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Biochemical Genetics Jul 2024Laryngeal cancer is a common malignancy of the larynx with a generally poor prognosis. This study systematically assessed the functional role of lncRNA BBOX1-AS1 in...
Laryngeal cancer is a common malignancy of the larynx with a generally poor prognosis. This study systematically assessed the functional role of lncRNA BBOX1-AS1 in laryngeal carcinoma progression and associated molecular regulatory mechanisms. The proliferation, migration, and invasion of laryngeal carcinoma cells were detected by Cell Counting Kit-8, wound healing, clonal formation, and transwell assays. In addition, the interaction between BBOX1-AS1, Serine/Arginine Splicing Factor 1 (SRSF1), and Ephrin-B2 (EFNB2) mRNA was examined employing RNA immunoprecipitation and RNA pull-down experiments. Furthermore, western blotting, and RT-qPCR assays were adopted to detect the expression levels of BBOX1-AS1, SRSF1, and EFNB2. The impact of BBOX1-AS1 and SRSF1 on EFNB2 mRNA stability was examined using the RNA stability assay. BBOX1-AS1 was highly expressed in human laryngeal carcinoma tissues and cell lines. BBOX1-AS1 knockdown suppressed the growth, proliferation, migration, and invasion of laryngeal carcinoma cells. BBOX1-AS1 maintained the stability of EFNB2 mRNA in laryngeal carcinoma cells by recruiting SRSF1. EFNB2 knockdown inhibited the growth and metastatic function of laryngeal carcinoma cells in vitro. EFNB2 overexpression reversed the influence of BBOX1-AS1 knockdown on laryngeal cancer tumorigenesis. BBOX1-AS1 maintained EFNB2 mRNA stability by recruiting SRSF1, thereby aggravating laryngeal carcinoma malignant phenotypes. BBOX1-AS1 might be a new theoretical target for the treatment of laryngeal carcinoma.
PubMed: 38965134
DOI: 10.1007/s10528-024-10879-2 -
Journal of Thrombosis and Thrombolysis Jul 2024Plasma protein carbonylation that reflects oxidative stress has been demonstrated to be associated with the prothrombotic fibrin clot phenotype. However, the role of...
INTRODUCTION
Plasma protein carbonylation that reflects oxidative stress has been demonstrated to be associated with the prothrombotic fibrin clot phenotype. However, the role of protein carbonyls (PC) in predicting ischemic stroke in atrial fibrillation (AF) is largely unknown. This study aimed to investigate whether PC increase the risk of stroke in anticoagulated AF patients during follow-up.
METHODS
In 243 AF patients on anticoagulation (median age 69 years; median CHADS-VASc of 4), we measured plasma PC using the assay by Becatti, along with plasma clot permeability (K), clot lysis time (CLT), thrombin generation, and fibrinolytic proteins, including plasminogen activator inhibitor type 1 (PAI-1) and thrombin activatable fibrinolysis inhibitor (TAFI). Ischemic stroke, major bleeding, and mortality were recorded during a median follow-up of 53 months.
RESULTS
Plasma PC levels (median, 3.16 [2.54-3.99] nM/mg protein) at baseline showed positive associations with age (P < 0.001), CHADS-VASc (P = 0.003), and N-terminal B-type natriuretic peptide (P = 0.001), but not with type of AF or comorbidities except for heart failure (P = 0.007). PC levels were correlated with CLT (r = 0.342, P < 0.001), endogenous thrombin potential (r = 0.217, P = 0.001) and weakly with Ks (r = -0.145, P = 0.024), but not with fibrinogen, PAI-1, or TAFI levels. Stroke was recorded in 20 patients (1.9%/year), who had at baseline 36% higher PC levels (P < 0.001). Elevated PC (P = 0.003) at baseline were independently associated with stroke risk.
CONCLUSION
Our findings suggest that in patients with AF enhanced protein carbonylation is associated with increased "residual" risk of stroke despite anticoagulation, which is at least in part due to unfavorably altered fibrin clot phenotype.
