In vitro preclinical study results for a phase I adoptive immunotherapy trial using cetuximab with allogeneic NK cells in KRAS oncogene-mutated metastatic colorectal cancer patients.

Abstract

e14057 Background: Prognosis for metastatic colorectal-cancer (mCRC) patients is unfavorable. Futhermore mCRC patients with tumors harboring a mutation in the KRAS gene do not derive benefit from the administration of cetuximab. We hypothesized that KRAS mutated CRC cells may be susceptible to cetuximab-induced ADCC mediated by healthy donor NK cells. Methods: Purified allogeneic NK cells or preincubated with IL-2 or IL-15 were used as effectors. Seven different CRC cell lines (DLD1, HCT-116, HT-29, LOVO, SW620, SW48, SW480) labeled with 51Cr were used as target cells alone or preincubated with cetuximab for ADCC assay. Results: All but one (SW620) cell lines express EGFR and MHC molecules; 4/7 harbor KRAS mutation (DLD 1, HCT-116, SW620, SW480). All cell lines were susceptible to unstimulated NK cells lysis (mean 20% ± 4.57, at ratio 25:1; mean 15% ± 3.65, at ratio 12:1) and lytic activity is strongly enhanced by IL-2 or IL-15 (mean 48% ± 5.8, at ratio 25:1; mean 38% ± 4.3, at ratio 12:1) (p < 0.05). No difference in lytic activity was observed between IL-2 vs. IL-15 and thawed vs. fresh NK cells. The SW48, a KRAS wilde-type cell line was killed by NK cells from 38% of lysis without cetuximab to 50% with cetuximab (E/T ratio of 50:1, p < 0.05). Similarly the HCT-116, a KRAS mutated cell line was killed by NK cells from 20% without cetuximab to 36% with cetuximab (E/T ratio of 50:1, p < 0.05). When IL-2 or IL-15 activated NK cells were used as effectors, both targets lysis increased from 50% without cetuximab to 71% with cetuximab (E/T ratio of 100:1, p < 0.05). When SW620 (EGFR expression 0%) was used as target cell line, pre-coating with cetuximab did not enhanced unstimulated or pre-activated NK lysis capacity. Conclusions: allogeneic NK cells are cytotoxic versus all tested cell lines. Moreover lytic activity of thawed and fresh NK cells is similar. In ADCC assay NK cell lytic activity is significantly enhanced by cetuximab; surprisingly cetuximab mediated ADCC activity is independent from CRC cell lines KRAS oncogene mutational status and require EGFR membrane expression. Moreover lytic activity of IL-2 and IL-15 activated NK cells is increased by cetuximab. No significant financial relationships to disclose.