hdaci panobinostat  (MedChemExpress)

Journal: Molecular Therapy. Nucleic Acids

Article Title: Combining anti-gene γPNA with small molecules and RNA inhibitors: A strategy to enhance anti-tumor efficacy

doi: 10.1016/j.omtn.2025.102804

Figure Lengend Snippet: Design of anti-transcription γPNA and combination treatments to target the c-Myc oncogene (A) Schematic representation of gamma peptide nucleic (γPNA)-mediated inhibition of human c-Myc transcription and target site (NCBI database RefSeq: NG_007161.2 ). (B) Design of γPNA conjugated with nuclear localization signal (NLS) to target the indicated sites. ScR-γPNA2 is the scramble control. (C) Graphic representation of combination treatments with anti-transcription, γPNA1. The combination treatments include histone deacetylase inhibitors (HDACis), MYC/MAX inhibitors, small interfering RNA (siRNA), and small molecules targeting other pathways. (D) Polymerase chain reaction (PCR)-based amplicon assay to confirm binding of γPNA1 to the target site in U2932 cells. Amplicon assay after treatment of γPNA1 and ScR-γPNA2 with HDACi. (E) The graph represents quantification of γPNA1 and ScR-γPNA2 amplicon in combination with HDACi. (F) The graph represents the quantification of amplicon assay from class I HDACi with γPNA1. Results are presented as mean ± SEM. One-way ANOVA was used to determine the statistically significant difference between groups.

Article Snippet: U2932 cells were treated with HDACi: panobinostat (MedchemExpress, #HY-10224; 30 nM), belinostat (MedchemExpress, #HY-10225; 2.5 μM), entinostat (MedchemExpress, #HY-12163; 10 μM, romidepsin (Sigma-Aldrich, #SML1175; 10 nM), and vorinostat (Selleckchem, #S1047; 2.5 μM) for 24 h followed by PBS, γPNA1, and ScR-γPNA2 at 3 μM for 48 h cotreatment.

Techniques: Inhibition, Control, Histone Deacetylase Assay, Small Interfering RNA, Polymerase Chain Reaction, Amplification, Binding Assay

Journal: Molecular Therapy. Nucleic Acids

Article Title: Combining anti-gene γPNA with small molecules and RNA inhibitors: A strategy to enhance anti-tumor efficacy

doi: 10.1016/j.omtn.2025.102804

Figure Lengend Snippet: Anti-transcription activity of γPNA1 with HDACi in lymphoma cells Relative fold change of c-Myc levels in U2932 cells measured by real-time PCR on day 2 after treatment with (A) γPNA1 and (B) ScR-γPNA2 in combination with romidepsin, entinostat, vorinostat, panobinostat, and belinostat. Results are presented as mean ± SEM and two-way ANOVA was used to determine the statistically significant difference between groups. Western blot analysis representing the change in c-MYC protein on day 2 after treatment with γPNA1 and ScR-γPNA2 in combination with (C) romidepsin, (D) entinostat, (E) vorinostat, (F) panobinostat, and (G) belinostat. ∗∗(C–F) Cyclophilin B was used as an endogenous control, and the same blots are presented in C–S3G. c-MYC, EZH2, and cyclophilin B were probed from the same blot. Results are presented as mean ± SEM, and the p value between groups was determined using one-way ANOVA.

Article Snippet: U2932 cells were treated with HDACi: panobinostat (MedchemExpress, #HY-10224; 30 nM), belinostat (MedchemExpress, #HY-10225; 2.5 μM), entinostat (MedchemExpress, #HY-12163; 10 μM, romidepsin (Sigma-Aldrich, #SML1175; 10 nM), and vorinostat (Selleckchem, #S1047; 2.5 μM) for 24 h followed by PBS, γPNA1, and ScR-γPNA2 at 3 μM for 48 h cotreatment.