PubMed: 38965130
DOI: 10.1007/s11239-024-03003-z -
European Urology Oncology Jul 2024Salvage radiation therapy (SRT) is a mainstay of treatment for biochemical relapse following radical prostatectomy; however, few studies have examined genomic biomarkers...
BACKGROUND
Salvage radiation therapy (SRT) is a mainstay of treatment for biochemical relapse following radical prostatectomy; however, few studies have examined genomic biomarkers in this context.
OBJECTIVE
We characterized the prognostic impact of previously identified deleterious molecular phenotypes-loss of PTEN, ERG expression, and TP53 mutation-for patients undergoing SRT.
DESIGN, SETTING, AND PARTICIPANTS
We leveraged an institutional database of 320 SRT patients with available tissue and follow-up. Tissue microarrays were used for genetically validated immunohistochemistry assays.
INTERVENTION
All men underwent SRT with or without androgen deprivation therapy OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Univariable and multivariable Cox-proportional hazard models assessed the association of molecular phenotypes with biochemical recurrence-free (bRFS) and metastasis-free (MFS) survival after SRT.
RESULTS AND LIMITATIONS
Loss of PTEN (n = 123, 43%) and ERG expression (n = 118, 39%) were common in this cohort, while p53 overexpression (signifying TP53 missense mutation) was infrequent (n = 21, 7%). In univariable analyses, any loss of PTEN portended worse bRFS (hazard ratio [HR] 1.86; 95% confidence interval 1.36-2.57) and MFS (HR 1.89; 1.21-2.94), with homogeneous PTEN loss being associated with the highest risk of MFS (HR 2.47; 1.54-3.95). Similarly, p53 overexpression predicted worse bRFS (HR 1.95; 1.14-3.32) and MFS (HR 2.79; 1.50-5.19). ERG expression was associated with worse MFS only (HR 1.6; 1.03-2.48). On the multivariable analysis adjusting for known prognostic features, homogeneous PTEN loss remained predictive of adverse bRFS (HR 1.82; 1.12-2.96) and MFS (HR 2.08; 1.06-4.86). The study is limited by its retrospective and single-institution design.
CONCLUSIONS
PTEN loss by immunohistochemistry is an independent adverse prognostic factor for bRFS and MFS in prostate cancer patients treated with SRT. Future trials will determine the optimal approach to treating SRT patients with adverse molecular prognostic features.
PATIENT SUMMARY
Loss of the PTEN tumor suppressor protein is associated with worse outcomes after salvage radiotherapy, independent of other clinical or pathologic patient characteristics.
PubMed: 38964997
DOI: 10.1016/j.euo.2024.06.008 -
Zhonghua Xue Ye Xue Za Zhi = Zhonghua... May 2024To construct a novel chimeric antigen receptor T (CAR-T) cell targeting CD138 and to investigate its cytotoxicity against myeloma cells. The hybridoma strain that can...
