Fig. 3.

Chemoradiation and MEDI0457 is associated with decreased PD-L1 expressing T cells and macrophages at the treated tumor site. (A) Multiplexed immunofluorescence for PD-L1 expression on CD8⁺ T cells, CD68⁺ macrophages, and panCK⁺ epithelial cells in cervical biopsies taken before chemoradiation (pre-CRT; n = 6), after chemoradiation (post-CRT; n = 5), and after chemoradiation and MEDI0457 (post-CRT + MEDI0457; n = 4). Inset in middle panels demonstrated PD-L1 costaining with CD8 or CD68. (B) Multiplexed immunofluorescence for CD8⁺ T cells (purple), Granzyme B (red), PD-1 (green), and the proliferative marker Ki-67 (yellow) from cervical biopsy specimens taken before chemoradiation and after chemoradiation and MEDI0457. Scale bar represents 400 μM for 5× magnification and 100 μM for 20× magnification. Post-CRT + MEDI0457 biopsies were taking at 63 and 122 days after the first MEDI0457 dose. (C) Percentages of panCK⁺ cells decrease whereas CD8⁺ T cells or CD68⁺ macrophages remain unchanged in pre-CRT, post-CRT, and post-CRT + MEDI0457 specimens. (D) Compared with pre-CRT and post-CRT time points, PD-L1⁺CD8⁺ T cells and PD-L1⁺CD68⁺ macrophages are decreased at post-CRT + MEDI0457 time points whereas PD-L1⁺PanCK⁺ epithelial cells remain unchanged. (E) CD8⁺PD-1⁺ cells are decreased whereas CD8⁺Granzyme B⁺ cells are increased after chemoradiation and MEDI0457. Calculated as percent change from prechemoradiation baseline. (C-E) represent quantitation of all available patient samples in the study. The asterisks (*) represents P < .05 as determined by Student t test. Error bars represent standard error of the mean. Abbreviations: CRT = chemoradiotherapy; DAPI = 4′,6-diamidino-2-phenylindole; PD-1 = programmed cell death protein 1; PD-L1 = programmed death-ligand 1; Vac = vaccine.