Techniques: Activity Assay, Real-time Polymerase Chain Reaction, Western Blot, Control

Journal: Molecular Therapy. Nucleic Acids

Article Title: Combining anti-gene γPNA with small molecules and RNA inhibitors: A strategy to enhance anti-tumor efficacy

doi: 10.1016/j.omtn.2025.102804

Figure Lengend Snippet: Cell viability of Histone deacetylase inhibitors in combination with γPNA1 (A) Cell viability of U2932 cells treated with increasing doses of HDACi (romidepsin, entinostat, vorinostat, panobinostat, and belinostat) alone and in combination with γPNA1 and ScR-γPNA2 (8 μM) for 48 h. Results are presented as mean ± SEM. (B) The IC 50 ± SEM values of HDACi and combination treatment of HDACi with γPNA1 in U2932 cells. (C) Cell viability of Raji cells treated with increasing doses of HDACi (romidepsin, entinostat, vorinostat, panobinostat, and belinostat) alone and in combination with γPNA1 and ScR-γPNA2 (8 μM) for 48 h. Results are presented as mean ± SEM. (D) The IC 50 ± SEM values of HDACi and combination treatment of HDACi with γPNA1 in Raji cells.

Article Snippet: U2932 cells were treated with HDACi: panobinostat (MedchemExpress, #HY-10224; 30 nM), belinostat (MedchemExpress, #HY-10225; 2.5 μM), entinostat (MedchemExpress, #HY-12163; 10 μM, romidepsin (Sigma-Aldrich, #SML1175; 10 nM), and vorinostat (Selleckchem, #S1047; 2.5 μM) for 24 h followed by PBS, γPNA1, and ScR-γPNA2 at 3 μM for 48 h cotreatment.

Techniques: Histone Deacetylase Assay

Journal: International Journal of Molecular Sciences

Article Title: Synergistic Cytotoxicity of Histone Deacetylase and Poly-ADP Ribose Polymerase Inhibitors and Decitabine in Breast and Ovarian Cancer Cells: Implications for Novel Therapeutic Combinations

doi: 10.3390/ijms25179241

Figure Lengend Snippet: Dose–response curves for the studied drugs in breast and ovarian cancer cell lines. Cells were seeded in 96-well plates overnight and exposed to different concentrations of individual drugs for 3 days as described in Methods. Rate of cell proliferation was determined relative to control by MTT assay ( A , B ). Model-adjusted means are shown with 95% confidence intervals for the non-zero doses modeled, and solid points indicate a significant difference from the first non-zero dose . Each cell line of each drug was modeled independently. IC 50 values were determined using CalcuSyn 2.0 software. Pano: panobinostat; SAHA: vorinostat; TLZ: talazoparib; Ola: olaparib; DAC: decitabine.

Article Snippet: The HDACis panobinostat and vorinostat, the PARPis talazoparib and olaparib, and the demethylating agent decitabine were obtained from Selleck Chemicals (Houston, TX, USA).

Techniques: Control, MTT Assay, Software

Journal: International Journal of Molecular Sciences

Article Title: Synergistic Cytotoxicity of Histone Deacetylase and Poly-ADP Ribose Polymerase Inhibitors and Decitabine in Breast and Ovarian Cancer Cells: Implications for Novel Therapeutic Combinations

doi: 10.3390/ijms25179241

Figure Lengend Snippet: Synergistic cytotoxicity of HDACi, PARPi, and decitabine. Cells were seeded in 96-well plates overnight and exposed to drugs individually, or in three-drug combinations at a constant concentration ratio, and cell proliferation was analyzed after 3 days. The relationships between the calculated combination indexes ( Y -axis) and fractions affected ( X -axis) are shown. A combination index < 1.0 indicates synergism. The graphs are representative of two independent experiments.. SAHA: vorinostat; TLZ: talazoparib; DAC: decitabine; PANO: panobinostat; OLA: olaparib.

Article Snippet: The HDACis panobinostat and vorinostat, the PARPis talazoparib and olaparib, and the demethylating agent decitabine were obtained from Selleck Chemicals (Houston, TX, USA).

