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: 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: MYC/MAX inhibitors in combination with anti-transcription γPNA1 Cell viability of (A) U2932 and (B) Raji cells treated with increasing doses of MYC/MAX inhibitors (Myci975, EN4, 10058-F4, and sAJM589) alone and in combination with γPNA1 and ScR-γPNA2 (8 μM) for 72 h. Results are presented as mean ± SEM. The IC 50 (95% CI) values of MYC/MAX inhibitors alone and combination treatment of MYC/MAX with γPNA1 in (C) U2932 and (D) Raji cells. (E) Cell viability of γPNA1-treated U2932 and Raji cells at 8 μM concentration. Western blot analysis representing the change in c-MYC protein 72 h after treatment with γPNA1 and ScR-γPNA2 in combination with (F) Myci975, (G) EN4, (H) 10058-F4, and (I) sAJM589. ∗∗(F–I) Cyclophilin B was used as an endogenous control, and the same blots are presented in A–S7D. c-MYC, EZH2, and cyclophilin B were probed from the same blot. Results are presented as mean ± SEM, p value for 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: Concentration Assay, 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: Efficacy of small molecules targeting other pathways with anti-transcription γPNA1 (A) Cell viability of U2932 and Raji cells treated with increasing doses JQ1 (BRD4 inhibitor) alone and with γPNA1 and ScR-γPNA2 for 48 h. (B) Cell viability of U2932 and Raji cells treated with increasing doses of sapnisertid (mTOR inhibitor) alone and with γPNA1 and ScR-γPNA2 for 48 h. (C) Cell viability of MDA-MB-231 cells treated with increasing doses of dinaciclib (CDK inhibitor) alone and with γPNA1 and ScR-γPNA2 for 48 h. (D) The IC 50 (95% CI) values of small molecule inhibitors alone and in combination with γPNA1. (A–C) Results are presented as mean ± SEM.

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:

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: Scientific Reports

Article Title: A novel human acute myeloid leukemia cell line SDEY-AML1 with KMT2A: MLLT3, IKZF1: EVX1 fusions exhibits high tumorigenicity in NSG mice

doi: 10.1038/s41598-026-39353-2

Figure Lengend Snippet: Drug sensitivity experiment of SDEY-AML1 cell. ( A )Testing the sensitivity of Chidamide, LBH589, S63845 , Olverembatinib, Cladribine, Afatinib to SDEY-AML1.

Article Snippet: Chidamide, LBH589, S63845 , Olverembatinib, Cladribine, Afatinib were purchased by MedChemExpress.

Techniques:

Journal: Science Advances

Article Title: DLX2 acts as a pioneer factor and drives Msx1 + ectomesenchyme formation from embryonic stem cells

doi: 10.1126/sciadv.aea0685

Figure Lengend Snippet: ( A ) Schematic of FLAG-tagged DLX2-binding partner identification and validation. m/z , mass/charge ratio; WB, Western blot. ( B ) Liquid chromatography–mass spectrometry (LC-MS) heatmap of day-2 Dlx2 -OE (FLAG +Dox) EBs. Proteins were labeled by UniProt entry names, with FC indicated by color intensity (red). ( C ) Immunostaining of Dlx2 -OE cells. DAPI for nuclear staining. Scale bars, 50 μm (EBs) and 10 μm (single cells). ( D and E ) Co-IP of DLX2 and LAP2 isoforms in day-2 Dlx2 -OE EBs. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as the loading control. ( F ) Schematic of DLX2 truncation mutants: WT and four mutants with deletions in the N-terminal (ΔN), C-terminal (ΔC), homeodomain (ΔHD), or 38 amino acids in the homeodomain, retaining the NLS. aa, amino acids. ( G ) Co-IP of LAP2α, LAP2β, and FLAG-DLX2 mutants in day-2 EBs using anti-FLAG antibody. ( H ) Predicted DLX2-LAP2α protein complex structure. ( I ) Flow cytometry quantification of Msx1 + cells in day-8 EBs. Data shown as mean ± SD from three independent experiments. Statistics: one-way ANOVA with Dunnett’s post hoc test. ( J ) RT-qPCR of ectomesenchymal marker genes in day-8 EBs. Data shown as mean ± SD from three independent experiments. Statistics: one-way ANOVA with Dunnett’s post hoc test. ( K ) RT-qPCR of Tmpo KD efficiency in day-8 Dlx2 -OE EBs, with two shRNA mix delivered into cells. Data shown as mean ± SD from three independent experiments. Statistics: two-tailed unpaired Student’s t test. ( L ) Flow cytometry quantification of Msx1 + cells in day-8 Dlx2 -OE EBs with Tmpo KD. Data shown as mean ± SD from four independent experiments. Statistics: one-way ANOVA with Dunnett’s post hoc test. ( M ) RT-qPCR of ectomesenchymal marker genes in day-8 Dlx2 -OE EBs with Tmpo KD. Data shown as mean ± SD from three independent experiments. Statistics: two-tailed unpaired Student’s t test. ** P < 0.01, *** P < 0.001, and **** P < 0.0001. n.s., not significant.

