Nanochannel-based biosensor for ultrasensitive and label-free detection of thymidine kinase activity.

Thymidine Kinase 1 (TK1) is a pivotal enzyme in fundamental biochemistry and molecular diagnosis, but recognition and molecule detection is a challenging task. Here, we constructed a DNA-integrated hybrid nanochannel sensor for TK1 activity and inhibition assay. Single-stranded DNA containing thymidine was used as a substrate to functionalize the nanochannels, restricting the ion current through channels. With kinase, the thymidine at the termini of the substrate DNA is phosphorylated, elevating surface charge density and mitigating the pore-obstruction effect by increasing transmembrane ion current. The kinase-induced distinctness can be accurately monitored by this hybrid nanodevice, which benefits from its high sensitivity to the change of surface charge. The excellent analytical performance in both kinase enzyme activity and inhibition analysis resulted in efficient and selective evaluation in human serum. Furthermore, compared to current approaches, it greatly simplifies and offers a direct method of analysis, making it a promising sensor technology for cancer management as well as the activities of multiple types of nucleic acid kinases.

Predictive value of spectral dual-detector computed tomography for PD-L1 expression in stage I lung adenocarcinoma: development and validation of a novel nomogram.

Background: Programmed death ligand-1 (PD-L1) expression serves a predictive biomarker for the efficacy of immune checkpoint inhibitors (ICIs) in the treatment of patients with early-stage lung adenocarcinoma (LA). However, only a limited number of studies have explored the relationship between PD-L1 expression and spectral dual-layer detector-based computed tomography (SDCT) quantification, qualitative parameters, and clinical biomarkers. Therefore, this study was conducted to clarify this relationship in stage I LA and to develop a nomogram to assist in preoperative individualized identification of PD-L1-positive expression. Methods: We analyzed SDCT parameters and PD-L1 expression in patients diagnosed with invasive nonmucinous LA through postoperative pathology. Patients were categorized into PD-L1-positive and PD-L1-negative expression groups based on a threshold of 1%. A retrospective set (N=356) was used to develop and internally validate the radiological and biomarker features collected from predictive models. Univariate analysis was employed to reduce dimensionality, and logistic regression was used to establish a nomogram for predicting PD-L1 expression. The predictive performance of the model was evaluated using receiver operating characteristic (ROC) curves, and external validation was performed in an independent set (N=80). Results: The proportions of solid components and pleural indentations were higher in the PD-L1-positive group, as indicated by the computed tomography (CT) value, CT at 40 keV (CT40keV; a/v), electron density (ED; a/v), and thymidine kinase 1 (TK1) exhibiting a positive correlation with PD-L1 expression. In contrast, the effective atomic number (Zeff; a/v) showed a negative correlation with PD-L1 expression [r=-0.4266 (Zeff.a), -0.1131 (Zeff.v); P<0.05]. After univariate analysis, 18 parameters were found to be associated with PD-L1 expression. Multiple regression analysis was performed on significant parameters with an area under the curve (AUC) >0.6, and CT value [AUC =0.627; odds ratio (OR) =0.993; P=0.033], CT40keV.a (AUC =0.642; OR =1.006; P=0.025), arterial Zeff (Zeff.a) (AUC =0.756; OR =0.102; P<0.001), arterial ED (ED.a) (AUC =0.641; OR =1.158, P<0.001), venous ED (ED.v) (AUC =0.607; OR =0.864; P<0.001), TK1 (AUC =0.601; OR =1.245; P=0.026), and diameter of solid components (Dsolid) (AUC =0.632; OR =1.058; P=0.04) were found to be independent risk factors for PD-L1 expression in stage I LA. These seven predictive factors were integrated into the development of an SDCT parameter-clinical nomogram, which demonstrated satisfactory discrimination ability in the training set [AUC =0.853; 95% confidence interval (CI): 0.76-0.947], internal validation set (AUC =0.824; 95% CI: 0.775-0.874), and external validation set (AUC =0.825; 95% CI: 0.733-0.918). Decision curve analyses also revealed the highest net benefit for the nomogram across a broad threshold probability range (20-80%), with a clinical impact curve (CIC) indicating its clinical validity. Comparisons with other models demonstrated the superior discriminatory accuracy of the nomogram over any individual variable (all P values <0.05). Conclusions: Quantitative parameters derived from SDCT demonstrated the ability to predict for PD-L1 expression in early-stage LA, with Zeff.a being notably effective. The nomogram established in combination with TK1 showed excellent predictive performance and good calibration. This approach may facilitate the improved noninvasive prediction of PD-L1 expression.