To construct a novel chimeric antigen receptor T (CAR-T) cell targeting CD138 and to investigate its cytotoxicity against myeloma cells. The hybridoma strain that can stably secrete the CD138 monoclonal antibody (mAb) was prepared and obtained through monoclonal antibody screening technology. The hybridoma strain cells were intraperitoneally injected into mice to produce ascites containing monoclonal antibodies, which were then collected and purified to obtain pure CD138 mAb. Further examinations were performed to assess the biological characteristics of CD138 mAb. The variable region sequence of this antibody was amplified through reverse transcription polymerase chain reaction and was used as the antigen recognition domain of CD138 CAR, which was subsequently expressed on the surface of T cells by lentiviral infection. Flow cytometry was employed to assess the phenotype of CD138 CAR-T cells. In vitro cytotoxicity and degranulation assays were performed to evaluate their antitumor effects. ① We successfully prepared anti-human CD138 antibody hybridoma cell lines and screened a hybridoma cell strain, 5G2, which could persistently and stably secrete the anti-CD138 antibody. ② The purified CD138 (5G2) mAb can especially recognize CD138(+) cells with a binding affinity constant (K(D)) of 6.011×10(-9) mol/L and showed no significant binding activity with CD138(-) cells. ③The variable region sequence of the CD138 (5G2) antibody was obtained using molecular cloning technology, and CD138 (5G2) CAR was successfully constructed and expressed on T cells through lentivirus infection and, concurrently, demonstrated effective binding to recombinant human CD138 protein.④ The proliferation of T cells transduced with the CD138 (5G2) CAR was highly efficient. The phenotype analysis revealed that CD138 (5G2) CAR-T cells exhibited a greater tendency to differentiate into central memory T cells and memory stem T cells, with a reduced proportion of terminally differentiated effector memory subsets. ⑤CD138 (5G2) CAR-T cells demonstrated specific cytotoxicity against CD138(+) myeloma cell line H929, whereas CD138(-) cell line K562 remained unaffected. The percentage of residual H929 cells was (12.92±8.02) % after co-culturing with CD138 (5G2) CAR-T cells, while (54.25±15.79) % was left in the Vector-T group (E∶T=1∶2; <0.001). ⑥Results of degranulation assays demonstrated a significant activation of CD138 (5G2) CAR-T cells after co-culture with the H929 cell line, whereas no significant activation was observed in Vector-T cells [ (25.78±3.35) % (6.13±1.30) %, <0.001]. ⑦After co-culturing with CD138(+) cells, CD138 (5G2) CAR-T cells exhibited a significant increase in cytokine secretion compared to the Vector-T group [interleukin-2: (1 697.52±599.05) pg/ml (5.07±1.17) pg/ml, <0.001; interferon-γ: (3 312.20±486.38) pg/ml (9.28±1.46) pg/ml, <0.001; and tumor necrosis factor-α: (1 837.43±640.49) pg/ml (8.75±1.65) pg/ml, <0.001]. However, no significant difference was observed in cytokine secretion levels between the two groups after co-culturing with CD138(-) cells. This study successfully prepared a novel monoclonal antibody against CD138, and CAR-T cells constructed with the antigen recognition domain derived from this 5G2 mAb demonstrated effective antitumor activity against myeloma cells. This can be used as a new option for the detection of the CD138 antigen and proposes a novel strategy for multiple myeloma immunotherapy.
Topics: Multiple Myeloma; Receptors, Chimeric Antigen; Mice; Animals; Humans; Syndecan-1; T-Lymphocytes; Hybridomas; Immunotherapy, Adoptive; Cell Line, Tumor; Receptors, Antigen, T-Cell; Antibodies, Monoclonal
PubMed: 38964917
DOI: 10.3760/cma.j.cn121090-20240131-00047 -
Zhonghua Gan Zang Bing Za Zhi =... Jun 2024To explore the genotype-phenotype relationship of Wilson's disease (WD) and further study the mutation spectrum in the ATP7B gene. The clinical data and genetic test...
