Journal: bioRxiv

Article Title: PDS5A and TOP2B cooperate for chromatin recruitment via CTCF

doi: 10.64898/2026.04.02.715958

Figure Lengend Snippet: (A) Correlation of TOP2B versus PDS5A expression by immunohistochemistry in human gliomas. All tumor grades in adults are shown including low-grade gliomas (grade 2 in yellow, grade 3 in orange) and GBM (grade 4 in red). Light yellow region indicates malignant meningioma. The correlation is determined by Pearson coefficient r=0.6054, p-value < 0.0001 in n=208 samples that consist of meningioma, astrocytoma and glioblastoma. Brain tumor tissue array was stained with TOP2B or PDS5A antibodies. Visiopharm software was used to detect DAB chromogen (3,3’-diaminobenzidine) and count the positive cells as percentage. (B) Heatmap of the percentage of positive cells in the brain tumor tissue array shown in panel ( A ), indicating the correlative expression of TOP2B and PDS5A detected by immunohistochemistry. Brain tumor tissue array was stained with TOP2B or PDS5A antibodies. Visiopharm software was used for analysis. (C) Immunohistochemistry of the representative brain or tumor specimens showing correlative levels of TOP2B and PDS5A in normal brain, low-grade glioma (grade 2 and 3), and GBM (grade 4). (D) Correlation of TOP2B versus PDS5A gene expression by RNA-seq in human gliomas. All tumor grades in adult patient samples including low-grade gliomas and GBM are from CGGA GlioVis portal. The correlation is determined by Pearson coefficient r=0.72, p-value < 0.001 in n=1019 samples. (E) Box plot of PDS5A expression by RNA-seq in human gliomas. All tumor grades in adults including low-grade gliomas and GBM are from GlioVis portal (n=1019 samples). (F) Correlation of TOP2B versus PDS5A protein abundance determined by Pearson coefficient r=0.66, p-value=6.91e-14. The datasets of adult patient samples across all tumor grades are from cBioportal. (G) Expression of TOP2B, PDS5A, CTCF, RAD21, GAPDH, and histone H3 in glioma cell lines (U87MG, BT142, and A172), and Human Embryonic Kidney cell line (HEK293) by WB. GAPDH and H3 served as loading controls. IB: Immunoblot.

Article Snippet: The identity of the BT142 IDH1 R132H cell line from ATCC (ACS-1018) was confirmed by short tandem repeat (STR) and the cells were cultured with NeuroCult NS-A Proliferation kit (Catalog No. 5751, Stem Cell Technologies), 20 ng/mL recombinant human Epidermal Growth Factor (EGF, Catalog No. 100-15, PeproTech), 100 ng/mL recombinant human Platelet-Derived Growth Factor-AA (PDGF-AA, Catalog No. 100-13A, PeproTech), 20 ng/mL recombinant human Fibroblast Growth Factor (R&D Systems, Catalog No. 233-FB), 2 μg/mL heparan sulfate (Catalog No. H3149, Sigma), and antibiotics (1X) .

Techniques: Expressing, Immunohistochemistry, Staining, Software, Gene Expression, RNA Sequencing, Quantitative Proteomics, Western Blot

Journal: bioRxiv

Article Title: PDS5A and TOP2B cooperate for chromatin recruitment via CTCF

doi: 10.64898/2026.04.02.715958

Figure Lengend Snippet: (A) Heat maps showing CTCF, PDS5A, and TOP2B ChIP-seq read densities at CTCF-bound chromatin sites in BT142 cells under DMSO versus ETO treatment conditions. ChIP-seq data is from one representative of two biological replicates for CTCF and TOP2B, and one replicate for PDS5A. One replicate of TOP2B ChIP-seq has been re-analyzed . (B-C) Venn diagrams showing CTCF, PDS5A, and TOP2B binding in BT142 under DMSO versus ETO treatment conditions. (D) Average density profile plots showing CTCF (left panel), PDS5A (middle panel), and TOP2B (right panel) ChIP-seq read densities under conditions of DMSO versus ETO treatment at CTCF-bound sites in BT142 cells. (E) Heat maps showing CTCF, PDS5A, and TOP2B ChIP-seq read densities at TOP2B-bound sites in BT142 cells under DMSO versus ETO treatment conditions. (F-G) Normalized ChIP-seq densities for CTCF, PDS5A, and TOP2B at PDGFRA ( F ) and MYC ( G ) loci under DMSO versus ETO treatment conditions in BT142 cells. These loci represent promoters ( F ) and gene bodies/intronic sites ( G ), respectively. (H) Motif analysis of TOP2B-binding regions in BT142 cells. Motifs associated with TOP2B peaks were identified by de novo MEME motif analysis, and the corresponding top matches ( i.e. CTCF, MAZ, PATZ1, and VEZF1) are detailed in fig. S9 by Tomtom motif comparison tool. Motif search in MEME was performed de novo until 1000 sites were reached, and the corresponding e-values are depicted under the motifs.

