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Nature Communications May 2023Cancer cells utilize the main de novo pathway and the alternative salvage pathway for deoxyribonucleotide biosynthesis to achieve adequate nucleotide pools....
Cancer cells utilize the main de novo pathway and the alternative salvage pathway for deoxyribonucleotide biosynthesis to achieve adequate nucleotide pools. Deoxycytidine kinase is the rate-limiting enzyme of the salvage pathway and it has recently emerged as a target for anti-proliferative therapies for cancers where it is essential. Here, we present the development of a potent inhibitor applying an iterative multidisciplinary approach, which relies on computational design coupled with experimental evaluations. This strategy allows an acceleration of the hit-to-lead process by gradually implementing key chemical modifications to increase affinity and activity. Our lead compound, OR0642, is more than 1000 times more potent than its initial parent compound, masitinib, previously identified from a drug repositioning approach. OR0642 in combination with a physiological inhibitor of the de novo pathway doubled the survival rate in a human T-cell acute lymphoblastic leukemia patient-derived xenograft mouse model, demonstrating the proof-of-concept of this drug design strategy.
Topics: Mice; Humans; Animals; Drug Repositioning; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Nucleotides; Drug Design; Disease Models, Animal
PubMed: 37248212
DOI: 10.1038/s41467-023-38668-2 -
Genes Oct 2022Hepatocellular carcinoma (HCC) originates from the hepatocytes and accounts for 90% of liver cancer. The study intends to identify novel prognostic biomarkers for...
BACKGROUND
Hepatocellular carcinoma (HCC) originates from the hepatocytes and accounts for 90% of liver cancer. The study intends to identify novel prognostic biomarkers for predicting the prognosis of HCC patients based on TCGA and GSE14520 cohorts.
METHODS
Differential analysis was employed to obtain the DEGs (Differentially Expressed Genes) of the TCGA-LIHC-TPM cohort. The lasso regression analysis was applied to build the prognosis model through using the TCGA cohort as the training group and the GSE14520 cohort as the testing group. Next, based on the prognosis model, we performed the following analyses: the survival analysis, the independent prognosis analysis, the clinical feature analysis, the mutation analysis, the immune cell infiltration analysis, the tumor microenvironment analysis, and the drug sensitivity analysis. Finally, the survival time of HCC patients was predicted by constructing nomograms.
RESULTS
Through the lasso regression analysis, we obtained a prognosis model of ten genes including (baculoviral IAP repeat containing 5), (cyclin-dependent kinase 4), (deoxycytidine kinase), (heat shock protein family A member 4), (heat shock protein 90 α family class A member 1), (Proteasome 26S Subunit Ubiquitin Receptor, Non-ATPase 2), (interleukin 1 receptor antagonist), (placental growth factor), (secreted phosphoprotein 1), and (stanniocalcin 2). First, we found that the risk score is an independent prognosis factor and is related to the clinical features of HCC patients, covering AFP (α-fetoprotein) and stage. Second, we observed that the mutation was the most obvious mutation between the high-risk and low-risk groups. Third, we also discovered that the risk score is related to some immune cells, covering B cells, T cells, dendritic, macrophages, neutrophils, etc. Fourth, the high-risk group possesses a lower TIDE score, a higher expression of immune checkpoints, and higher ESTIMATE score. Finally, nomograms include the clinical features and risk signatures, displaying the clinical utility of the signature in the survival prediction of HCC patients.
CONCLUSIONS
Through the comprehensive analysis, we constructed an immune-related prognosis model to predict the survival of HCC patients. In addition to predicting the survival time of HCC patients, this model significantly correlates with the tumor microenvironment. Furthermore, we concluded that these ten immune-related genes (, , , , , , , , , and ) serve as novel targets for antitumor immunity. Therefore, this study plays a significant role in exploring the clinical application of immune-related genes.
