ABSTRACT
PURPOSE: We have been developing a new type of miniscrew to specifically withstand orthodontic torque load. This study aimed to investigate the effect of thread depth and thread pitch on the primary stability of these miniscrews if stressed with torque load. METHODS: Finite element analysis (FEA) was used to evaluate the primary stability of the miniscrews. For thread depth analysis, the thread depth was set to 0.1-0.4â¯mm to construct 7 models. For thread pitch analysis, the thread pitch was set to 0.4-1.0â¯mm to construct another 7 models. A torque load of 6â¯Nmm was applied to the miniscrew, and the other parameters were kept constant for the analyses. Maximum equivalent stress (Max EQV) of cortical bone and maximum displacement of the miniscrews (Max DM) were the indicators for primary stability of the miniscrew in the 14 models. RESULTS: In the thread depth analysis, Max DM increased as the miniscrew thread depth increased, while Max EQV was smallest in model 3 (thread depthâ¯= 0.2, Max EQVâ¯= 8.91â¯MPa). In the pitch analysis, with an increase of the thread pitch, Max DM generally exhibited a trend to increase, while Max EQV of cortical bone showed a general trend to decrease. CONCLUSION: Considering the data of Max DM and Max EQV, the most appropriate thread depth and thread pitch of the miniscrews in our model was 0.2 and 0.7â¯mm, respectively. This knowledge may effectively improve the primary stability of newly developed miniscrews.
Subject(s)
Bone Screws , Orthodontic Anchorage Procedures , Torque , Stress, Mechanical , Finite Element AnalysisABSTRACT
Provoked by sterile/nonsterile insults, prolonged monocyte mobilization and uncontrolled monocyte/macrophage activation can pose imminent or impending harm to the affected organs. Curiously, folate receptor beta (FRß), with subnanomolar affinity for the vitamin folic acid (FA), is upregulated during immune activation in hematopoietic cells of the myeloid lineage. This phenomenon has inspired a strong interest in exploring FRß-directed diagnostics/therapeutics. Previously, we have reported that FA-targeted aminopterin (AMT) therapy can modulate macrophage function and effectively treat animal models of inflammation. Our current investigation of a lead compound (EC2319) leads to discovery of a highly FR-specific mechanism of action independent of the root causes against inflammatory monocytes. We further show that EC2319 suppresses interleukin-6/interleukin-1ß release by FRß+ monocytes in a triple co-culture leukemic model of cytokine release syndrome with anti-CD19 chimeric antigen receptor T cells. Because of its chemical stability and metabolically activated linker, EC2319 demonstrates favorable pharmacokinetic characteristics and cross-species translatability to support future pre-clinical and clinical development.
Subject(s)
Aminopterin/pharmacology , Cytokine Release Syndrome/prevention & control , Folate Receptor 2/genetics , Folic Acid Antagonists/pharmacology , Folic Acid/metabolism , Macrophages/drug effects , Animals , Antigens, CD19/genetics , Antigens, CD19/immunology , CHO Cells , Cricetulus , Cytokine Release Syndrome/genetics , Cytokine Release Syndrome/immunology , Cytokine Release Syndrome/pathology , Female , Folate Receptor 1/antagonists & inhibitors , Folate Receptor 1/genetics , Folate Receptor 1/immunology , Folate Receptor 2/antagonists & inhibitors , Folate Receptor 2/immunology , Humans , Interleukin-1beta/genetics , Interleukin-1beta/immunology , Interleukin-6/genetics , Interleukin-6/immunology , Macrophage Activation/drug effects , Macrophages/immunology , Macrophages/pathology , Mice , Models, Biological , Monocytes/drug effects , Monocytes/immunology , Monocytes/pathology , RAW 264.7 Cells , Rats , Rats, Inbred Lew , Receptors, Chimeric Antigen/genetics , Receptors, Chimeric Antigen/immunology , T-Lymphocytes/drug effects , T-Lymphocytes/immunology , T-Lymphocytes/pathologyABSTRACT
Since activated macrophages express a functional folate receptor ß (FRß), targeting this macrophage population with folate-linked drugs could increase selectivity to treat inflammatory diseases. Using a macrophage-mediated anti-glomerular basement membrane (anti-GBM) glomerulonephritis (GN) in WKY rats, we investigated the effect of a novel folic acid-aminopterin (AMT) conjugate (EC2319) designed to intracellularly deliver AMT via the FR. We found that treatment with EC2319 significantly attenuated kidney injury and preserved renal function. Kidney protection with EC2319 was blocked by a folate competitor, indicating that its mechanism of action was specifically FRß-mediated. Notably, treatment with methotrexate (MTX), another folic acid antagonist related to AMT, did not protect from kidney damage. EC2319 reduced glomerular and interstitial macrophage infiltration and decreased M1 macrophage recruitment but not M2 macrophages. The expression of CCL2 and the pro-fibrotic cytokine TGF-ß were also reduced in nephritic glomeruli with EC2319 treatment. In EC2319-treated rats, there was a significant decrease in the deposition of collagens. In nephritic kidneys, FRß was expressed on periglomerular macrophages and macrophages present in the crescents, but its expression was not observed in normal kidneys. These data indicate that selectively targeting the activated macrophage population could represent a novel means for treating anti-GBM GN and other acute crescentic glomerulonephritis.
