15353-j-veluchamy

4 NK cell ADCC enhances treatment efficacy in colorectal cancer | 101 cetuximab to see if sensitizing target cells with cetuximab could contribute to improved NK cell killing through ADCC. The results showed that COLO320, Caco-2 and SW620 cells are more sensitive for NK killing compared to SW480 and HT-29 cells (Figure 4A). However, while co-incubation with cetuximab did not result in a significant increase in COLO320 tumor cell death, a significant increase in killing was seen in four EGFR + cell lines independent of RAS and BRAF status. With respect to Caco-2, SW480, SW620 and HT-29, the cytotoxic effect could be increased by cetuximab (Figure 4A), this enhancement of the cytolytic NK cell response was also evident from an increased rate of degranulation in the condition where NK cells and cetuximab were added in combination to EGFR + colon cancer cell lines (Figure 4B). Interestingly, we also assessed tumor cell expression of HLA-E and NK cell expression of NKG2A as HLA-E is a known ligand for the NK cell inhibitory receptor NKG2A. As can be seen in the supplementary figure 3A and B, while NKG2A levels were comparable on NK cells in all donors that were tested, HLA-E expression levels were strikingly different and correlated with the overall susceptibility of tumor cells to NK cell lysis as shown in figure 4, where cell lines with low HLA-E levels (COLO320, Caco-2, SW620) were more sensitive to NK cell killing compared to higher HLA-E expressing cell lines (SW480 and HT-29).These data suggest that NK cells have the potential to improve anti-EGFR mAb therapy efficacy even in situations where tumors carry RAS mut , BRAF mut or are EGFR - . Figure 3: Evaluation of NK cell CD16a polymorphism on cetuximab induced ADCC Differences in cetuximab induced ADCC between NK cells expressing V/V and V/F polymorphic versions of CD16 were analyzed. Natural cytotoxicity and NK cell CD16a mediated degranulation in the absence and presence of cetuximab coated A431 cells are shown in figure A and B. Columns are mean of triplicate values; with bars showing SD. Data are from independent experiments performed in triplicates from 3 V/V and 3 V/F donors. Mean ± SD for each significant condition are represented as p = <0.05 *, <0.01 **, <0.005 ***, <0.001 ****.