RENAL TRANSPLANTATION INTO HIGH RISK, HIGHLY SENSITIZED RECIPIENTS: A SINGLE CENTER EXPERIENCE

1
RENAL TRANSPLANTATION INTO HIGH RISK, HIGHLY
SENSITIZED RECIPIENTS A SINGLE CENTER EXPERIENCE

• Randy Hennigar PhD, MD
• Director, Nephropathology and Electron Microscopy
• Emory University Hospital
• Atlanta ,GA

2
Incidence of C4d in Renal Transplant Population
Emory University Hospital (EUH)

• Objective To gain more information about the
  role of antibody mediated rejection in the renal
  transplant population _at_ EUH.
• Method From Nov 2003 to Mar 2005, a total of
  313 consecutive biopsies (252 tx patients) were
  screened for C4d deposition. Bxs were performed
  for renal dysfunction.

3
Immunoperoxidase Staining for C4d
4
Incidence of C4d in Various Renal Tx Populations
Modified from Bohmig Regele, Transpl Int
16773, 2003
5
Incidence of C4d in Renal Transplant Population _at_
EUH

• Results 23 of 252 pts (9) were positive, using
  the criteria of Nickeleit and Mihatsch (Nephrol
  Dial Transpl 18 2232-2239, 2003).
• Conclusion The incidence of C4d deposition (and
  presumably antibody-mediated rejection) among the
  kidney transplant population at EUH appears less
  prevalent than that reported in the literature.

6
????
7
Emory University Hospital Renal Transplant
Center Activity (2004)

• Deceased donor txs 111 (74)
• Living donor txs 39 (26)
• Total 150
• Tx rate among waitlist pts 0.3
• From The Scientific Registry of Transplant
  Recipients

8
Emory University HospitalTransplant Recipient
Characteristics (2004)

• Ethnicity/race of waitlist pts (end of 2004)
• EUH() USA average()
• African-American 63 36
• White 32 39
• Hispanic/Latino 2 16
• Asian 3 8
• Other lt1 1
• From The Scientific Registry of
  Transplant Recipients

9
Emory University HospitalTransplant Recipient
Characteristics (2004)

• Ethnicity/race of tx patients (deceased donors)
• EUH() USA average()
• African-American 52 30
• White 45 49
• Hispanic/Latino 1 14
• Asian 2 6
• Other 0 2
• From The Scientific Registry
  of Transplant Patients

10
Panel Reactive Antibodies (PRA)

• A screening mechanism to determine the HLA
  antibody profile of potential transplant
  recipients.

• Periodic screening (monthly/quarterly) of
  recipient sera with a panel of HLA-typed cells.

• Sensitization of the recipient is expressed as
  the percentage of serum reactivity with the total
  panel. Typically, high PRA is indicative of a
  highly sensitized recipient- one who is at risk
  for early graft loss.

11
Deceased Donor Renal Transplants (1999 2004)
12
Emory University Hospital Peak PRA Prior to
Deceased Donor Renal Tx (2004)
From The Scientific Registry of Transplant
Recipients
13
Cadaveric Renal Allograft Survival (1998 2003)
99
100
97
Emory N gt500
90
94
93
90
UNOS N 20791
80
81
Graft Survival
70
60
50
3 mos
0
1
2
3
Years
UNOS/SRTR 2003
14
(No Transcript)
15
(No Transcript)
16
Evolution of HLA Antibody Detection
Cytotoxicity Enhanced
Cytotoxicity Flow Cytometry
Bray et al Immunol Res. 2941, 2004
17
From Gebel et al. Am J Transpl 31488-1500, 2003
18
From Gebel et al. Am J Transpl 31488-1500, 2003
19
Impact of HLA Antibodies Detected Only by Flow
Cytometric Crossmatch (Regrafts)Gebel et al. Am
J Transpl 31488-1500, 2003
20
In 2002, of the gt150 labs participating in the
ASHI-CAP class I crossmatch surveys (MX1-A, B,
C), only 6870 reported AHG augmented CDC and
4752 flow-based crossmatches.
21
From Gebel et al. Am J Transpl 31488-1500, 2003
22
Perceived Pitfalls of Flow Cytometry
Crossmatching (FCXM)

• Too sensitive
• Detection of low titer and noncomplement-fixing
  antibodies of little or no clinical relevance
• Would inappropriately deny a patient access to
  transplantion
• Does not reliably predict poor clinical outcomes

23
IgG FCXMRenal Allograft StudyFrequency of
rejection in a single center
44
40
n 56
n 41
81 vs 83 1 yr survival
rejection
FCXMs ARE IRRELEVANT!
IgG
Kerman et al Transplantation 681855-1858, 1999
24
In 2002, of the gt150 labs participating in the
ASHI-CAP class I crossmatch surveys (MX1-A, B,
C), only 6870 reported AHG augmented CDC and
4752 flow-based crossmatches.
25
Panel Reactive Antibodies (PRA)

• A screening mechanism to determine the HLA
  antibody profile of potential transplant
  recipients.

• Periodic screening (monthly/quarterly) of
  recipient sera with a panel of HLA typed cells.

