Additional file 2: of Mutated Von Hippel-Lindau-renal cell carcinoma (RCC) promotes patients specific natural killer (NK) cytotoxicity

Figure S1. MHC Class-I profile in K562, CAKI-1, SN12C, A498 and 786-O cells. The expression of MHC-I was evaluated in RCC target cells by flow cytometry (FITC-conjugated major histocompatibility complex (MHC) class I–specific antibody (IgG2a, W6/32, CBL139F, Cymbus Biotech, Hants, UK). Figure S2. NKs from VHL-MUT-RCC patients display higher cytotoxicity toward human renal cancer cells VHL mutated A498 and 786-O. NK cell function was evaluated through CD107a cell-surface expression in response to ex vivo stimulation with K562, CAKI-1 (VHL-WT), A498 and 786-O (VHL-MUT) cells. Degranulation (CD107a) was evaluated after gating on CD3-CD56+ cells. CD107a+NK cells in 5 VHL-MUT (A) and 9 VHL-WT (B) RCC patients versus K562, CAKI-1, A498 and 786-O renal cell lines. Figure S3. NCAM-1, DNAM-1, FcγRIIIa, NKp44 and NKG2D are slightly overexpressed in VHL-MUT RCC tumors. RNA from 34 RCC tumors (17 VHL-WT and 17 VHL-MUT). Transcript levels are presented as mean±SEM. Statistical significance was calculated by unpaired Student t test (p < 0.05 ). Figure S4. NCAM-1, DNAM-1, FcγRIIIa, NKp30, NKp46, NKp44, NKG2D expression are upregulated in VHL-MUT tumors as compared to VHL-WT tumors. 17 VHL-MUT (A) and 17 VHL-WT (B). RT-PCR was performed on total RNA isolated from 34 tumors and relative peritumoral tissues (17 VHL-WT and 17 VHL-MUT). Relative gene expression levels were normalized to GUSB. Statistical significance was calculated by unpaired Student t test (p < 0.05). Figure S5. Expression of DNAM-1 ligand (PVR) in CAKI-1 and A498 cell lines. The expression of PVR (CD155) was evaluated in RCC target cells by flow cytometry (PE anti-human CD155/PVR, ( clone SKIL.4, Biolegend, Cat No 337609). (ZIP 561 kb)