Fig. 6

Pep R54 in combination with nivolumab inhibits PES43 human melanoma cell growth, signaling and tumor growth. a. PD-1/CD279 (Clone HA2-7B1), PD-L1 (Clone MIH1) and CXCR4 (Clone 12G5) expression in PES43 by flow cytometry. b. PES43 cell growth in the presence of nivolumab or pembrolizumab [10 μM], non-specific IgG4[10 μM], PD-L1 [2μg/mL]. Growth curve graph (mean Number of Viable PES43 Cells ± SEM). c. Immunoblotting for ERK1/2, P38, AKT, 4EBP1 phosphorylated protein in PES43, PD-L-1 [2 μg/mL] plus nivolumab [10 μM] (6 h incubation); representative data from one of three experiments. d. Athymic mice were subcutaneously inoculated with 2.5 × 10⁶ PES43 human melanoma cells and treated for 3 weeks with Peptide R54 (2 mg/kg ip, 5 days per week), nivolumab (5 mg/kg ip, twice weekly) and combination. Tumor volume mm³ ± SEM: untreated 622.72 ± 119; nivolumab 503.47 ± 107; Pep R54 567 ± 214; nivolumab + Pep R54 410.33 ± 105). (untreated mice n = 8; nivolumab n = 8; Pep R54 n = 8; nivolumab + Pep R54 n = 9); e. Lung disseminated cells (DTC) PES43 cells were detected through flow cytometry as % hu- %MSCP APC positive cells (untreated mice n = 6; nivolumab n = 4; Pep R54 n = 5; nivolumab + Pep R54 n = 4) (empty dot = sample; black dot mean)