Memory vs' De Novo HLA Antibodies: Relative Impact and Prevalence in Primary Renal Allografts with N

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Memory vs. De Novo HLA AntibodiesRelative
Impact and Prevalencein Primary Renal
Allograftswith Negative Pre-transplant AHG-CDC
Crossmatches
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Utility of FCXM in Renal Transplantation
Effect of T FCXM
T FCXM
T FCXM
CDC-XM
Population
Early Loss 22 vs. 7
18
40 channels
CDC
Primary
1987 Cook (n196)
Early Loss 33 vs. 7 1 year 67 vs. 85
18
40 channels
AHG
Primary
1990 Mahoney (n 67)
Early Loss 20 vs. 7 1 year 75 vs. 82
18
50 channels
CDC
Primary
1993 Ogura (n841)
1 year 75 vs. 86
7
50 channels
AHG
Primary
1996 LeFor (n214)
Rejection 67 vs. 51 1 year 86 vs. 98
18
40 channels
Amos
Primary
1997 Pelletier (n102) (No Difference)
Rejection 51 vs. 25 1 year 44 vs. 97
14
40 channels
Amos
Primary
1998 Kimball (n157)
Rejection 44 vs. 40 1 year 81 vs. 83
80 channels
AHG
Primary (Cadaveric)
1999 Kerman (n 97) (No Difference)
Early Loss 33 vs. 11
40 channels
AHG
Primary
2001 Karpinski (n 143)
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AHG-CDC -ve but FCXM ve Associated with Renal
Graft Loss when FlowPRA Detects HLA Antibodies
Graft Survival
H Gebel and R Kerman ATC Wash DC 2002
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Donor Reactive HLA Antibody Pre-TransplantManitob
a Experience ( 303 Primary Transplants Jun 92 -
May 03 )

• Donor Reactive
• HLA Antibody
• I (17)
• II (10)
• Controls
• (No Donor Reactive Ab)
• HLA Ab (19)

Ab Mediated Graft Loss 5 (29) 3 (30) 0 (
0)
Rejection lt 4 weeks 15 (88) 7 (80)
4 (21)
Time to Graft Loss 3 (1-12) 5 (2-9)
Time to Rejection 6 (1-19) 5 (2-9) 13
(8-19)
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Mechanisms of Antibody Mediated Rejection
Endothelium
Donor HLA
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Case I

• 65 yr Female, 1st renal transplant (living
  related (Son))
• History of pregnancies (x 3)
• Current / Historical Sera AHG PRA 0
• AHG-CDC CXM T-cell -ve B-cell -ve

• Flow CXM
• Current T-cell -ve B-cell -ve
• Remote T-cell -ve B-cell -ve
• Auto Flow CXM T-cell -ve B-cell -ve
• Flow PRA 0 class I 5 class II

• HLA Recipient A 26 32 B 27 56 DRB1 10 13
  DR52
• HLA Donor A 68 32 B 18 56 DRB1 15 13 DR51
  DR52

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Case I
FK506 MMF Prednisone
Biopsy
Creatinine (mmol/L)
FK Trough Levels
14.2
27.3
17.0
11.7
24.7
12.4
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C4d (Dr H. Regele, Vienna)
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C4d (Dr H. Regele, Vienna)
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Case I Bx 1

• 6 days post-Tx
• elevation of sCr from 79 to 160
• anti-donor HLA class II Ab (titre 1256)
• acute tubular injury
• mild glomerulitis (g1,i0,t0,v0)
• mild peritubular capillaritis
• C4d positive (Dr H. Regele, Vienna)
• interstitial hemorrhage in medulla
• focal ah1/2 (donor disease)
• (Bx marginal, 5 glomeruli, 2 arteries)

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Case I Bx 2

• clinicopathological diagnosis at day 6 post-Tx
  was AbAR
• treated with IVIg and pulse steroid
• sCr returned to baseline
• 5 weeks post-Tx protocol Bx
• anti-donor Ab titre now very low / undetectable
• graft function stable

