Journal: bioRxiv

Article Title: A gel-top model for characterizing mesenchymal features of glioblastoma cells

doi: 10.1101/2025.04.03.647072

Figure Lengend Snippet: A-B . GSEA analysis of differentially expressed (DE) genes after treatment by CK2 inhibitors on the set of differential signature genes of mesenchymal versus proneural subtype (A) and on the hallmark epithelial to mesenchymal transition gene set (B). C . GO enrichment analysis of DE genes after CK2 inhibitors showed significant terms on extracellular matrix organization and response to hypoxia, both of which are associated with increased mesenchymal feature of mGBM cells as show in . D . Bar graphs showing vimentin and mesenchymal-associated cell-ECM interaction genes are repressed by CK2 inhibitors. p-values were calculated by DEseq2. N = 3.

Article Snippet: CX-4945 (MCE cat. HY-50855) and SGC-CK2-1 (MCE cat. HY-139004) was purchased from MedChemExpress.

Techniques:

Journal: bioRxiv

Article Title: A gel-top model for characterizing mesenchymal features of glioblastoma cells

doi: 10.1101/2025.04.03.647072

Figure Lengend Snippet: A . Bright filed images showing the morphology of networks formed by MGG31 cells onBME treated by either DMSO or CK2 inhibitors. Scale bar = 500 µm. B . Quantification of branch length of networks formed by MGG31 cells after CK2 inhibition. p-values were calculated by t-test. N = 5. C . CK2 inhibitor treatment on the co-culture of microglia with MGG31 on BME. Scale bar = 500 µm for top panel and 200 µm for middle and bottom panels. D-E . Bar graph showing maintained network connectedness after drug treatment (D), but recruitment of microglia cells is repressed by CK2 inhibitors(E). p-values were calculated by t-test. N=3 (D) or 26 (E).

Article Snippet: CX-4945 (MCE cat. HY-50855) and SGC-CK2-1 (MCE cat. HY-139004) was purchased from MedChemExpress.

Techniques: Inhibition, Co-Culture Assay

Journal: Cells

Article Title: Contribution of the CK2 Catalytic Isoforms α and α’ to the Glycolytic Phenotype of Tumor Cells

doi: 10.3390/cells10010181

Figure Lengend Snippet: CK2 expression and activity in SK-N-BE and U2OS cells. ( A ) Titration of the antibody reactivity towards CK2 catalytic subunits. The indicated amounts of recombinant CK2 catalytic subunits (myc-α’ or α) were loaded on SDS-PAGE, and either blotted for the WB (western blot) analysis or stained by Colloidal Coomassie Blue; ( B ) CK2 expression and activity in w.t. and KO clones of the cells used for this study. 10 μg proteins from cell lysates were analyzed by WB with the indicated antibodies. The last two right lanes belong to an independent experiment. As a reporter of CK2 endogenous activity, pS129 Akt signal has been quantified, normalized to total Akt signal, and reported in the bar graph as % of w.t. cells. At least three independent experiments were performed; representative western blots are shown, while quantification in the bar graphs reports the mean values ± SEM of all experiments and of the two clones of the same KO. Statistical significance refers to w.t. cells. (*) p < 0.05, (**) p < 0.01, (***) p < 0.001

Article Snippet: The CK2 inhibitor CX-4945 (5-[(3-Chlorophenyl)amino]-benzo[c]-2,6-naphthyridine-8-carboxylic) was purchased from MedChemExpress and the chemical inducer of hypoxia DMOG (dimethyloxallyl glycine) was from Sigma.

Techniques: Expressing, Activity Assay, Titration, Recombinant, SDS Page, Western Blot, Staining, Clone Assay

Journal: Cells

Article Title: Contribution of the CK2 Catalytic Isoforms α and α’ to the Glycolytic Phenotype of Tumor Cells

doi: 10.3390/cells10010181

Figure Lengend Snippet: Lactate production in response to CK2 knock out or pharmacological inhibition. ( A ) The extracellular lactate secreted in the culture medium in 8 h by the indicated cells was analyzed by means of a luminescence assay. Values for each KO are the means ± SEM obtained with the different clones of the same KO. At least three independent experiments were performed, in duplicate. Significance refers to w.t. cells ( B ) Cells were analyzed for the extracellular lactate secreted in the culture medium in 6 h, in the absence or in the presence of the indicated concentrations of CX-4945. Three independent experiments were performed, in duplicate. Results (means ± SEM) are reported as % compared to the respective w.t. cells, in the absence of CX-4945. Significance refers to untreated control. (*) p < 0.05, (**) p < 0.01, (***) p < 0.001, (****) p < 0.0001.