Techniques: Concentration Assay

Journal: International Journal of Molecular Sciences

Article Title: Synergistic Cytotoxicity of Histone Deacetylase and Poly-ADP Ribose Polymerase Inhibitors and Decitabine in Breast and Ovarian Cancer Cells: Implications for Novel Therapeutic Combinations

doi: 10.3390/ijms25179241

Figure Lengend Snippet: Colony formation assay. Cells were seeded in six-well plates overnight and exposed to individual drugs or three-drug combinations for 1–2 weeks and stained as described in Methods ( A ). Colony formation is presented relative to control ( B ). For the double- and triple-drug combinations, the drug concentrations were the same as indicated in the single-drug concentrations. Model-adjusted means are shown with 95% confidence intervals, and solid points indicate a significant synergistic difference from all of the component drugs (see ). Each cell line of each drug was modeled independently. Number of replicates = 3; Pano/P: panobinostat; SAHA/S: vorinostat; TLZ/T: talazoparib; Ola/O: olaparib; DAC/D: decitabine.

Article Snippet: The HDACis panobinostat and vorinostat, the PARPis talazoparib and olaparib, and the demethylating agent decitabine were obtained from Selleck Chemicals (Houston, TX, USA).

Techniques: Colony Assay, Staining, Control

Journal: International Journal of Molecular Sciences

Article Title: Synergistic Cytotoxicity of Histone Deacetylase and Poly-ADP Ribose Polymerase Inhibitors and Decitabine in Breast and Ovarian Cancer Cells: Implications for Novel Therapeutic Combinations

doi: 10.3390/ijms25179241

Figure Lengend Snippet: Drug-mediated inhibition of cell proliferation and PARylation and effects on survival and apoptosis protein markers. Cells were seeded in T25 flasks overnight and exposed to individual drugs or three-drug combinations for 3 days, harvested, and analyzed for cell proliferation by MTT assay ( A ) and Western blotting ( B ). For the double- and triple-drug combinations, the drug concentrations were the same as indicated in the single-drug concentrations. Model-adjusted means are shown with 95% confidence intervals, and solid points indicate a significant synergistic difference from all of the component drugs . Each cell line of each drug was modeled independently. Number of replicates = 3; Pano/P: panobinostat; SAHA/S: vorinostat; TLZ/T: talazoparib; Ola/O: olaparib; DAC/D: decitabine.

Article Snippet: The HDACis panobinostat and vorinostat, the PARPis talazoparib and olaparib, and the demethylating agent decitabine were obtained from Selleck Chemicals (Houston, TX, USA).

Techniques: Inhibition, MTT Assay, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: Synergistic Cytotoxicity of Histone Deacetylase and Poly-ADP Ribose Polymerase Inhibitors and Decitabine in Breast and Ovarian Cancer Cells: Implications for Novel Therapeutic Combinations

doi: 10.3390/ijms25179241

Figure Lengend Snippet: Effects of drugs on the levels of various proteins involved in DNA repair/DNA damage response. Cells were exposed to the indicated drug concentrations for 3 days prior to analysis by Western blotting. For the double- and triple-drug combinations, the drug concentrations were the same as indicated in the single-drug concentrations. Number of replicates = 3; DSB: double-strand break; HR: homologous recombination; NHEJ: non-homologous DNA end-joining; NuRD: nucleosome remodeling and deacetylase. Pano: panobinostat; SAHA: vorinostat; TLZ: talazoparib; Ola: olaparib; DAC: decitabine.

Article Snippet: The HDACis panobinostat and vorinostat, the PARPis talazoparib and olaparib, and the demethylating agent decitabine were obtained from Selleck Chemicals (Houston, TX, USA).

