15353-j-veluchamy

5 Towards UCB-NK cells treatment in colorectal cancer | 123 Enhanced in vitro cytotoxicity of colon cancer cells mediated by UCB-NK cells In order to explore novel therapies to replace dysfunctional NK cells in patients with advanced CRC, we tested two different sources of allogeneic NK cell products (A-PBNK and UCB-NK), that could eventually be used for adoptive transfer strategies. We next compared the activity of A-PBNK cells (age range 22-37 years) and UCB-NK cells using a flow-based NK cell cytotoxicity assay based on detection of 7-AAD accumulation in tumor cells. Three different cell lines of colon cancer origin were compared, i.e. COLO320 (EGFR - RAS wt ), SW480 (EGFR + RAS mut ) and HT-29 (EGFR + RAS wt , BRAF mut ). As expected, addition of cetuximab to EGFR - RAS wt COLO320 cells did not result in increased killing. Of interest, lysis was consistently and significantly higher (p<0.01) using UCB-NK compared to A-PBNK. As reported previously, the combination of cetuximab and A-PBNK resulted in increased killing Figure 1: Low prevalence and functionally impaired NK cells in CRC patients (A) Frequency of NK cells within PBMC from healthy controls and from mCRC patients at baseline and after the first cycle of chemotherapy. (B) NK cell degranulation in healthy controls and mCRC patients after a 4 hr. co-culture of resting NK cells with A431 cells in the presence (open symbols) or absence (closed symbols) of cetuximab at an E:T ratio of 1:1. (C) Expression levels of resting NK cell CD16 and (D) NKp44 in healthy controls and in mCRC patients before and after 1 cycle of chemotherapy. Data represent mean ± SEM from 10 mCRC patients and 10 age and sex matched healthy controls. *P < 0.05, **P < 0.01, ***P < 0.005, calculated with one-way ANOVA, multiple comparison between column means.