Article Snippet: The CK2 inhibitor CX-4945 (5-[(3-Chlorophenyl)amino]-benzo[c]-2,6-naphthyridine-8-carboxylic) was purchased from MedChemExpress and the chemical inducer of hypoxia DMOG (dimethyloxallyl glycine) was from Sigma.

Techniques: Knock-Out, Inhibition, Luminescence Assay, Clone Assay, Control

Journal: Cancers

Article Title: Mechanistic Basis for In Vivo Therapeutic Efficacy of CK2 Inhibitor CX-4945 in Acute Myeloid Leukemia

doi: 10.3390/cancers13051127

Figure Lengend Snippet: CK2 overexpression in AML cells causes IKAROS phosphorylation which is reversed by CK2 inhibitor, CX-4945. ( A ) Baseline protein levels of CK2α, CK2α’, CK2β, IKAROS and BCL-XL in the myeloid leukemia cell panel [U937, THP-1, K562, and primary AML cells (labelled AML-1) were measured by western blot and compared to CD34+ Hematopoietic Stem Cells (HSC). Multiple IKAROS bands represent Ikaros isoforms 1 and 2. ( B ) qRT-PCR showing mRNA level of CK2α and BCL-XL in various AML cells compared to CD34+ HSC. **** ( p < 0.0001). ( C ) Radio-blot showing increased Phosphorylated IKAROS (p-IKAROS) in AML cells compared to HSC. ( D ) Radio-immunoblot showing dose-dependent decrease in Phospho-IKAROS level following CX-4945 treatment. U937 and THP-1 were treated with 10 µM and AML-1 was treated with 5μM (IC50–3.791 μM) CX-4945 for 48 h. IC50 values of CX-4945 treated cells are shown in . ( E ) Western blot showing decrease in amount of phosphorylated CK2 substrates with molecular masses of approximately 175, 120, 80, 70 and 56 kDa as indicated by arrows (left panel) and western blot showing phosphorylation extent of specific CK2 substrate, AKT1 at Ser129 (right panel).

Article Snippet: The CK2 inhibitor CX-4945 sodium salt was a gift from Senhwa Biosciences.

Techniques: Over Expression, Phospho-proteomics, Western Blot, Quantitative RT-PCR

Journal: Cancers

Article Title: Mechanistic Basis for In Vivo Therapeutic Efficacy of CK2 Inhibitor CX-4945 in Acute Myeloid Leukemia

doi: 10.3390/cancers13051127

Figure Lengend Snippet: CK2 inhibition suppresses BCL-XL expression in AML cells and xenograft model. ( A ) U937 and AML-1 were treated with 5 and 10 µM concentration of CX-4945 for 48 h before RNA and protein extraction. mRNA level of BCL-XL was measured by qRT-PCR. ( B ) U937 and K562 cells were treated with 5 and 10 µM of CX-4945 for 48 hours and protein was extracted. BCL-XL protein level was measured by western blot. ( C ) Downregulation of CK2α in U937 was achieved using shRNA. Validation of CK2α protein knockdown in U937-shCK2α cell line is shown in . qRT-PCR shows mRNA level of CK2α and BCL-XL in CK2α shRNA treated U937 cells. ( D ) Overexpression of CK2α was achieved by retroviral transduction of U937 cells. Validation of CK2α protein overexpression is shown in . qRT-PCR showed increased BCL-XL mRNA level in CK2α overexpressed U937 (left panel) and THP-1 cells (right panel). ( E ) WST assay showing increased cell viability in CK2α overexpressing U937 and THP-1 cells. U937 cells transduced with retroviral vector CK2α-gfp-luc were transplanted into NRG-S mice. 25,000 cells were injected via the tail vein. Bioluminescence imaging using IVIS 100 was obtained weekly following transplant. ( F ) Bioluminescence signal in xenograft mice engrafting U937-gfp-luc cell or control (U937-ctl-gfp-luc) shown at week 2 and 3 post transplantation. ( G ) Kaplan Maier plot showing survival probability in U937-CK2-gfp-luc cells vs control. P -value summaries are as follows: p > 0.05 (ns-not significant); p < 0.01 (**); p < 0.001 (***); p < 0.0001 (****).

