su dhl 10 acc 576  (DSMZ)

Journal: ImmunoTargets and Therapy

Article Title: The Dual Mechanism of Action of CO-005 Overcomes CD20 Resistance in Diffuse Large B-Cell Lymphoma

doi: 10.2147/ITT.S572396

Figure Lengend Snippet: CO-005 triggers receptor capping and intracellular signaling events associated with type II anti-CD20 antibodies. (A) NU-DUL-1, (B) ULA, (C) , SU-DHL-10 and (D) OCI-LY19 cells treated with 1 µg/mL CO-005, rituximab (RTX), or obinutuzumab (OBZ) for 3 or 24 hours exhibit distinct patterns of CD47 and CD20 clustering. Receptor distribution was visualized by confocal microscopy following immunofluorescence staining. Scale bar 10 µm. Cellular stress profiling was measured as following (E) Intracellular calcium flux in lymphoma cells treated with 1 µg/mL of the indicated antibodies or isotype control for 3 hours, measured by Fluo-4 AM and flow cytometry. (F) Reactive oxygen species (ROS) levels following antibody treatment (1 µg/mL, 3 hours), assessed by CellROX Green and flow cytometry. (G) Mitochondrial membrane potential (MMP) after CO-005 treatment (0.1 or 1 µg/mL) for 3 hours, determined by TMRM staining. CCCP served as a positive control for mitochondrial depolarization. (H) Effect of actin polymerization inhibition on CO-005 – induced cell death following cytochalasin pretreatment (20 µM, 15 min), measured by Annexin V and 7-AAD staining. Data represent the mean ± SD. Significance was determined by ANOVA analysis *, P < 0.01 compared to the sample treated with CO-005 or as indicated in the figure; ns indicates not significant. All experiments were performed independently three times (n = 3). MFI, mean fluorescence intensity.

Article Snippet: ULA (ACC-267), Riva (ACC-585), OCI-LY18 (ACC-699), OCI-LY19 (ACC-528), and SU-DHL-10 (ACC-576) were obtained from the Leibniz Institute DSMZ–German Collection of Microorganisms and Cell Cultures (DSMZ).

Techniques: Confocal Microscopy, Immunofluorescence, Staining, Control, Flow Cytometry, Membrane, Positive Control, Inhibition, Fluorescence

Journal: ImmunoTargets and Therapy

Article Title: The Dual Mechanism of Action of CO-005 Overcomes CD20 Resistance in Diffuse Large B-Cell Lymphoma

doi: 10.2147/ITT.S572396

Figure Lengend Snippet: CO-005 shifts the phagocytic balance by enhancing “eat me” and promoting macrophage- mediated clearance. ( A ) Surface Calreticulin exposure was assessed in a panel of lymphoma cell lines (NU-DUL-1, OCI-LY18, ULA, SU-DHL-6, SU-DHL-10 and Daudi) following treatment with CO-005, Rituximab (RTX), or Obinutuzumab (OBZ) for 3 hours; Doxorubicin served as a positive control. Calreticulin expression is shown relative to isotype control. (B) Phagocytosis activity of CO-005 or RTX was measured using CellTrace-labeled NU-DUL-1 and OCI-LY19 cells co-cultured with DiO-labeled RAW264.7 macrophages for 2 hours. Phagocytosed cells were quantified as %CellTrace+DiO+ by flow cytometry. Data represent the mean ± SD. Significance was determined by ANOVA analysis *, P < 0.01 compared to the sample treated with CO-005; ns indicates not significant. All experiments were performed independently three times (n = 3).

Article Snippet: ULA (ACC-267), Riva (ACC-585), OCI-LY18 (ACC-699), OCI-LY19 (ACC-528), and SU-DHL-10 (ACC-576) were obtained from the Leibniz Institute DSMZ–German Collection of Microorganisms and Cell Cultures (DSMZ).