Pharmacokinetics and Safety of a 1:1 Mixture of Doxecitine and Doxribtimine: Open-label Phase 1 Single Ascending Dose and Food Effect Studies in Healthy Adults.

PURPOSE: Doxecitine (deoxycytidine [dC]) and doxribtimine (deoxythymidine [dT]) powder for oral solution is a 1:1 mixture consisting of equal weights 2'-deoxycytidine (dC) and 2'-deoxythymidine (dT). Doxecitine and doxribtimine (referred to as study drug) is being developed as treatment for people with thymidine kinase 2 deficiency (TK2d). TK2d is an ultra-rare mitochondrial DNA depletion and multiple deletion syndrome characterized by progressive muscle weakness and premature death. Here, we report the pharmacokinetics (PK), the effect of food, and the tolerability of 2 study drug formulations, evaluated in 2 studies (Study MT-1621-103 and Study MT-1621-105). METHODS: A sequential, ascending 1:1 dose ratio was used for both studies (n = 14 healthy volunteer adult participants/study). After a 28-day (Study MT-1621-103) or 35-day (Study MT-1621-105) screening period, participants fasted overnight and sequentially received 86.6, 173.4, and 266.6 mg/kg study drug with a 48-hour PK assessment period and 48-hour washout period between doses. After 48 additional hours, participants were fed a high-fat meal and received 266.6 mg/kg study drug. Plasma and urine were collected before dosing and throughout the 48-hour PK period. dC and dT concentrations were analyzed by validated liquid chromatography mass spectrometry methods. Safety was evaluated throughout the study and at 2-week follow-up. FINDINGS: Plasma levels of dC and dT increased rapidly and dose-dependently above endogenous levels for both formulations, with a median Tmax of 1 to 2 hours under fasting conditions. Post-dose plasma dC and dT concentrations declined to nearly pre-dose (baseline) concentrations after 8 to 12 hours, suggesting rapid elimination. Peak and extent of plasma exposure (baseline-corrected Cmax and AUC0-t) tended to increase less than dose-proportionally for plasma dC and greater than dose-proportionally for plasma dT. PK variability of dC and dT was moderate-to-high (>30%). Administration with food delayed Tmax to a median of 2 to 4 hours and increased plasma exposure: baseline-corrected plasma dC Cmax and AUC0-t increased by ∼79% to 96% and 137% to 250%, respectively, and dT Cmax and AUC0-t increased by 27% to 29% and 74% to 89%, respectively, indicating a significant food effect. Renal clearance played a minor role in the elimination of systemically available intact dC and dT (Fe<0.3%). The study drug was generally well tolerated; most frequent study-drug-related adverse events (AEs) were diarrhea (n = 4/29, 14%) and dizziness (n = 3/29, 10%). Most AEs were mild-to-moderate in severity. IMPLICATIONS: Doxecitine and doxribtimine are orally bioavailable in the intended clinical dose range. The PK profile supports a formulation consisting of equal doses of doxecitine and doxribtimine, a 3-times-daily dosing regimen, and administration with food.

Increased levels of thymidine kinase 1 in malignant cell-derived extracellular vesicles.