Article Snippet: The CK2 inhibitor CX-4945 sodium salt was a gift from Senhwa Biosciences.

Techniques: Inhibition, Expressing, Concentration Assay, Protein Extraction, Quantitative RT-PCR, Western Blot, shRNA, Biomarker Discovery, Knockdown, Over Expression, Retroviral, Transduction, WST Assay, Plasmid Preparation, Injection, Imaging, Control, Transplantation Assay

Journal: Cancers

Article Title: Mechanistic Basis for In Vivo Therapeutic Efficacy of CK2 Inhibitor CX-4945 in Acute Myeloid Leukemia

doi: 10.3390/cancers13051127

Figure Lengend Snippet: CX-4945 treatment shows therapeutic efficacy in AML Patient Derived Xenograft (PDX). ( A ) Schema showing AML PDX generation and treatment. AML-1 PDX was developed by injecting NRG-S mice with one million cells per mouse via the tail vein. Treatment was started when 2–5% human CD45+ cells were detected in peripheral blood (PB). Mice received 100 mg/kg of CX-4945 via gavage twice daily for 21 days. ( B ) Percent AML (human CD45+, CD13+,CD33+) in bone marrow (BM) and spleen of treated and untreated AML-1 PDX mice by flow cytometry. ( C ) Kaplan Maier plot showing survival probability of treated and untreated AML-1 PDX mice. Cell line derived xenograft (CDX) mouse models was developed using luciferase, and green fluro-protein (gfp) labeled U937 cells (U937-gfp-luc) or THP-1 cells (THP1-gfp-luc) transplanted into immunocompromised NRG-S mice as described in methods. Mice were treated with vehicle or CX-4945 at a dose of 100 mg/kg twice daily via gavage for up to 7 days. ( D ) Engraftment was monitored using bioluminescence imaging shown as the mean of the total flux in photons/second of mice in each group. ( E ) Following treatment, mice were sacrificed, and bone marrow mononuclear cells were collected. Human CD45+ cells in bone marrow were measured using flow cytometry. Flow cytometry using conjugated BCL-XL antibody was used to quantify intracellular BCL-XL protein level in FACS enriched human CD45+cells in the bone marrow of treated and untreated mice. shows histogram of BCL-XL and CK2α protein level in vehicle and CX-4945 treated U937 and THP-1 CDX. ( F ) Mean fluorescence intensity(MFI) is graphed, showing decreased BCL-XL protein level. P -value summaries are as follows: p > 0.05 (ns-not significant); p ≤ 0.05 (*); p < 0.01 (**); p < 0.001 (***).

Article Snippet: The CK2 inhibitor CX-4945 sodium salt was a gift from Senhwa Biosciences.

Techniques: Drug discovery, Derivative Assay, Flow Cytometry, Luciferase, Labeling, Imaging, Fluorescence

Journal: Cancers

Article Title: Mechanistic Basis for In Vivo Therapeutic Efficacy of CK2 Inhibitor CX-4945 in Acute Myeloid Leukemia

doi: 10.3390/cancers13051127

Figure Lengend Snippet: CK2 inhibitor increases IKAROS DNA-binding to BCL-XL . Chromatin immunoprecipitation (ChIP) followed by next-generation sequencing (ChIP-seq) and analysis of genome-wide occupancy of IKAROS was performed on U937 following the CX-4945 treatment at 10 µM concentration for 72 h. A change in IKAROS binding to promoter regions of the BCL-XL gene was analyzed following the CX-4945 treatment. ( A ) A chIP-seq signal map for IKAROS binding to the BCL-XL/BCL- 2L1 promoter region in U937-untreated labeled as WT (wild-type) (top panel) and CX-4945 treated U937 (bottom panel). Y-axis represents the log-2-fold change enrichment of IKAROS binding (** p < 0.01). ( B ) U937 and primary AML cells (AML-1) cells were treated with 10 and 5 μM of CX-4945, respectively (based on the IC50 value) for 48 and 72 h. IKAROS binding to the BCL-XL promoter region was confirmed using the qChIP assay in WT and CX-4945 treated cells. Binding at 72 h was not significantly increased compared to the 48 h treatment (not shown in the graph). Results are the mean +/– SD of three independent experiments. P-value summaries are as follows: p > 0.05 (ns-non significant); p < 0.001 (***); p < 0.0001 (****).

