A 35-year-old woman presented for cardiac surgery with a history of 2 prior DHTRs at age 30 and 32. Her blood type is D+C–E–c+e+, K–, Fy(a–b–), Jk(a+b–), S–s+. Prior to the 2 DHTRs, she had previously developed anti-C, K, and auto–anti-e. At age 30, DHTR was triggered by development of anti-S and anti-Lua with prophylactic transfusion during pregnancy of crossmatch-compatible RBCs (C–E–k–Fya–Jkb–). At age 32, DHTR was triggered by transfusion for abortion with development of anti-E despite provision of E-negative and crossmatch-compatible RBCs (C–E–k–Fya–Jkb–S–). In both instances, she was admitted to the intensive care unit (ICU) with severe pain, hemoglobinuria, and evidence of hemolysis, with an lactate dehydrogenase (LDH) increase from 320 to 1550 and 2058 IU/L, respectively. Her Hb dropped 2 g/dL between days 2 and 10 after transfusion in the first DHTR, and 3 g/dL between days 2 and 12 in the second DHTR.
For the cardiac surgery, she received 2 1000-mg doses of rituximab prophylaxis at 1 month and 15 days before surgery. She received 2 units of crossmatch-compatible RBCs (C–E–k–Fya–Jkb–S–), with a day 1 posttransfusion Hb of 7.7 g/dL and HbA% of 62. Her clinical course was uneventful, with Hb 8.6 g/dL and HbA% 37 at day 25 and no new antibodies in follow-up (3 months). The patient had previously received antipneumococcal vaccine and received prophylactic antibiotics for 3 months.
This high-DHTR-risk case illustrates how an immunization prevention strategy likely prevented development of DHTR.
How to prevent alloimmunization?
What do we know about alloimmunization in SCD?
The incidence of alloimmunization is high in SCD,13 in part because of higher prevalence of C, E, Fya, Jkb, and S polymorphic blood group antigens in donors of primarily European descent than patients of African origin.14⇓⇓⇓-18 In addition, transfusion burden and exposure, as measured by cumulative number of transfusions, is higher in SCD and linearly correlates with SCD alloimmunization.19
The higher alloimmunization in SCD may also be because of the inflammatory nature of this condition. Studies in experimental models, although not seen in mouse SCD models,20 indicate that recipient inflammatory state increases the risk of alloimmunization. In a retrospective study, SCD patients transfused for acute, and therefore inflamed, complications had increased alloimmunization risk.21
Some patients, so-called responders, become alloimmunized early during transfusion therapy, whereas others never become immunized, invoking the concept of genetic predisposition to alloimmunization.19 Genetic studies have also identified gene polymorphisms encoding immune modulatory proteins including TRIM21, CD81, and CTLA-4, as well as certain HLA alleles associated with SCD alloimmunization.22⇓⇓-25 A genetic variant in the Fcγ receptor gene was recently associated with decreased risk of SCD alloimmunization, although interestingly, this variant did not protect against alloimmunization to the highly immunogenic Rh and K antigens.26 Once validated in larger cohorts, such genetic markers may help risk stratify SCD patients, ensuring that use of extended and genotype-matched units are reserved for patients at highest risk of alloimmunization. Functional immune studies comparing alloantibody responders with nonresponders have unraveled aberrant molecular pathways potentially associated with alloimmunization, including innate immune response to heme, follicular helper T-cell subset signaling, and regulatory T-cell suppressive pathways.27⇓⇓⇓⇓⇓-33 Prospective studies, following patients with SCD as they become alloimmunized, are needed to determine whether these immunological alterations are the cause or effect of alloimmunization. Although the latter may help identify novel targeted therapies to reverse or prevent alloimmunization, the former is essential for identifying alloimmunization risk biomarkers.
Antigen matching: what and how much to match?
Transfusing SCD patients with RBCs matched for Rh (D, C, E, c, e) and K antigens is the standard of care in many centers.34⇓-36 Individuals of African descent have a high degree of genetic variation in the RH locus,37,38 and thus may have RH variants encoding so-called partial Rh antigens, which lack certain immunogenic epitopes of the normal antigen. Such patients, even when receiving Rh serologic-phenotype matched units from minority donors, may develop anti-Rh antibodies against those missing epitopes when transfused with RBCs carrying the “normal” antigen (see supplemental Figure 1 for examples of partial D antigens). In a pediatric population receiving Rh (D, C, E, c, e) and K matched units, 45% of the chronically and 12% of the episodically transfused recipients still became Rh alloimmunized,37 with similar results in a patient cohort in France.38 In SCD patients, many partial D, C, and e antigens are described, and importantly, the resulting antibodies are associated with DHTR cases.37,39,40 Such alloimmunized patients should be transfused with antigen-negative units (D-negative RBCs for a partial D patient). Although serologic tools can identify some Rh variant antigens, molecular techniques are more specific in identifying and distinguishing between RH alleles encoding partial and other variant antigens.41 However, until costs decrease, we propose that RH genotyping to identify partial Rh antigens be performed only in patients already immunized with clinically significant alloantibodies and/or autoantibodies whom we consider high antibody responders, and in patients who develop an Rh antibody despite conventional Rh matching. We also propose that Rh partial antigen matching as an alloimmunization prevention strategy should be based on antigen-negative unit availability and reserved for the high responders because not all patients with Rh variants become alloimmunized.
