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Molecular Metabolism Oct 2023Readily accessible human pancreatic beta cells that are functionally close to primary adult beta cells are a crucial model to better understand human beta cell...
OBJECTIVES
Readily accessible human pancreatic beta cells that are functionally close to primary adult beta cells are a crucial model to better understand human beta cell physiology and develop new treatments for diabetes. We here report the characterization of EndoC-βH5 cells, the latest in the EndoC-βH cell family.
METHODS
EndoC-βH5 cells were generated by integrative gene transfer of immortalizing transgenes hTERT and SV40 large T along with Herpes Simplex Virus-1 thymidine kinase into human fetal pancreas. Immortalizing transgenes were removed after amplification using CRE activation and remaining non-excized cells eliminated using ganciclovir. Resulting cells were distributed as ready to use EndoC-βH5 cells. We performed transcriptome, immunological and extensive functional assays.
RESULTS
Ready to use EndoC-βH5 cells display highly efficient glucose dependent insulin secretion. A robust 10-fold insulin secretion index was observed and reproduced in four independent laboratories across Europe. EndoC-βH5 cells secrete insulin in a dynamic manner in response to glucose and secretion is further potentiated by GIP and GLP-1 analogs. RNA-seq confirmed abundant expression of beta cell transcription factors and functional markers, including incretin receptors. Cytokines induce a gene expression signature of inflammatory pathways and antigen processing and presentation. Finally, modified HLA-A2 expressing EndoC-βH5 cells elicit specific A2-alloreactive CD8 T cell activation.
CONCLUSIONS
EndoC-βH5 cells represent a unique storable and ready to use human pancreatic beta cell model with highly robust and reproducible features. Such cells are thus relevant for the study of beta cell function, screening and validation of new drugs, and development of disease models.
Topics: Humans; Insulin-Secreting Cells; Insulin Secretion; Cell Line; Insulin; Transcription Factors; Glucose
PubMed: 37442376
DOI: 10.1016/j.molmet.2023.101772 -
European Journal of Cancer (Oxford,... Jun 2023Thymidine kinase 1 (TK1) is an enzyme downstream of the CDK4/6 pathway, with a critical role in DNA synthesis; serum TK1 activity (sTKa) is a novel liquid biopsy...
Serum thymidine kinase activity in patients with HR-positive/HER2-negative advanced breast cancer treated with ribociclib plus letrozole: Results from the prospective BioItaLEE trial.
BACKGROUND
Thymidine kinase 1 (TK1) is an enzyme downstream of the CDK4/6 pathway, with a critical role in DNA synthesis; serum TK1 activity (sTKa) is a novel liquid biopsy biomarker of tumour cell proliferation.
METHODS
The phase IIIb, BioItaLEE trial (NCT03439046) collected sera from postmenopausal patients with hormone receptor-positive (HR+), HER2-negative (HER2-) advanced breast cancer (ABC) treated with first-line ribociclib plus letrozole at baseline, day 15 of cycle 1 (C1D15), day 1 of cycle 2 (C2D1), and at first imaging. Associations between sTKa assessed at different time points or sTKa dynamic patterns, and progression-free survival (PFS) were evaluated using multivariate Cox models.
RESULTS
Overall, 287 patients were enroled. Median follow-up was 26.9 months. High sTKa (>median) at baseline was associated with higher risk of progression (hazard ratio [HR], 2.21; 95% confidence interval [95% CI], 1.45, 3.37; P = 0.0002); similar results were observed for patients with high sTKa levels at C1D15 and C2D1. Early sTKa dynamic patterns were strongly predictive of PFS. The pattern with high sTKa levels at C2D1 following initial decrease at C1D15 was associated with higher risk of progression versus the pattern with low sTKa levels at both time points (HR, 2.89; 95% CI, 1.57, 5.31; P = 0.0006), while the pattern with high sTKa levels at C1D15 was associated with the shortest PFS (HR, 5.65; CI: 2.84, 11.2; P < 0.0001). Baseline and dynamic sTKa changes provided independent information.
CONCLUSIONS
sTKa appears to be a new promising prognostic and pharmacodynamic biomarker in patients with HR+/HER2- ABC treated with ribociclib plus letrozole as first-line therapy.