Extracellular vesicles (EVs), whose main subtypes are exosomes, microparticles, and apoptotic bodies, are secreted by all cells and harbor biomolecules such as DNA, RNA, and proteins. They function as intercellular messengers and, depending on their cargo, may have multiple roles in cancer development. Thymidine kinase 1 (TK1) is a cell cycle-dependent enzyme used as a biomarker for cell proliferation. TK1 is usually elevated in cancer patients' serum, making the enzyme a valuable tumor proliferation biomarker that strongly correlates with cancer stage and metastatic capabilities. Here, we investigated the presence of TK1 in EVs derived from three prostate cancer cell lines with various p53 mutation statuses (LNCaP, PC3, and DU145), EVs from the normal prostate epithelial cell line RWPE-1 and EVs isolated from human seminal fluid (prostasomes). We measured the TK1 activity by a real-time assay for these EVs. We demonstrated that the TK1 enzyme activity is higher in EVs derived from the malignant cell lines, with the highest activity from cells deriving from the most aggressive cancer, compared to the prostasomes and RWPE-1 EVs. The measurement of TK1 activity in EVs may be essential in future prostate cancer studies.

The Active Tumor Vaccination in Combination With CDK4/6 Inhibitor Treatment: A Case Report.

BACKGROUND: Current standard treatment for metastatic breast cancer (MBC) involves cyclin-dependent kinase 4/6 (CDK4/6) inhibitors with endocrine therapy, showing potential in enhancing anti-tumor immune responses. CASE REPORT: This report details a clinical case of MBC where palbociclib was co-administered with letrozole. The integration of allogeneic tumor vaccination to this treatment led to heightened interferon-γ production, expansion of CD8+ and NK cell populations, and positive delayed-type hypersensitivity reactions, indicating successful development of anti-tumor immunity. The induced production of interferon-γ by tumor vaccination was associated with manageable modulation of sensitivity to palbociclib-letrozole therapy. Administration of the BioNTech/Pfizer Covid-19 vaccine compromised the anti-tumor immune response by reducing cytotoxic cell populations and increasing immunosuppressive cytokine production. The patient undergoing combined treatment achieved a progressive-free survival of 42 months. CONCLUSION: Incorporating active tumor vaccination with CDK4/6 inhibitor therapy presents a feasible approach for metastatic breast cancer. The precise regulation of the microenvironment emerges as a crucial factor and warrants careful consideration.

Strand displacement-triggered FRET nanoprobe tracking TK1 mRNA in living cells for ratiometric fluorimetry of nucleic acid biomarker.

A precisely designed dual-color biosensor has realized a visual assessment of thymidine kinase 1 (TK1) mRNA in both living cells and cell lysates. The oligonucleotide probe is constructed by hybridizing the antisense strand of the target and two recognition sequences, in which FAM serves as the donor and TAMRA as the acceptor. Once interacting with the target, two recognition strands are replaced, and then the antisense complementary sequence forms a more stable double-stranded structure. Due to the increasing spatial distance between two dyes, the FRET is attenuated, leading to a rapid recovery of FAM fluorescence and a reduction of TAMRA fluorescence. A discernible color response from orange to green could be observed by the naked eye, with a limit of detection (LOD) of 0.38 nM and 5.22 nM for spectrometer- and smartphone-based assays, respectively. The proposed ratiometric method transcends previous reports in its capacities in visualizing TK1 expression toward reliable nucleic acid biomarker analysis, which might establish a general strategy for ratiometric biosensing via strand displacement.

Evaluation of Serum Cytokeratines, Thymidine Kinase, and Growth Factors as Cancer Biomarkers in Colorectal Cancer.

BACKGROUND/AIM: Classical serum cancer biomarkers, such as carcinoembryonic antigen (CEA) and cancer antigen 19-9 (CA 19-9), remain important tools in colorectal cancer (CRC) management for disease follow up. However, their sensitivity and specificity are low for diagnostic and prognostic evaluation. The aim of this study was to evaluate the potential of biomarkers reflecting biological activity of tumors - tissue polypeptide specific antigen (TPS), cytokeratin fragment 19 (CYFRA 21-1), thymidine kinase (TK), insulin-like growth factor 1 (IGF-1) and insulin-like growth factor binding protein 3 (IGF-BP3) - together with the CEA and CA 19-9 in CRC diagnosis and prognosis. PATIENTS AND METHODS: This is a retrospective study including 148 CRC patients and 68 age-matched healthy subjects. Serum biomarkers were measured in pre-operative serum samples using immunoanalytical methods. The end-point for the diagnostic evaluation was the area under the receiving operating characteristic curve (AUC ROC) of the biomarkers. The end-point for the prognostic evaluation was overall survival. RESULTS: Serum levels of CEA, CA 19-9, TPS, and TK were significantly increased in CRC early-stage patients compared with healthy controls. Each of the studied biomarkers had AUC between 0.6 and 0.7. Analysis of survival demonstrated that the patients with CEA, CA 19-9, cytokeratin, and TK above optimal cut offs had significantly shorter survival. A multivariate analysis performed on all the study biomarkers resulted in the selection of CYFRA 21-1 as the best performing biomarker with hazard ratio 10.413. CONCLUSION: The combination of cytokeratins and thymidine kinase with classical cancer biomarkers enables the prediction of tumor aggressiveness and long-term prognosis.

The Histogenetic Origin of Malignant Cells Predicts Their Susceptibility towards Synthetic Lethality Utilizing the TK.007 System.

BACKGROUND: Remarkable differences exist in the outcome of systemic cancer therapies. Lymphomas and leukemias generally respond well to systemic chemotherapies, while solid cancers often fail. We engineered different human cancer cells lines to uniformly express a modified herpes simplex virus thymidine kinase TK.007 as a suicide gene when ganciclovir (GCV) is applied, thus in theory achieving a similar response in all cell lines. METHODS: Fifteen different cell lines were engineered to express the TK.007 gene. XTT-cell proliferation assays were performed and the IC50-values were calculated. Functional kinome profiling, mRNA sequencing, and bottom-up proteomics analysis with Ingenuity pathway analysis were performed. RESULTS: GCV potency varied among cell lines, with lymphoma and leukemia cells showing higher susceptibility than solid cancer cells. Functional kinome profiling implies a contribution of the SRC family kinases and decreased overall kinase activity. mRNA sequencing highlighted alterations in the MAPK pathways and bottom-up proteomics showed differences in apoptotic and epithelial junction signaling proteins. CONCLUSIONS: The histogenetic origin of cells influenced the susceptibility of human malignant cells towards cytotoxic agents with leukemias and lymphomas being more sensitive than solid cancer cells.

CRISPR/Cas9-mediated genome editing of the thymidine kinase gene in a clinical HSV-1 isolate identifies F289S as novel acyclovir-resistant mutation.

Herpes simplex virus type 1 (HSV-1) is a neurotropic alphaherpesvirus that establishes a lifelong infection in sensory neurons of infected individuals, accompanied with intermittent reactivation of latent virus causing (a)symptomatic virus shedding. Whereas acyclovir (ACV) is a safe and highly effective antiviral to treat HSV-1 infections, long-term usage can lead to emergence of ACV resistant (ACVR) HSV-1 and subsequently ACV refractory disease. Here, we isolated an HSV-1 strain from a patient with reactivated herpetic eye disease that did not respond to ACV treatment. The isolate carried a novel non-synonymous F289S mutation in the viral UL23 gene encoding the thymidine kinase (TK) protein. Because ACV needs conversion by viral TK and subsequently cellular kinases to inhibit HSV-1 replication, the UL23 gene is commonly mutated in ACVR HSV-1 strains. The potential role of the F289S mutation causing ACVR was investigated using CRISPR/Cas9-mediated HSV-1 genome editing. Reverting the F289S mutation in the original clinical isolate to the wild-type sequence S289F resulted in an ACV-sensitive (ACVS) phenotype, and introduction of the F289S substitution in an ACVS HSV-1 reference strain led to an ACVR phenotype. In summary, we identified a new HSV-1 TK mutation in the eye of a patient with ACV refractory herpetic eye disease, which was identified as the causative ACVR mutation with the aid of CRISPR/Cas9-mediated genome engineering technology. Direct editing of clinical HSV-1 isolates by CRISPR/Cas9 is a powerful strategy to assess whether single residue substitutions are causative to a clinical ACVR phenotype.