Subject(s)
Folate Receptor 2/metabolism , Glomerulonephritis/drug therapy , Glomerulonephritis/metabolism , Inflammation/drug therapy , Inflammation/metabolism , Macrophages/metabolism , Aminopterin/chemistry , Aminopterin/therapeutic use , Animals , Fibrosis/drug therapy , Fibrosis/metabolism , Folic Acid/chemistry , Folic Acid/therapeutic use , Macrophages/drug effects , Methotrexate/therapeutic use , RatsABSTRACT
PURPOSE: The biological roles and clinical significance of RNA-binding proteins (RBPs) in oral squamous cell carcinoma (OSCC) are not fully understood. We investigated the prognostic value of RBPs in OSCC using several bioinformatic strategies. MATERIALS AND METHODS: OSCC data were obtained from a public online database, the Limma R package was used to identify differentially expressed RBPs, and functional enrichment analysis was performed to elucidate the biological functions of the above RBPs in OSCC. We performed protein-protein interaction (PPI) network and Cox regression analyses to extract prognosis-related hub RBPs. Next, we established and validated a prognostic model based on the hub RBPs using Cox regression and risk score analyses. RESULTS: We found that the differentially expressed RBPs were closely related to the defense response to viruses and multiple RNA processes. We identified 10 prognosis-related hub RBPs (ZC3H12D, OAS2, INTS10, ACO1, PCBP4, RNASE3, PTGES3L-AARSD1, RNASE13, DDX4, and PCF11) and effectively predicted the overall survival of OSCC patients. The area under the receiver operating characteristic (ROC) curve (AUC) of the risk score model was 0.781, suggesting that our model exhibited excellent prognostic performance. Finally, we built a nomogram integrating the 10 RBPs. The internal validation cohort results showed a reliable predictive capability of the nomogram for OSCC. CONCLUSION: We established a novel 10-RBP-based model for OSCC that could enable precise individual treatment and follow-up management strategies in the future.
ABSTRACT
BACKGROUND: Activated macrophages in the experimental model of multiple sclerosis (MS) express folate receptor-ß (FR-ß), representing a promising target for the treatment of MS. Here, we both evaluated the efficacy of a novel folate-aminopterin construct (EC2319) in a rat focal model of multiple sclerosis (MS) and investigated the utility of 68Ga-labeled 1,4,7-triazacyclononane-1,4,7-triacetic acid-conjugated folate (68Ga-FOL) for assessing inflammatory lesions. In addition, we investigated whether FR-ß is expressed in the brain of patients with MS. METHODS: Focal delayed-type hypersensitivity experimental autoimmune encephalomyelitis (fDTH-EAE) was induced in 40 Lewis rats; 20 healthy Lewis rats were used as controls. Rats were divided into six groups according to the duration of disease (control, acute, or chronic) and intervention (vehicle versus EC2319). 68Ga-FOL analyses, histology, and immunofluorescence of the brain were performed to evaluate the efficacy of subcutaneously administered EC2319 on lesion development. Immunofluorescence was used to assess FR-ß expression in postmortem brain samples from 5 patients with MS and 5 healthy controls. RESULTS: Immunofluorescence and histological analyses revealed significant reductions in FR-ß expression (P < 0.05) and lesion size (P < 0.01), as well as improved inducible nitric oxide synthase/mannose receptor C type 1 ratios (P < 0.01) in macrophages and microglia during the chronic but not acute phase of fDTH-EAE in EC2319-treated rats. The uptake of IV-injected 68Ga-FOL in the brain was low and did not differ between the groups, but the in vitro binding of 68Ga-FOL was significantly lower in EC2319-treated rats (P < 0.01). FR-ß positivity was observed in chronically active lesions and in normal-appearing white matter in MS brain samples. CONCLUSIONS: EC2319 was well tolerated and attenuated inflammation and lesion development in a rat model of a chronic progressive form of MS. Human MS patients have FR-ß-positive cells in chronically active plaques, which suggests that these results may have translational relevance.