Article Snippet: The identity of the BT142 IDH1 R132H cell line from ATCC (ACS-1018) was confirmed by short tandem repeat (STR) and the cells were cultured with NeuroCult NS-A Proliferation kit (Catalog No. 5751, Stem Cell Technologies), 20 ng/mL recombinant human Epidermal Growth Factor (EGF, Catalog No. 100-15, PeproTech), 100 ng/mL recombinant human Platelet-Derived Growth Factor-AA (PDGF-AA, Catalog No. 100-13A, PeproTech), 20 ng/mL recombinant human Fibroblast Growth Factor (R&D Systems, Catalog No. 233-FB), 2 μg/mL heparan sulfate (Catalog No. H3149, Sigma), and antibiotics (1X) .

Techniques: ChIP-sequencing, Binding Assay, Comparison

Journal: bioRxiv

Article Title: PDS5A and TOP2B cooperate for chromatin recruitment via CTCF

doi: 10.64898/2026.04.02.715958

Figure Lengend Snippet: (A) BT142 cells were transfected with three different Tet-ON shPDS5A for inducible PDS5A KD as determined by WB of PDS5A, TOP2B, PDS5B, CTCF, and histone H3 (n=2). H3 serves as a loading control. IB: Immunoblot. (B) Venn diagram indicating the overlap of CTCF (black), PDS5A (orange), and TOP2B (blue) ChIP-seq binding sites in BT142 cells. (C) Heat maps showing PDS5A, TOP2B, and CTCF ChIP-seq read densities at PDS5A-TOP2B-CTCF common binding sites (n=19878) in BT142 cells in WT versus PDS5A KD conditions. (D-F) Average density profile plots showing PDS5A ( D ), TOP2B ( E ), and CTCF ( F ) ChIP-seq read densities at PDS5A-bound sites in BT142 cells in WT versus PDS5A KD conditions. (G) Normalized ChIP-seq read densities of CTCF, PDS5A, and TOP2B in WT versus PDS5A KD conditions at the IDH2 locus, which is associated with tumor progression in BT142 cells. ChIP-seq data is from one representative biological replicate for each condition. (H) RNA-seq MA plot showing the impact of treatment with TOP2B inhibitor (ICRF-193, 15 μM, 6 hr) in BT142 glioma cells (TOP2B ICRF-193 inhibition re-analyzed data ). DMSO treatment serves as control. Gray dots indicate all genes, and black dots indicate DEGs across the conditions compared. RNA-seq data for control (DMSO) and ICRF-193 treated conditions are from five and seven biological replicates, respectively. (I) RNA-seq MA plot of BT142 cells carrying inducible Tet-ON shPDS5A under WT (-Dox) vs PDS5A KD (+Dox) conditions from four biological replicates. Gray dots indicate all genes and black dots indicate DEGs across the conditions compared. (J) The cell growth curves of WT vs PDS5A KD BT142 cells indicating that PDS5A KD did not affect cell growth. BT142 cells carrying Tet-ON shPDS5A, either unexpressed [WT (-Dox)] or expressed [PDS5A KD (+Dox)], were gauged for cell growth for 10 days in three biological replicates and showed no statistical differences. (K-L) PDS5A mediates sensitivity to TOP2-drugs. Shown are the surviving fraction of BT142 cells carrying Tet-ON shPDS5A, either unexpressed [WT (-Dox)] or expressed [PDS5A KD (+Dox)], and treated for 72 hr with TOP2-drugs in four biological replicates: ( K ) Etoposide (0 and 0.25 μM) or ( L ) Doxorubicin (0, 2.5, 5, and 7.5 μM). Two-sided Student’s t-test without multiple test correction was used (***p < 0.001, **p < 0.01).

