quantitative real-time polymerase chain reaction analysis, western blot analysis, hematoxylin and eosin staining, flow cytometry, luciferase assay, gene interference, and overexpression. RESULTS: MiR-582-5p was found to be differentially upregulated from the transcriptome sequencing data. The elevated levels of miR-582-5p were verified in MIRI mice and hypoxia/reperfusion (H/R)-induced HL-1 cells. Functional experiments revealed that miR-582-5p promoted apoptosis of H/R-induced HL-1 cells via downregulating cAMP-response element-binding protein 1 (Creb1). The inhibiting action of miR-582-5p inhibitor on H/R-induced apoptosis was partially reversed after Creb1 interference. CONCLUSIONS: Collectively, the research findings reported that upregulation of miR-582-5p promoted H/R-induced cardiomyocyte apoptosis by inhibiting Creb1. The potential diagnostic and therapeutic strategies targeting miR-582-5p and Creb1 could be beneficial for the MIRI treatment.

Integrated bioinformatics analysis for novel miRNAs markers and ceRNA network in diabetic retinopathy.

In order to seek a more outstanding diagnosis and treatment of diabetic retinopathy (DR), we predicted the miRNA biomarkers of DR and explored the pathological mechanism of DR through bioinformatics analysis. Method: Based on public omics data and databases, we investigated ncRNA (non-coding RNA) functions based on the ceRNA hypothesis. Result: Among differentially expressed miRNAs (DE-miRNAs), hsa-miR-1179, -4797-3p and -665 may be diagnosis biomarkers of DR. Functional enrichment analysis revealed differentially expressed mRNAs (DE-mRNAs) enriched in mitochondrial transport, cellular respiration and energy derivation. 18 tissue/organ-specific expressed genes, 10 hub genes and gene cluster modules were identified. The ceRNA networks lncRNA FBXL19-AS1/miR-378f/MRPL39 and lncRNA UBL7-AS1/miR-378f/MRPL39 might be potential RNA regulatory pathways in DR. Conclusion: Differentially expressed hsa-miR-1179, -4797-3p and -665 can be used as powerful markers for DR diagnosis, and the ceRNA network: lncRNA FBXL19-AS1/UBL7-AS1-miR-378f-MRPL39 may represent an important regulatory role in DR progression.

CircRIC8B regulates the lipid metabolism of chronic lymphocytic leukemia through miR199b-5p/LPL axis.

OBJECTIVE: Circular RNAs (circRNAs) play a critical role in the modulation of tumor metabolism. However, the expression patterns and metabolic function of circRNAs in chronic lymphocytic leukemia (CLL) remain largely unknown. This study aimed to elucidate the role of circRNAs in the lipid metabolism of CLL. METHODS: The expression and metabolic patterns of circRNAs in a cohort of 53 patients with CLL were investigated using whole transcriptome sequencing. Cell viability, liquid chromatography with tandem mass spectrometry (LC-MS/MS) analysis, lipid analysis, Nile red staining as well as triglyceride (TG) assay were used to evaluate the biological function of circRIC8B in CLL. The regulatory mechanisms of circRIC8B/miR-199b-5p/lipoprotein lipase (LPL) axis were explored by luciferase assay, RNA immunoprecipitation (RIP), qRT-PCR, and fluorescence in situ hybridization (FISH). CCK-8 and flow cytometry were used to verify the inhibition role of cholesterol absorption inhibitor, ezetimibe, in CLL cells. RESULTS: Increased circRIC8B expression was positively correlated with advanced progression and poor prognosis. Knockdown of circRIC8B significantly suppressed the proliferation and lipid accumulation of CLL cells. In contrast, the upregulation of circRIC8B exerted opposite effects. Mechanistically, circRIC8B acted as a sponge of miR-199b-5p and prevented it from decreasing the level of LPL mRNA, and this promotes lipid metabolism alteration and facilitates the progression of CLL. What's more, ezetimibe suppressed the expression of LPL mRNA and inhibited the growth of CLL cells. CONCLUSIONS: In this study, the expressional and metabolic patterns of circRNAs in CLL was illustrated for the 1st time. Our findings revealed that circRIC8B regulates the lipid metabolism abnormalities in and development of CLL through the miR-199b-5p/LPL axis. CircRIC8B may serve as a promising prognostic marker and therapeutic target, which enhances the sensitivity to ezetimibe in CLL.

Identification and Validation of Immune-Related Biomarker Gene and Construction of ceRNA Networks in Septic Cardiomyopathy.

Septic cardiomyopathy (SCM) is a cardiac dysfunction caused by severe sepsis, which greatly increases the risk of heart failure and death, and its molecular mechanism is unclear. The immune response has been reported to be an important process in septic cardiomyopathy and is present in the cardiac tissue of patients with sepsis, suggesting that the immune response may be an underlying mechanism of myocardial injury in SCM. Therefore, we explored the role of immune-related genes (IRGs) in SCM and aimed to identify pivotal immune-related targets with the aim of identifying key immune-related targets in SCM and potential therapeutic mechanisms involved in the pathological process of SCM. To explore the regulatory mechanisms of immune responses in SCM, we identified differentially expressed genes (DEGs) shared in the SCM datasets GSE179554 and GSE40180 by bioinformatics analysis and then obtained hub genes from the DEGs. Then, we obtained the immune-related hub genes (IRHGs) by intersecting the hub genes with IRGs and performed quantitative reverse transcription polymerase chain reaction to confirm the abnormal expression of IRHGs. Finally, we further constructed an immune-related lncRNA-miRNA-IRHG ceRNA regulatory network. In this study, we identified an IRHG that may be involved in the pathogenesis of SCM, which helps us to further elucidate the role of immune response in SCM and gain insights into the molecular mechanisms and potential therapeutic targets of SCM.