Subject(s)
Aminopterin/pharmacology , Encephalomyelitis, Autoimmune, Experimental/pathology , Folate Receptor 2/metabolism , Folic Acid Antagonists/pharmacology , Folic Acid/pharmacology , Animals , Humans , Multiple Sclerosis/metabolism , Rats , Rats, Inbred LewABSTRACT
This study aimed to investigate the effect of miniscrews thread shape on the stress distribution receiving a torque load. Seven thread shapes (S,V1,V2,B1,B2,R1,R2) models were constructed and a 6 Nmm-torque load was applied. The order of maximum equivalent stress (EQV) value was V1 > V2 > B1 > R1 > R2 > B2 > S. The order of maximum displacement of miniscrew (Max DM) value was S > B2 > R1 = V1 > B1 > V2 > R2. Model R2 may be the most appropriate thread shape affording a torque force.
Subject(s)
Bone Screws , Finite Element Analysis , Orthodontic Appliance Design , Stress, Mechanical , Torque , Biomechanical Phenomena , Cortical Bone/pathology , Dental Stress Analysis , HumansABSTRACT
Hepatitis B virus (HBV) infection plays an important role in hepatocarcinogenesis, especially in hepatocellular carcinoma (HCC). Long non-coding RNAs (lncRNAs) have emerged as crucial biomarkers and regulators in many cancers. Novel lncRNAs involved in the initiation and progression of HBV-related hepatocellular carcinoma (HCC) need to be investigated. Here, we report that the long non-coding RNA LINC01352 is markedly downregulated by HBV/HBx (HBV X protein) in HCC cells and clinical samples. The LINC01352 expression level in HCC is an independent prognostic factor for survival. We found that HBx suppresses LINC01352 promoter activity by forming a complex with the estrogen receptor (ERα). Furthermore, using a combination of in vitro and in vivo studies, we confirmed that HBx promotes HCC cell growth and metastasis by inhibiting LINC01352 expression. Further investigation revealed that the downregulation of LINC01352, which acts as an endogenous sponge, increases the expression of miR-135b, leading to the reduced production of adenomatous polyposis coli (APC), consequently activating Wnt/ß-catenin signalling to facilitate tumour progression. These findings strongly suggest that the LINC01352-miR-135b-APC axis regulated by the HBx/ERα complex acts as an important pathogenic factor for tumour progression, which may help provide a theoretical basis for the identification of new therapeutic targets for HBV-related HCC.
Subject(s)
Adenomatous Polyposis Coli Protein/genetics , Carcinoma, Hepatocellular/genetics , Liver Neoplasms/genetics , MicroRNAs/genetics , RNA, Long Noncoding/genetics , Aged , Carcinoma, Hepatocellular/pathology , Carcinoma, Hepatocellular/virology , Cell Line, Tumor , Estrogen Receptor alpha/genetics , Female , Hepatitis B/complications , Hepatitis B/genetics , Hepatitis B/pathology , Hepatitis B/virology , Hepatitis B virus/genetics , Hepatitis B virus/pathogenicity , Humans , Liver Neoplasms/pathology , Liver Neoplasms/virology , Male , Middle Aged , Promoter Regions, Genetic/genetics , Signal Transduction/genetics , Trans-Activators/genetics , Viral Regulatory and Accessory Proteins/geneticsABSTRACT
Lymphomatoid granulomatosis (LYG) is an extremely rare angio-centric and angio-destructive B-cell lymphoproliferative disease. Driven by Epstein-Barr virus (EBV), LYG predominantly involves the bilateral lungs. Commonly presenting as multiple nodules in the lung, pulmonary LYG can masquerade as various infectious diseases, vasculitis, lung cancer, or other metastatic neoplasm. It is difficult to be diagnosed and is always neglected by clinicians. No standardized therapeutic regimens for LYG has been established yet now. Hemophagocytic lymphohistiocytosis (HLH), a life-threatening condition caused by abnormal activation of macrophages and T-cells, is characterized by fever, hepatosplenomegaly, pancytopenia, hypercytokinemia, and the presence of hemophagocytosis within the bone marrow, liver, spleen, or other lymphatic tissue. We herein report a 55-year-old man with recurrent fever, severe jaundice, and multiple high-density opacities and nodules in both lungs, who was finally diagnosed with pulmonary LYG (Grade 3) manifested with secondary HLH. Administration of HLH-1994 protocol led to the rapid control of the symptoms caused by HLH. Rituximab-based combination therapy was useful yet LYG (Grade 3) progressed rapidly. This case demonstrates that tissue biopsy is essential for early pathological diagnosis and effective treatment of LYG.
ABSTRACT
OBJECTIVE: To explore the effect and mechanism of miR-30b on cisplatin-resistance of human NK/T cell lymphoma lines SNK-6 and YTS cells. METHODS: Normal NK cells, SNK-6 and YTS cells were cultured, the expression levels of miR-30b and macrophage-derived chemokine (CCL22) were detected by real-time PCR assay, and the CCL22 expression was detected by Western blot. The SNK-6 and YTS cells were transfected with miR-30b mimics and inhibitor respectively, then the effect of cisplatin resistance in SNK-6 and YTS cells was measured by MTT assay, the activity of caspase-3 was detected by caspase-3 assay kit, and the cell apoptosis was analyzed by flow cytometry. Dual-luciferase reporter gene assay was used to determine the targeting relationship between miR-30b and CCL22. Furthermore, the effect of CCL22 on cisplatin-resistance and caspase-3 actirity was also evaluated. RESULTS: Compared with the normal NK cells, the expression levels of miR-30b significantly decreased in both SNK-6 and YTS cells (Pï¼0.01), but CCL22 mRNA expression increase in both cells (Pï¼0.01). MiR-30b mimics decreased the cell activity (Pï¼0.05), down-regulated the cisplatin-resistance (Pï¼0.05), and increased cell apoptosis and caspase-3 activity (Pï¼0.05). The effects of miR-30b inhibitor were contrary to the mimics. Up-regulation of miR-30b expression significantly decreased the luciferase activity in CCL22 3'-UTR-transfected NK cells, but not in Mut-CCL22 3'UTR group, suggesting that CCL22 could act as a direct target of miR-30b. The expressions of CCL22 pathway proteins were down-regulated after SNK-6 cells transfected with miR-30b mimics (Pï¼0.05), while this effect was restored by overexpression of CCL22. Moreover, CCL22 overexpression also increased the cell activity and decreased caspase-3 activity when SNK-6 cells were transfected with miR-30b mimics. CONCLUSION: MiR-30b inhibits cisplatin-resistance of human NK/TCL SNK-6 and YTS cells by targeting CCL22.
Subject(s)
Lymphoma, T-Cell/genetics , Apoptosis , Cell Line, Tumor , Cell Proliferation , Chemokine CCL22 , Cisplatin , Gene Expression Regulation, Neoplastic , Humans , Killer Cells, Natural , MicroRNAs , T-LymphocytesABSTRACT
MicroRNAs (miRNAs) have been recently found in the mitochondria, and were named "mitomiRs", but their function has remained elusive. Here, we aimed to assess the presence and function(s) of mitomiRs in tongue squamous cell carcinoma (TSCC). Methods: miRNA microarray was performed in paired TSCC cell lines, Cal27 and its chemoresistant counterpart, Cal27-re. Decreased expression of mitomiRs in chemoresistant cells was characterized. The functions of mitomiRs were investigated by a series of in vitro and in vivo experiments. Results: Differential microarray analysis identified downregulation of mitomiR-5787 in Cal27-re cells. We knocked down mitomiR-5787 in parental cells and upregulated its expression in cisplatin-resistant cells. The sensitivity of TSCC cells to cisplatin was regulated by miR-5787. The glucose metabolism assay suggested that reduced expression of miR-5787 changed the balance of glucose metabolism by shifting it from oxidative phosphorylation to aerobic glycolysis. Xenograft experiments in BALB/c-nu mice further verified the in vitro results. Reduced expression of miR-5787 contributes to chemoresistance in TSCC cells by inhibiting the translation of mitochondrial cytochrome c oxidase subunit 3 (MT-CO3). The prognostic analysis of 126 TSCC patients showed that the patients with low expression of miR-5787 and/or MT-CO3 had poor cisplatin sensitivity and prognosis. Conclusions: Mitochondrial miR-5787 could regulate cisplatin resistance of TSCC cells and affect oxidative phosphorylation and aerobic glycolysis. Downregulation of miR-5787 inhibited the translation of MT-CO3 to regulate cisplatin resistance of TSCC. Mitochondrial miR-5787 and MT-CO3 can be used as predictive biomarkers or therapeutic targets for cisplatin chemotherapy resistance.
Subject(s)
Carcinoma, Squamous Cell/genetics , Cytochrome c Group/metabolism , MicroRNAs/metabolism , Mitochondria/metabolism , Tongue Neoplasms/genetics , Animals , Apoptosis/drug effects , Apoptosis/genetics , Carcinoma, Squamous Cell/drug therapy , Carcinoma, Squamous Cell/metabolism , Cell Line, Tumor , Cisplatin/therapeutic use , Drug Resistance, Neoplasm/genetics , Female , Gene Expression Regulation, Neoplastic/drug effects , Gene Expression Regulation, Neoplastic/genetics , Glucose/metabolism , Glycolysis/drug effects , Glycolysis/genetics , Humans , Male , Mice , Mice, Inbred BALB C , MicroRNAs/genetics , Oxidative Phosphorylation , Tongue Neoplasms/drug therapy , Tongue Neoplasms/metabolism , Xenograft Model Antitumor AssaysABSTRACT
Folate-based small molecule drug conjugates (SMDCs) are currently under development and have shown promising preclinical and clinical results against various cancers and polycystic kidney disease. Two requisites for response to a folate-based SMDC are (i) folate receptor alpha (FRα) protein is expressed in the diseased tissues, and (ii) FRα in those tissues is accessible and functionally competent to bind systemically administered SMDCs. Here we report on the development of a small molecule reporter conjugate (SMRC), called EC2220, which is composed of a folate ligand for FRα binding, a multilysine containing linker that can cross-link to FRα in the presence of formaldehyde fixation, and a small hapten (fluorescein) used for immunohistochemical detection. Data show that EC2220 produces a far greater IHC signal in FRα-positive tissues over that produced with EC17, a folate-fluorescein SMRC that is released from the formaldehyde-denatured FRα protein. Furthermore, the extent of the EC2220 IHC signal was proportional to the level of FRα expression. This EC2220-based assay was qualified both in vitro and in vivo using normal tissue, cancer tissue, and polycystic kidneys. Overall, EC2220 is a sensitive and effective reagent for evaluating functional and accessible receptor expression in vitro and in vivo.
Subject(s)
Folate Receptor 1/metabolism , Molecular Targeted Therapy/methods , Neoplasms/drug therapy , Polycystic Kidney Diseases/drug therapy , A549 Cells , Animals , Doxycycline/pharmacology , Fluorescein-5-isothiocyanate/chemistry , Fluorescein-5-isothiocyanate/metabolism , Folate Receptor 1/analysis , Folic Acid/analogs & derivatives , Folic Acid/chemistry , Folic Acid/metabolism , HeLa Cells , Humans , Lysine/analogs & derivatives , Lysine/chemistry , Lysine/metabolism , Mice , Mice, Inbred BALB C , Mice, Nude , Mice, Transgenic , Neoplasms/metabolism , Oligopeptides/chemistry , Oligopeptides/metabolism , Polycystic Kidney Diseases/chemically induced , Polycystic Kidney Diseases/metabolism , Protein Kinase C/genetics , Tissue Distribution , Trityl Compounds/chemistry , Trityl Compounds/metabolism , Xenograft Model Antitumor AssaysABSTRACT
BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most aggressive malignant tumors, with a poor long-term prognosis worldwide. The functional deregulations of global transcriptome were associated with the genesis and development of HCC, but lacks systematic research and validation. METHODS: A total of 519 postoperative HCC patients were included. We built an interactive and visual competing endogenous RNA network. The prognostic signature was established with the least absolute shrinkage and selection operator algorithm. Multivariate Cox regression analysis was used to screen for independent prognostic factors for HCC overall survival. RESULTS: In the training set, we identified a four-gene signature (PBK, CBX2, CLSPN, and CPEB3) and effectively predicted the overall survival. The survival times of patients in the high-score group were worse than those in the low-score group (p = 0.0004), and death was also more likely in the high-score group (HR 2.444, p < 0.001). The results were validated in internal validation set (p = 0.0057) and two external validation cohorts (HR 2.467 and 2.6). The signature (AUCs of 1, 2, 3 years were 0.716, 0.726, 0.714, respectively) showed high prognostic accuracy in the complete TCGA cohort. CONCLUSIONS: In conclusion, we successfully built a more extensive ceRNA network for HCC and then identified a four-gene-based signature, enabling prediction of the overall survival of patients with HCC.
Subject(s)
Carcinoma, Hepatocellular/genetics , Genes/genetics , Liver Neoplasms/genetics , RNA/genetics , Adaptor Proteins, Signal Transducing/genetics , Carcinoma, Hepatocellular/diagnosis , Carcinoma, Hepatocellular/mortality , Female , Gene Expression Regulation, Neoplastic/genetics , Humans , Liver Neoplasms/diagnosis , Liver Neoplasms/mortality , Male , Middle Aged , Mitogen-Activated Protein Kinase Kinases/genetics , Polycomb Repressive Complex 1/genetics , Prognosis , Proportional Hazards Models , RNA-Binding Proteins/genetics , Real-Time Polymerase Chain Reaction , Survival AnalysisABSTRACT
Although chimeric antigen receptor (CAR) T cell therapies have demonstrated considerable success in treating hematologic malignancies, they have simultaneously been plagued by a cytokine release syndrome (CRS) that can harm or even kill the cancer patient. We describe a CAR T cell strategy in which CAR T cell activation and cancer cell killing can be sensitively regulated by adjusting the dose of a low molecular weight adapter that must bridge between the CAR T cell and cancer cell to initiate tumor eradication. By controlling the concentration and dosing schedule of adapter administration, we document two methods that can rapidly terminate (<3 h) a pre-existing CRS-like toxicity and two unrelated methods that can pre-emptively prevent a CRS-like toxicity that would have otherwise occurred. Because all four methods concurrently enhance CAR T cell potency, we conclude that proper use of bispecific adapters could potentially avoid a life-threatening CRS while enhancing CAR T cell tumoricidal activity.
Subject(s)
Immune System Diseases/prevention & control , Immunotherapy, Adoptive/adverse effects , Neoplasms/therapy , Receptors, Chimeric Antigen/immunology , T-Lymphocytes/immunology , Animals , Cell Engineering/methods , Cell Line, Tumor , Cytokines/immunology , Fluorescein/metabolism , Folate Receptors, GPI-Anchored/metabolism , Folic Acid/metabolism , Humans , Immune System Diseases/etiology , Immunotherapy, Adoptive/methods , Lymphocyte Activation/immunology , Mice , Neoplasms/immunology , Receptors, Chimeric Antigen/metabolism , Single-Chain Antibodies/immunology , Single-Chain Antibodies/metabolism , Syndrome , T-Lymphocytes/metabolism , T-Lymphocytes/transplantation , Xenograft Model Antitumor AssaysABSTRACT
Chimeric antigen receptor (CAR)-T cell therapy has transformed pediatric oncology by producing high remission rates and potent effects in CD19+ B-cell malignancies. This scenario is ideal as CD19 expression is homogeneous and human blood provides a favorable environment for CAR-T cells to thrive and destroy cancer cells (along with normal B cells). Yet, CAR-T cell therapies for solid tumors remain challenged by fewer tumor targets and poor CAR-T cell performances in a hostile tumor microenvironment. For acute myeloid leukemia and childhood solid tumors such as osteosarcoma, the primary treatment is systemic chemotherapy that often falls short of expectation especially for relapsed and refractory conditions. We aim to develop a CAR-T adaptor molecule (CAM)-based therapy that uses a bispecific small-molecule ligand EC17, fluorescein isothiocyanate (FITC) conjugated with folic acid, to redirect FITC-specific CAR-T cells against folate receptor (FR)-positive tumors. As previously confirmed in rodents as well as in human clinical studies, EC17 penetrates solid tumors within minutes and is retained due to high affinity for the FR, whereas unbound EC17 rapidly clears from the blood and from receptor-negative tissues. When combined with a rationally designed CAR construct, EC17 CAM was shown to trigger CAR-modified T cell activation and cytolytic activity with a low FR threshold against tumor targets. However, maximal cytolytic potential correlated with (i) functional FR levels (in a semi-log fashion), (ii) the amount of effector cells present, and (iii) tumors' natural sensitivity to T cell mediated killing. In tumor-bearing mice, administration of EC17 CAM was the key to drive CAR-T cell activation, proliferation, and persistence against FR+ pediatric hematologic and solid tumors. In our modeling systems, cytokine release syndrome (CRS) was induced under specific conditions, but the risk of severe CRS could be easily mitigated or prevented by applying intermittent dosing and/or dose-titration strategies for the EC17 CAM. Our approach offers the flexibility of antigen control, prevents T cell exhaustion, and provides additional safety mechanisms including rapid reversal of severe CRS with intravenous sodium fluorescein. In this paper, we summarize the translational aspects of our technology in support of clinical development.
