Presentation on theme: "HLA Typing for Blood Bankers"— Presentation transcript:
1 HLA Typing for Blood Bankers Kaaron Benson, M.D.Professor and Senior MemberDepartments of Oncologic Science, and Pathology and Cell BiologyH. Lee Moffitt Cancer Center
2 Introduction to the HLA System “HLA” – Human Leukocyte AntigensKey role in immunologic functionPart of the major histocompatibility complex (MHC)HLA antigens encoded for by genes from 6 primary loci on chromosome 6
3 Structure of HLA Class I and Class II Molecules HLA typing targetsN Engl J Med 2000;343:702.
9 New Nomenclature: April 2010 HLA-Cw becomes HLA-CCw*0202 now C*02:02, represents Cw2 AgColons (:) to delimit separate fieldsA*0301 becomes A*03:01A*2601 becomes A*26:01A*02 and A*92 to changeA*9201 becomes A*02:101A*9202 becomes A*02:102B*15 and B*95 to changeB*9501 becomes B*15:101B*9502 becomes B*15:102
10 Nomenclature Confusion: One Example HLA-DRB1*03:01HLA-DRB1*03HLA-DR17HLA-DR3This one allele could be written these four different ways.All four do not refer to the same thing.
11 GENES OF THE HUMAN MHC SYSTEM Chromosome 6Class II Class III Class IDP DQ DR B C AB1 A B1 A B1 B3/4/5 A C4 TNFDRB1*0401DRB1*0402DRB1*0403DRB1*0404DRB1*0405DRB1*0406B*0702B*0801B*1301B*1401B*1501A*0101A*0201A*0301
20 All dead = Red Score =8All alive = GreenScore = 1
21 HLA Typing by Molecular Methods ProsMore accurate and precise than serologye.g. DRB1 >300 alleles but only 17 serotypesBetter matching between pt and donorLess sample required, nonviable cellsWider variety of samples can be usedBecoming easier + automatedConsDoes not account for genes that are present but not expressed: Genotype ≠ PhenotypeRare alleles = growing list of ambiguities
30 SSO Pros and Cons Pros Cons Fairly rapid High volume High(er) resolutionReasonable cost/testConsSingle typing is more expensiveRequires more DNAExpensive equipment
31 Sequence-Based Typing (SBT) Gold standard for HLA typing Detects novel alleles Expensive Sequencers are costly (but other uses)Requires highly skilled technologistsCis/trans polymorphisms - ambiguities result - require additional testing
32 DNA Typing Resolution Low Intermediate High serologic equivalent DRB1*04, DRB1*13IntermediateDRB1*04:01 or *04:03 or *04:04DRB1*04:01/03/04Highallele levelDRB1*04:01DRB1*04:01/36/45/52/56
33 DNA Typing Resolution Low Intermediate High serologic equivalent DRB1*04, DRB1*13IntermediateDRB1*04:01 or *04:03 or *04:04DRB1*04:01/03/04Highallele levelDRB1*04:01DRB1*04:01/36/45/52/56ambiguities
34 High Resolution Typing Type to a single “common” alleleRare allele definition:For Class I < 1:50,000 allelesFor DRB1 < 1:100,000 allelesB*15:01, 35:01B*15:01/29/33/34, 35:01/40N/42
38 Confirmatory HLA Typing New sample must be collectedPatient: prior to final donor selectionDonor: prior to stem cell collectionBoth MRD and MUD transplantsNMDP donor typing counts as one typingOnly one typing must be at high resolutionLevel of typing (LR, IR, HR) decided by programMCC: IR/HR typing for original and repeat
39 Clinical Applications of HLA Typing Population studiesDisease associationsPharmacogenomicsPlatelet transfusionTRALI risk reductionTransplantationhematopoietic stem cellsolid organ
40 Population Studies: HLA Antigen Frequencies (%)
42 HLA-B27 and Ankylosing Spondylitis A.S.>90%B27+HLA-B*27~8% of population~2% of B*27+ dev AS
43 Ingelman-Sundberg M. Pharmacogenomic Biomarkers for Prediction of Severe Adverse Drug Reactions NEJM 2008;358:
44 Platelet Transfusion Platelet refractoriness: non-immune vs immune Non-immune refractorinessinfection, splenomeg, BMT, DIC, bleed, medsImmune platelet refractorinessHLA Abs (other Abs: plt-specific, drug-induced, ABO)HLA alloimmunization: due to allogeneic WBC exposure via prior transfusion or pregnancy (WBCs – cl. I and II, PLTs – cl. I only)Management: more PLTs vs. histocompat. PLTsPrevention: WBC-reduced blood
55 Solid Organ Transplantation ABO compatibility essentialOrgan size requirementsCold ischemia time: organ to recipientMedical urgency / time on waiting listHLA matching for pt Ab / donor Ag essentialHLA flow cytometric XM = standard of careRecipient serum and donor lymphsHLA compatibility beneficialRequired for renal transplants matching: graft survival, meds
56 Solid Organ Transplantation United Network for Organ Sharing (UNOS): federal contractHLA typing must be by molecular +/- serologic methodsHLA-A, B, C, DRB1, DRB3/4/5, DQB1 typingHLA-A, B, Bw4/6, Cw, DR51/52/53, DQ Ags reported+/-HLA-DPB1 typing for heart and/or lungsZero Ag MM (6/6) for HLA-A, B, DR Ags only“Zero Ag MM” for HLA-C, DQB1, DPB1
57 Transplant HLA and ABO Matching HLA ABOKidney No* YesLiver No YesHeart No YesLung No YesPancreas No* YesCornea No NoStem cell Yes No*HLA matching preferred but not required
58 Cadaver Kidney Transplants HLA-A+B+DR Mismatches (MM) HLA DNA Typing Review and Transplantation. Immunity, Vol. 14, 347–356, April, 2001
60 HSCT: Donor Selection HLA compatibility essential ABO compatibility not requiredSiblings: best chance for identity25% chance of matching any one sib~30% of patients have a matched sibMonozygotic twin not preferred?Other family members may match ptNo MRD? Search for MUD ASAP!