To explore the genotype-phenotype relationship of Wilson's disease (WD) and further study the mutation spectrum in the ATP7B gene. The clinical data and genetic test results of 115 cases with WD diagnosed in the First Affiliated Hospital of Zhengzhou University from 2015 to 2022 were retrospectively analyzed. The rank sum test was used for quantitative data comparison, and (2) test was used for count data comparison. Multivariate logistic regression was used to analyze the relationship between patients' genotype and phenotype. The onset of liver manifestations (hepatic type) accounted for 60.9%, neurological symptoms (cerebral type) for 13.0%, and mixed hepato-cerebral symptoms for 26.1%. Presymptomatic individuals (hepatic types) accounted for 62.9%. Next-generation sequencing- diagnosed WD cases accounted for 87.8%. Combined multiplex ligation-dependent probe amplification assay-diagnosed WD cases accounted for 89.6%. A single case with a detected pathogenic locus accounted for 10.4%. The diagnostic rate of WD by genetic testing combined with clinical data was 100%. A total of 76 ATP7B mutations were detected, and the top three mutation frequencies were c.2333G>T (p.Arg778Leu) (30.7%), c.2975C>T (p.Pro992Leu) (7.3%), and c.2621C>T (p.Ala874Val) (6.4%). The mutations were mainly distributed in exons 8, 11-13, and 15-18, accounting for more than 90% of the total mutations. Eight new mutations were found, including c.3724G>A (p.Glu1242Lys), c.3703G>C (p.Gly1235Arg), c.3593T>C (p.Val1198Ala), c.2494A>C (p.Lys832Gln), c.1517T>A (p.Ile506Lys), c.484G>T (p.Glu162Ter), c.1870-49A>G, and the missing of exons 10-21. Liver histopathology showed cellular edema, degeneration, inflammation, and necrosis, as well as a 42.8% copper staining positive rate. Genotype-phenotype analysis showed that the p.Arg778Leu mutation had higher alanine aminotransferase (ALT) levels than those carrying other mutations (=0.024), while the homozygous mutation of p.Arg778Leu was associated with cerebral-type patients (=0.027). Genetic testing plays an important role in the diagnosis of WD. p.Arg778Leu is the first high-frequency mutation in the Chinese population, and patients carrying it have higher ALT levels. The p.Arg778Leu homozygous mutation is prone to causing cerebral-type WD. This study expands the ATP7B gene mutation spectrum.
Topics: Humans; Hepatolenticular Degeneration; Copper-Transporting ATPases; Mutation; Genotype; Phenotype; Retrospective Studies; Female; Male; Cation Transport Proteins; Genetic Association Studies; Adult; Adenosine Triphosphatases; Young Adult; Adolescent; Child; Genetic Testing; Middle Aged; High-Throughput Nucleotide Sequencing
PubMed: 38964899
DOI: 10.3760/cma.j.cn501113-20240416-00208 -
Zhonghua Gan Zang Bing Za Zhi =... Jun 2024To investigate the clinical and genetic characteristics and predictive role of the severe liver disease phenotype in patients with hepatolenticular degeneration (HLD)....
To investigate the clinical and genetic characteristics and predictive role of the severe liver disease phenotype in patients with hepatolenticular degeneration (HLD). Inpatients with HLD confirmed at Xinhua Hospital affiliated with Shanghai Jiao Tong University School of Medicine from January 1989 to December 2022 were selected as the research subjects. Clinical classification was performed according to the affected organs. Patients with liver disease phenotypes were classified into the liver disease group and further divided into the severe liver disease group and the ordinary liver disease group. The clinical characteristics and genetic variations were compared in each group of patients. The predictive indicators of patients with severe liver disease were analyzed by multiple regression. Statistical analysis was performed using the -test, Mann-Whitney test, or (2) test according to different data. Of the 159 HLD cases, 142 were in the liver disease group (34 in the severe liver disease group and 108 in the ordinary liver disease group), and 17 were in the encephalopathy group. The median age of onset was statistically significantly different between the liver disease group and the encephalopathy group [12.6 (7.0, 13.3) years versus 16.9 (11.0, 21.5) years, <0.01]. 156 gene mutation sites were found in 83 cases with genetic testing results, of which 54 cases carried the p.Arg778Leu gene mutation (allele frequency 46.2%). Compared with patients with other types of gene mutations (=65), patients with homozygous p.Arg778Leu mutations (=18) had lower blood ceruloplasmin and albumin levels, a higher prognostic index, Child-Pugh score, an international normalized ratio, and prothrombin time (<0.05). Hemolytic anemia, corneal K-F ring, homozygous p.Arg778Leu mutation, and multiple laboratory indexes in the severe liver disease group were statistically significantly different from those in the ordinary liver disease group (<0.05). Multivariate logistic regression analysis showed that the predictive factors for severe liver disease were homozygous p.Arg778Leu mutation, total bilirubin, and bile acids (=16.512, 1.022, 1.021, 95% : 1.204-226.425, 1.005-1.039, and 1.006-1.037, respectively, <0.05). The drawn ROC curve demonstrated a cutoff value of 0.215 3, an AUC of 0.953 2, and sensitivity and specificity of 90.91% and 92.42%, respectively. Liver disease phenotypes are common in HLD patients and have an early onset. Total bilirubin, bile acids, and the homozygous p.Arg778Leu mutation of is related to the severity of liver disease in HLD patients, which aids in predicting the occurrence and risk of severe liver disease.
