C1q-Binding HLA Antibodies Do Not Predict Platelet Transfusion Failure in TRAP Study Participant

Introduction: In the Trial to Reduce Alloimmunization to Platelets (TRAP) study, 101 of the 530 subjects became clinically refractory (CR) to platelet transfusions in the absence of detectable antibodies against HLA as measured by the lymphocytotoxicity assay (LCA). Using more sensitive bead-based detection methods we have previously demonstrated that while many of these LCA^- patients do have anti-HLA antibodies, that these low to moderate level antibodies do not predict refractoriness. In addition to being less sensitive then bead based methods, the LCA screen only detects complement-binding antibodies. As these antibodies could be important for platelet rejection, we assessed if previously undetected complement-binding antibodies could account for some of the refractoriness seen in LCA^- patients.

Methods: 169 LCA- (69 CR, 100 non-CR) and 20 LCA+ (10 CR, 10 non-CR) subjects were selected from the TRAP study. Anti-class I HLA IgG and C1q binding antibodies were measured in serum or plasma using two different multi-analyte, semi-quantitative, bead-based fluorescent antibody detection assays: the LabScreen mixed Luminex assay, and the LabScreen single antigen class I assay with or without added EDTA (One Lambda). Groups were compared using an unpaired t-test, a=0.05, and correlation between variables was also assessed.

Results: New measurements of anti-class I HLA IgG antibodies reliably reproduced earlier data with a strong correlation between the old and new measurements (R²=0.9736, p<0.0001). While some of the LCA^- subjects did have detectable C1q-binding anti-class I HLA antibodies, and some LCA⁺ subjects did not, levels of these antibodies were significantly higher among LCA⁺ subjects (p<0.0001). Levels of C1q-binding anti-class I HLA antibodies were not significantly different between CR and non-CR among either the LCA^- or LCA⁺ subjects.

Conclusions: While complement-binding anti-class I HLA antibody levels were higher in the LCA⁺ subjects, higher levels of these antibodies were not seen in CR LCA⁺ patients as compared with non-CR LCA⁺ patients. Complement-binding anti-class I HLA antibodies do not account for refractoriness seen among the LCA^- TRAP subjects. This work confirms that low to mid level anti-class I antibodies do not drive refractoriness to platelets, and suggests that antibody-independent mechanisms cause refractoriness in patients lacking higher levels of anti-HLA antibodies.