Topics: Female; Humans; alpha-Fetoproteins; Biomarkers, Tumor; Carcinoma, Hepatocellular; Cyclin-Dependent Kinase 4; Deoxycytidine Kinase; Heat-Shock Proteins; Kaplan-Meier Estimate; Liver Neoplasms; Osteopontin; Placenta Growth Factor; Proteasome Endopeptidase Complex; Receptors, Interleukin-1; Tumor Suppressor Protein p53; Ubiquitins
PubMed: 36292719
DOI: 10.3390/genes13101834 -
Immunology Jan 2023Multiple sclerosis (MS) is an autoimmune disease driven by lymphocyte activation against myelin autoantigens in the central nervous system leading to demyelination and...
Multiple sclerosis (MS) is an autoimmune disease driven by lymphocyte activation against myelin autoantigens in the central nervous system leading to demyelination and neurodegeneration. The deoxyribonucleoside salvage pathway with the rate-limiting enzyme deoxycytidine kinase (dCK) captures extracellular deoxyribonucleosides for use in intracellular deoxyribonucleotide metabolism. Previous studies have shown that deoxyribonucleoside salvage activity is enriched in lymphocytes and required for early lymphocyte development. However, specific roles for the deoxyribonucleoside salvage pathway and dCK in autoimmune diseases such as MS are unknown. Here we demonstrate that dCK activity is necessary for the development of clinical symptoms in the MOG and MOG experimental autoimmune encephalomyelitis (EAE) mouse models of MS. During EAE disease, deoxyribonucleoside salvage activity is elevated in the spleen and lymph nodes. Targeting dCK with the small molecule dCK inhibitor TRE-515 limits disease severity when treatments are started at disease induction or when symptoms first appear. EAE mice treated with TRE-515 have significantly fewer infiltrating leukocytes in the spinal cord, and TRE-515 blocks activation-induced B and T cell proliferation and MOG -specific T cell expansion without affecting innate immune cells or naïve T and B cell populations. Our results demonstrate that targeting dCK limits symptoms in EAE mice and suggest that dCK activity is required for MOG -specific lymphocyte activation-induced proliferation.
Topics: Animals; Mice; Multiple Sclerosis; Deoxycytidine Kinase; Encephalomyelitis, Autoimmune, Experimental; Lymphocytes; Disease Models, Animal; Mice, Inbred C57BL
PubMed: 35986643
DOI: 10.1111/imm.13569 -
Molecular Pharmaceutics Feb 2013Gemcitabine, an anticancer agent which acts against a wide range of solid tumors, is known to be rapidly deaminated in blood to the inactive metabolite... (Review)
Review
Gemcitabine, an anticancer agent which acts against a wide range of solid tumors, is known to be rapidly deaminated in blood to the inactive metabolite 2',2'-difluorodeoxyuridine and to be rapidly excreted by the urine. Moreover, many cancers develop resistance against this drug, such as loss of transporters and kinases responsible for the first phosphorylation step. To increase its therapeutic levels, gemcitabine is administered at high doses (1000 mg/m(2)) causing side effects (neutropenia, nausea, and so forth). To improve its metabolic stability and cytotoxic activity and to limit the phenomena of resistance many alternatives have emerged, such as the synthesis of prodrugs. Modifying an anticancer agent is not new; paclitaxel or ara-C has been subjected to such changes. This review summarizes the various chemical modifications that can be found in the 4-(N)- and 5'-positions of gemcitabine. They can provide (i) a protection against deamination, (ii) a better storage and (iii) a prolonged release in the cell, (iv) a possible use in the case of deoxycytidine kinase deficiency, and (v) transporter deficiency. These new gemcitabine-based sysems have the potential to improve the clinical outcome of a chemotherapy strategy.
Topics: Animals; Antimetabolites, Antineoplastic; Deoxycytidine; Humans; Kaplan-Meier Estimate; Models, Biological; Phosphorylation; Prodrugs; Gemcitabine
PubMed: 22978251
DOI: 10.1021/mp300370t -
PloS One 2014Efficient and adequate generation of deoxyribonucleotides is critical to successful DNA repair. We show that ataxia telangiectasia mutated (ATM) integrates the DNA...