Techniques: Western Blot, Homologous Recombination, Histone Deacetylase Assay

Journal: Cancer Research Communications

Article Title: HDAC Inhibition Induces CD26 Expression on Multiple Myeloma Cells via the c-Myc/Sp1-mediated Promoter Activation

doi: 10.1158/2767-9764.CRC-23-0215

Figure Lengend Snippet: Induction of CD26 expression on myeloma cell lines by HDACi. A, CD26 expression in plasma cells of bone marrow tissues of patients with multiple myeloma. Analysis of primary BM samples from several patients with multiple myeloma revealed that CD138 pos plasma cells were rarely stained with CD26 (gray, CD138; red, CD26; original magnification, × 200). B, Flow cytometry with anti-CD26 (rat clone)-fluorescein (FITC) or isotype control IgG 1 was performed in five myeloma cell lines KMS11, 26, 27, 28, and RPMI8226. Overlay histograms show CD26 expression on myeloma cell lines before and after treatment with one of nine HDACi for the indicated times (24, 48, 72, and 48 hours after subsequent removal of each HDACi) at the indicated doses. HDACi elicited exposure time-dependent upregulation of CD26 expression on myeloma cells, whereas subsequent removal of HDACi resulted in a decline of CD26 expression to the decreased or near-pretreatment levels. C, KMS27 and KMS28 was incubated with titrated concentrations of panobinostat (0.5, 5.0, 50, and 100 nmol/L), RG2833 (5, 50 nmol/L, 0.5, and 5.0 µmol/L) and entinostat (0.5, 5.0, 50, and 500 µmol/L) for 48 hours, after which cells were harvested to analyze the levels of surface CD26 expression in myeloma cells by flow cytometry. Overlay histogram shows CD26 expression on each myeloma cell before and after 48 hours of treatment with each HDACi at the indicated doses. HDACi elicited a dose-dependent upregulation of CD26 expression on myeloma cells, whereas 5.0 µmol/L of RG2833 and 500 µmol/L of entinostat did not further/significantly enhance CD26 expression on myeloma cells, compared with 0.5 µmol/L of RG2833 and 50 µmol/L of entinostat. D, Myeloma cell lines KMS11, 26, 27, 28, and RPMI8226 were immunohistochemically stained for CD26 before and after treatment with one of nine HDACi. All tested myeloma cell lines cultured alone without each HDACi were either slightly stained for CD26 or completely lacked CD26 expression. In contrast, cell lines treated with each HDACi for 48 hours revealed moderate to intense CD26 expression (CD26, brown stain; original magnification, × 200). E, Thereafter, removal of the HDACi for 48 hours resulted in CD26 expression to the decreased or near-pretreatment levels again (CD26; brown-stained; original magnification, × 200). F, Expression levels of CD26 mRNA in myeloma cell lines KMS11, 26, 27, 28, and RPMI8226 before and after the treatment of one of nine HDACi for 48 hours were analyzed using real-time quantitative RT-PCR assay with specific primers for CD26 . The CD26 mRNA transcription levels in myeloma cell lines treated with each HDACi revealed a significant increase, compared with those of untreated myeloma cells. Results are shown as ratio of CD26mRNA/GAPDH mRNA. Bar diagrams represent the mean values ± SE. n = 3; *, P < 0.05; **, P < 0.01. G, The levels of DPPⅣ activity in supernatants derived from myeloma cell lines KMS11, 27, 28, and RPMI8226, cultured in the presence or absence of one of nine HDACi for 48 hours were determined by ELISA. The DPPⅣ levels in supernatants of myeloma cells, which were incubated in the presence of each HDACi were significantly elevated, compared with those of control IgG 1 . The data represent the mean ± SE of triplicate wells from the representative of three independent experiments. The error bars represent the range, *, P < 0.05.

Article Snippet: HDAC inhibitors; pan HDACi: panobinostat_50 μmol/L, vorinostat_1.0 μmol/L, isoform-selective HDACi: romidepsin (HDAC1i)_0.125 μmol/L, BG45 (HDAC1, 3i) 1.0 μmol/L, entinostat (HDAC1, 3i)_50 μmol/L, RG2833 (HDAC1, 3i)_0.5 μmol/L, nexturastat A (HDAC6i)_0.125 μmol/L, tubastatin A (HDAC6i)_25 μmol/L, ricolinostat (HDAC1, 3, 6i)_0.5 μmol/L were purchased from Selleck Chemical Co. LTD. for use as therapeutic agents.

Techniques: Expressing, Clinical Proteomics, Staining, Flow Cytometry, Control, Incubation, Cell Culture, Quantitative RT-PCR, Activity Assay, Derivative Assay, Enzyme-linked Immunosorbent Assay

Journal: Cancer Research Communications

Article Title: HDAC Inhibition Induces CD26 Expression on Multiple Myeloma Cells via the c-Myc/Sp1-mediated Promoter Activation

doi: 10.1158/2767-9764.CRC-23-0215

Figure Lengend Snippet: Transcriptomic profiles in three myeloma cell lines, either treated or untreated with HDACi or HDACi plus CD26mAb. A, Schema of transcriptomic analysis of three myeloma cell lines KMS11, 27, and RPMI8226, treated with one of two HDACi by panobinostat (50 µmol/L) or RG2833 (0.5 µmol/L) or HDACi plus CD26mAb in combination. B, Three myeloma cell lines were treated with panobinostat (50 µmol/L) or RG2833 (0.5 µmol/L) for 48 hours, followed by additional incubation with CD26mAb for 24 hours and transcriptomic profiles in each myeloma cell line were analyzed at each timepoints using microarray analysis. The heat maps show normalized relative mRNA expression differences, based on log 2 fold change, with a cut-off P value <0.05. The color scale of the heat map from blue to red indicates low to high expression. C, Venn diagrams showing overlap in the most significantly upregulated genes (log 2 fold change >10, with a cut-off P value <0.05) among three myeloma cell lines treated with either panobinostat (50 µmol/L) or RG2833 (0.5 µmol/L) for 48 hours compared with the isotype control IgG 1 . D, Venn diagrams showing overlap in the most significantly downregulated genes (log 2 fold change <−10, with a cut-off P value <0.05) among three myeloma cell lines treated with either panobinostat (50 µmol/L) or RG2833 (0.5 µmol/L) for 48 hours compared with the isotype control IgG 1 . E, Among the 36 downregulated genes common to all three myeloma cell lines with the treatment of either panobinostat or RG2833, overlapped 23 genes excluding 13 noncoding genes were identified (log 2 fold change <−10, with a cut-off P value<0.05). The values show the fold changes of mRNA expression of transcripts in each myeloma cell line treated with each HDACi or either HDACi plus CD26mAb compared with control IgG 1 . The color scale of the heat map from blue to red indicates low to high expression. Similarly, among the 16 genes, commonly upregulated in all three myeloma cell lines on treatment with each HDACi, overlapped 14 genes excluding noncoding genes were indicated. F, Left, c-Myc gene signal (log 2 ) in KMS11, KMS27, and RPMI8226, treated with isotype control IgG 1 , HDACi; panobinostat or RG2833 and HDACi plus CD26mAb were shown. Myc is one of the genes, significantly downregulated in common, excluding noncoding genes in three myeloma cell lines following treatment with each HDACi or either HDACi plus CD26mAb. Right, A three-dimenisonal MAP was constructed by principal component analysis, indicating gene expression patterns based on transcriptome analysis.

Article Snippet: HDAC inhibitors; pan HDACi: panobinostat_50 μmol/L, vorinostat_1.0 μmol/L, isoform-selective HDACi: romidepsin (HDAC1i)_0.125 μmol/L, BG45 (HDAC1, 3i) 1.0 μmol/L, entinostat (HDAC1, 3i)_50 μmol/L, RG2833 (HDAC1, 3i)_0.5 μmol/L, nexturastat A (HDAC6i)_0.125 μmol/L, tubastatin A (HDAC6i)_25 μmol/L, ricolinostat (HDAC1, 3, 6i)_0.5 μmol/L were purchased from Selleck Chemical Co. LTD. for use as therapeutic agents.

Techniques: Incubation, Microarray, Expressing, Control, Construct, Gene Expression

Journal: Cancer Research Communications

Article Title: HDAC Inhibition Induces CD26 Expression on Multiple Myeloma Cells via the c-Myc/Sp1-mediated Promoter Activation

doi: 10.1158/2767-9764.CRC-23-0215

Figure Lengend Snippet: Epigenetic modification at the CD26 promoter of myeloma cells on treatment with HDACi. A, The effects of HDACi on histone H3 acetylation on the CD26 promoter of myeloma cells are shown. Myeloma cell lines KMS11, 26, 27, 28, and RPMI8226, treated with either panobinostat (50 µmol/L), RG2833 (0.5 µmol/L), or tubastatinA (2.5 µmol/L) or control IgG 1 for 48 hours were investigated by ChIP assay using anti-histone 3 on lysine 27 acetylated (H3K27Ac) antibody or rabbit IgG and then, the DNAs of immunoprecipitated chromatin were amplified and quantified by real-time qPCR with the primer pairs for the CD26 promoter shown in . NoAb means samples prepared without antibodies as a control and INPUT indicates that PCR was performed with genomic DNA. Increased levels of acetylation at H3K27 was observed on the CD26 promoter of myeloma cells treated with each HDACi, compared with control IgG 1 . Values represent percentage of IP/INPUT. Bars represent the mean ± S.D. of three independent experiments done in triplicate. B, qRT-PCR for c-Myc expression in myeloma cell lines KMS11, 26, 27, 28, and RPMI8226 treated with one of nine HDACi, namely, BG45 (1 µmol/L), romidepsin (0.125 µmol/L), ricolinostat (0.5 µmol/L), panobinobinostat (50 µmol/L), entinostat (50 µmol/L), nexturastatA (0.125 µmol/L), vorinostat (1 µmol/L), tubastatinA (25 µmol/L), RG2833 (0.5 µmol/L), or control IgG for 48 hours. The expression of c-Myc mRNA was significantly reduced in each myeloma cell line after exposure to each HDACi for 48 hours (*, P < 0.01). Furthermore, the expression levels of c-Myc mRNA in KMS11, 27, and RPMI8226 were examined at the indicated times (3.0, 6.0, 12, 24 hours) following treatment with panobinostat or RG2833 (*, P < 0.05; **, P < 0.01). The expression levels of c-Myc were normalized to that of GAPDH and quantified by the 2 −△△Ct method. Data are shown as the ratio of c-Myc mRNA/GAPDH mRNA and represent the means ± S.D. of three independent experiments. C, Expression levels of c-Myc and c-MycK323ac protein in KMS11, 27, and RPMI8226 were examined in the presence or absence of panobinostat (50 µmol/L) or RG2833 (0.5 µmol/L) at the indicated times (0.5, 1.0, 3.0, 6.0, 12, 24 hours) by immunoblotting. Levels of these proteins in normal MNCs, incubated with panobinostat, RG2833 or control IgG 1 for 24 hours were also analyzed. D, To investigate the binding of c-Myc and c-MycK323ac to the CD26 promoter of myeloma cells, KMS11, 27, and RPMI8226 were treated with panobinostat (50 µmol/L), RG2833 (0.5 µmol/L), or control IgG 1 for the indicated times (3.0, 6.0, 12, 24 hours) and then, ChIP assays were conducted in each myeloma cell using anti-c-Myc or c-MycK323ac antibody or rabbit IgG. Thereafter, the DNAs of each immunoprecipitated chromatin suspension were amplified and quantified by real-time qPCR with specific primers for the CD26 promoter via the proximal G-C box . The recovery of ChIP's DNAs was calculated as the percentages of IP/INPUT. The time-dependent decrease of the binding of c-Myc to the promoter, concomitant with a time-dependent increase in the binding of c-MycK323ac to the promoter, was observed in each myeloma cell line treated with each HDACi, compared with control IgG 1 . Bars represent the mean ± S.D. of three independent experiments done. The amplified products were also visualized by MIDORI green Direct staining following 1.5% agarose gel electrophoresis. Representative data of 40 cycles are shown. INPUTs show that PCR was conducted with genomic DNA. The actin signal shows equal loading as a control. E, The binding of c-Myc to the transcriptional factor, Sp1 in myeloma cell lines is shown after exposure to each HDACi. The binding of c-Myc to the Sp1 on the CD26 promoter via the proximal G-C box of KMS11, 27, and RPMI8226 after exposure to panobinostat (50 µmol/L) or RG2833 (0.5 µmol/L) was examined by ChIP and re-ChIP assay using antibodies for c-Myc and Sp1, followed by real-time qPCR using specific primers to amplify the CD26 promoter, including the proximal G-C box. The recovery of ChIP's DNAs was calculated as the percentages of IP/INPUT for each sample. In the absence of HDACi, c-Myc binds to the Sp1 on the proximal G-C box of the CD26 promoter in each myeloma cell line, whereas in the presence of HDACi, this binding was time-dependently detached. Bars represent the mean ± S.D. of three independent experiments. F, Schema of HDACi-regulated CD26 induction in myeloma cells.