• Sensitization of the recipient is expressed as
  the percentage of serum reactivity with the total
  panel. Typically, high PRA is indicative of a
  highly sensitized recipient- one who is at risk
  for early graft loss.

• Historically, PRA has been antigen-nonspecific.

26
METHODS FOR ANTIBODY EVALUATION
Complement-dependent Cytotoxicity (CDC) -
Direct CDC (Standard CDC) -
Modifications Washes Extended Incubation
Anti-human globulin (AHG-CDC) DTT /
DTE Flow Cytometry (cells) - T cell / B
cell - Pronase
ELISA - Yes / No - PRA (I II)
- Specificity (I II) FlowPRA Flow
cytometry using microparticles (beads) -
PRA (I and II ) - Specificity (I
II) Multi-plex - Suspension Arrays -
Protein Chips
27
Flow Microparticles
One Lambda www.onelambda.com
28
Solid Phase, Antigen-Specific Assays
Extract and Purify HLA Antigens
B cells EBV
Class I or II Phenotype or Individual Molecule
Flow Cytometry
Microparticles
Purified HLA Antigens
ELISA
29
Microparticles
ELISA
Coated with 30 HLA I or 30 HLA II antigens
30
Table 6. Flow PRA versus AHG-CDC PRA (n 203)
Flow PRA-Negative Flow PRA-Positive AHG-CDC
PRA gt10 2 7 AHG-CDC PRA lt10
160 34
31
PRA ANALYSIS BY DIFFERING METHODLOGIES
POSITIVE NEGATIVE
CDC 102 162
AHG-CDC 116 (13) 148
ELISA 127 (10) 137
FlowPRA 139 (10) 125
Gebel and Bray, Transplantation 691370-1374,
2000.
32
Positive FCXM are associated with graft loss when
FlowPRA detects high levels of HLA antibodies
8
30
20
Graft Survival
7
12
20
Bray RA, Nickerson PW, Kerman RH, Gebel HM.
Immunol Res. 2941, 2004
33
Renal Transplantation (DD) into High vs. Low PRA
Patients with Negative FCXM
N 372
N 492
N 120
Submitted for publication
34
Antibody Paradigms - 2005
Screening Crossmatch
Low Risk
High Risk
35
PRA

• PRA can be a qualitative and/or quantitative
• assessment of alloimmunization in transplant
• patients.
• Optimally, PRA testing should identify the
• specificity of an antibody and provide the
• transplantability index of a patient.
• More succinctly, PRA testing should correlate
• with the final crossmatch.

36
CLASS II DONOR SPECIFIC ANTIBODIES ARE PATHOGENIC
IN PRIMARY RENAL ALLOGRAFTS Nickerson et al AJT
4(8) 257, 2004
Impact of Donor Reactive HLA Antibodies
Rejection Time to Ab
mediated Time to




First Month Rejection
Graft Loss Graft Loss
Donor Reactive Class I 14/15 (93)
6 (1-17) 4 (27)
4 (1-14) Donor Reactive Class II 8/10
(80) 5 (2-7) 3 (30)
5 (2-9) HLA Ab (non-donor)
3/21 (14) 13 (13-19)
0 (0) NA
37
77 of positive B cell crossmatches ARE NOT DUE
to HLA antibodies!
38
Approaches
Pharmacological
Biological
Desensitization IVIG PP / IVIG Rituxan
Identical Sibling Xenotransplantation Acceptable
Mismatch - Detailed Antibody Analysis -
Comprehensive PRA - Virtual Crossmatch
39
Acceptable Mismatches
Putative Recipient A1, A30 B7, B8
DR11, 15 Antibodies - A2, 23, 24,
68 Potential Donor A25, A33 B42,
B18 DR12, DR13
40
Strategic Approaches
- Based on recognition that matching is not for
everyone- 85 of DD Txs are mismatched. -
Focus on appropriate mismatching rather than
looking for an HLA match. - Requires
detailed evaluation of the patients HLA
antibodies. - Shifts emphasis to antibody
evaluation and away from
crossmatching to identify acceptable
mismatches.
41
Desensitization Protocols Arent For Everyone
- High Titer HLA Antibodies
gt512 - Refractory Specificities DR52,
DR53 - Fragile Patients - Restricted to
Living Donors - s
42

• Recommendations to define the non-sensitized
  patient
• Validate patient history for the lack of
  sensitizing
• events.
• Confirm that a patient is nonsensitized using a
  solid
• phase assay documented to be more sensitive
  than
• CDC assays.

43

• Recommendations to evaluate the sensitized
  patient
• To optimize detection of low titer HLA
  antibodies,
• monitoring should be performed using sensitive
• solid-phase assays.
• Monitoring should include evaluation for both
• antibodies to class I and class II HLA
  antigens.
• A crossmatch test must be performed before
• transplantation using, as a minimum, an
  enhanced CDC
• technique.
• The final crossmatch technique should be of
  equal
• sensitivity to the solid-phase assay used to
  screen for the
• presence of HLA antibody.
• A B-cell crossmatch should be included in the
  final
• crossmatch.
• Peak sera should be included in the final
  crossmatch.
• Auto-crossmatches should be utilized to aid in
  the
• interpretation of allo-crossmatches.

44
END OF LECTURE