Journal: Oncogene

Article Title: LAMP5 modulates IRF4 stability and nuclear transport: a critical mechanism in myeloma progression and therapy

doi: 10.1038/s41388-025-03513-x

Figure Lengend Snippet: A , B LAMP5 mRNA expression in myeloma and other indicated cancer cell lines or other tissues, according to the Cancer Cell Line Encyclopedia (CCLE) dataset. C LAMP5 mRNA levels in plasma cells from myeloma patients ( n = 559) compared to normal plasma cells from healthy donors ( n = 22) (GEO: GSE2658 and GSE5900 ) (left). LAMP5 mRNA levels in plasma cells from myeloma patients ( n = 73) compared to normal plasma cells from healthy donors ( n = 14) (GEO: GSE6477 ) (right). P value was determined by unpaired two-tailed t test. D Quantitative real-time PCR analysis shows LAMP5 expression in normal plasma cells (Healthy, n = 10), malignant plasma cells (Pt MM, n = 20), and human myeloma cell lines (MM cell lines, n = 6). GAPDH served as loading control. P values were determined using one-way ANOVA. E Representative two patients of UMAP plot of CD138 (left) and LAMP5 (right) expression in scRNA-seq data ( GSE161801 ). F Immunofluorescent staining of healthy of myeloma patients bone marrow samples with DAPI and antibodies against LAMP5 and CD138 ( n = 5). Scale bar, 10 μm. G Overall survival (OS) of patient’s myeloma cells with high LAMP5 (High, n = 200) and low LAMP5 (Low, n = 200) expression in MMRF CoMMpass study IA15.

Article Snippet: The CD138 antibody (#AF2780) was purchased from R&D Systems.

Techniques: Expressing, Clinical Proteomics, Two Tailed Test, Real-time Polymerase Chain Reaction, Control, Staining

Journal: Oncogene

Article Title: LAMP5 modulates IRF4 stability and nuclear transport: a critical mechanism in myeloma progression and therapy

doi: 10.1038/s41388-025-03513-x

Figure Lengend Snippet: A Analysis of immunoprecipitates of HA-tagged LAMP5 in IM-9 cells with mass spectrometry. The proteins identified are indicated. B Co-immunoprecipitation of LAMP5 with IRF4 in ARP-1 or IM-9 cells. C Co-immunoprecipitation of LAMP5 with IRF4 in HEK293T cells co-transfected with LAMP5 and IRF4 plasmids. D Pull-down of HA-LAMP5 with IRF4 in HEK293T cells. E Immunofluorescent staining of ARP-1 or IM-9 cells with DAPI and antibodies against LAMP5 and IRF4. Scale bar, 10 μm. F Immunofluorescent staining of myeloma patients bone marrow samples with DAPI and antibodies against CD138, LAMP5 and IRF4 ( n = 5). Scale bar, 10 μm. G Schematic of the truncations including ΔDBD and ΔIAD fragments. H Western blotting showing different truncations of IRF4 (ΔDBD and ΔIAD) in HEK293T cells. I Pull-down of LAMP5 with different truncations of IRF4 (ΔDBD and ΔIAD) in HEK293T cells. J – M The expression of IRF4 or c-MYC in myeloma cells with reduced (sh LAMP5 ) or increased ( LAMP5 ) LAMP5 expression. Nontargeted shRNA (sh Ctrl ) or control vector ( Vec ) served as control. N , O The expression of LDHA or KLF2 in myeloma cells with reduced (sh LAMP5 ) LAMP5 expression. Nontargeted shRNA (sh Ctrl ) served as control. P , Q Immunofluorescent staining of ARP-1 or IM-9 cells (sh Ctrl and sh LAMP5 ) with DAPI and antibody against LAMP5 and IRF4. Scale bar, 10 μm. R Immunofluorescent staining of tumor tissues in Fig. with DAPI and antibodies against LAMP5 and IRF4. Scale bar, 20 μm. S Correlation coefficient of the mRNA levels of LAMP5 and mRNA levels of IRF4 or c-MYC in myeloma patients ( n = 559). The correlations were evaluated using Pearson coefficient. r, correlation coefficient. B – D , H , I , and M are representative of two independent experiments. E , J –L, and N – R are representative of three independent experiments. Data are averages ±SD. P value was determined by Pearson coefficient. J , K , L , N , O P values were determined using one-way ANOVA.

Article Snippet: The CD138 antibody (#AF2780) was purchased from R&D Systems.

Techniques: Mass Spectrometry, Immunoprecipitation, Transfection, Staining, Western Blot, Expressing, shRNA, Control, Plasmid Preparation

Journal: Oncogene

Article Title: LAMP5 modulates IRF4 stability and nuclear transport: a critical mechanism in myeloma progression and therapy

doi: 10.1038/s41388-025-03513-x

Figure Lengend Snippet: A Workflow of high-throughput virtual screening (HTVS) for LAMP5 inhibitors. B Docking scores of the top 4 candidates obtained from HTVS. C Chemical structure and in silico docking of pyrazofurin into the active pocket of human LAMP5 protein. D Proliferation of myeloma cells (ARP-1 or IM-9) treated with or without pyrazofurin (Pyr.) (0.1 or 1 μM) over time. PBS buffer treated group served as control. E Antiproliferative activities of pyrazofurin were evaluated on human myeloma cell lines ARP-1 or IM-9. F Co-immunoprecipitation of LAMP5 with IRF4 in ARP-1 or IM-9 cells treated with or without pyrazofurin (1 μM). G Co-immunoprecipitation of LAMP5 with IRF4 in HEK293T cells co-transfected with HA-LAMP5 and IRF4 plasmids. H IRF4 protein levels in myeloma cells treated with 1 μM pyrazofurin and 2.5 μM MG-132 or 2.5 μM leupeptin for 7 h. 6-week-old male C57BL/6 J mice were intravenously injected vk12598 cells ( n = 9 mice/group), followed by intraperitoneal administration of pyrazofurin (5 mg/kg bodyweight) three times per week for a duration of two weeks, beginning two weeks after cell injection. Shown are the experimental schematic ( I ). J ELISA analysis shown the concentrations of IgG2b in mouse sera. K Immunofluorescent staining of I mentioned spleen tumor cells of vk12598 mice with DAPI and antibodies against CD138 or IRF4. Scale bar, 40 μm. L Photographic images of tumors in NSG mice after implantation of ARP-1 or IM-9 cells ( n = 3 mice/group), and intraperitoneal administration with or without pyrazofurin (5 mg/kg bodyweight). Shown are time course of tumor volume ( M ). N , O Immunofluorescent staining of Ki67 or IRF4 expression in tumor tissues. D and E are representative of three independent experiments. F – H are representative of two independent experiments. Data are averages ±SD. J P value was determined by unpaired two-tailed t test; D , M P values were determined using two-way ANOVA.

Article Snippet: The CD138 antibody (#AF2780) was purchased from R&D Systems.

Techniques: High Throughput Screening Assay, In Silico, Control, Immunoprecipitation, Transfection, Injection, Enzyme-linked Immunosorbent Assay, Staining, Expressing, Two Tailed Test