NeuroMab™ Anti-SIRPA BBB Shuttle Antibody, Clone 3F9

Figure 1 shows FACS histograms of selected SIRPA antibodies bound to rodent Chinese hamster ovary cell lines (CHO) expressing human SIRPA (HuSIRPA) or mouse SIRPA (MuSIRPA) Left panel

Figure 2 shows FACS histograms of binding of selected SIRPA antibodies to primary human macrophages.

Figure 3 shows the binding of increasing concentrations of anti-SIRPA antibodies to human SIRPA overexpressed on CHO cells. Calculate EC50 values by fitting the data to a sigmoid curve using Graph Pad Prism.

Figure 4 shows the ability of CD47-blocking and CD47-non-blocking anti-SIRPA antibodies to affect HuSIRPA-dependent luciferase expression in a cell-based reporter assay.

BWZ-HuSIRPA cells were seeded on wells with or without plate-bound CD47 protein. All CD47-blocking antibodies (1B3, 12D6, 1H11, 5F7) potently suppress luminescence signal. Two CD47-non-blocking anti-SIRPA antibodies did not reduce luciferase expression. Results are expressed as fold over background. The background level is set to 1 on y-axis.

Figure 5 shows induction of human SIRPA-dependent or human SIRPB1-dependent luciferase expression in a cell-based reporter assay.

Figure 6 shows SIRPA receptor downregulation in primary human macrophages in response to antibody stimulation.

Cells were treated with either soluble full-length isotype control or soluble full-length anti-SIRPA antibodies and subsequently stained with a DyLight650-conjugated anti-SIRPA reference antibody (SA56-DyL650) that binds to a distinct epitope bin.

Figure 7 shows enhanced phagocytic activity of macrophages treated with CD47 blocking anti-SIRPA antibodies.

Macrophages were cultured overnight in 2.5% FBS RPMI media with 5 μg/mL of 12D6, 9C5, 1H11, 5F7, 1B3, 3F9 (CD47-non-blocker) or isotype control.

Figure 8 Graph showing mean tumor volumes of NSG mice transplanted with human immune stem cells from different cord blood donors (donors 5031, 5048, 129).

Humanized mice were implanted subcutaneously with the human breast cancer cell line MDA-MB-231 and randomized into treatment or control groups based on tumor volume at Day −1, huCD34+ stem cell donor, body weight before randomization, and huC45+ engraftment rate before randomization. Mice were dosed with i.p. injections of either mouse IgG1 or 3F9 at 40 mg/kg every 4 days or Keytruda at 10 mg/kg every 5 days.