Article Snippet: After preclearing with magnetic beads, the lysates were incubated overnight at 4°C with anti-FLAG, anti-pan LAP2, anti-H3K27ac, or control IgG antibodies, followed by a 4-hour incubation with Protein G or Protein A/G magnetic beads (Invitrogen, catalog no. 10004D; MedChemExpress, catalog no. HY-K0202).

Techniques: Binding Assay, Biomarker Discovery, Western Blot, Liquid Chromatography, Mass Spectrometry, Liquid Chromatography with Mass Spectroscopy, Labeling, Immunostaining, Staining, Co-Immunoprecipitation Assay, Control, Flow Cytometry, Quantitative RT-PCR, Marker, shRNA, Two Tailed Test

Journal: Science Advances

Article Title: DLX2 acts as a pioneer factor and drives Msx1 + ectomesenchyme formation from embryonic stem cells

doi: 10.1126/sciadv.aea0685

Figure Lengend Snippet: ( A ) Schematic of integrative analysis combining RNA-seq and CUT&Tag data. ( B ) DEG analysis between Dlx2 - and GFP-OE control cells. DEGs were identified with DESeq2 (log 2 FC ≥ 2 and P < 0.05) and included multiple ectomesenchymal markers. ( C ) GO analysis of up-regulated genes in Dlx2 -OE cells. The top 20 GO terms displayed in the dot plot. ( D ) Heatmap of a genome-wide overview of DLX2-binding sites identified by CUT&Tag. Each row represented a 3-kb window centered on the peak midpoint. ( E ) Pie chart showing the genomic distribution of DLX2-binding sites. 3′UTR, 3′ untranslated region. ( F ) HOMER analysis of known DNA sequence motifs enriched at significant DLX2-binding peaks. ( G ) Venn diagram showing overlap of promoter-proximal DLX2- and LAP2α-binding targets with DEGs from RNA-seq. ( H ) Representative Integrative Genomics Viewer tracks of DLX2 and LAP2α signals at promoter regions of target genes. ( I ) KD efficiency of target genes assessed by RT-qPCR in day-8 Dlx2 -OE EBs, with three shRNA mix delivered via the PB transposon system. Error bars represented data as mean ± SD from three independent experiments. Statistics: two-tailed unpaired Student’s t test. ( J and K ) Flow cytometry analysis of Msx1 + cell proportions in day-8 EBs after KD of target genes, showing representative plots (J) and quantitative FC in Msx1 low and Msx1 high populations relative to scramble control (K). Error bars represented data as mean ± SD from three independent experiments. Statistics: one-way ANOVA with Dunnett’s post hoc test. ( L ) RT-qPCR of ectomesenchymal marker gene expression in day-8 Dlx2 -OE EBs with Dlx6 , Hmga2 , Msx2 , or Gata4 KD. Heatmap showed relative expression levels compared to scramble control: blue (<1), and pink (>1). Data represented mean ± SD from three independent experiments. * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001. n.s., not significant.

Article Snippet: After preclearing with magnetic beads, the lysates were incubated overnight at 4°C with anti-FLAG, anti-pan LAP2, anti-H3K27ac, or control IgG antibodies, followed by a 4-hour incubation with Protein G or Protein A/G magnetic beads (Invitrogen, catalog no. 10004D; MedChemExpress, catalog no. HY-K0202).

Techniques: RNA Sequencing, Control, Genome Wide, Binding Assay, Sequencing, Quantitative RT-PCR, shRNA, Two Tailed Test, Flow Cytometry, Marker, Gene Expression, Expressing

Journal: Science Advances

Article Title: DLX2 acts as a pioneer factor and drives Msx1 + ectomesenchyme formation from embryonic stem cells

doi: 10.1126/sciadv.aea0685

Figure Lengend Snippet: Through the formation of a DLX2-LAP2α-nucleosome complex that remodeled chromatin, DLX2 drove ectomesenchymal specification in mESCs, generating Msx1 + progenitors with craniofacial regeneration potential.

Article Snippet: After preclearing with magnetic beads, the lysates were incubated overnight at 4°C with anti-FLAG, anti-pan LAP2, anti-H3K27ac, or control IgG antibodies, followed by a 4-hour incubation with Protein G or Protein A/G magnetic beads (Invitrogen, catalog no. 10004D; MedChemExpress, catalog no. HY-K0202).

Techniques: