Elimination of Anti-CD47 drug interference in serological testing by antigen masking

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CPH and CS are employees of imusyn GmbH & Co. KG. CS is a designated inventor of the technology presented here.

INTRODUCTION: CD47 is a self-marker on cells that is overexpressed in several malignancies. Its interaction with SIRPα serves as a "don"t eat me" signal to macrophages. Among other effects, blocking this signal transduction to increase the rate of phagocytosis of cancer cells is the rationale behind the development of therapeutic anti-CD47 drugs such as the monoclonal antibody magrolimab. However, magrolimab induces strong agglutination of red blood cells (RBCs) not only in the indirect antiglobulin test (IAT) but also in the neutral test, impairing the determination of irregular antibodies and serological typing. Even though all trials with magrolimab have been discontinued, there are still a number of other anti-CD47 drugs under clinical investigation. It is unclear to which degree these interfere with serological testing.

METHODS: We tested the anti-CD47 drugs ontorpacept (TTI-621), ligufalimab (AK 117), letaplimab (IBI-188) and lemzoparlimab (TJ011133/TJC4) in the IAT, in the neutral test and in the Capture Assay in comparison with magrolimab. Furthermore, we tested if MagroEx, an optimized soluble SIRPα variant under development, is able to eliminate the interference by antigen masking.

RESULTS: With the exception of the SIRPα IgG1 Fc fusion protein ontorpacept, all anti-CD47 drugs tested were able to cross-link the RBCs without addition of anti-human globulin (AHG). In the IAT, all of the tested anti-CD47 drugs induced agglutination and all but ontorpacept resulted in positive reactions in the Capture Assay. MagroEx completely mitigated the interference in the neutral card and in the Capture Assay, and at least partially mitigated the interference in the IAT by antigen masking.

CONCLUSION: Anti-CD47 drugs will become a major problem in serological testing if they are approved for anti-cancer therapy. Antigen masking with SIRPα has proven to be an effective and easy method to mitigate their interference.