Histocompatibility Evaluation for Sensitized Patients: Maximizing the Patient’s Benefit.

1. Histocompatibility Evaluation for Sensitized Patients: Maximizing the Patient’s Benefit. Marcelo Pando UNOS Region 5 Collaborative March, 2017

2. Disclosure Relevant Financial Relationship • None Off Label Usage • None

3. Learning Objectives • Understand the complexity of the HLA system. • Discuss some of the methodologies we use to detect antibodies against HLA. • Understand the current HLA practice in solid organ transplantation. • Discuss some educational cases.

4. The Human Leukocyte Antigen In Medicine: In Nature: Ag presentation Organ Transplantation DIVERSITY! Pregnancy Disease association HLA Interaction with KIR Drug hypersensitivity Mate selection!

5. Then… • HLA in Transplantation.

6. The First Crossmatch

7. Crossmatch by complement-dependent cytotoxicity (CDC): Donor cells + CFDA Patient serum Complement EtBr

8. Crossmatch by Complement-Dependent Cytotoxicity (CDC): NEGATIVE POSITIVE

9. The HLA System: Antigens and alleles • HLA-A • Serology: A2 • Molecular: A*02:01 • Alleles of A2: A*02:01, A*02:02, A*02:03, A*02:04…… A*02:336

10. HLA Epitopes  AA Pos.                    10         20         30         40         50         60         70         80         90        100  A*01:01:01:01      GSHSMRYFFT SVSRPGRGEP RFIAVGYVDD TQFVRFDSDA ASQKMEPRAP WIEQEGPEYW DQETRNMKAH SQTDRANLGT LRGYYNQSED GSHTIQIMYG  A*02:01:01:01      ---------- ---------- ---------- ---------- ---R------ ---------- -G---KV--- ---H-VD--- ---------A ----V-R---  AA Pos.                   110        120        130        140        150        160        170        180        190        200  A*01:01:01:01      CDVGPDGRFL RGYRQDAYDG KDYIALNEDL RSWTAADMAA QITKRKWEAV HAAEQRRVYL EGRCVDGLRR YLENGKETLQ RTDPPKTHMT HHPISDHEAT  A*02:01:01:01      ----S-W--- ---H-Y---- ------K--- ---------- -T--H----A -V---L-A-- --T--EW--- ---------- ---A------ --AV------  AA Pos.                   210        220        230        240        250        260        270        280        290        300  A*01:01:01:01      LRCWALGFYP AEITLTWQRD GEDQTQDTEL VETRPAGDGT FQKWAAVVVP SGEEQRYTCH VQHEGLPKPL TLRWELSSQP TIPIVGIIAG LVLLGAVITG  A*02:01:01:01      ------S--- ---------- ---------- ---------- ---------- --Q------- ---------- -----P---- ---------- ---F------  AA Pos.                   310        320        330        340  A*01:01:01:01      AVVAAVMWRR KSSDRKGGSY TQAASSDSAQ GSDVSLTACK V  A*02:01:01:01      ---------- ---------- S--------- ---------- -  AA Pos    10         20         30         40         50A*01      GSHSMRYFFT SVSRPGRGEP RFIAVGYVDD TQFVRFDSDA ASQKMEPRAPA*02      ---------- ---------- ---------- ---------- ---R------ AA Pos.   110        120        130        140        150A*01      CDVGPDGRFL RGYRQDAYDG KDYIALNEDL RSWTAADMAA QITKRKWEAVA*02      ----S-W--- ---H-Y---- ------K--- ---------- -T--H----A AA Pos.   210        220        230        240        250A*01      LRCWALGFYP AEITLTWQRD GEDQTQDTEL VETRPAGDGT FQKWAAVVVPA*02      ------S--- ---------- ---------- ---------- ---------- AA Pos.   310        320        330        340 A*01      AVVAAVMWRR KSSDRKGGSY TQAASSDSAQ GSDVSLTACK V A*02      ---------- ---------- S--------- ---------- - 1 5 4 3 2 6 α1 β1 β2 α2

11. HLA Epitopes: The Bw4/Bw6 Case AA Pos. 10 20 30 40 50 60 70 80 90 100 B*07:02:01 GSHSMRYFYT SVSRPGRGEP RFISVGYVDD TQFVRFDSDA ASPREEPRAP WIEQEGPEYW DRNTQIYKAQ AQTDRESLRN LRGYYNQSEA GSHTLQSMYG B*13:01:01 ---------- AM-------- ---T------ ---------- T---MA---- ---------- --E---S-TN T--Y--N--T ALR------- ---II-R--- B*13:02:01 ---------- AM-------- ---T------ ---------- T---MA---- ---------- --E---S-TN T--Y--N--T ALR------- ----W-T--- B*15:13:01 ---------- AM-------- ---A------ ---------- ----MA---- ---------- ------S-TN T--Y--N--I ALR------- ---II-R--- B*15:16:01 ---F------ AM-------- ---A------ ---------- ----MA---- ---------- --E-RNM--S ---Y--N--I ALR------- ----W-R--- B*44:02:01:01 ---------- AM-------- ---T------ -L-------- T---K----- ---------- --E---S-TN T--Y--N--T ALR------- ---II-R--- B*14:01:01 ---------- ---------- ---------- ---------- ---------- ---------- ------C-TN T--------- ---------- ------W--- B*14:02:01 ---------- A--------- ---------- ---------- ---------- ---------- ------C-TN T--------- ---------- ------W--- B*15:01:01:01 ---------- AM-------- ---A------ ---------- ----MA---- ---------- --E---S-TN T--Y------ ---------- ------R--- B*15:02:01 ---------- AM-------- ---A------ ---------- ----MA---- ---------- ------S-TN T--Y------ ---------- ---II-R--- B*15:03:01 ---------- AM-------- ---------- ---------- ---------- ---------- --E---S-TN T--Y------ ---------- ------R--- B*15:10:01 ---------- AM-------- ---------- ---------- ---------- ---------- ------C-TN T--Y------ ---------- ------R--- B*15:11:01 ---------- AM-------- ---A------ ---------- ----MA---- ---------- --------TN T--Y------ ---------- ------R--- B*15:11:02 *--------- AM-------- ---A------ ---------- ----MA---- ---------- --------TN T--Y------ ---------- ------R--- B*15:11:03 *--------- AM-------- ---A------ ---------- ----MA---- ---------- --------TN T--Y------ ---------- ------R---

