Extended Figure 4. Glutaminyl cyclase inhibition leads to reduced binding of recombinant hSIRPγ.

a, Cell surface binding of αhCD47-2D3, αhCD47-CC2C6 and hSIRPα-Fc to control (DMSO)-treated (-) or SEN177-treated (+) lung cancer (A549), colorectal (DLD1), HAP1, rectal carcinoma (RKO) and breast cancer (SKBR3) cells, as determined by flow cytometry. Data represent n=3 biological replicates and mean ± s.d. of triplicates. ***P≤0.000695 by unpaired two-sided t-test. b, Cell surface binding of αhCD47-2D3, αhCD47-CC2C6 and hSIRPα-Fc to control (DMSO)-treated (-), SEN177-treated, and PQ912-treated melanoma (A375) cells, as determined by flow cytometry. Data represent n=3 biological replicates and mean ± s.d. of triplicates. ***P=0.0001 by one-way ANOVA with multiple comparison correction. c, Flow cytometry plot of surface binding of αhCD47-B6H12 and αhCD47-CC2C6 to control-treated and PQ912-treated melanoma (A375) cells. Data are representative of two independent experiments with similar results (n=3 biological replicates per experiment). d, Cell surface binding of αhCD47-2D3, αhCD47-CC2C6 and hSIRPα-Fc to control (DMSO)-treated (-) and SEN177-treated (+) wild-type and QPCTL-knockout epidermoid carcinoma (A431) and lung cancer (A549) cells, as determined by flow cytometry. Data represent n=3 biological replicates and mean ± s.d. of triplicates. e, Cell surface binding of secondary antibody alone, hSIRPα-Fc (followed by secondary antibody) or hSIRPα-Fc in the presence of the CD47 blocking antibody αhCD47-B6H12 (followed by secondary antibody) to control (DMSO)-treated (-) or SEN177-treated lung cancer (A549) cells, as determined by flow cytometry. Data represent n=3 biological replicates and mean ± s.d. of triplicates. Values indicate MFI relative to WT cells stained with the same reagent (a, b, d) or MFI (e).

Data are representative of one (e) or at least two independent experiments (a-d)