Article Snippet: The identity of the BT142 IDH1 R132H cell line from ATCC (ACS-1018) was confirmed by short tandem repeat (STR) and the cells were cultured with NeuroCult NS-A Proliferation kit (Catalog No. 5751, Stem Cell Technologies), 20 ng/mL recombinant human Epidermal Growth Factor (EGF, Catalog No. 100-15, PeproTech), 100 ng/mL recombinant human Platelet-Derived Growth Factor-AA (PDGF-AA, Catalog No. 100-13A, PeproTech), 20 ng/mL recombinant human Fibroblast Growth Factor (R&D Systems, Catalog No. 233-FB), 2 μg/mL heparan sulfate (Catalog No. H3149, Sigma), and antibiotics (1X) .

Techniques: Transfection, Control, Western Blot, ChIP-sequencing, Binding Assay, RNA Sequencing, Inhibition

Journal: bioRxiv

Article Title: PDS5A and TOP2B cooperate for chromatin recruitment via CTCF

doi: 10.64898/2026.04.02.715958

Figure Lengend Snippet: . (A) Heat maps showing PDS5A, TOP2B, and CTCF ChIP-seq read densities at PDS5A bound sites in BT142 cells in WT versus PDS5A KD conditions. (B-D) Average density profile plots showing PDS5A ( B ), TOP2B ( C ), and CTCF ( D ) ChIP-seq read densities at PDS5A bound sites in BT142 cells in WT versus PDS5A KD conditions. (E-G) Venn diagrams indicating the PDS5A ( E ), CTCF ( F ) and TOP2B ( G ) ChIP-seq binding sites in WT versus PDS5A KD conditions in BT142 cells. (H-I) Venn diagrams indicating the overlap of PDS5A (orange) and TOP2B ChIP-seq binding sites (blue) in WT condition ( H ), and PDS5A KD condition ( I ) in BT142 cells. ChIP-seq data is from one representative biological replicate for each condition. (J-K) TOP2B and PDS5A interaction in BT142 cells under WT ( J ) versus PDS5A KD ( K ) conditions using chromatin fractions isolated from cells treated with DMSO and ETO, respectively, as determined by IP followed by WB (n=2).

Article Snippet: The identity of the BT142 IDH1 R132H cell line from ATCC (ACS-1018) was confirmed by short tandem repeat (STR) and the cells were cultured with NeuroCult NS-A Proliferation kit (Catalog No. 5751, Stem Cell Technologies), 20 ng/mL recombinant human Epidermal Growth Factor (EGF, Catalog No. 100-15, PeproTech), 100 ng/mL recombinant human Platelet-Derived Growth Factor-AA (PDGF-AA, Catalog No. 100-13A, PeproTech), 20 ng/mL recombinant human Fibroblast Growth Factor (R&D Systems, Catalog No. 233-FB), 2 μg/mL heparan sulfate (Catalog No. H3149, Sigma), and antibiotics (1X) .

Techniques: ChIP-sequencing, Binding Assay, Isolation

Journal: bioRxiv

Article Title: PDS5A and TOP2B cooperate for chromatin recruitment via CTCF

doi: 10.64898/2026.04.02.715958

Figure Lengend Snippet: . (A) Gene ontology analysis of DEGs upon TOP2B inhibition (ICRF-193 treatment) using RNA-seq in BT142 glioma cells. RNA-seq data for control (DMSO) and ICRF-193 treated conditions are from five and seven biological replicates, respectively . (B) Gene ontology analysis of DEGs upon PDS5A KD using RNA-seq in BT142 glioma cells from four biological replicates.

Article Snippet: The identity of the BT142 IDH1 R132H cell line from ATCC (ACS-1018) was confirmed by short tandem repeat (STR) and the cells were cultured with NeuroCult NS-A Proliferation kit (Catalog No. 5751, Stem Cell Technologies), 20 ng/mL recombinant human Epidermal Growth Factor (EGF, Catalog No. 100-15, PeproTech), 100 ng/mL recombinant human Platelet-Derived Growth Factor-AA (PDGF-AA, Catalog No. 100-13A, PeproTech), 20 ng/mL recombinant human Fibroblast Growth Factor (R&D Systems, Catalog No. 233-FB), 2 μg/mL heparan sulfate (Catalog No. H3149, Sigma), and antibiotics (1X) .