A putative design for the electromagnetic activation of split proteins for molecular and cellular manipulation.

The ability to manipulate cellular function using an external stimulus is a powerful strategy for studying complex biological phenomena. One approach to modulate the function of the cellular environment is split proteins. In this method, a biologically active protein or an enzyme is fragmented so that it reassembles only upon a specific stimulus. Although many tools are available to induce these systems, nature has provided other mechanisms to expand the split protein toolbox. Here, we show a novel method for reconstituting split proteins using magnetic stimulation. We found that the electromagnetic perceptive gene (EPG) changes conformation due to magnetic field stimulation. By fusing split fragments of a certain protein to both termini of the EPG, the fragments can be reassembled into a functional protein under magnetic stimulation due to conformational change. We show this effect with three separate split proteins: NanoLuc, APEX2, and herpes simplex virus type-1 thymidine kinase. Our results show, for the first time, that reconstitution of split proteins can be achieved only with magnetic fields. We anticipate that this study will be a starting point for future magnetically inducible split protein designs for cellular perturbation and manipulation. With this technology, we can help expand the toolbox of the split protein platform and allow better elucidation of complex biological systems.

Development of a gold nanoparticle-based lateral flow immunochromatographic assay for the rapid and quantitative detection of thymidine kinase 1 in human serum.

Thymidine kinase 1 (TK1) is a marker of cell proliferation that can be used for early screening, treatment monitoring, and evaluating the prognosis of patients with tumors. The main purpose of this study was to develop clinically applicable TK1 antibodies, establish an appropriate detection method, and provide material and technical support for the research and clinical application for different types of tumors. Experimental mice were immunized with the C-terminal 31 peptide of human TK1 to screen monoclonal cell lines capable of stably secreting specific antibodies. Monoclonal antibodies were then prepared, purified and screened for optimal pairing following the identification of purity and isotype. Finally, based on the principles adopted by the double-antibody sandwich detection method, we constructed a lateral flow immunochromatographic assay (LFIA) to quantify the concentration of TK1 in serum samples when using a gold nanoparticle-labeled anti-TK1 monoclonal antibody as a probe. The limit of detection for TK1 in serum was 0.31 pmol/L with a detection range of 0.31-50 pmol/L. The spiked recoveries ranged from 97.7% to 109.0% with an analytical precision of 5.7-8.2%; there was no cross-reactivity with common proteins in the serum. The established LFIA also exhibited good consistency with commercially available chemiluminescent immunoassay kits for the detection of clinical samples. The LFIA developed in this study has the advantages of high sensitivity, accuracy, reproducibility and strong specificity, and provides a new technical tool for the quantitative detection of TK1.

Enzymes as indispensable markers in disease diagnosis.

Enzymes have been used for disease diagnosis for many decades; however, advancements in technology like ELISA and flow cytometry-based detection have significantly increased their use and have increased the sensitivity of detection. Technological advancements in recombinant enzyme production have increased enzymatic stability, and the use of colorimetric-based and florescence-based assays has led to their increased use as biomarkers for disease detection. Enzymes like acid phosphatase, cathepsin, lactate dehydrogenase, thymidine kinase and creatine kinase are indispensable markers for diagnosing cancer, cardiovascular diseases and others. This minireview summarizes various enzymes used in disease diagnosis, their metabolic role, market value and potential as disease markers across various metabolic and other disorders.

[Box: see text].

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.

Clinical Significance of the TK1-Specific Activity in the Early Detection of Ovarian Cancer.

