Journal: Oncotarget

Article Title: CD30-targeted oncolytic viruses as novel therapeutic approach against classical Hodgkin lymphoma

doi: 10.18632/oncotarget.24191

Figure Lengend Snippet: ( A ) Schematic genome organization of the applied oncolytic viruses. Asterisks indicate the mutated residues in H protein to achieve blinding for the natural MV receptors CD46 and SLAM. The coding sequence for the CD30-specific scFv together with a C-terminal hexa-His-tag (H6) is fused to H mut . In VSV-CD30 the glycoprotein (G) reading frame was replaced with those of MV-F and H mut -CD30scFv ( B ) Immunoblot of virus stocks using rabbit-α-VSV serum (left panel) or antibodies recognizing the cytoplasmic tail of MV-H (center and right panel top row) or MV-F (center and right panel bottom row). ( C ) Multi-step growth curves of CD30-targeted viruses and untargeted parental viruses on Vero-αHis cells of cell-associated (lysate) and supernatant (SN) virus after infection at a multiplicity of infection (MOI) of 0.3, respectively. Titers were determined as 50% tissue culture infective dose (TCID50). n = 2, error bars: mean ± SD.

Article Snippet: Expression of human CD30 was detected by a phycoerythrin (PE)-labeled mouse CD30 antibody (clone: Ki-2, Miltenyi Biotech, Germany).

Techniques: Sequencing, Western Blot, Virus, Infection

Journal: Oncotarget

Article Title: CD30-targeted oncolytic viruses as novel therapeutic approach against classical Hodgkin lymphoma

doi: 10.18632/oncotarget.24191

Figure Lengend Snippet: ( A ) CHO-cells stably expressing SLAM, CD46 or CD30 were infected with CD30-targeted viruses or untargeted parental viruses at an MOI of 1 and analyzed by fluorescence microscopy 24 h (VSV-MV, VSV-CD30) or 72 h (MV, MV-CD30) post infection, respectively. Scale bar = 200 μm. ( B ) HT1080-RFP and HT1080-CD30 were co-cultured at a ratio of 70:30 and infected with CD30-targeted viruses or untargeted parental viruses at an MOI of 1 and analyzed by fluorescence microscopy 24 h (VSV-MV, VSV-CD30) or 72 h (MV, MV-CD30) post infection, respectively. An overlay of red and green fluorescence is shown. Scale bar = 200 μm. (C) VSV-MV and VSV-CD30 were pre-incubated for 30 min with increasing amounts of CD30-Fc protein. Subsequently, HT1080-CD30 cells were inoculated and the percentages of infected cells were determined 72 h later. Relative infectivities were calculated by normalizing the values to mock treated controls. n = 3, error bars: mean ± SD.

Article Snippet: Expression of human CD30 was detected by a phycoerythrin (PE)-labeled mouse CD30 antibody (clone: Ki-2, Miltenyi Biotech, Germany).

Techniques: Stable Transfection, Expressing, Infection, Fluorescence, Microscopy, Cell Culture, Incubation

Journal: Oncotarget

Article Title: CD30-targeted oncolytic viruses as novel therapeutic approach against classical Hodgkin lymphoma

doi: 10.18632/oncotarget.24191

Figure Lengend Snippet: ( A ) FACS analysis of the human cHL cell lines L-428, L-1236 and KM-H2 for expression of CD30, SLAM and CD46. ( B ) The cHL cell lines L-428, L-1236 and KM-H2 were infected with the CD30-targeted or the untargeted parental viruses at an MOI of 1, respectively, and analyzed by fluorescence microscopy 24 h (VSV-MV, VSV-CD30) or 72 h (MV, MV-CD30) post infection. Scale bar = 200 μm. ( C ) Cell viability was determined using the RealTime-Glo MT Cell Viability Assay. Human KM-H2 cells were infected with MV-CD30 or VSV-CD30 an MOI of 1, respectively, and viability was determined until 48 h post infection. n = 3, Error bars: mean ± SD.

Article Snippet: Expression of human CD30 was detected by a phycoerythrin (PE)-labeled mouse CD30 antibody (clone: Ki-2, Miltenyi Biotech, Germany).

Techniques: Expressing, Infection, Fluorescence, Microscopy, Viability Assay

Journal: Oncotarget

Article Title: CD30-targeted oncolytic viruses as novel therapeutic approach against classical Hodgkin lymphoma

doi: 10.18632/oncotarget.24191

Figure Lengend Snippet: KM-H2 cells were implanted subcutaneously into NSG mice. 13 days post cell implantation, mice received three intratumoral injections over a period of five days (dotted lines) covering a total dose of 3 × 10 6 TCID50 (MV-CD30, VSV-CD30 low dose) or 3 × 10 8 TCID50 (VSV-CD30 high dose). ( A ) Tumor growth curves of MV-CD30 (blue circles), VSV-CD30 low dose (purple triangles), VSV-CD30 high dose (red triangles, +) or mock (black rectangles) treated mice. ( B ) Calculated area under the curve (AUC) values for tumor growth data shown in (A). One-way ANOVA test (Multiple comparisons), **** p < 0.0001. ( C ) Kaplan-Meier plot survival analysis. Logrank test (Bonferroni adjusted), *** p < 0.001, ** p < 0.01. Measles virus (MV)-CD30, n = 7; vesicular stomatitis virus (VSV)-CD30 low dose, n = 8; VSV-CD30 high dose, n = 8; mock, n = 8. Error bars: mean ± SEM.

Article Snippet: Expression of human CD30 was detected by a phycoerythrin (PE)-labeled mouse CD30 antibody (clone: Ki-2, Miltenyi Biotech, Germany).

Techniques: Virus

Journal: Oncotarget

Article Title: CD30-targeted oncolytic viruses as novel therapeutic approach against classical Hodgkin lymphoma

doi: 10.18632/oncotarget.24191

Figure Lengend Snippet: ( A ) KM-H2 cells were implanted subcutaneously into NSG mice. 14 days post cell implantation, a total dose of 3 × 10 8 TCID50 of VSV-CD30 split in three aliquots was systemically injected weekly (dotted lines). Tumor growth curves of VSV-CD30 (red triangles) or mock (black rectangles) treated mice are shown. ( B ) Calculated area under the curve (AUC) values for tumor growth data shown in (A). Unpaired t test, **** p < 0.0001. Vesicular stomatitis virus (VSV)-CD30, n = 7; mock, n = 8. Error bars: mean ± SEM. ( C ) KM-H2-luc cells were injected intravenously into NSG mice. Based on the luciferase signal intensity on day 14, mice received three intravenous injections of VSV-CD30 covering a total dose of 3 × 10 8 TCID50 on day 18, 21 and 28 post cell administration. Quantified luciferase signals on day 46 are shown as logarithm of the total flux (p/sec). Unpaired T test, p = 0.07. VSV-CD30, n = 8; mock, n = 6.

Article Snippet: Expression of human CD30 was detected by a phycoerythrin (PE)-labeled mouse CD30 antibody (clone: Ki-2, Miltenyi Biotech, Germany).

Techniques: Injection, Virus, Luciferase