AbstractSignal regulatory protein alpha (SIRPα) is the receptor for cluster of differentiation (CD)47, a potent “don't eat me” signal for macrophages. Disruption of CD47‐SIRPα signaling in the presence of prophagocytic signals can lead to enhanced phagocytosis of tumor cells, resulting in a direct antitumor effect; agents targeting this pathway have shown efficacy in non‐Hodgkin lymphoma (NHL) and other tumor types. GS‐0189 is a novel anti‐SIRPα humanized monoclonal antibody. Here we report: (1) clinical safety, preliminary activity, and pharmacokinetics of GS‐0189 as monotherapy and in combination with rituximab from a phase 1 clinical trial in patients with relapsed/refractory NHL (NCT04502706, SRP001); (2) in vitro characterization of GS‐0189 binding to SIRPα; and (3) in vitro phagocytic activity. Clinically, GS‐0189 was well tolerated in patients with relapsed/refractory NHL with evidence of clinical activity in combination with rituximab. Receptor occupancy (RO) of GS‐0189 was highly variable in NHL patients; binding affinity studies showed significantly higher affinity for SIRPα variant 1 than variant 2, consistent with RO in patient and healthy donor samples. In vitro phagocytosis induced by GS‐0189 was also SIRPα variant–dependent. Although clinical development of GS‐0189 was discontinued, the CD47‐SIRPα signaling pathway remains a promising therapeutic target and should continue to be explored.