Topics: Humans; Female; Breast Neoplasms; Letrozole; Proportional Hazards Models; Thymidine Kinase; Prospective Studies; Antineoplastic Combined Chemotherapy Protocols; Aminopyridines; Aromatase Inhibitors; Biomarkers; Receptor, ErbB-2
PubMed: 37003098
DOI: 10.1016/j.ejca.2023.03.001 -
Heliyon Nov 2023In the past, multiple studies have offered incremental evidence that indicates that competitive endogenous RNA (ceRNA) regulatory networks are involved in tumor growth...
BACKGROUND
In the past, multiple studies have offered incremental evidence that indicates that competitive endogenous RNA (ceRNA) regulatory networks are involved in tumor growth and present novel therapeutic targets. Herein, we investigated the impact of thymidine kinase 1 ()-related ceRNA networks on the prognosis of non-small cell lung cancer (NSCLC).
METHODS
expression data in NSCLC and normal tissue samples were retrieved from the Cancer Genome Atlas (TCGA) database and were then compared. Thereafter, the findings of the immunohistochemical staining experiments and clinical follow-up data derived from patients with NSCLC were used for conducting prognostic analysis. The starBase database was searched to determine -targeted microRNAs and long non-coding RNAs, and clinical data from TCGA were used for survival analysis to construct a ceRNA network associated with expression and prognosis. Finally, the roles played by methylation and immunity in the prognosis and treatment of NSCLC were analyzed.
RESULTS
Our findings revealed that the cancer tissues expressed significantly higher levels than normal tissues, and the follow-up clinical data revealed that the prognosis was generally worse in the high-expression patients than in the low-expression patients. In addition, clinical data collected from the starBase and TCGA databases showed that the LINC00665/has-let-7b-5p/ network could influence the growth and prognosis of NSCLC. It was also noted that the methylation site was correlated with the prognosis of NSCLC, and immunoprognostic analysis further indicated that patients with higher expression levels displayed a worse prognosis.
CONCLUSION
When the regulatory network of LINC00665/has-let-7b-5p/ was assessed, it was observed that elevated levels may affect the prognosis of NSCLC. Therefore, it could be considered a prognostic biomarker and a probable therapeutic target for predicting NSCLC prognosis.
PubMed: 37954276
DOI: 10.1016/j.heliyon.2023.e21328 -
Viruses Aug 2023The thymidine kinase (TK) and DNA polymerase (pol) genes of the herpes simplex viruses type 1 (HSV-1) and type 2 (HSV-2) are two important genes involved in antiviral...
The thymidine kinase (TK) and DNA polymerase (pol) genes of the herpes simplex viruses type 1 (HSV-1) and type 2 (HSV-2) are two important genes involved in antiviral resistance. We investigated the genetic polymorphisms of the HSV-TK and pol genes in clinical isolates from Korean HSV-infected patients using next-generation sequencing (NGS) for the first time in Korea. A total of 81 HSV-1 and 47 HSV-2 isolates were examined. NGS was used to amplify and sequence the TK and pol genes. Among the 81 HSV-1 isolates, 12 and 17 natural polymorphisms and 9 and 23 polymorphisms of unknown significance in TK and pol were found, respectively. Two HSV-1 isolates (2.5%) exhibited the E257K amino acid substitution in TK, associated with antiviral resistance. Out of 47 HSV-2 isolates, 8 natural polymorphisms were identified in TK, and 9 in pol, with 13 polymorphisms of unknown significance in TK and 10 in pol. No known resistance-related mutations were observed in HSV-2. These findings contribute to our understanding of the genetic variants associated with antiviral resistance in HSV-1 and HSV-2 in Korea, with frequencies of known antiviral resistance-related mutations of 2.5% and 0% in HSV-1 and HSV-2, respectively.
Topics: Humans; Acyclovir; Antiviral Agents; DNA-Directed DNA Polymerase; Herpesvirus 1, Human; Herpesvirus 2, Human; Mutation; Republic of Korea; Thymidine Kinase; Drug Resistance, Viral
PubMed: 37632051
DOI: 10.3390/v15081709 -
International Journal of General... 2023Construction of a nomogram model based on Thymidine kinase 1 (TK1) in combination with inflammatory indicators and tumor markers to predict the probability of recurrence...
An Energy-Efficient Test and Predictive Model for Recurrence After Radiotherapy in Localized Intermediate and Advanced Cervical Cancer Were Created Using Thymidine Kinase 1 in Conjunction with Inflammatory Markers and Tumor Markers.
PURPOSE
Construction of a nomogram model based on Thymidine kinase 1 (TK1) in combination with inflammatory indicators and tumor markers to predict the probability of recurrence in mid- to late-stage cervical cancer.
METHODS
One hundred fourteen instances of intermediate and advanced cervical cancer admitted to our hospital's radiotherapy department between June 2017 and January 2023 were retrospectively studied. Logistic regression analysis includes variables relevant for univariate analysis. Meaningful indications from multifactor analysis were included in the nomogram model, the model's correctness was evaluated using the C-index, and the model's effectiveness was assessed using calibration curves, clinical decision curves, and clinical impact curves.
RESULTS
A nomogram model was created due to the logit regression analysis that revealed the squamous cell carcinoma antigen (SCC) and TK1 as independent recurrence predictors following cervical cancer radiation (P<0.05). The C index and Area Under the Curve (AUC) were 0.79 (95% CI 0.67-0.91). The AUC and C-index were both more extraordinary than those of TNM staging alone (C-index 0.57, 95% CI 0.43-0.71) and SCC alone (C-index 0.67, 95% CI 0.51-0.82). Calibration curves, Decision Curve Analysis (DCA), and clinical impact curves (CIC) indicate that the model predicts probabilities more accurately.
CONCLUSION
The nomogram model based on TK1 combined with inflammatory markers and tumor markers is more reliable than the TNM staging and SCC systems alone for forecasting recurrence after radiotherapy in intermediate- and advanced-stage cervical cancer. It is also a cheap, practical, and simple-to-obtain model that can supplement the TNM staging system for forecasting prognosis and significantly enhances clinicians' decision-making.
PubMed: 38089716
DOI: 10.2147/IJGM.S442389 -
Molecular Therapy : the Journal of the... Oct 2023Gliomas are the most prevalent and devastating primary malignant brain tumors in adults. Despite substantial advances in understanding glioma biology, there have been no... (Review)
Review
Gliomas are the most prevalent and devastating primary malignant brain tumors in adults. Despite substantial advances in understanding glioma biology, there have been no regulatory drug approvals in the US since bevacizumab in 2009 and tumor treating fields in 2011. Recent phase III clinical trials have failed to meet their prespecified therapeutic primary endpoints, highlighting the need for novel therapies. The poor prognosis of glioma patients, resistance to chemo-radiotherapy, and the immunosuppressive tumor microenvironment underscore the need for the development of novel therapies. Gene therapy-based immunotherapeutic strategies that couple the ability of the host immune system to specifically kill glioma cells and develop immunological memory have shown remarkable progress. Two adenoviral vectors expressing Ad-HSV1-TK/GCV and Ad-Flt3L have shown promising preclinical data, leading to FDA approval of a non-randomized, phase I open-label, first in human trial to test safety, cytotoxicity, and immune-stimulatory efficiency in high-grade glioma patients (NCT01811992). This review provides a thorough overview of immune-stimulatory gene therapy highlighting recent advancements, potential drawbacks, future directions, and recommendations for future implementation of clinical trials.
Topics: Animals; Humans; Brain Neoplasms; Rodentia; Adenoviridae; Glioma; Genetic Therapy; Thymidine Kinase; Genetic Vectors; Tumor Microenvironment
PubMed: 37574780
DOI: 10.1016/j.ymthe.2023.08.009 -
Breast Cancer Research and Treatment Apr 2024Thymidine kinase 1 (TK1) plays a pivotal role in DNA synthesis and cellular proliferation. TK1 has been studied as a prognostic marker and as an early indicator of... (Randomized Controlled Trial)
Randomized Controlled Trial
The role of serum thymidine kinase 1 activity in neoadjuvant-treated HER2-positive breast cancer: biomarker analysis from the Swedish phase II randomized PREDIX HER2 trial.