12. The Dogma of AMR • Antibody to the Donor • Antibody Mediated Rejection Assumptions: • Antibody is specific to the donor • The test detects clinical relevant antibodies • There is enough antibody to trigger IR

13. The Dogma of AMR DSA AMR

14. Tests for Antibody Screening

15. Luminex: Two different targets • PRA Beads • Phenotype: A2, A3; B7, B44; Cw4, Cw7 • Less sensitive • More stable • SAB • SA: A2 • High sensitivity • Less stable = denature Ags = False Positives

16. SAB

17. Phenotype Beads (PRA Beads) AECI294 AECI294

18. Calculated Panel of Reactive Antibodies (cPRA) • cPRA: • the percentage of donors that will be excluded • probability of having a positive crossmatch • likelihood of getting an incompatible organ • 100-cPRA = probability of getting a compatible organ

19. Examples of cPRA • Patient with 20% cPRA • 20% of donors will be excluded • 20% of crossmatches will be positive • 80% probability of getting a compatible organ

20. Maximizing Successful Offers The CPRA game Likelihood of Compatible Donor

21. Allele Specific Antibody DRB3*02

22. CPRA

23. Single Antigen Bead (SAB) test results:

24. Case 1: 100% cPRA patient

25. Case 2: 100% cPRA patient

26. Unacceptable antigens • Case 1: • A1 A2 A3 A11 A23 A24 A25 A26 A29 A36 A34 A43 A66 A68 A69 A80 • B13 B27 B2708 B44 B58 B60 B61 B7 B73 B76 B8 B81 B57 • DR4 • DQ7 DQ8 DQ9 • Case 2: • A1 A3 A11 A2403 A25 A26 A29 A30 A32 A33 A34 A36 A43 A66 A68 A69 A74 A80 • B7 B8 B13 B27 B2708 B42 B45 B49 B50 B56 B57 B58 B62 B63 B67 B71 B72 B73 B75 B76 B77 B81 • Cw1 Cw5 Cw6 Cw7 Cw8 Cw12 Cw14 Cw16 Cw17 Cw18 • DR1 DR4 DR7 DR8 DR10 DR103 DR11 DRB1*13:03 DRB1*14:02 DR15 DR16 • DRw51 DRw52 • DQB1*06:03 DQ2 DQ5 DQ7 DQA1*04:01 • DPB1*01:01 DPB1*04:01 DPB1*05:01 DPB1*19:01

27. cPRA • Case 1: • A1 A2 A3 A11 A23 A24 A25 A26 A29 A36 A34 A43 A66 A68 A69 A80 • B13 B27 B2708 B44 B58 B60 B61 B7 B73 B76 B8 B81 B57 • DR4 • DQ7 DQ8 DQ9 • Case 2: • A1 A3 A11 A2403 A25 A26 A29 A30 A32 A33 A34 A36 A43 A66 A68 A69 A74 A80 • B7 B8 B13 B27 B2708 B42 B45 B49 B50 B56 B57 B58 B62 B63 B67 B71 B72 B73 B75 B76 B77 B81 • Cw1 Cw5 Cw6 Cw7 Cw8 Cw12 Cw14 Cw16 Cw17 Cw18 • DR1 DR4 DR7 DR8 DR10 DR103 DR11 DRB1*13:03 DRB1*14:02 DR15 DR16 • DRw51 DRw52 • DQB1*06:03 DQ2 DQ5 DQ7 DQA1*04:01 • DPB1*01:01 DPB1*04:01 DPB1*05:01 DPB1*19:01 100% 99.5069% 100% 99.9971%

28. cPRA • Case 1: • Case 2: Probability of 1: 203 donors 100% 99.5069% 100% 99.9971% Probability of 1: 34,483 donors

29. Maximizing Successful Offers Unacceptable Ags A2: 48% CPRA

30. Summary • An accurate definition of a antibody profile can give more opportunities to waiting list patients. • Determining the list of HLA antigens to avoid is patient specific and depends on several factors. • Highly sensitized patients, particularly the 100% cPRA, needs especial assessment to maximize their opportunities. • These patients, although a minority in the waiting list, have enormous differences in the opportunity of getting a compatible organ.

31. Thanks!