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Case I Bx 2

• 5 weeks post-Tx protocol Bx
• anti-donor Ab titre now very low / undetectable
• graft function stable
• Banff borderline change (g0,i1,t1,v0)
• mild / moderate peritubular capillaritis
• C4d now unequivocally negative (Dr H. Regele,
  Vienna)

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Case I Bx 3

• 7 weeks post-Tx Bx
• clinically deydrated
• very mild Banff borderline change (g0,i1,t1,v0)
• no features of AbAR, peritubular capillaries
  unremarkable

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Lessons from Case I

• C4d went from unequivocally ve to unequivocally
  ve within 4 weeks, after IVIg therapy
• PTC inflammatory changes were still present when
  the C4d had turned ve
• PTC inflammatory changes settled down several
  weeks after the class II Ab was no longer detected

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Case II

• 32 yr Female, 1st renal transplant (living
  related)
• History of pregnancies (x 3)
• HLA Recipient A 24 31, B 27 51, DRB1 4 9
  DR53
• HLA Donor A 24 3, B 62 , DRB1 4 14
  DR52 DR53
• Current / Historical Sera AHG PRA 0
• AHG-CDC CXM T-cell -ve B-cell -ve

• Flow crossmatch
• Current T-cell -ve B-cell -ve (Sept 2002)
• Remote T-cell -ve B-cell ve (Mar 2000)
• Auto Flow CXM T-cell -ve B-cell -ve
• Flow PRA remote 20 class I 73 class II
• Flow Specificity A2, B57, B58 DR52

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Case II
FK506 MMF Prednisone
Simulect
Simulect
Creatinine (mmol/L)
10.4
6.6
14.9
11.3
8.9
10.4
10.4
6.5
FK Trough Levels
Days Post-Transplant
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Case II Bx 1

• 1 week post-Tx
• donor specific anti HLA class II Ab (1128)
• sCr mildly elevated
• ATN with regeneration
• acute Tx glomerulitis (g1) with thrombotic
  microangiopathy
• Banff borderline changes (i2,t1,v0)
• moderate peritubular capillaritis with PMNLs
• C4d unequivocally ve (Dr H. Regele)

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Case II Bx 2

• clinicopathological diagnosis at day 8 post-Tx
  was AbAR
• good response to IVIg and steroid
• 1 month post-Tx protocol Bx
• anti-donor Ab titre now down to 116
• graft function stable

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Case II Bx 2

• 1 month post-Tx protocol Bx
• anti-donor Ab titre now down to 116
• graft function stable
• Banff borderline change (g0,i2,t1,v0)
• minimal peritubular capillary changes
• C4d ve
• new-onset mild-to-moderate chronic
  tubulointerstitial damage (ci1,ct1)

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Case II Bx 3

• 2.5 months post-Tx
• mild elevation of sCr

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Case II Bx 3

• 2.5 months post-Tx
• mild elevation of sCr
• Banff borderline changes (g0,i0,t1,v0)
• severe peritubular capillaritis
• C4d ve
• chronic allograft nephropathy (grade I,
  ci1,ct1)

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Case II Bx 4

• 4 months post-Tx protocol Bx
• graft function stable

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Case II Bx 4

• 4 months post-Tx protocol Bx
• graft function stable
• Banff borderline changes (g0,i2,t1,v0)
• moderate peritubular capillaritis
• C4d ve
• chronic allograft nephropathy (grade I/II,
  ci2,ct2)

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Case II Bx 5

• 7 months post-Tx protocol Bx
• graft function stable

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Case II Bx 5

• 7 months post-Tx protocol Bx
• graft function stable
• Banff borderline changes (g0,i1,t1,v0)
• moderate peritubular capillaritis
• C4d ve
• chronic allograft nephropathy (grade I,
  ci1,ct1)

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Lessons from Case II

• by clinical, serologic and histologic criteria,
  this is AbAR due to donor-specific anti-HLA class
  II Ab
• C4d ve throughout the series of Bxs, ?
  alternate immunopathogenesis of AbAR
• anti-donor HLA class II Ab can cause early
  chronic allograft nephropathy
• PTC inflammatory changes may be the predominant
  histologic lesion in AbAR