Techniques: Positive Control, Expressing, Control, Activity Assay, Labeling, Cell Culture, Flow Cytometry

Journal: bioRxiv

Article Title: OSTM1 is a ubiquitin E3 ligase that suppresses B-cell malignancy by activating the cAMP/PKA/CREB pathway

doi: 10.64898/2026.01.23.701155

Figure Lengend Snippet: (A) Venn diagram showing overlap of B cell-specific CREB1 and CREBBP ChIP-seq targets. All targets identified in each dataset were used to determine overlap, and the top 1,500 common targets were selected for downstream analyses. (B) RNA expression data from the TCGA MDACC B-cell malignancies cohort. Heatmaps showing correlations between OSTM1, CREB1, or PDE3B mRNA levels and the top 1,500 CREB1/CREBBP co-occupied target genes identified in ( A ). (C) Venn diagrams showing overlap among genes significantly correlated with OSTM1, CREB1, or PDE3B within the top 1,500 CREB1/CREBBP targets. As OSTM1 negatively regulates PDE3B, genes positively correlated with OSTM1 and CREB1 expression and negatively correlated with PDE3B expression showed substantial overlap (78.2%). Conversely, genes negatively correlated with OSTM1 and CREB1 expression and positively correlated with PDE3B expression also overlapped significantly (66.8%). (D) CRISPR gene-effect scores from the CCLE database for OSTM1 and PDE3B across BCL lines, showing generally negative effects upon PDE3B silencing and positive effects upon OSTM1 silencing. (E) Pde3b was silenced by sgRNA in sg Ostm1 clone #3 Ba/F3 cells. PDE3B protein levels were determined by IB, and sg Ostm1/Pde3b double knockout (DKO) Ba/F3 cells were selected for subsequent experiments. (F) sgControl, sg Ostm1 , and DKO Ba/F3 cells were cutured in the presense or absence of IL3. Pde3b silencing reversed IL3-independence in sg Ostm1 cells. (G) GFP-expressing sg Ostm1 or DKO Ba/F3 cells were transplanted into nude mice via i.p. injection. Mice were harvested at the endpoint of the sg Ostm1 cohort. Spleens and livers were photographed and weighed. P values were calculated using Student’s t-test. GFP-positive tumor cells were detected only in sg Ostm1 recipients, but not in DKO recipients. (H) qRT-PCR in sgControl and sg Ostm1 Ba/F3 cells showing that Ostm1 silencing reduced expression of PKA/CREB/CREBBP target genes. (I) sgCtrl, sg Ostm1 , and DKO Ba/F3 cells were probed for phospho-PKA substrates and phospho-CREB (Ser133). (J) OSTM1 was silenced using two independent sgRNAs in SU-DHL-5 cells. Left: qPCR validation of OSTM1 knockout using on-target primers. Right: IB showing stablization of PDE3B and downregulation of cAMP/PKA signaling upon OSTM1 deletion. (K) OSTM1 was silenced in ARH-77 cells. IB of two clones shows increased PDE3B protein levles and decreased cAMP/PKA signaling upon OSTM1 silencing. (L) PDE3B-His was stably expressed in SU-DHL-10 cell line, which suppressed cAMP/PKA signaling. (M) OSTM1-Flag or OSTM1Δ31-Flag was stably expressed in OPM2 and RPMI-8226 cells. IB shows that OSTM1Δ31, but not the full-length OSTM1, promoted PDE3B degradation and enhanced cAMP/PKA signaling. (N) IB of whole-spleen lysates from indicated age-matched mice collected at the endpoints of O +/- ;C -/- or DKO cohorts. Phosphorylation levels of CREB and PKA substrates were generally reduced in the O -/- , O +/- C -/- , and DKO mice. (O) Bulk RNA-seq of purified splenic B cells from the indicated genotypes (as in ). Genes up-or down-regulated in DKO versus C -/- mice were intersected with the CREB1/CREBBP ChIP-seq targets identified in (A) . (P) Spleens from two-month-old C -/- and DKO mice were harvested, and 10,000 cells per mouse were analyzed by scRNA-seq. UMAP plots of B-cell subpopulations are shown by genotype. Bar graphs show the relative proportions of B-cell subsets. (Q) Genes down-regulated in DKO vs C -/- B cells, as identified by both bulk RNA-seq of splenic B cells and scRNA-seq of follicular B cells, were intersected with CREB1/CREBBP targets. Eight genes were commonly identified across all 4 datasets.

Article Snippet: Human DLBCL cell lines SU-DHL-4 (CRL-2957), SU-DHL-5 (CRL-2958), and SU-DHL-10 (CRL-2963), Burkitt’s lymphoma cell lines Daudi and Ramos, as well as mantle cell lymphoma cell lines Z-138 and JeKo-1 were obtained from American Type Culture Collection (ATCC, Manassas, VA).

Techniques: ChIP-sequencing, RNA Expression, Expressing, CRISPR, Double Knockout, Injection, Quantitative RT-PCR, Biomarker Discovery, Knock-Out, Clone Assay, Stable Transfection, Phospho-proteomics, RNA Sequencing, Purification