Topics: Humans; Hepatolenticular Degeneration; Phenotype; Male; Female; Adolescent; Young Adult; Child; Mutation; Adult; Liver Diseases; Middle Aged
PubMed: 38964898
DOI: 10.3760/cma.j.cn501113-20230926-00122 -
Journal of Medical Genetics Jul 2024variants have been reported to be associated with immune-related disorders with/without seizures. It is unknown whether variants are associated with common epilepsy...
BACKGROUND
variants have been reported to be associated with immune-related disorders with/without seizures. It is unknown whether variants are associated with common epilepsy without acquired causes and the mechanism underlying phenotypic variation remains elusive.
METHODS
Trio-based whole-exome sequencing was performed on patients with febrile seizures or epilepsy with antecedent febrile seizures. Previously reported variants were systematically reviewed to investigate genotype-phenotype associations.
RESULTS
Two de novo heterozygous and three biallelic missense variants were identified in five patients with generalised epilepsy with antecedent febrile seizures. The variants were predicted to be damaging by in silico tools and were associated with hydrogen bonding changes to neighbouring amino acids or decreased protein stability. Patients exhibited an early onset age and became seizure-free with favourable outcome. Further analysis revealed that de novo missense variants located in the Hel region resulted in seizures with multiple neurological abnormalities, while those in the pincer domain or C-terminal domain led to seizures with normal neurodevelopment, suggesting a sub-molecular effect. Biallelic missense variants, which were inherited from unaffected parents and presented low allele frequencies in general populations, were associated with seizures without neurological abnormalities. Truncation variants were related to refractory epilepsy and severe developmental delay, suggesting a genotype-phenotype correlation. is predominantly expressed in the neonatal stage and decreases dramatically in the adulthood, which is consistent with the early onset age and favourable outcome of the patients.
CONCLUSIONS
variants are potentially associated with generalised epilepsy with antecedent febrile seizures. The sub-molecular implication and genotype-phenotype association help explain phenotype variations of variants.
PubMed: 38964834
DOI: 10.1136/jmg-2024-109950 -
Journal For Immunotherapy of Cancer Jul 2024We aimed to investigate the distinct immunological characteristics of the tumor immune microenvironment in epithelial ovarian cancer (EOC) according to mutations status...
BACKGROUND
We aimed to investigate the distinct immunological characteristics of the tumor immune microenvironment in epithelial ovarian cancer (EOC) according to mutations status and differential PD-1 expression levels.
METHODS
Tumor-infiltrating lymphocytes (TILs) were collected from patients with newly diagnosed advanced-stage EOC (YUHS cohort, n=117). This YUHS cohort was compared with The Cancer Genome Atlas (TCGA) data for ovarian serous cystadenocarcinoma (n=482), in terms of survival outcomes and immune-related gene profiles according to status. We used multicolor flow cytometry to characterize the immune phenotypes and heterogeneity of TILs with or without mutations. functional assays were conducted to evaluate the reinvigorating ability of CD8 TILs on anti-PD-1 treatment.
RESULTS
We found that EOC patients with mutations (mt) exhibited better survival outcomes and significantly higher tumor mutation burden (TMB), compared with non-mutated (wt) patients. Furthermore, CD8 TILs within mt tumors displayed characteristics indicating more severe T-cell exhaustion than their wt counterparts. Notably, the capacity for anti-PD-1-mediated reinvigoration of CD8 TILs was significantly greater in wt tumors compared with mt tumors. Additionally, within the wt group, the frequency of PD-1CD8 TILs was positively correlated with the reinvigoration capacity of CD8 TILs after anti-PD-1 treatment.
CONCLUSION
Our results highlight unique immune features of CD8 TILs in EOC and a differential response to anti-PD-1 treatment, contingent on mutation status. These findings suggest that immune checkpoint blockade may be a promising frontline therapeutic option for selected wt EOC patients.