Efficient and adequate generation of deoxyribonucleotides is critical to successful DNA repair. We show that ataxia telangiectasia mutated (ATM) integrates the DNA damage response with DNA metabolism by regulating the salvage of deoxyribonucleosides. Specifically, ATM phosphorylates and activates deoxycytidine kinase (dCK) at serine 74 in response to ionizing radiation (IR). Activation of dCK shifts its substrate specificity toward deoxycytidine, increases intracellular dCTP pools post IR, and enhances the rate of DNA repair. Mutation of a single serine 74 residue has profound effects on murine T and B lymphocyte development, suggesting that post-translational regulation of dCK may be important in maintaining genomic stability during hematopoiesis. Using [(18)F]-FAC, a dCK-specific positron emission tomography (PET) probe, we visualized and quantified dCK activation in tumor xenografts after IR, indicating that dCK activation could serve as a biomarker for ATM function and DNA damage response in vivo. In addition, dCK-deficient leukemia cell lines and murine embryonic fibroblasts exhibited increased sensitivity to IR, indicating that pharmacologic inhibition of dCK may be an effective radiosensitization strategy.
Topics: Animals; Ataxia Telangiectasia Mutated Proteins; B-Lymphocytes; Cell Line, Tumor; DNA Damage; DNA Repair; Deoxycytidine; Deoxycytidine Kinase; Deoxyribonucleosides; Genomic Instability; Hematopoiesis; Humans; Mice; Mice, Inbred BALB C; Mice, Knockout; Mutagenesis, Site-Directed; Phosphorylation; Protein Processing, Post-Translational; Substrate Specificity; T-Lymphocytes
PubMed: 25101980
DOI: 10.1371/journal.pone.0104125 -
Biochemical Pharmacology Feb 2020Deoxycytidine kinase (dCK) is an essential enzyme for production of nucleotides via the salvage pathway; DI-87 is a novel dCK inhibitor in preclinical development for...
BACKGROUND
Deoxycytidine kinase (dCK) is an essential enzyme for production of nucleotides via the salvage pathway; DI-87 is a novel dCK inhibitor in preclinical development for use in anticancer therapy. The current study utilizes PET imaging to evaluate PK-PD relationships and to determine optimal dosing of the drug.
METHODS
NSG mice bearing CEM tumors had plasma and tumor PK assessed using mass spectrometry following oral administration of DI-87. dCK inhibition was assessed after a single dose of oral DI-87 followed by a [F]CFA PET probe and PET imaging. Tumor growth inhibition was assessed by orally administering DI-87 with concurrent intraperitoneal thymidine.
RESULTS
DI-87 had an in vitro EC of 10.2 nM with low protein binding. Peak DI-87 concentrations were observed between 1-3 h and 3-9 h in plasma and tumor, respectively, with tumor concentrations less than one third of plasma. Full dCK inhibition, as evaluated by PET imaging, was observed as early as 3 h following 25 mg/kg dosing and was maintained for 12 h, with full recovery of enzyme activity after 36 h. When DI-87 was administered as repeated doses in combination with thymidine, full dCK inhibition was maintained at 12 h (25 mg/kg twice daily dose) and led to maximal tumor growth inhibition.
CONCLUSIONS
DI-87 is a promising new compound for use in combination therapy against tumors expressing dCK. Utilizing a [F]CFA PET probe targeting the pathway of interest allowed for efficient and accurate identification of the optimal dose for growth inhibition.
Topics: Animals; Antineoplastic Agents; Cell Line; Cell Proliferation; Deoxycytidine Kinase; Dose-Response Relationship, Drug; Drug Therapy, Combination; Gene Expression Regulation, Enzymologic; Humans; Male; Mice; Molecular Structure; Neoplasms, Experimental
PubMed: 31812677
DOI: 10.1016/j.bcp.2019.113742 -
Experimental Hematology & Oncology 2014The combination of rituximab and 2-CdA is an effective therapy for B-cell tumors. However, the molecular mechanisms and enzymatic pathways involved in the interaction...
Modulation of deoxycytidine kinase (dCK) and glycogen synthase kinase (GSK-3β) by anti-CD20 (rituximab) and 2-chlorodeoxyadenosine (2-CdA) in human lymphoid malignancies.
BACKGROUND
The combination of rituximab and 2-CdA is an effective therapy for B-cell tumors. However, the molecular mechanisms and enzymatic pathways involved in the interaction between the two agents are not fully understood. In this study, we provide molecular evidence for positive interaction between these two agents with resultant therapeutic benefit.