Exploring the significance of PAK1 through chromosome conformation signatures in ibrutinib-resistant chronic lymphocytic leukaemia.

Ibrutinib exerts promising anticancer effects in chronic lymphocytic leukaemia (CLL). However, acquired resistance occurs during treatment, necessitating the exploration of underlying mechanisms. Although three-dimensional genome organization has been identified as a major player in the development and progression of cancer, including drug resistance, little is known regarding its role in CLL. Therefore, we investigated the molecular mechanisms underlying ibrutinib resistance through multi-omics analysis, including high-throughput chromosome conformation capture (Hi-C) technology. We demonstrated that the therapeutic response to ibrutinib is associated with the expression of p21-activated kinase 1 (PAK1). PAK1, which was up-regulated in CLL and associated with patients' survival, was involved in cell proliferation, glycolysis and oxidative phosphorylation. Furthermore, the PAK1 inhibitor IPA-3 exerted an anti-tumour effect and its combination with ibrutinib exhibited a synergistic effect in ibrutinib-sensitive and -resistant cells. These findings suggest the oncogenic role of PAK1 in CLL progression and drug resistance, highlighting PAK1 as a potential diagnostic marker and therapeutic target in CLL including ibrutinib-resistant CLL.

Single-cell profiling-guided combination therapy of c-Fos and histone deacetylase inhibitors in diffuse large B-cell lymphoma.

BACKGROUND: Diffuse large B-cell lymphoma (DLBCL) is the most common subtype of non-Hodgkin lymphoma. Histone deacetylase inhibitors (HDACis) have been widely applied in multiple tumours, but the expected efficacy was not observed in DLBCL. Therefore, this study is aimed to explore superior HDACis and optimise a relative combinational therapeutic strategy. METHODS: The antitumour effects of the drug were evaluated by Cell Counting Kit-8 (CCK-8) assay and apoptosis analysis. Single-cell RNA sequencing (scRNA-Seq) was used to analyse the intratumoural heterogeneity of DLBCL cells. Whole-exome sequencing and RNA sequencing were performed to analyse the genetic and transcriptional features. Western blotting, qRT-PCR, protein array, immunohistochemistry, and chromatin immunoprecipitation assays were applied to explore the involved pathways. The antitumour effects of the compounds were assessed using subcutaneous xenograft tumour models. RESULTS: LAQ824 was screened and confirmed to kill DLBCL cells effectively. Using scRNA-Seq, we characterised the heterogeneity of DLBCL cells under different drug pressures, and c-Fos was identified as a critical factor in the survival of residual tumour cells. Moreover, we demonstrated that combinatorial treatment with LAQ824 and a c-Fos inhibitor more potently inhibited tumour cells both in vitro and in vivo. CONCLUSION: Altogether, we found an HDACi, LAQ824, with high efficacy in DLBCL and provided a promising HDACi-based combination therapy strategy.

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[Effect and Involved Mechanism of RSL3-induced Ferroptosis in Acute Leukemia Cells MOLM13 and Drug-resistant Cell Lines].

OBJECTIVE: To investigate the effect and involved mechanism of RSL3 on ferroptosis action in acute leukemia cells MOLM13 and its drug-resistant cells. METHODS: After MOLM13 treated with RSL3, CCK-8 assay was performed to detect cell viability, flow cytometry was used to detect the reactive oxygen species (ROS) level of the cells, RT-qPCR and Western blot were used to detect the expression of glutathione peroxidase 4 (GPX4). After MOLM13/IDA and MOLM13/Ara-C, the drug-resistant cell lines were constructed, the ferroptosis induced by RSL3 was observed. Bone marrow samples were collected from patients with acute monocytic leukemia. RT-qPCR and Western blot were performed to detect the expression of related genes and proteins involved in ferroptosis pathway. RESULTS: RSL3 significantly inhibited the cell viability of MOLM13 and increased the intracellular ROS level, which were partially reversed by ferrostatin-1. The mRNA and protein expression of GPX4 decreased in MOLM13 treated with RSL3. RSL3 inhibited the viability of MOLM13/IDA and MOLM13/Ara-C cells more strongly than that of non-drug resistant cells, also increased the intracellular ROS level . The cytotoxic effects were partially reversed by ferrostatin-1. The mRNA and protein expressions of GPX4 in MOLM13/IDA and MOLM13/Ara-C cells were higher than those in non-drug resistant cells. The mRNA and protein levels of GPX4 in bone marrow of relapsed/refractory acute mononuclear leukemia patients were higher than those of ordinary acute mononuclear leukemia patients. CONCLUSION: RSL3 can induce non-drug resistant cells MOLM13 ferroptosis by inhibiting GPX4 activity. MOLM13/IDA and MOLM13/Ara-C are more sensitive to RSL3 compared with non-drug resistant cells MOLM13, which may be caused by the differences in GPX4 expression. The expressions of GPX4 mRNA and protein in relapsed/refractory acute mononuclear leukemia are higher than those in ordinary acute mononuclear leukemia.