INTRODUCTION: Ovarian cancer is the eighth most common cause of cancer death in women. One of the major concerns is almost two-thirds of cases are typically diagnosed in the late stage as the symptoms are unspecific in the early stage of ovarian cancer. It is known that the combination of TK1 protein with CA 125 or HE4 showed better performance than either of them alone. That is why, the aim of the study was to investigate whether the TK1-specific activity (TK1 SA) could function as a complement marker for early-stage diagnosis of ovarian cancer. METHODS: The study included a set of 198 sera consisting of 134 patients with ovarian tumors (72 benign and 62 malignant) and 64 healthy age-matched controls. The TK1 SA was determined using TK1 activity by TK-Liaison and TK1 protein by AroCell TK 210 ELISA. Further, CA 125, HE4, as well as risk of ovarian malignancy algorithm index were also determined in the same set of clinical samples. RESULTS: The TK1 SA was significantly different between healthy compared to ovarian cancer patients (p < 0.0001). Strikingly, TK1 SA has higher sensitivity (55%) compared to other biomarkers in the detection of benign ovarian tumors. Further, the highest sensitivity was achieved by the combination of TK1 SA with CA 125 and HE4 for the detection of benign tumors as well as malignant ovarian tumors (72.2% and 88.7%). In addition, TK1 SA could significantly differentiate FIGO stage I/II from stage III/IV malignancies (p = 0.026). Follow-up of patients after surgery and chemotherapy showed a significant difference compared to TK1 SA at the time of diagnosis. CONCLUSIONS: These results indicate that TK1 SA is a promising blood-based biomarker that could complement CA 125 and HE4 for the detection of early stages of ovarian cancer.

In-vitro and in-silico anti-HSV-1 activity of a marine steroid from the jellyfish Cassiopea andromeda venom.

In this study, we investigated the potential in vitro anti-HSV-1 activities of the Cassiopea andromeda jellyfish tentacle extract (TE) and its fractions, as well as computational work on the thymidine kinase (TK) inhibitory activity of the identified secondary metabolites. The LD50, secondary metabolite identification, preparative and analytical chromatography, and in silico TK assessment were performed using the Spearman-Karber, GC-MS, silica gel column chromatography, RP-HPLC, LC-MS, and docking methods, respectively. The antiviral activity of TE and the two purified compounds Ca2 and Ca7 against HSV-1 in Vero cells was evaluated by MTT and RT-PCR assays. The LD50 (IV, mouse) values of TE, Ca2, and Ca7 were 104.0 ± 4, 5120 ± 14, and 197.0 ± 7 (µg/kg), respectively. They exhibited extremely effective antiviral activity against HSV-1. The CC50 and MNTD of TE, Ca2, and Ca7 were (125, 62.5), (25, 12.5), and (50, 3.125) µg/ml, respectively. GC-MS analysis of the tentacle extract revealed seven structurally distinct chemical compositions. Four of the seven compounds had a steroid structure. According to the docking results, all compounds showed binding affinity to the active sites of both thymidine kinase chains. Among them, the steroid compound Pregn-5-ene-3,11-dione, 17,20:20,21 bis [methylenebis(oxy)]-, cyclic 3-(1,2-ethane diyl acetal) (Ca2) exhibited the highest affinity for both enzyme chains, surpassing that of standard acyclovir. In silico data confirmed the experimental results. We conclude that the oxosteroid Ca2 may act as a potent agent against HSV-1.

Serum thymidine kinase 1 protein concentrations and presence of its autoantibody as biomarkers for screening dogs with malignant tumors.