Article Snippet: HDAC inhibitors; pan HDACi: panobinostat_50 μmol/L, vorinostat_1.0 μmol/L, isoform-selective HDACi: romidepsin (HDAC1i)_0.125 μmol/L, BG45 (HDAC1, 3i) 1.0 μmol/L, entinostat (HDAC1, 3i)_50 μmol/L, RG2833 (HDAC1, 3i)_0.5 μmol/L, nexturastat A (HDAC6i)_0.125 μmol/L, tubastatin A (HDAC6i)_25 μmol/L, ricolinostat (HDAC1, 3, 6i)_0.5 μmol/L were purchased from Selleck Chemical Co. LTD. for use as therapeutic agents.

Techniques: Modification, Control, Immunoprecipitation, Amplification, Quantitative RT-PCR, Expressing, Western Blot, Incubation, Binding Assay, Suspension, Staining, Agarose Gel Electrophoresis

Journal: Cancer Research

Article Title: HAND2 Assists MYCN Enhancer Invasion to Regulate a Noradrenergic Neuroblastoma Phenotype

doi: 10.1158/0008-5472.can-22-2042

Figure Lengend Snippet: Figure 6. Targeting both MYCN and CRC TFs. (A) Heatmaps show the percentage of cell viability after different dose of HDAC inhibitor LBH589 and Aurora A kinase inhibitor alisertib treatment in NB cell lines. Cell viability is measured by CellTiter-Glo Cell Viability Assay after 72 h drug treatment. (B) SynergyFinder online tool is used for bliss synergistic analysis to evaluate the synergistic effect of the combination treatment in MYCN-amplified cell lines shown in (A). (C) IncuCyte cell confluence assays show the synergistic effect of the alisertib (Ali) + LBH589 (LBH) treatment on cell proliferation (% confluency) over time. The red arrow is the time point of adding compounds. (D) Western blot analysis shows the protein levels of MYCN and CRC TFs in NB cells treated with LBH589 (LBH, 7.5 nM for IMR32, 7.5 nM for IMR5 and 5 nM for KCNR ), alisertib (Ali, 2 nM for IMR32, 5 nM for IMR5 and 5 nM for KCNR) alone or in combination for 24 h and 48 h. (E) Schematic diagram to show the strategy of drug treatment in orthotopic IMR5-GFP-Luc implanted xenografts. IVIS: in vivo imaging system. (F) Tumor weight measurement shows a significant decrease of the tumor weight of the drug treatment groups compared to the tumor weight of the vehicle treatment group. The p-value indicated is calculated in one-way ANOVA.

Article Snippet: A uthor M anuscript A uthor M anuscript A uthor M anuscript A uthor M anuscript Monitoring of synergistic effects of drug combinations The therapeutic effect of Aurora kinase A inhibitor (AURKAi) alisertib (MedChem Express, HY-10971) and HDACs inhibitor (HDACi) LBH589 (Medchem express, HY-10224) in NB cell lines was determined in a checkerboard fashion.

Techniques: Viability Assay, Amplification, Western Blot, In Vivo Imaging