NK Cell Modulation of Anti-PD-L1 Antibody Effects

Example 4

This example provides a showing of the effect of NK cells on the disclosed anti-PD-L1 antibodies on mediated inhibition of proliferation. With the anti-PD-L1 antibodies showing a preferential binding to NK cells, the significance of this in the inhibition of proliferation was tested. By cell sorting using a FACS Aria (Becton Dickinson, San Jose, CA) purified population of CD4+, CD8+, CD56+ (NK) and monocytes were obtained. As a base culture, 1.5×10⁵ CD4+ cells and 3×10⁴ monocytes were stimulated with anti-CD3 (1 ng/ml) with or without H10 anti-PD-L1 antibody (10 μg/ml). In separate cultures, either CD8+ cells or NK cells (both at 3×10⁴) were added to this base culture. After three days of culture, cells were stained for expression of CD25 as a measure of lymphocyte activation as measured by flow cytometry. The results shown in FIG. 4 were compared to those obtained using whole, unfractionated PBMC (1.5×10⁵). The anti-PD-L1 antibody inhibited the activation of lymphocytes in the cultures containing whole PBMC and those where NK cells were added, but not in the absence of NK cells.