BACKGROUND: An accurate and easily accessible method for diagnosing malignancies in local veterinary clinics has not yet been established. OBJECTIVES: To investigate the usefulness of serum thymidine kinase 1 (TK1) protein and its autoantibody as tumor biomarkers in dogs. ANIMALS: Serum samples from 1702 dogs were collected from local animal hospitals and referral animal medical centers in South Korea. METHODS: TK1 protein OD value and TK1 autoantibody ratio (TK1 autoantibody OD/total IgG OD) in serum samples of dogs classified into healthy controls, group with nontumor disease, group with benign and group with malignant tumors were measured using lateral flow immunochromatographic assay methods. RESULTS: TK1 autoantibody levels were significantly higher in malignant tumor group (median 0.71) than in healthy controls (median 0.34), group with nontumor disease (median 0.34), and group with benign tumor (median 0.32, Welch t test, P < .0001). They were also significantly different among dogs with carcinomas (median 0.77), hematopoietic tumors (median 0.71), and sarcomas (median 0.56) than in healthy controls (median 0.34, post hoc Games-Howell test, P < .0001). In the receiver operating characteristic curve of TK1 protein, AUC was 0.633 (95% CI: 0.592-0.675, P < .0001). The AUC of TK1 autoantibody ratio was 0.758 (95% CI: 0.723-0.793, P < .0001). CONCLUSIONS AND CLINICAL IMPORTANCE: TK1 autoantibody is a potentially useful biomarker for differentiating between healthy and tumor-bearing dogs, better than TK1 protein measurement. However, both were inadequate when used as single biomarkers for screening dogs to discover occult malignant tumors.

Relationship between the expression of serum TK1 and DKK1 and the prognosis of advanced non-small cell lung cancer patients / 国际检验医学杂志

Objective To observe the levels of serum thymidine kinase 1(TK1)and secreted protein Dikkopf-1(DKK1)in patients with advanced non-small cell lung cancer(NSCLC),and analyze the relation-ship between serum TK1,DKK1 and the prognosis of NSCLC.Methods This study adopted a prospective co-hort study method,a total of 91 chemotherapy patients with advanced NSCLC admitted in Jinshan Branch of Shanghai Sixth People's Hospital from January 2020 to June 2021 were enrolled as the research objects.All patients received the detection of serum TK1 and DKK1 on admission,completed 4 chemotherapy cycles in Jinshan Branch of Shanghai Sixth People's Hospital and were followed up for 3 months.The disease remission rate was evaluated according to the relevant standards.The patients with complete remission and partial re-mission were included in the good prognosis group,and those with the stable and progressive lesions were in-cluded in the poor prognosis group.The levels of serum TK1 and DKK1 were compared between the two groups.Logistic regression was used to analyze the relationship between the levels of serum TK1 and DKK1 and the prognosis of patients with advanced NSCLC.Results Among 91 patients with advanced NSCLC who received chemotherapy,threre were 58 cases(63.74％)in the good prognosis group,and 33 cases(36.26％)in the poor prognosis group.The levels of serum carcinoembryonic antigen(CEA),TK1 and DKK1 in the poor prognosis group were higher than those in the good prognosis group(P＜0.05).Logistic regression anal-ysis showed that the high levels of serum TK1 and DKK1 were the influencing factors of poor prognosis in pa-tients with advanced NSCLC(OR＞1,P＜0.05).The receiver operating characteristic curve was drawn,and the results showed that the area under the curve of serum TK1,DKK1 alone and combined for predicting the poor prognosis in patients with advanced NSCLC was＞0.700,all of which had certain predictive value,and the predictive value of the combined detection was the highest.Conclusion The abnormal increase of serum TK1 and DKK1 levels may indicate a high risk of poor prognosis in patients with advanced NSCLC.Early monito-ring of serum TK1 and DKK1 levels in patients has certain positive significance for predicting and evaluating the prognosis of patients.