Article Snippet: The CK2 inhibitor CX-4945 sodium salt was a gift from Senhwa Biosciences.

Techniques: Binding Assay, Chromatin Immunoprecipitation, Next-Generation Sequencing, ChIP-sequencing, Genome Wide, Concentration Assay, Labeling

Journal: Cancers

Article Title: Mechanistic Basis for In Vivo Therapeutic Efficacy of CK2 Inhibitor CX-4945 in Acute Myeloid Leukemia

doi: 10.3390/cancers13051127

Figure Lengend Snippet: CK2 and IKAROS regulate sensitivity towards daunorubicin. CK2 overexpressing U937 ( A , B ) and THP-1 ( C , D ) were treated with 10 nM or 50 nM of daunorubicin for 48 hand stained with 7-AAD and Annexin V for flow cytometry to determine apoptosis. Flow plots ( A , C ) and percent apoptotic cells (early + late apoptosis) are shown in ( B , D ). Flow plots showing representative results from three replicates. The percentage of cells in the right upper and lower quadrant of each flow chart represents the percentage of late and early apoptotic cells, respectively. Q1-dead, Q2-late apoptosis, Q3-early apoptosis, Q4-live. ( E ) Cytotoxicity and drug response measured by MTT assay after treating CK2 overexpressing U937 and THP-1 cells and respective controls with various concentrations of daunorubicin for 48 h. p > 0.05 (ns); p < 0.05 (*); p < 0.001 (***); p < 0.0001 (****).

Article Snippet: The CK2 inhibitor CX-4945 sodium salt was a gift from Senhwa Biosciences.

Techniques: Staining, Flow Cytometry, MTT Assay

Journal: Cancers

Article Title: Mechanistic Basis for In Vivo Therapeutic Efficacy of CK2 Inhibitor CX-4945 in Acute Myeloid Leukemia

doi: 10.3390/cancers13051127

Figure Lengend Snippet: CK2 inhibition potentiates daunorubicin-induced apoptosis in AML cells. Cells were treated with 10 µM of CX-4945 or a combination of 10 µM CX-4945 with 10 nM of daunorubicin for up to 48 h. Cells were stained with 7-AAD and Annexin V for flow cytometry to assess apoptosis. ( A ) Flow plots showing representative results from three replicate experiments. The percentage of cells in the right upper and lower quadrant of each flow chart represents the percentage of late and early apoptotic cells, respectively, in U937 (top row), THP-1 (middle row), and AML-1 cells (bottom row) Q1: Dead, Q2: Late apoptosis, Q3: Early apoptosis, Q4: Live. Graphed in ( B ) are the mean +/−SD of triplicates from two independent experiments showing the percent of apoptosis cells following the drug treatment, as indicated above. ( C ) CK2α silencing in U937 cells achieved using ShCK2α and sorted for GFP after 24 h. Sorted cells were treated with 10 nM of DNR for 24 h before staining for Annexin V and 7AAD to assess apoptosis. The graph shows the combined percent apoptotic cells (early + late) in each group with and without the DNR treatment. ( D ) Cytotoxic drug response measured by the MTT assay after treating CK2α ShRNA treated U937 cells and the respective controls with various daunorubicin concentrations for 24 h. ( E ) Cells were pretreated as above for 48 h and were plated in a Methocult medium. Colonies were counted under an inverted light microscope. Colonies that contained around 50 cells or more were counted for analysis. Graphed in E is the number of colonies after 14 days as the mean of three replicates +/− SD of two independent experiments. ( F ) The qRT-PCR showing decreases in the mRNA level in U937 cells treated with daunorubicin alone (10 nM) or a combination of CX-4945 and daunorubicin. p -value summaries are as follows: p > 0.05 (ns); p < 0.01 (**); p < 0.001 (***); p < 0.0001 (****).