BACKGROUND
Thymidine kinase 1 (TK1) plays a pivotal role in DNA synthesis and cellular proliferation. TK1 has been studied as a prognostic marker and as an early indicator of treatment response in human epidermal growth factor 2 (HER2)-negative early and metastatic breast cancer (BC). However, the prognostic and predictive value of serial TK1 activity in HER2-positive BC remains unknown.
METHODS
In the PREDIX HER2 trial, 197 HER2-positive BC patients were randomized to neoadjuvant trastuzumab, pertuzumab, and docetaxel (DPH) or trastuzumab emtansine (T-DM1), followed by surgery and adjuvant epirubicin and cyclophosphamide. Serum samples were prospectively collected from all participants at multiple timepoints: at baseline, after cycle 1, 2, 4, and 6, at end of adjuvant therapy, annually for a total period of 5 years and/or at the time of recurrence. The associations of sTK1 activity with baseline characteristics, pathologic complete response (pCR), event-free survival (EFS), and disease-free survival (DFS) were evaluated.
RESULTS
No association was detected between baseline sTK1 levels and all the baseline clinicopathologic characteristics. An increase of TK1 activity from baseline to cycle 2 was seen in all cases. sTK1 level at baseline, after 2 and 4 cycles was not associated with pCR status. After a median follow-up of 58 months, 23 patients had EFS events. There was no significant effect between baseline or cycle 2 sTK1 activity and time to event. A non-significant trend was noted among patents with residual disease (non-pCR) and high sTK1 activity at the end of treatment visit, indicating a potentially worse long-term prognosis.
CONCLUSION
sTK1 activity increased following neoadjuvant therapy for HER2-positive BC but was not associated with patient outcomes or treatment benefit. However, the post-surgery prognostic value in patients that have not attained pCR warrants further investigation.
TRIAL REGISTRATION
ClinicalTrials.gov, NCT02568839. Registered on 6 October 2015.
Topics: Humans; Female; Breast Neoplasms; Neoadjuvant Therapy; Sweden; Receptor, ErbB-2; Biomarkers, Tumor; Antineoplastic Combined Chemotherapy Protocols; Trastuzumab; Ado-Trastuzumab Emtansine; Thymidine Kinase
PubMed: 38175448
DOI: 10.1007/s10549-023-07200-x -
Frontiers in Immunology 2023The incidence of thyroid carcinoma (THCA), the most common endocrine tumor, is continuously increasing worldwide. Although the overall prognosis of THCA is good,...
BACKGROUND
The incidence of thyroid carcinoma (THCA), the most common endocrine tumor, is continuously increasing worldwide. Although the overall prognosis of THCA is good, patients with distant metastases exhibit a mortality rate of 5-20%.
METHODS
To improve the diagnosis and overall prognosis of patients with thyroid cancer, we screened specific candidate neoantigen genes in early- and late-stage THCA by analyzing the transcriptome and somatic cell mutations in this study.
RESULTS
The top five early-stage neoantigen-related genes (NRGs) were G protein-coupled receptor 4 [], chondroitin sulfate proteoglycan 4 [], teneurin transmembrane protein 1 [], protein S 1 [], and thymidine kinase 1 [], whereas the top five late-stage NRGs were cadherin 6 [], semaphorin 6B [], dysferlin [], xenotropic and polytropic retrovirus receptor 1 [], and ABR activator of RhoGEF and GTPase []. Subsequently, we used machine learning models to verify their ability to screen NRGs and analyze the correlations among NRGs, immune cell types, and immune checkpoint regulators. The use of candidate antigen genes resulted in a better diagnostic model (the area under the curve [AUC] value of the early-stage group [0.979] was higher than that of the late-stage group [0.959]). Then, a prognostic model was constructed to predict NRG survival, and the 1-, 3- and 5-year AUC values were 0.83, 0.87, and 0.86, respectively, which were closely related to different immune cell types. Comparison of the expression trends and mutation frequencies of NRGs in multiple tumors revealed their potential for the development of broad spectrum therapeutic drugs.
CONCLUSION
In conclusion, the candidate NRGs identified in this study could potentially be used as therapeutic targets and diagnostic biomarkers for the development of novel broad spectrum therapeutic agents.