Techniques: Inhibition, RNA Sequencing, Control

Journal: Inflammation Research

Article Title: IDH1 Mutant Glioma Favors Group 3 Innate Lymphoid Cells and Is Resistant to Immune Checkpoint Expression

doi: 10.1007/s00011-026-02223-8

Figure Lengend Snippet: The frequency of ILC1 is decreased, whereas ILC3 is increased, in IDH1-mutant human glioma tissues compared with the IDH1-wild-type group. A Representative flow-cytometry gating strategies for ILCs in human tonsil (control) tissue, IDH1-wildtype, and IDH1-mutant glioma tissue (FSC-A = forward-scatter area; SSC-A = side-scatter area). B Comparison of total ILC percentages in tonsil controls (n = 11), IDH1-wild-type (n = 9), and IDH1-mutant (n = 3) glioma tissues (top center); comparison of ILC1 percentages (bottom left), ILC2 percentages (bottom center), and ILC3 percentages (bottom right) across the same groups. C Comparison of mean fluorescence intensity (MFI) of KLRG1 (top left), PD-1 (top right), and CTLA-4 (bottom center) expression on ILC1s in tonsil control, IDH1-wild-type, and IDH1-mutant groups. D Comparison of MFI of KLRG1 (top left), PD-1 (top right), and CTLA-4 (bottom center) on ILC2s. E Comparison of MFI of KLRG1 (top left), PD-1 (top right), and CTLA-4 (bottom center) on ILC3s. Error bars represent ± SD. Statistical significance was determined by one-way ANOVA (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001)

Article Snippet: Human glioma U87-MG and its isogenic IDH1 (R132H)-mutant U87-MG (#HTB14IG) cell lines were obtained from the American Type Culture Collection (ATCC).

Techniques: Mutagenesis, Flow Cytometry, Control, Comparison, Fluorescence, Expressing

Journal: Inflammation Research

Article Title: IDH1 Mutant Glioma Favors Group 3 Innate Lymphoid Cells and Is Resistant to Immune Checkpoint Expression

doi: 10.1007/s00011-026-02223-8

Figure Lengend Snippet: The frequency of ILC3 is increased in the peripheral blood of IDH1-mutant glioma patients compared with the IDH1-wild-type group. A Representative flow-cytometry gating strategies for ILCs in peripheral blood of healthy controls, IDH1-wild-type glioma patients, and IDH1-mutant glioma patients (FSC-A = forward-scatter area; SSC-A = side-scatter area). B Comparison of total ILC percentages in blood samples from healthy controls (HC, n = 20), IDH1-wild-type (n = 12), and IDH1-mutant (n = 7) groups (top center); comparison of ILC1 (bottom left), ILC2 (bottom center), and ILC3 (bottom right) frequencies across the same groups. C Comparison of MFI of KLRG1 (top left), PD-1 (top right), and CTLA-4 (bottom center) on ILC1s in HC, IDH1-wild-type, and IDH1-mutant groups. D Comparison of MFI of KLRG1 (top left), PD-1 (top right), and CTLA-4 (bottom center) on ILC2s. E Comparison of MFI of KLRG1 (top left), PD-1 (top right), and CTLA-4 (bottom center) on ILC3s. Error bars represent ± SD. Statistical significance was determined by one-way ANOVA (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001)

Article Snippet: Human glioma U87-MG and its isogenic IDH1 (R132H)-mutant U87-MG (#HTB14IG) cell lines were obtained from the American Type Culture Collection (ATCC).

Techniques: Mutagenesis, Flow Cytometry, Comparison

Journal: Inflammation Research

Article Title: IDH1 Mutant Glioma Favors Group 3 Innate Lymphoid Cells and Is Resistant to Immune Checkpoint Expression

doi: 10.1007/s00011-026-02223-8

Figure Lengend Snippet: Co-culture of tonsil-derived ILCs with U87-MG significantly increased surface PD-1, KLRG1, and CTLA-4 compared with IDH1-mutant U87-MG. A – D Human tonsil-derived ILCs were cultured alone or co-cultured with U87-MG or IDH1-mutant U87-MG glioma cell lines, either with cytokine supplementation (recombinant human IL-2 [5 ng/mL], IL-7 [50 ng/mL], IL-12 [50 ng/mL], IL-1β [50 ng/mL], IL-23 [50 ng/mL]) or without cytokines for four days. Data points represent two independent experiments (ILC n = 3; U87-MG + ILC n = 6; IDH1-mutant U87-MG + ILC n = 6; technical replicates) A Representative flow-cytometry gating strategy for human tonsil-derived ILCs. B Flow-cytometry contour plots showing CTLA-4, KLRG1, and PD-1 surface expression percentages on ILCs. C Quantification of CTLA-4, KLRG1, and PD-1 expression percentages on ILCs. D Mean fluorescence intensity (MFI) of CTLA-4, KLRG1, and PD-1 expression on ILCs. E – F ILCs were cultured alone or exposed to glioma-conditioned medium (GCM) from U87-MG or IDH1-mutant U87-MG cell lines under the same cytokine conditions for four days. Data points represent two independent experiments (each with three technical replicates). E Percentages of CTLA-4, KLRG1, and PD-1–expressing ILCs following GCM exposure. F Corresponding MFI of CTLA-4, KLRG1, and PD-1 expression on ILCs. G – H Proliferation of CFSE-labeled tonsil ILCs co-cultured with U87-MG or IDH1-mutant U87-MG cells was analyzed after four days. G Representative flow-cytometry plots of CFSE dilution. H Quantification of proliferating CFSE-labeled ILCs from three independent experiments (each with four technical replicates). Error bars represent ± SD. Statistical analyses were performed using one-way ANOVA (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001)

