Extended Data Fig. 2: Comparison between Cmab-hCCL5 and hCCL5-Cmab and characterization of OV-Cmab-hCCL5.
(a, b) Detection of purified Cmab-hCCL5 or hCCL5-Cmab bound to U251T2 cells, measured by flow cytometry after staining Cmab-hCCL5- or hCCL5-Cmab-incubated tumor cells with anti-Fc-APC (a) or anti-hCCL5-APC (b). Cmab-hCCL5 and hCCL5-Cmab were purified from lentivirus-infected CHO cells. IgG1 isotype served as control. (c, d) Detection of Cmab-hCCL5 in the supernatant, collected from OV-Cmab-hCCL5-infected U251T2 GBM cell culture, that was associated with U251T2 cells, measured by flow cytometry after staining supernatant-incubated U251T2 cells with anti-Fc-APC (c) or anti-hCCL5-APC (d). (e) Cmab-hCCL5 produced in the supernatant of OV-Cmab-hCCL5-infected U251T2 cells at different MOIs (n = 3 independent experiments). (f) Cell viabilities of OV-Cmab-hCCL5 infection at three indicated MOIs were measured at 24 hours post infection (hpi), 48 hpi, and 72 hpi (n = 3 independent experiments). (g) The ability of OV-Q1 and OV-Cmab-hCCL5 to induce oncolysis of GBM cells, measured by real-time cell analysis (RTCA) The experiment was performed twice with similar data. (h) U251T2 cells were infected with OV-Q1 or OV-Cmab-hCCL5 at an MOI of 2. The supernatant was harvested at the indicated time points to assess viral production using a plaque assay with Vero cells (n = 3 independent experiments). Experiments in a-d were representative of three independent experiments with similar results. Error bars indicate the standard deviations (s.d.), and data are presented as mean ± s.d. (e, f, h). Statistical analyses were performed by one-way ANOVA with P values corrected for multiple comparisons by Bonferroni method (e).