Article Snippet: The CK2 inhibitor CX-4945 sodium salt was a gift from Senhwa Biosciences.

Techniques: Inhibition, Staining, Flow Cytometry, MTT Assay, shRNA, Light Microscopy, Quantitative RT-PCR

Journal: Cancers

Article Title: Mechanistic Basis for In Vivo Therapeutic Efficacy of CK2 Inhibitor CX-4945 in Acute Myeloid Leukemia

doi: 10.3390/cancers13051127

Figure Lengend Snippet: Model illustration of the regulation of apoptosis in AML by CK2 and IKAROS via repression of BCL-XL.

Article Snippet: The CK2 inhibitor CX-4945 sodium salt was a gift from Senhwa Biosciences.

Techniques:

Journal: Cells

Article Title: Regulation of TMEM16A by CK2 and Its Role in Cellular Proliferation

doi: 10.3390/cells9051138

Figure Lengend Snippet: CK2 controls membrane expression of TMEM16A in CFBE airway epithelial cells. ( A ) Expression of double-tagged (eGFP and extracellular HA-tag) TMEM16A in CFBE airway epithelial cells. Membrane localized TMEM16A (Alexa647 positivity) was detected by an extracellular anti-HA-Alexa647-conjugated antibody. ( B , C ) RT-PCR and densitometric analysis indicating successful knockdown of CK2α’, #significant inhibition (unpaired t -test; p = 0.01). ( D , E ) Immunocytochemistry of TMEM16A expressed endogenously in CFBE cells. Membrane expression was reduced by knockdown of CK2α’, #significant inhibition (unpaired t -test; p = 0.000000002). Mean ± SEM. In parentheses are numbers of experiments.

Article Snippet: The CK2 inhibitors CX-4945 (silmitasertib) and TBB (4,5,6,7-Tetrabromobenzotriazole) were purchased from Cayman Chemicals and Sigma, respectively.

Techniques: Membrane, Expressing, Reverse Transcription Polymerase Chain Reaction, Knockdown, Inhibition, Immunocytochemistry

Journal: Cells

Article Title: Regulation of TMEM16A by CK2 and Its Role in Cellular Proliferation

doi: 10.3390/cells9051138

Figure Lengend Snippet: Inhibitors of CK2 inhibit TMEM16A in CFBE airway epithelial cells. ( A – F ) Whole cell current overlay recorded in patch clamp experiments and current/voltage relationships. ATP (100 µM) activated TMEM16A whole cell Cl − currents that were strongly inhibited by the CK2-inhibitors TBB (10 µM; #significant inhibition, unpaired t -test; p = 0.01, ( A , B )) and CX4945 (20 µM; #significant inhibition, unpaired t -test; p = 0.02; ( C , D )), and siRNA-knockdown of CK2α’ (#significant inhibition, unpaired t -test; p = 0.0001; ( E , F )). ( G , H ) Plasma membrane (PM) expression of endogenous TMEM16A in CFBE cells and inhibition of PM expression by the CK2-inhibitor CX4945 (#significant inhibition, unpaired t -test; p = 0.000000000007). Mean ± SEM #significant inhibition ( p < 0.05; unpaired t -test). In parentheses are numbers of experiments.

Article Snippet: The CK2 inhibitors CX-4945 (silmitasertib) and TBB (4,5,6,7-Tetrabromobenzotriazole) were purchased from Cayman Chemicals and Sigma, respectively.

Techniques: Patch Clamp, Inhibition, Knockdown, Clinical Proteomics, Membrane, Expressing

Journal: Cells

Article Title: Regulation of TMEM16A by CK2 and Its Role in Cellular Proliferation

doi: 10.3390/cells9051138

Figure Lengend Snippet: Role of CK2 for plasma membrane expression of TMEM16A in Cal33 head and neck cancer cells. ( A , B ) RT-PCR and densitometric analysis indicating successful knockdown of CK2α’ by siRNA for CK2α’ in Cal33 head and neck cancer cells (#significant inhibition, unpaired t -test; p = 0.01). Knockdown of CK2α’ did not inhibit transcription of TMEM16A. ( C , D ) Western blot analysis indicating successful knockdown of CK2α’ but unaffected expression of TMEM16A. ( E , F ) Plasma membrane (PM) expression of TMEM16A expressed endogenously in Cal33 cells and inhibition of PM expression by knockdown of CK2α’ (#significant inhibition, unpaired t -test; p = 0.00000002). ( G ) Current/voltage relationships of ATP-activated TMEM16A whole cells currents, indicating inhibition of TMEM16A by knockdown of CK2α’ (#significant inhibition, unpaired t -test; p = 0.01). Mean ± SEM. In parentheses are numbers of experiments.

