International validation of a dithiothreitol (DTT)-based method to resolve the daratumumab interference with blood compatibility testing.

BACKGROUND: Daratumumab (DARA) consistently interferes with routine blood bank serologic testing by directly binding to CD38 expressed on reagent red blood cells (RBCs). Treating RBCs with dithiothreitol (DTT) eliminates the DARA interference. We conducted an international, multicenter, blinded study aimed at validating the DTT method for use by blood bank laboratories worldwide. STUDY DESIGN AND METHODS: Paired plasma sample unknowns were sent to 25 participating blood bank laboratories. Sample 1 was spiked with DARA only (10 µg/mL), and Sample 2 with DARA plus a clinically significant RBC antibody (anti-D [n = 6], anti-Fya [n = 9], or anti-s [n = 10]). Sites were instructed to perform an antibody screen with and without DTT-treated RBCs and to use a DTT-treated RBC panel for antibody identification. Qualitative data about the DTT method were collected by online survey. The primary outcome was the proportion of study sites able to identify the antibody unknown in the presence of DARA. RESULTS: All sites observed the DARA interference with the antibody screen. The DARA interference was seen with all testing methods (gel, tube, or solid phase). Using the DTT method, 25 of 25 sites (100%) successfully identified the antibody unknown in the presence of DARA. Feedback on the DTT method was positive, with 17 of 19 (90%) sites responding to the survey indicating that they planned to use the DTT method to test clinical samples from DARA-treated patients. CONCLUSION: The DTT method is robust and reproducible and can be implemented by transfusion services worldwide to help provide safe blood products to patients treated with DARA.

Resolving the daratumumab interference with blood compatibility testing.

BACKGROUND: Daratumumab (DARA), a promising novel therapy for multiple myeloma, is an IgG1κ monoclonal antibody that recognizes CD38 on myeloma cells. During routine compatibility testing, we observed that the plasma of five of five DARA-treated patients demonstrated a positive antibody screen and panreactivity on red blood cell (RBC) panel testing. We hypothesized that the observed panreactivity reflected DARA binding to CD38 on reagent RBCs, and we investigated methods to prevent this binding. STUDY DESIGN AND METHODS: DARA binding to CD38+ or CD38- HL60 cells was assessed by flow cytometry. To remove cell surface CD38, cells were incubated with dithiothreitol (DTT) or trypsin. Soluble CD38 or anti-DARA was used to neutralize DARA in solution. Routine blood bank serologic methods were used to test samples from DARA-treated patients and normal plasma samples spiked with DARA and/or alloantibodies. RESULTS: Normal plasma samples spiked with DARA (0.1-10 µg/mL) and incubated with reagent RBCs recapitulated the interference observed with samples from DARA-treated patients. Flow cytometry experiments confirmed DARA binding to CD38+ HL60 cells, but not to CD38- controls. DTT treatment of CD38+ HL60 cells reduced DARA binding by 92% by denaturing cell surface CD38. Treating DARA-containing plasma with soluble CD38 or anti-DARA idiotype also inhibited DARA binding. CONCLUSION: DARA causes panreactivity in vitro by binding to CD38 on reagent RBCs. Treating reagent RBCs with DTT is a robust method to negate the DARA interference, enabling the safe provision of blood to DARA-treated patients. Because DTT denatures Kell antigens, K- units are provided to these patients.