METHODS
Efficacy of the R-2CdA regimen was evaluated in thirteen patients with B-cell tumors (9 CLL; 3 WM and 1 FL), in vitro against 3 lymphoma cell lines and in a xenograft mouse model. Treatment-induced changes involving phenotype, kinase activity and protein expression were assessed in vitro and in the mouse xenograft tumors. The interaction between RTX and 2-CdA was analyzed using the multiple comparison method, Tukey's honestly significant difference (HSD). For the clinical and animal data, survival functions were estimated using the Kaplan-Meier method and compared by the log-rank test. P-values <0.05 were considered statistically significant. All statistical analyses were evaluated using GraphPad Prism 4 (San Diego, CA).
RESULTS
9 of 12 (75%) evaluable patients responded to the R-2-CdA regimen with median duration of response of 34 months. Median survival of patients from diagnosis and from completion of R-2-CdA treatment was 13.3 and 7.9 years, respectively. In vitro, the combination was effective in all 3 cell lines of lymphomas but with higher sensitivity in the follicular lymphoma cell line. The combination was also effective in the WSU-WM-SCID xenograft model with dose-dependent response and synergistic benefit. All animals were tumor-free for up to 120 days post 2 cycles of this regimen. Rituximab induced activation of deoxycytidine kinase (dCK), p38 mitogen activated protein kinase (p38MAPK) and glycogen synthase kinase-3β (GSK-3β) in the xenograft WSU-WM tumors. Chemical inhibition of p38MAPK led to inhibition of the GSK-3β phosphorylation suggesting that GSK-3β is regulated by p38MAPK in this model.
CONCLUSION
Collectively, our studies show concordance between the activity of R-2-CdA in vitro, in human and in WSU-WM xenograft model attesting to the validity of this model in predicting clinical response. Modulation of dCK and GSK-3β by rituximab may contribute to the positive therapeutic interaction between rituximab and 2-CdA.
PubMed: 25937997
DOI: 10.1186/2162-3619-3-31 -
Leukemia & Lymphoma Apr 2013Clofarabine is a second-generation purine nucleoside analog that has been synthesized to overcome the limitations and incorporate the best qualities of fludarabine and... (Review)
Review
Clofarabine is a second-generation purine nucleoside analog that has been synthesized to overcome the limitations and incorporate the best qualities of fludarabine and cladribine. Clofarabine acts by inhibiting ribonucleotide reductase and DNA polymerase, thereby depleting the amount of intracellular deoxynucleoside triphosphates available for DNA replication. Compared to its precursors, clofarabine has an increased resistance to deamination and phosphorolysis, and hence better stability as well as higher affinity to deoxycytidine kinase (dCyd), the rate-limiting step in nucleoside phosphorylation. Since the initiation of the first phase I study of clofarabine in 1993 in patients with hematologic and solid malignancies, clofarabine has demonstrated single-agent antitumor activity in adult acute leukemia, including acute myeloid leukemia (AML). Due to its unique properties of biochemical modulation when used in combination with other chemotherapy drugs, mainly cytarabine, combination regimens containing clofarabine have been evaluated. A review of the English literature was performed that included original articles and related reviews from the MEDLINE (PubMed) database and from abstracts based on the publication of meeting materials. This review describes the development, pharmacology and clinical activity of clofarabine, as well as its emerging role in the treatment of adult patients with AML and myelodysplastic syndrome.
Topics: Adenine Nucleotides; Age Factors; Animals; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Arabinonucleosides; Clofarabine; Drug Evaluation, Preclinical; Humans; Leukemia, Myeloid, Acute
PubMed: 22957815
DOI: 10.3109/10428194.2012.726722 -
Proceedings of the National Academy of... Dec 1996Three of the four deoxynucleoside kinases required for growth of Lactobacillus acidophilus R-26 exist as heterodimeric pairs specific for deoxyadenosine (dAK) and...