Article Snippet: The CK2 inhibitors CX-4945 (silmitasertib) and TBB (4,5,6,7-Tetrabromobenzotriazole) were purchased from Cayman Chemicals and Sigma, respectively.

Techniques: Clinical Proteomics, Membrane, Expressing, Reverse Transcription Polymerase Chain Reaction, Knockdown, Inhibition, Western Blot

Journal: Cells

Article Title: Regulation of TMEM16A by CK2 and Its Role in Cellular Proliferation

doi: 10.3390/cells9051138

Figure Lengend Snippet: Inhibition of proliferation by knockdown of CK2α’ and TMEM16A. ( A ) Cell proliferation assessed in MTT assays and shown as absorbance. Both siRNA-knockdown of CK2α’ and TMEM16A inhibited cell proliferation (#significant inhibition, unpaired t -tests; p = 0.0001). Simultaneous knockdown of CK2α’ and TMEM16A had a more pronounced inhibitory effect on cell proliferation (#significant inhibition, unpaired t -test; p = 0.0015). ( B ) Inhibition of cell proliferation by the CK2-inhibitor CX4945 (20 µM) and additional inhibitory effect of TMEM16A-knockdown (#significant inhibition, unpaired t -test; p = 0.0001). Mean ± SEM. In parentheses are numbers of experiments.

Article Snippet: The CK2 inhibitors CX-4945 (silmitasertib) and TBB (4,5,6,7-Tetrabromobenzotriazole) were purchased from Cayman Chemicals and Sigma, respectively.

Techniques: Inhibition, Knockdown

Journal: Cells

Article Title: Regulation of TMEM16A by CK2 and Its Role in Cellular Proliferation

doi: 10.3390/cells9051138

Figure Lengend Snippet: Blockers of CK2 and TMEM16A inhibit proliferation of Cal33 and BHY head and neck cancer cells. ( A ) Blocking CK2 by CX4945 (20 µM) and blocking TMEM16A by niclosamide (0.5 µM) inhibited proliferation of Cal33 cells. Simultaneous application of both blockers had an additive effect (#significant inhibition, unpaired t -tests; p = 0.0001). ( B ) Enhanced cell proliferation of BHY cells induced by the TMEM16A-activator, Eact. Blocking CK2 by CX4945 (20 µM) and blocking TMEM16A by niclosamide (0.5 µM) inhibited proliferation of BHY cells. Simultaneous application of both blockers had an additive effect (#significant inhibition, unpaired t -tests; p = 0.000015). Mean ± SEM. In parentheses are numbers of experiments.

Article Snippet: The CK2 inhibitors CX-4945 (silmitasertib) and TBB (4,5,6,7-Tetrabromobenzotriazole) were purchased from Cayman Chemicals and Sigma, respectively.

Techniques: Blocking Assay, Inhibition

Journal: Cells

Article Title: Regulation of TMEM16A by CK2 and Its Role in Cellular Proliferation

doi: 10.3390/cells9051138

Figure Lengend Snippet: Blockers of CK2 and TMEM16A inhibit receptor-mediated Ca 2+ signaling. ( A , B ) Original recordings and summaries for basal and ATP-induced intracellular Ca 2+ concentrations in Cal33 cells. Increase of intracellular Ca 2+ by 10 and 100 µM ATP, respectively. Both CX4945 (20 µM; #significant inhibition, ANOVA; p = 0.0004) and niclosamide (1 µM; #significant inhibition, ANOVA; p = 0.0002) largely reduced ATP-induced Ca 2+ increase. Mean ± SEM. In parentheses are numbers of experiments.

Article Snippet: The CK2 inhibitors CX-4945 (silmitasertib) and TBB (4,5,6,7-Tetrabromobenzotriazole) were purchased from Cayman Chemicals and Sigma, respectively.

Techniques: Inhibition