Effects of Fuzheng Quxie Prescription Combined with Neoadjuvant Chemotherapy on Tumor Recurrence,Serum Thymidine Kinase 1 Level and Immune Function in Patients with Triple-Negative Breast Cancer / 广州中医药大学学报

Objective To investigate the effects of Fuzheng Quxie Prescription(mainly with the actions of supporting healthy qi and dispelling pathogens)combined with neoadjuvant chemotherapy on tumor recurrence,serum thymidine kinase 1(TK1)level and immune function in patients with triple-negative breast cancer(TNBC).Methods Eighty patients with TNBC of qi and yin deficiency type were randomly divided into a combination group and a control group,with 40 patients in each group.The control group was treated with AC-T sequential chemotherapy(Doxorubicin combined with Cyclophosphamide plus sequential Docetaxel),and the combination group was treated with Fuzheng Quxie Prescription on the basis of treatment for the control group.One course of treatment covered 21 days,and the two groups were treated for 4 consecutive courses.The changes of traditional Chinese medicine(TCM)syndrome scores,Karnofsky Performance Status(KPS)score,levels of tumor markers of carbohydrate antigen 125(CA125),carbohydrate antigen 153(CA153)and TK1,and T lymphocyte subset levels in the two groups were observed before and after the treatment.Moreover,the clinical efficacy and tumor metastasis and recurrence in the two groups were compared.Results(1)After 4 courses of treatment,the total effective rate of the combination group was 87.50%(35/40),and that of the control group was 67.50%(27/40),and the intergroup comparison(tested by chi-square test)showed that the efficacy of the combination group was significantly superior to that of the control group(P＜0.05).(2)After treatment,the TCM syndrome scores in the two groups were significantly decreased compared with those before treatment(P＜0.05),and the KPS scores were significantly increased compared with those before treatment(P＜0.05),and the decrease of TCM syndrome scores and the increase of KPS scores in the combination group were significantly superior to that in the control group(P＜0.05 or P＜0.01).(3)After treatment,the serum CA125,CA153 and TK1 levels of patients in the two groups were significantly decreased compared with those before treatment(P＜0.05),and the decrease of serum CA125,CA153 and TK1 levels in the combination group was significantly superior to that in the control group(P＜0.01).(4)After treatment,the T lymphocyte subset CD3+,CD4+ levels and CD4+/CD8+ ratio in the two groups were significantly increased compared with those before treatment(P＜0.05),and the CD8+ level was significantly decreased compared with that before treatment(P＜0.05).The post-treatment intergroup comparison showed that the increase of the T lymphocyte subset CD3+,CD4+ levels and CD4+/CD8+ ratio as well as the decrease of the CD8+ level in the combination group was all significantly superior to that in the control group(P＜0.05 or P＜0.01).(5)The one-year follow-up showed that the tumor recurrence rate and tumor metastasis rate in the combination group were 7.50%(3/40)and 12.50%(5/40)respectively,significantly lower than 25.00%(10/40)and 35.00%(14/40)in the control group,and the differences were statistically significant when comparing between the two groups(P＜0.05).Conclusion The combination of neoadjuvant chemotherapy with Fuzheng Quxie Prescription has a better therapeutic effect on TNBC patients with qi and yin deficiency syndrome,which can effectively improve the immune function of the patients,decrease the level of serum tumor markers,improve the quality of life of the patients,and reduce the incidence of tumor recurrence and metastasis.

Quantitative Structure-Activity Relationship in the Series of 5-Ethyluridine, N2-Guanine, and 6-Oxopurine Derivatives with Pronounced Anti-Herpetic Activity.

A quantitative analysis of the relationship between the structure and inhibitory activity against the herpes simplex virus thymidine kinase (HSV-TK) was performed for the series of 5-ethyluridine, N2-guanine, and 6-oxopurines derivatives with pronounced anti-herpetic activity (IC50 = 0.09 ÷ 160,000 µmol/L) using the GUSAR 2019 software. On the basis of the MNA and QNA descriptors and whole-molecule descriptors using the self-consistent regression, 12 statistically significant consensus models for predicting numerical pIC50 values were constructed. These models demonstrated high predictive accuracy for the training and test sets. Molecular fragments of HSV-1 and HSV-2 TK inhibitors that enhance or diminish the anti-herpetic activity are considered. Virtual screening of the ChEMBL database using the developed QSAR models revealed 42 new effective HSV-1 and HSV-2 TK inhibitors. These compounds are promising for further research. The obtained data open up new opportunities for developing novel effective inhibitors of TK.