Article Snippet: Human glioma U87-MG and its isogenic IDH1 (R132H)-mutant U87-MG (#HTB14IG) cell lines were obtained from the American Type Culture Collection (ATCC).

Techniques: Co-Culture Assay, Derivative Assay, Mutagenesis, Cell Culture, Recombinant, Flow Cytometry, Expressing, Fluorescence, Labeling

Journal: Inflammation Research

Article Title: IDH1 Mutant Glioma Favors Group 3 Innate Lymphoid Cells and Is Resistant to Immune Checkpoint Expression

doi: 10.1007/s00011-026-02223-8

Figure Lengend Snippet: IL-17 and IFN-γ production is increased in tonsil-derived ILCs exposed to glioma-conditioned medium (GCM) from U87-MG and IDH1-mutant U87-MG cell lines. A – E Tonsil-derived ILCs were cultured alone or with glioma-conditioned medium (GCM) obtained from U87-MG or IDH1-mutant U87-MG cell lines for four days, with or without cytokine supplementation (recombinant human IL-2 [5 ng/mL], IL-7 [50 ng/mL], IL-12 [50 ng/mL], IL-1β [50 ng/mL], IL-23 [50 ng/mL]). Golgi Stop was added during the final nine hours of incubation. A Percentages (left) and mean fluorescence intensity (MFI; right) of TNF-α–producing ILCs. B Percentages (left) and MFI (right) of IL-2–producing ILCs. C Percentages (left) and MFI (right) of IL-17–producing ILCs. D Percentages (left) and MFI (right) of IFN-γ–producing ILCs. E Percentages (left) and MFI (right) of GM-CSF–producing ILCs. Data represents two independent experiments (each with three technical replicates). Error bars indicate ± SEM. Statistical significance was determined by one-way ANOVA (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001)

Article Snippet: Human glioma U87-MG and its isogenic IDH1 (R132H)-mutant U87-MG (#HTB14IG) cell lines were obtained from the American Type Culture Collection (ATCC).

Techniques: Derivative Assay, Mutagenesis, Cell Culture, Recombinant, Incubation, Fluorescence

Journal: Inflammation Research

Article Title: IDH1 Mutant Glioma Favors Group 3 Innate Lymphoid Cells and Is Resistant to Immune Checkpoint Expression

doi: 10.1007/s00011-026-02223-8

Figure Lengend Snippet: D-2-HG levels are comparable in-patient plasma but elevated in glioma-conditioned medium (GCM) from IDH1-mutant U87-MG + ILC co-cultures compared with U87-MG + ILC. A Quantification of D-2-HG (OD₄₅₀) in plasma samples from healthy controls (HC, n = 12), IDH1-wild-type glioma patients (n = 12), and IDH1-mutant glioma patients (n = 7). B Quantification of D-2-HG (OD₄₅₀) in glioma-conditioned medium (GCM) collected from cultures of ILCs alone, U87-MG, IDH1-mutant U87-MG cell lines, and tonsil ILCs co-cultured with U87-MG or IDH1-mutant U87-MG cells under cytokine-supplemented (recombinant human IL-2 [5 ng/mL], IL-7 [50 ng/mL], IL-12 [50 ng/mL], IL-1β [50 ng/mL], IL-23 [50 ng/mL]) and cytokine-free conditions for four days. Data points represent two independent experiments (ILC n = 4, U87-MG + ILC n = 2, IDH1-mutant U87-MG + ILC n = 2 technical replicates). Error bars indicate ± SEM. (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001)

Article Snippet: Human glioma U87-MG and its isogenic IDH1 (R132H)-mutant U87-MG (#HTB14IG) cell lines were obtained from the American Type Culture Collection (ATCC).

Techniques: Clinical Proteomics, Mutagenesis, Cell Culture, Recombinant