Topics: Humans; Female; Carcinoma, Ovarian Epithelial; Lymphocytes, Tumor-Infiltrating; Mutation; CD8-Positive T-Lymphocytes; BRCA1 Protein; Middle Aged; BRCA2 Protein; Programmed Cell Death 1 Receptor; Ovarian Neoplasms; Immune Checkpoint Inhibitors; Adult; Tumor Microenvironment; Aged
PubMed: 38964784
DOI: 10.1136/jitc-2024-009058 -
The European Respiratory Journal Jul 2024The clinical classification of pulmonary hypertension (PH) has guided diagnosis and treatment of patients with PH for several decades. Discoveries relating to underlying... (Review)
Review
The clinical classification of pulmonary hypertension (PH) has guided diagnosis and treatment of patients with PH for several decades. Discoveries relating to underlying mechanisms, pathobiology, and responses to treatments for PH have informed the evolution in this clinical classification to describe the heterogeneity in PH phenotypes. In more recent years, advances in imaging, computational science, and multi-omic approaches have yielded new insights into potential phenotypes and sub-phenotypes within the existing clinical classification. Identification of novel phenotypes in pulmonary arterial hypertension (PAH) with unique molecular profiles, for example, could lead to new precision therapies. Recent phenotyping studies have also identified groups of patients with PAH that more closely resemble patients with left heart disease (group 2 PH) and lung disease (group 3 PH), which has important prognostic and therapeutic implications. Within group 2 and group 3 PH, novel phenotypes have emerged that reflect a persistent and severe pulmonary vasculopathy that is associated with worse prognosis but still distinct from PAH. In group 4 PH (chronic thromboembolic pulmonary disease) and sarcoidosis (group 5 PH) the current approach to patient phenotyping integrates clinical, hemodynamic and imaging characteristics to guide treatment but applications of multi-omic approaches to sub-phenotyping in these areas are sparse. The next iteration of the PH clinical classification is likely to reflect several emerging PH phenotypes and improve the next generation of prognostication tools, clinical trial design, and improve treatment selection in clinical practice.
PubMed: 38964779
DOI: 10.1183/13993003.01633-2023 -
Molecular and Cellular Endocrinology Jul 2024Obesity is characterized by excessive accumulation of adipose tissue (mainly visceral). The morphology and function of mitochondria are crucial for regulating adipose...
Obesity is characterized by excessive accumulation of adipose tissue (mainly visceral). The morphology and function of mitochondria are crucial for regulating adipose browning and weight loss. Research suggests that the SGLT2 inhibitor canagliflozin may induce weight loss through an unknown mechanism, particularly targeting visceral adipose tissue. While Krueppel-Like Factor 4 (KLF4) is known to be essential for energy metabolism and mitochondrial function, its specific impact on visceral adipose tissue remains unclear. We administered canagliflozin to db/db mice for 8 weeks, or exposed adipocytes to canagliflozin for 24 hours. The expression levels of browning markers, mitochondrial dynamics, and KLF4 were assessed. Then we validated the function of KLF4 through overexpression in vivo and in vitro. Adenosine monophosphate-activated protein kinase (AMPK) agonists, inhibitors, and KLF4 si-RNA were employed to elucidate the relationship between AMPK and KLF4. The findings demonstrated that canagliflozin significantly decreased body weight in db/db mice and augmented cold-induced thermogenesis. Additionally, canagliflozin increased the expression of mitochondrial fusion-related factors while reducing the levels of fission markers in epididymal white adipose tissue. These consistent findings were mirrored in canagliflozin-treated adipocytes. Similarly, overexpression of KLF4 in both adipocytes and db/db mice yielded comparable results. In all, canagliflozin mitigates obesity in db/db mice by promoting the brown visceral adipocyte phenotype through enhanced mitochondrial fusion via AMPK/KLF4 signaling.
PubMed: 38964727
DOI: 10.1016/j.mce.2024.112320