ABSTRACT
PURPOSE: The overall biological roles and clinical significance of most long noncoding RNAs (lncRNA) in chemosensitivity are not fully understood. We investigated the biological function, mechanism, and clinical significance of lncRNA NR_034085, which we termed miRNA processing-related lncRNA (MPRL), in tongue squamous cell carcinoma (TSCC). EXPERIMENTAL DESIGN: LncRNA expression in TSCC cell lines with cisplatin treatment was measured by lncRNA microarray and confirmed in TSCC tissues. The functional roles of MPRL were demonstrated by a series of in vitro and in vivo experiments. The miRNA profiles, RNA pull-down, RNA immunoprecipitation, serial deletion analysis, and luciferase analyses were used to investigate the potential mechanisms of MPRL. RESULTS: We found that MPRL expression was significantly upregulated in TSCC cell lines treated with cisplatin and transactivated by E2F1. MPRL controlled mitochondrial fission and cisplatin sensitivity through miR-483-5p. In exploring the underlying interaction between MPRL and miR-483-5p, we identified that cytoplasmic MPRL directly binds to pre-miR-483 within the loop region and blocks pre-miR-483 recognition and cleavage by TRBP-DICER-complex, thereby inhibiting miR-483-5p generation and upregulating miR-483-5p downstream target-FIS1 expression. Furthermore, overexpression or knockdown MPRL altered tumor apoptosis and growth in mouse xenografts. Importantly, we found that high expression of MPRL and pre-miR-483, and low expression of miR-483-5p were significantly associated with neoadjuvant chemosensitivity and better TSCC patients' prognosis. CONCLUSIONS: We propose a model in which lncRNAs impair microprocessor recognition and are efficient of pre-miRNA cropping. In addition, our study reveals a novel regulatory network for mitochondrial fission and chemosensitivity and new biomarkers for prediction of neoadjuvant chemosensitivity in TSCC.These findings uncover a novel mechanism by which lncRNA determines mitochondrial fission and cisplatin chemosensitivity by inhibition of pre-miRNA processing and provide for the first time the rationale for lncRNA and miRNA biogenesis for predicting chemosensitivity and patient clinical prognosis.
Subject(s)
Cisplatin/pharmacology , MicroRNAs/genetics , Mitochondrial Dynamics/genetics , RNA, Long Noncoding/genetics , Squamous Cell Carcinoma of Head and Neck/drug therapy , Squamous Cell Carcinoma of Head and Neck/genetics , Tongue Neoplasms/drug therapy , Tongue Neoplasms/genetics , Animals , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Drug Resistance, Neoplasm , Female , Humans , Male , Mice , Mice, Inbred BALB C , Mice, Nude , Middle Aged , Prognosis , Squamous Cell Carcinoma of Head and Neck/pathology , Tongue Neoplasms/pathologyABSTRACT
Repulsive guidance molecules comprise a group of proteins that play an important role in carcinogenesis through interactions with their receptors, but their function in oral squamous cell carcinoma (OSCC) is unclear. Here, we investigated the potential role of the RGM family members in oral cancer pathogenesis. Our study showed that only RGMA was significantly downregulated in the OSCC tissues analyzed by TCGA and validated this finding in OSCC cells. The decreased expression of RGMA was strongly associated with the T stage and with poor prognosis. The ectopic expression of RGMA significantly inhibited the proliferation of OSCC cells both in vitro and in vivo. Moreover, we confirmed that RGMA was a target of miR-210-3p in OSCC and miR-210-3p overexpression contributed to the acceleration of OSCC growth. Further experiments revealed that HIF1A specifically interacted with the promoter of miR-210-3p and enhanced its expression. In summary, our research indicates that RGMA is regulated by the HIF1A/miR-210-3p axis and inhibits OSCC cell proliferation; thus, in the future, the development of therapies that target the HIF1A/miR-210-3p/RGMA axis may aid in the treatment of aggressive cancers.
Subject(s)
Carcinoma, Squamous Cell/metabolism , Cell Proliferation/physiology , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , MicroRNAs/metabolism , Mouth Neoplasms/metabolism , Nerve Tissue Proteins/metabolism , Adult , Animals , Carcinoma, Squamous Cell/pathology , Cell Line, Tumor , Cohort Studies , Down-Regulation/physiology , Female , GPI-Linked Proteins/metabolism , Humans , Male , Mice , Mice, Inbred BALB C , Mice, Nude , Middle Aged , Mouth Neoplasms/pathology , Xenograft Model Antitumor Assays/methodsABSTRACT
T cells are involved in bone marrow failure in aplastic anemia (AA). MEG3 is a long, non-coding RNA that can modulate target gene expression and T cell differentiation by acting as a microRNA sponge. Our previous study showed that T cell immunoglobulin and immunoreceptor tyrosine-based inhibition motif (ITIM) domain (TIGIT) plays a critical role in regulating CD4 + T cell functions. In this study, we found that MEG3 expression was significantly downregulated in CD4 + T cells derived from AA patients. MEG3 modulated CD4 + T cell proliferation and IFN-γ and TNF-α levels, as well as TIGIT, T-bet, and orphan nuclear receptor (RORγt) expression. Furthermore, MEG3 overexpression sequestered miR-23a and prompted TIGIT expression in CD4 + T cells. CD4 + T cells with MEG3 overexpression impeded expansion of Th1 and Th17 cells, restored the decreased red blood cell count, attenuated the increase in serum INF-γ and TNF-α levels, and lengthened median survival time, as well as upregulated mRNA levels of CD34, stem cell factor (SCF), and granulocyte/macrophage-colony-stimulating factor (GM-CSF) in bone marrow mononuclear cells of a mouse model. In conclusion, our study provides evidence that MEG3 regulated TIGIT expression and CD4 + T cell activation by absorbing miR-23a. These findings provide novel insight into autoimmune-mediated AA.