64 Case 2: Extended Family Typing Pt: A 1, 24; B 8, 48; DR 3, 7Sib 1: A 1, 2; B 8, 35; DR 2, 3Sib 2: A 2, 24; B35,48; DR 2, 7Sib 3: A 2, 24; B35,48; DR 2, 7
65 Case 2: Extended Family Typing Pt: A 1, 24; B 8, 48; DR 3, 7Sib 1: A 1, 2; B 8, 35; DR 2, 3Sib 2: A 2, 24; B35,48; DR 2, 7Sib 3: A 2, 24; B35,48; DR 2, 7
66 Case 2: Extended Family Typing Pt Sib Sib Sib 3
67 Case 2: Extended Family Typing Parent Parent 2Pt Sib Sib Sib 3
68 Case 2: Extended Family Typing perfect matchParent Parent 2Pt Sib Sib Sib 3
69 Unrelated Donor Searches HLA matched unrelated donors = MRDAvailable donors:NMDP: > 9.5 million volunteer donorsWorldwide: ~20 million totalCord blood: > 550,000Chance of finding A, B, DR match:~60-80% with one million donorsChance best:Cauc> Am. Indian> Hisp/Asian> Afr-AmerFebruary 2012
74 Molecular vs Serologic HLA Typing: Benefits of Better Matching Improved rate of engraftmentDecreased incidence/severity aGVHDDecreased incidence/severity cGVHDImproved rate of overall survivalSerologic typing sufficient for MRD?
75 Acceptable Mismatches? Locus important?A vs B vs C vs DRB1 > DQB1Marrow: A+DR MM worse than B+CPBSC: C MM worse than othersCord: C Ag MM increases TRMSpecific mismatches important?A*02:01 vs 02:02 vs 02:03No preformed anti-donor-specific HLA Ab (DSA)
76 HLA Typing Guidelines for HSC Transplantation Type patient for A, B, C, DRB1 +/- DQB1Type siblings for A, B, C, DRB1 +/- DQB1 (may screen with HLA-LR class I or II)No sibling donor (MRD)? Consider MUDNo MUD? Consider cord or MMUDMolecular methods preferredHigh resolution (allele matching)
77 HLA Typing Guidelines for HSC Transplantation Using URD Search all donor registries worldwideMatched URDmolecular HLA typing > serologychoose young, male / nonparous femaleconsider CMV status, donor/pt size, ABO/RhMismatched URDmismatch rare allelesethnic group matching preferredConsider cord blood donor
78 HLA-DPB1: Need to Match? Studies have suggested that DPB1 matching does not impact overall survivalDPB1 match increases relapse riskDPB1 mismatch increases aGVHD and TRMLack of tight DPB1 linkage with other locidecreases the ease of finding a DPB1 matchOnly ~20% of 10 of 10 matched transplants will be matched for DPB1“Permissive” mismatches?
79 HLA Alloantibodies in HSCT Presence of recipient HLA alloantibodies are not predictive of graft failureDonor-specific HLA Abs (DSA) are predictive of graft failure (e.g., recipient anti-A*02 and A*02 donor pair)HLA antibody evaluations should be a part of the routine workup for unrelated stem cell transplantationThe detection of donor-directed, HLA-specificalloantibodies in recipients of unrelated HCT ispredictive of graft failure. Blood 2010;115:
80 Non-Inherited Maternal Antigens (NIMA) HLA-A*HLA-B*HLA-DRB1*Patient02, 2418, 3501:01, 11:04UCB donor unit02, 32UCB donor’s mother07, 3501:01, 13:01HLA-A*24 is not carried by UCB donor but is carried by UCB donor’s mother and the pt; this is a NIMA-matched UCBT.Van Rood JJ, et al. Proc Natl Acad Sci USA 2009;106:19952.Rocha V, et al. Biol Blood Marrow Transplant 2012; July 17 Epub.
81 NIMA Mismatch HLA-A* HLA-B* HLA-DRB1* Patient 02, 11 18, 35 01:01, 11:04UCB donor unit02, 32UCB donor’s mother02, 2407, 3501:01, 13:01HLA-A*11 is not carried by UCB donor or the UCB donor’s mother; this is a NIMA-mismatched UCBT.
84 HLA Typing Summary HLA typing nomenclature Low, intermediate, high resolution typingClinical application of HLA typingRole in TRALI risk mitigationImportant role in donor selection for solid organ transplant and HSCTOptimal matching between pt and donor