Topics: Humans; Exome; Transcriptome; Thyroid Neoplasms; Adenocarcinoma
PubMed: 37854594
DOI: 10.3389/fimmu.2023.1187160 -
Virus Research Oct 2023Avipoxvirus 282E4 strain was extensively applied into recombinant vaccine vector to prevent other infectious diseases. However, little information on the genomic...
Avipoxvirus 282E4 strain was extensively applied into recombinant vaccine vector to prevent other infectious diseases. However, little information on the genomic background, functional and genetic evolutionary of the isolate 282E4 strain was clarified. The results showed that the linear genome of avipoxvirus 282E4 was 308,826 bp, containing 313 open reading frames (ORFs) and 12 new predicted ORFs. The 282E4 strain appears to encode two novel thymidine kinase proteins and two TGF-beta-like proteins that may be associated with the suppression of the host's antiviral response. Avipoxvirus 282E4 also encodes 57 ankyrin repeat proteins and 5 variola B22R-like proteins, which composed 7% of the avipoxvirus 282E4 genome. GO and KEGG analysis further revealed that 12 ORFs participate in viral transcription process, 7 ORFs may function during DNA repair, replication and biological synthesis, and ORF 208 is involved in the process of virus life cycle. Interestingly, phylogenetic analysis based on concatenated sequences p4b and DNA polymerase of avipoxviruses gene demonstrates that avipoxvirus 282E4 strain is divergent from known FWPV isolates and is similar to shearwater poxvirus (SWPV-1) that belongs to the CNPV-like virus. Sequencing avipoxvirus 282E4 is a significant step to judge the genetic position of avipoxviruses within the larger Poxviridae phylogenetic tree and provide a new insight into the genetic background of avipoxvirus 282E4 and interspecies transmission of poxviruses, meanwhile, explanation of gene function provides theoretical foundation for vaccine design with 282E4 strain as skeleton.
PubMed: 37678517
DOI: 10.1016/j.virusres.2023.199218 -
PLoS Genetics Nov 2023A prominent source of mutation in cancer is single-stranded DNA cytosine deamination by cellular APOBEC3 enzymes, which results in signature C-to-T and C-to-G mutations...
A prominent source of mutation in cancer is single-stranded DNA cytosine deamination by cellular APOBEC3 enzymes, which results in signature C-to-T and C-to-G mutations in TCA and TCT motifs. Although multiple enzymes have been implicated, reports conflict and it is unclear which protein(s) are responsible. Here we report the development of a selectable system to quantify genome mutation and demonstrate its utility by comparing the mutagenic activities of three leading candidates-APOBEC3A, APOBEC3B, and APOBEC3H. The human cell line, HAP1, is engineered to express the thymidine kinase (TK) gene of HSV-1, which confers sensitivity to ganciclovir. Expression of APOBEC3A and APOBEC3B, but not catalytic mutant controls or APOBEC3H, triggers increased frequencies of TK mutation and similar TC-biased cytosine mutation profiles in the selectable TK reporter gene. Whole genome sequences from independent clones enabled an analysis of thousands of single base substitution mutations and extraction of local sequence preferences with APOBEC3A preferring YTCW motifs 70% of the time and APOBEC3B 50% of the time (Y = C/T; W = A/T). Signature comparisons with breast tumor whole genome sequences indicate that most malignancies manifest intermediate percentages of APOBEC3 signature mutations in YTCW motifs, mostly between 50 and 70%, suggesting that both enzymes contribute in a combinatorial manner to the overall mutation landscape. Although the vast majority of APOBEC3A- and APOBEC3B-induced single base substitution mutations occur outside of predicted chromosomal DNA hairpin structures, whole genome sequence analyses and supporting biochemical studies also indicate that both enzymes are capable of deaminating the single-stranded loop regions of DNA hairpins at elevated rates. These studies combine to help resolve a long-standing etiologic debate on the source of APOBEC3 signature mutations in cancer and indicate that future diagnostic and therapeutic efforts should focus on both APOBEC3A and APOBEC3B.
Topics: Humans; Female; Breast Neoplasms; Mutation; Cytidine Deaminase; Cell Line; DNA; Minor Histocompatibility Antigens; Cytosine
PubMed: 38033156
DOI: 10.1371/journal.pgen.1011043