Subject(s)
Anemia, Aplastic/metabolism , CD4-Positive T-Lymphocytes/metabolism , Lymphocyte Activation , MicroRNAs/metabolism , RNA, Long Noncoding/metabolism , Receptors, Immunologic/genetics , Adult , Anemia, Aplastic/genetics , Anemia, Aplastic/immunology , Animals , CD4-Positive T-Lymphocytes/immunology , Cells, Cultured , Disease Models, Animal , Female , Gene Expression Regulation , Humans , Male , Mice , Young AdultABSTRACT
BACKGROUND: Tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6) has been proved to play an important role in tumorigenesis, invasion, and metastasis. However, its precise role salivary adenoid cystic carcinoma (SACC) has not been determined. The aim of this study was to explore the role of TRAF6 in SACC including invasion and metastasis of SACC cells. MATERIALS AND METHODS: Immunohistochemistry and quantitative real-time PCR were performed in SACC tissues paired with their adjacent normal tissues to analyze the expression of TRAF6. Downstream proteins expression was explored when TRAF6 was knockdown by siRNA. RESULTS: The results show that TRAF6 is upregulated in SACC samples, especially in SACC with metastasis, which is closely correlated with an aggressive phenotype (P = .0073) and shorter life survival span (P = .0061) in SACC patients. Knockdown of TRAF6 can attenuate the promotion effect of SACC cell invasion induced by TGF-ß. Western blot results also showed that silencing TRAF6 expression can inhibit the activation of SMAD2, SMAD3, ERK, p38, and JNK induced by TGF-ß in SACC cells. CONCLUSION: These data suggested that TRAF6 regulates TGF-ß-mediated SACC progression through SMAD2/3-ERK-p38-JNK cascades.
Subject(s)
Carcinoma, Adenoid Cystic/metabolism , MAP Kinase Signaling System , Salivary Gland Neoplasms/metabolism , TNF Receptor-Associated Factor 6/metabolism , Transforming Growth Factor beta/pharmacology , Carcinogenesis , Carcinoma, Adenoid Cystic/genetics , Carcinoma, Adenoid Cystic/pathology , Cell Line, Tumor , Disease Progression , Female , Humans , Intracellular Signaling Peptides and Proteins , Male , Middle Aged , Neoplasm Invasiveness , Salivary Gland Neoplasms/genetics , Salivary Gland Neoplasms/pathology , Smad2 Protein/metabolism , Smad3 Protein/metabolism , TNF Receptor-Associated Factor 6/biosynthesis , TNF Receptor-Associated Factor 6/genetics , Transforming Growth Factor beta/metabolism , Up-Regulation , p38 Mitogen-Activated Protein Kinases/metabolismABSTRACT
Salivary adenoid cystic carcinoma (SACC) is a relatively uncommon epithelial-like malignancy that can occur in the head and neck region. Despite its slow growth, this aggressive salivary gland tumor frequently recurs and metastasizes to distant organs since lacking effective chemotherapy treatment. MicroRNAs are key regulators in tumor metastasis and progression, but their roles during SACC progression have not been illustrated. In current study, we demonstrate that miR-125a-5p is down-regulated in SACC and closely related to the metastasis and progression in human SACC specimens. In vitro, miR-125a-5p mimic can suppress SACC cell migration and invasion; while blocking miR-125a-5p can relieve the inhibition effect. By using dual-luciferase assay, we confirmed that miR-125a-5p directly targeted to p38 and tissue samples of patients indicated the negative correlation between miR-125a-5p and p38; clinical analysis also showed that low level expression of miR-125a-5p is closely associated with poor prognosis of SACC. Furthermore, down-regulation of miR-125a-5p triggered downstream p38/JNK/ERK activation. Taken together, our results indicate that down-regulation of miR-125a-5p promotes SACC progression through p38 signal pathway and miR-125a-5p can be a potential therapeutic target of SACC.