Extended matching to Fy, Jk, and Ss blood groups can further reduce alloimmunization36 but is not standard practice, in part because of insufficient supply of extended matched units for managing the routine transfusion needs of all patients with SCD. Furthermore, alloimmunization against Fy, Jk, and Ss occurs in only 5% to 15% of polytransfused patients.21,37,38 Therefore, extended matched units should be reserved only for patients in need, including those who develop the corresponding alloantibodies.
Why the need for an additional preventive strategy against RBC immunization?
Although detectable alloantibodies in SCD DHTRs are frequently against antigens such as Rh, K, Fy, Jk, and Ss, patients can also develop antibodies against many other RBC antigens, including low-frequency antigens as well as autoantibodies and nonspecific antibodies.15,42 Although clinical relevance of some of these antibodies in DHTR is not completely understood, severe DHTRs associated with autoantibodies or low-frequency alloantibodies have been reported.43,44 B-cell depletion therapy has therefore been empirically used to prevent RBC immunization in high-risk patients (those already immunized and with history of DHTR). In many cases, prophylactic rituximab has prevented alloimmunization and DHTRs.43,45,46 In 1 case study, however, its use was associated with fatal outcome,47 and in a case series involving 8 patients, all with favorable outcomes, mild DHTR still developed in 3 patients.45 Although case control studies are needed to more definitively establish efficacy, rituximab can be considered when a new transfusion is absolutely necessary in patients with a history of severe DHTR and evidence of being a high antibody responder. Patients receiving rituximab require antipneumococcal vaccination, which is already recommended for asplenic SCD patients, and antibiotics (twice a day, 1 MU penicillin V or 500 mg penicillin) until CD19+ B-cell counts recover. Corticosteroid is also typically administered with rituximab for prevention of hypersensitivity reactions; only a low dose (methylprednisolone, 10 mg) should be given to SCD patients, to avoid corticosteroid-induced VOE. Beyond limiting development of new RBC antibodies, rituximab prophylaxis is not indicated in prevention of alloantibody-negative DHTRs.
Proposed DHTR prevention strategies based on known risks of DHTR
Given that DHTRs are unpredictable and often life threatening, identifying risk factors by patient history and presentation is urgently needed in order to risk stratify the transfusion regimen.
Many studies suggest that 3 factors increase the risk of DHTR: (1) history of immunization, (2) previous history of DHTRs, and (3) transfusion for an acute complication. A lower cumulative number of transfused units (≤12 units) may also be a risk factor in adult patients.48 Odds ratios from a single-institution multivariate analysis of DHTR risks were used to create a scoring system to predict the probability of DHTR and accordingly adapt transfusion protocols (Figure 1).48
Patients with no history of DHTR on chronic transfusion have a lower risk of developing DHTR. In fact, in a recent prospective study of adults, DHTR was demonstrated exclusively in patients receiving episodic transfusions.48 Within that group, history of RBC immunization and a previous DHTR dramatically increased DHTR risk. These 3 risk factors (number of previous transfusions, history of immunization, and previous DHTRs) should thus be carefully considered when evaluating a patient for transfusion. If transfusion is necessary in high-risk patients, preventing alloimmunization should be prioritized because it may induce DHTR. For patients with anti-Fy, -Jk, or -Ss, we recommend extended matching (Fy, Jk, Ss) because the risk of producing additional antibodies increases with each new transfusion.19,38 Given their high likelihood of alloimmunization against any blood group antigen, we also recommend prophylaxis with rituximab prior to transfusion in this patient group.
Prevention of alloantibody-negative DHTR is challenging. Although there currently is no known DHTR preventive measure for this type of patient, one must rule out the possibility of actual involvement of antibodies, such as those against low-frequency antigens that require the use of specific RBCs for their detection or evanescent antibodies.49 Follow-up antibody screens at set intervals will maximize detection of any possible posttransfusion antibodies, which once identified, help selection of units for future transfusions to avoid restimulation. For patients who are nonalloimmunized but have a history of DHTR and few previous transfusions, we propose empiric extended matching (Fy, Jk, Ss) alone. Rituximab prophylaxis is not indicated unless antibody production was possibly missed, as described previously. The mechanism(s) that trigger RBC destruction in confirmed cases of antibody-negative DHTR are largely unknown, but possible mechanisms, which remain to be demonstrated, could involve heme-driven alternative complement activation pathways, suicidal death of transfused aged-stored RBCs, or destruction of G6PD-deficient RBCs.50⇓-52 Mechanistic insights to understand this clinical enigma may help guide development of optimal prevention strategies for such cases.
In low-risk patients, such as those heavily transfused with no DHTR history, or episodically transfused with a low risk score (Figure 1), matching for the highly immunogenic Rh (D, C, E, c, e) and K antigens should be the standard of care. The risk of alloimmunization to Rh variants, however, remains high in all patients,18,19 and the decision to match for these variants needs to be determined on a case-by-case basis. We consider unnecessary, however, prophylactic extended matching (Fy, Jk, Ss) in patients who have antibodies against only Rh (D, C, E, c, e) and K, and no other DHTR risk. The scoring system also provides recommendations for prophylactic matching for patients with intermediate risk based on DHTR history and number of previous transfusions.
This is the first published stratification system for DHTR risk and prevention and thus needs to be validated by other institutions and in different patient cohorts including SCD pediatric patients. It can be implemented only if the patient transfusion history is known, a not-so-trivial hurdle because patients may receive transfusions at multiple hospitals. This prevention strategy based on the above DHTR risk criteria has been successfully implemented at a single French facility, with ongoing efforts to expand it nationally.