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Case II SummaryPTC Inflammatory Changes

• 1 week g1,i2,t1,v0,cg0,mm0,ci0,ct0,cv0,ah0 g1
  with thrombotic microangiopathy, ATN, moderate
  PTC inflammation (ptc2)
• 1 month g0,i2,t1,v0,cg0,mm0,ci1,ct1,cv0,ah0
  mild PTC inflammation (ptc1)
• 2.5 months g0,i0,t1,v0,cg0,mm0,ci1,ct1,cv0,ah0
  severe PTC inflammation (ptc3)
• 4 months g0,i2,t1,v0,cg0,mm0,ci2,ct2,cv0,ah0
  moderate PTC inflammation (ptc2)
• 7 months g0,i1,t1,v0,cg0,mm0,ci1,ct1,cv0,ah0
  moderate PTC inflammation (ptc2)
• Proposal Banff type scoring of PTC inflammation,
  with objective criteria

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The Point of Banff

• Directing the eye / mind of the pathologist to
  the relevant areas of the Bx, which for acute
  rejection are
• glomerulitis g score
• interstitial inflammation i score
• lymphocytic tubulitis t score
• intimal arteritis v score
• By the same principle, in possible AbAR, a
  further relevant area is
• peritubular capillaritis - ? ptc score

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Proposal Banff ptc score

• ptc 0 no significant peritubular inflammatory
  changes
• ptc 1 peritubular capillary dilatation with 3
  to 4 luminal inflammatory cells
• ptc 2 peritubular capillary dilatation with 5
  to 10 luminal inflammatory cells
• ptc 3 peritubular capillary dilatation with gt10
  luminal inflammatory cells
• Inflammatory cells include PMNLs, mononuclear
  cells and macrophages

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Case II
g1i2t1 ptc2 ci0 ct0
i0t1 ptc3 ci1 ct1
i2t1 ptc1 ci1 ct1
i2t1 ptc2 ci2 ct2
i1t1 ptc2 ci1 ct1
Creatinine (mmol/L)
Days Post-Transplant
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What is the relative role of de novo Antibody
vs. Cellular effectors in mediating graft injury?
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METHODClinical Practice

• Pathologic Interpretation
• Protocol and Cause biopsies were interpreted
  using the Banff Schema
• Rejection was classified as Subclinical (lt10
  change baseline Cr) or Clinical
• C4d staining was performed as previously reported
  on paraffin sections (JASN 2001 122807) and read
  as positive when focal or diffuse linear staining
  of peri-tubular capillaries was present.

C4d staining of peri-tubular capillaries
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METHODStudy Populations

• AHG-CDC Cross-match -ve, Flow Cross-match ve
• 11 renal transplant patients who had evidence of
  donor specific antibody pre-transplant but who
  did not loose their grafts to accelerated
  rejection.
• Rationale This group was used to serve as a
  positive control group as they were clearly at
  risk for antibody mediated rejection
  post-transplant.

• AHG-CDC Cross-match -ve, Flow Cross-match -ve
• 17 renal transplant patients who had no evidence
  of donor specific antibody pre-transplant.
• Rationale This group was the group of interest
  in that any donor specific antibody detected via
  C4d staining post-transplant would most likely be
  de novo rather than a recall response.

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RESULTSPrevalence of C4d staining
Pre-Transplant Cross-match
Flow CXM ve
Flow CXM -ve

C4d ve Clinical/Pathologic Dx Normal Borderline
Subclinical Clinical
38 Focal/Diffuse C4dve 0/2 ( 0) 2/7
(29) 5/9 (56) 3/8 (38)
17 Focal/Diffuse C4dve 2/21 ( 9) 1/6
(17) 3/12 (25) 2/8 (25)
Plt0.05
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Conclusions

• Flow based techniques further emphasize the
  importance of memory in humoral rejection (both
  Class I and II)
• PTC inflammatory changes may, independent of C4d,
  reflect on-going humoral injury.
• Banff should consider a PTC score
• Significance of focal linear PTC C4d deposition
  in paraffin sections needs to be reassessed.