trampc2  (ATCC)

Journal: Cell Reports Medicine

Article Title: Optimized arenaviruses with tumor-tropic mutations promote safe anti-tumor efficacy via sustainable immune modulatory properties

doi: 10.1016/j.xcrm.2025.102411

Figure Lengend Snippet: Analysis of different combinations of fast evolution mutations (A) Infection assays (MOI = 0.1, 16 h) of different human cancer cells (lung adenocarcinoma: H1975 and A549; primary sarcoma: Gist-T1; melanoma metastasis: MaMel86a and MaMel51), myotubes and human skeletal muscle myoblasts (HSMM) ( n = 4–6; duplicates in 2–3 experimental replicates) with different recombinant viruses carrying a point mutation in the respective position. Notably, the GP mutation 492I always occurred alongside one single NP mutation. Hence, in all further analyses, the mention of mutation 492I mutation inherently includes the NP mutation, even if not explicitly stated. (B) Infection assays (MOI = 0.1, 16 h) of different murine cancer cells (oropharyngeal carcinoma: MOPC; Lewis lung carcinoma: LLC; prostate adenocarcinoma: TrampC2; colon adenocarcinoma: MC38) and human cancer cells (lung adenocarcinoma: H1975 and A549; melanoma metastasis: MaMel86a and MaMel51) ( n = 6; duplicates in 3 experimental replicates) using different recombinant viruses carrying specific point mutations. (C) Spider plots showing the factor of acceleration in propagation of the mutations tested in various human and murine tumor cells. The mean ratio for each mutated virus is given ( n = 4–6; duplicates in 2–3 experimental set ups). (D) Entry assay on human lung adenocarcinoma (A549) cells and A549-αDG knockout cells of recombination virus GP181M-185W-492I and control virus ( n = 6; duplicates in 3 independent experiments). (E) A549 lung adenocarcinoma cells were treated with CD164 blocking antibody or isotype control for 1 h and subsequently infected with the recombination virus GP181M-185W-492I (MOI 10) for 1, 5, and 15 min. The number of viral particles outside the cells per one cell is shown ( n = 6 cells analyzed per sample, ∗∗ p < 0.01). (F and G) Representative pictures for cells treated with isotype control (scale bars: left = 2 μm, right = 200 nm) (F) and CD164 blocking antibody (scale bars: left = 2 μm, right = 200 nm) (G) for 1 min of infection are shown. (H) Infection assays (MOI = 0.1, 16 h) of different human cancer cells (thyroid anaplastic carcinoma: Cal62, C643, 8305C, and 8505C; epidermoid carcinoma: A431; lung adenocarcinoma: KRAS-mutated: A549 and H23; EGFR-mutated: H1975, Alk-rearranged: H2228, WT/other: H1299, H1355, H1792, and H1373; small cell lung cancer: HCC-44; endocervical adenocarcinoma: HeLa; fibroblast liposarcoma: SW-872; colon adenocarcinoma: SW-480; bronchiole lung carcinoma: H358; hepatocellular carcinoma: HepG2) ( n = 6–8; duplicates in 3–4 experimental set ups) comparing WE, recombinant WE-CL13 and a recombinant virus carrying three point mutations as shown. For statistical analysis, WE-CL13-GP181M-185W-492I was compared to both WE and WE-CL13. (I) Hepatocytes ( n = 3; biological replicates; separate flasks), melanocytes ( n = 3; biological replicates; separate flasks), epithelial cells (InEpc, n = 3; biological replicates; separate flasks) and alveolar cells (ALI-cultures, n = 6; duplicates of 3 different patients) were infected with WE or WE-CL13-GP181M-185W-492I. Number of infected cells was determined 24 h with flow cytometry. For statistical analysis, the WE-CL13-GP181M-185W-492I virus was compared to the mock-infected control, but no significant difference was observed. However, a statistical difference between WE and WE-CL13-GP181M-185W-492I was detected. Data are presented as the mean ± SEM; ns = not significant, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001 by t test (D, E, H, and I).

Article Snippet: TrampC2 , ATCC , CRL-2731.

Techniques: Infection, Recombinant, Mutagenesis, Virus, Knock-Out, Control, Blocking Assay, Flow Cytometry

Journal: Cell reports

Article Title: The lactate-NAD + axis activates cancer-associated fibroblasts by downregulating p62

doi: 10.1016/j.celrep.2022.110792

Figure Lengend Snippet: (A) Experimental design and qPCR of the indicated genes in mPSC-GFP cells mixed with TRAMPC2 cells and cultured for 7 days (n = 3 biological replicates). (B and C) Experimental design, immunoblot analysis (B), and qPCR of SQSTM1 (C) in WPMY-1 cells co-cultured with PCa cells during 48 h (n = 3 biological replicates). (D and E) Experimental design, immunoblot analysis (D), and qPCR of SQSTM1 in WPMY-1 cells incubated 48 h with conditioned media (CM) from PCa cells (E) (n = 3 biological replicates). (F) SQSTM1 promoter-driven luciferase in WPMY-1 cells incubated 48 h with CM from PCa cells (n = 3 biological replicates). (G and H) Immunoblot analysis (G) and qPCR of SQSTM1 (H) in WPMY-1 cells incubated 48 h with fractionated CM from PC3 cells (n = 3 biological replicates). (I) SQSTM1 promoter-driven luciferase in WPMY-1 cells treated as in (G) (n = 3 biological replicates). (J and K) Immunoblot analysis (J) and qPCR of SQSTM1 (K) in WPMY-1 cells incubated 48 h with normal or boiled CM from PC3 cells (n = 3 biological replicates). (L) SQSTM1 promoter-driven luciferase in WPMY-1 cells treated as in (J) (n = 3 biological replicates). Results are shown as mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Article Snippet: Mouse TRAMPC2 , ATCC , Cat# CRL-2731, CVCL_3615.

Techniques: Cell Culture, Western Blot, Incubation, Luciferase

Journal: Cell reports

Article Title: The lactate-NAD + axis activates cancer-associated fibroblasts by downregulating p62

doi: 10.1016/j.celrep.2022.110792

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Mouse TRAMPC2 , ATCC , Cat# CRL-2731, CVCL_3615.

Techniques: Polymer, Virus, Recombinant, Transfection, Western Blot, Infection, Electroporation, Quantitation Assay, Control, Staining, Real-time Polymerase Chain Reaction, Plasmid Preparation, Software, Imaging, Spectrophotometry, Microscopy