Three of the four deoxynucleoside kinases required for growth of Lactobacillus acidophilus R-26 exist as heterodimeric pairs specific for deoxyadenosine (dAK) and deoxycytidine (dCK) or dAK and deoxyguanosine (dGK). However, only two tandem genes, dak/dgk, are found, and are expressed only as dAK/dGK in transformed Escherichia coli. Sequencing peptides spanning 63% of the native dCK subunit revealed a sequence identical to that deduced from dgk (beginning MTVIVL...), except that dCK lacks residues 2 and 3 (dCK is M..IVL; dGK is .TVIVL). Also, mass spectrometry indicates that native dCK and dGK subunits are identical in mass adjusted for the first three residues. Furthermore, the native enzymes have identical isoelectric pH values, indicating an equal number of charged residues. To enable E. coli to express peptide having the native dCK sequence, codons 2 and 3 were deleted from the dgk portion of the tandem genes, resulting in expression of protein having the specificities and regulatory properties of native dAK/dCK, including heterotropic stimulation of dAK activity by deoxycytidine or dCTP (not deoxyguanosine or dGTP) and end-product inhibition of the respective activities by dATP and dCTP. Subcloning normal and mutant dgk yielded homodimeric dGK and dCK, respectively. The dCK homodimer strongly resembles human dCK, with a low K(m) for deoxycytidine, the ability to phosphorylate deoxyadenosine and deoxyguanosine at much higher K(m) values, and end-product inhibition by dCTP. Thus two distinct and specific enzymes evidently are derived from a single Lactobacillus gene. The mechanism by which this occurs in vivo has yet to be elucidated.
Topics: Amino Acid Sequence; Cloning, Molecular; Deoxycytidine Kinase; Escherichia coli; Gene Expression Regulation, Bacterial; Genes, Bacterial; Humans; Lactobacillus acidophilus; Molecular Sequence Data; Phosphotransferases (Alcohol Group Acceptor)
PubMed: 8962060
DOI: 10.1073/pnas.93.25.14385 -
Molecular Cancer Therapeutics Aug 2020Although gemcitabine is the cornerstone of care for pancreatic ductal adenocarcinoma (PDA), patients lack durable responses and relapse is inevitable. While the...
Although gemcitabine is the cornerstone of care for pancreatic ductal adenocarcinoma (PDA), patients lack durable responses and relapse is inevitable. While the underlying mechanisms leading to gemcitabine resistance are likely to be multifactorial, there is a strong association between activating gemcitabine metabolism pathways and clinical outcome. This study evaluated casein kinase 1 delta (CK1δ) as a potential therapeutic target for PDA and bladder cancer, in which CK1δ is frequently overexpressed. We assessed the antitumor effects of genetically silencing or pharmacologically inhibiting CK1δ using our in-house CK1δ small-molecule inhibitor SR-3029, either alone or in combination with gemcitabine, on the proliferation and survival of pancreatic and bladder cancer cell lines and orthotopic mouse models. Genetic studies confirmed that silencing CK1δ or treatment with SR-3029 induced a significant upregulation of deoxycytidine kinase (dCK), a rate-limiting enzyme in gemcitabine metabolite activation. The combination of SR-3029 with gemcitabine induced synergistic antiproliferative activity and enhanced apoptosis in both pancreatic and bladder cancer cells. Furthermore, in an orthotopic pancreatic tumor model, we observed improved efficacy with combination treatment concomitant with increased dCK expression. This study demonstrates that CK1δ plays a role in gemcitabine metabolism, and that the combination of CK1δ inhibition with gemcitabine holds promise as a future therapeutic option for metastatic PDA as well as other cancers with upregulated CK1δ expression.
Topics: Animals; Antimetabolites, Antineoplastic; Apoptosis; Biomarkers, Tumor; Breast Neoplasms; Casein Kinase Idelta; Cell Proliferation; Deoxycytidine; Deoxycytidine Kinase; Drug Resistance, Neoplasm; Female; Gene Expression Regulation, Neoplastic; Humans; Mice; Mice, Nude; Pancreatic Neoplasms; Tumor Cells, Cultured; Urinary Bladder Neoplasms; Xenograft Model Antitumor Assays; Gemcitabine
PubMed: 32430484
DOI: 10.1158/1535-7163.MCT-19-0997