Transplantation of Immunologically High-risk

Recipients with Donor-specific Antibody but who are Crossmatch Negative

to Their Donors

Transplantation of Immunologically High-risk

Recipients with Donor-specific Antibody but who are Crossmatch Negative

to Their Donors

Ronald H. Kerman, PhDProfessor of Surgery

Director, Histocompatibility and Immune Evaluation Laboratory

Division of Immunology & Organ TransplantationThe University of Texas Medical School at Houston

We used to allocate donor organs

based upon the HLA antigen

match between donor and

recipient

We used to allocate donor organs

based upon the HLA antigen

match between donor and

recipient

We can now identify the presence of

HLA-Abs and their Ag specificities.

Can this information help in improving

pairing of donors to recipients?

Does the virtual XM work?

We can now identify the presence of

HLA-Abs and their Ag specificities.

Can this information help in improving

pairing of donors to recipients?

Does the virtual XM work?

Are All Antibodies Bad?Are All Antibodies Bad?

Pre-formed HLA Ab Adversely Affects:

Equity

• Prolongs wait-time for first or re-transplants• Disadvantages Women• Disadvantages African Americans

Survival

• Increased incidence of death while waiting

HLA antibodies are bad for transplant

recipients.

-Paul Terasaki

Non-HLA antibodies may also be bad

(anti-endothelial, vimentin, MICA, MICB

and others).

HLA antibodies are bad for transplant

recipients.

-Paul Terasaki

Non-HLA antibodies may also be bad

(anti-endothelial, vimentin, MICA, MICB

and others).

HLA antibodies instantly kill a kidney:

hyperacute rejection

State of preimmunization is detected by HLA antibodies

HLA antibodies are associated with acute early rejection

HLA antibodies instantly kill a kidney:

hyperacute rejection

State of preimmunization is detected by HLA antibodies

HLA antibodies are associated with acute early rejection

Lefaucheur et al. Am J Trans; 8:324, 2008Lefaucheur et al. Am J Trans; 8:324, 2008

8-Year Graft Survival

4-11

4-11 Banu Sis et al. ATC, 2010Banu Sis et al. ATC, 2010

DSA titer

SCr

FCXM

Rejection

DSA titer

SCr

FCXM

Rejection

Group 1

1:1024

1.4

Neg

No

Group 1

1:1024

1.4

Neg

No

Group 2

1:32

2.8

Pos

Yes

Group 2

1:32

2.8

Pos

Yes

Clinical Relevance of the XMClinical Relevance of the XM

Dolly Tyan (Stanford University)

MethodsC1q Assay Based on the Single Ag Bead Technology

MethodsC1q Assay Based on the Single Ag Bead Technology

Dolly Tyan (Stanford University)

High Specificity of C1q Assay

Are we trading the old allocation system

(with it’s problems) for a new allocation

system with problems of it’s own we do

not yet appreciate?

Are we trading the old allocation system

(with it’s problems) for a new allocation

system with problems of it’s own we do

not yet appreciate?

What is the real XM outcome for patients with identified donor-specific antigen that were ruled out because of the virtual crossmatch?

What is the frequency of these patients?

What is the real XM outcome for patients with identified donor-specific antigen that were ruled out because of the virtual crossmatch?

What is the frequency of these patients?

A positive (+) pretransplant (Tx) donor specific

crossmatch (XM) has been a contraindication to

transplant. Current testing methodologies allow for

performance of sensitive flow cytometry

crossmatches (FCXMs) and detection of IgG HLA

antibodies (Abs) and their antigen specificities.

Data obtained by these assays must be evaluated to

not only identify non-reactive recips but to determine

the clinical significance when recips display (+)

results.

A positive (+) pretransplant (Tx) donor specific

crossmatch (XM) has been a contraindication to

transplant. Current testing methodologies allow for

performance of sensitive flow cytometry

crossmatches (FCXMs) and detection of IgG HLA

antibodies (Abs) and their antigen specificities.

Data obtained by these assays must be evaluated to

not only identify non-reactive recips but to determine

the clinical significance when recips display (+)

results.

To understand the clinical correlation of these tests

we retrospectively evaluated Flow-PRA, FCXM, HLA

Ab specificities and Ab titers of 300 pre-Tx sera from

recips of deceased renal allograft donors

transplanted after negative (-)

AHG-XMs.

To understand the clinical correlation of these tests

we retrospectively evaluated Flow-PRA, FCXM, HLA

Ab specificities and Ab titers of 300 pre-Tx sera from

recips of deceased renal allograft donors

transplanted after negative (-)

AHG-XMs.

HLA Ab, Specificity, Titer and FCXMHLA Ab, Specificity, Titer and FCXMHLA AbHLA Ab

DSDS Non-DSNon-DS TiterTiter

FCXMFCXM

(+)(+) (-)(-)

1.1. 15*15* ++ ++ ≥256≥256 (+)(+)

4444 ++ ++ ≤16≤16 (-)(-)

1010 ++ ++ ≤16≤16 (+)(+)

2.2.

3.3.

2323 00 ++ ≥256≥256 (+)(+)5.5.

3535 00 ++ ≤16≤16 (-)(-)

3030 00 ++ ≤16≤16 (+)(+)

2222 00 ++ ≥256≥256 (-)(-)4.4.

6.6.

7.7.

9696 00 00 -- (-)(-)

2525 00 00 -- (+)(+)

8.8.

9.9.

NN

24 mo.GraftSurvival

24 mo.GraftSurvival

0%0%

91%91%

60%60%

74%74%

89%89%

74%74%

86%86%

95%95%

76%76%

2-yr GS of 91% for (-) vs 72% for (+) FCXM P<0.0012-yr GS of 91% for (-) vs 72% for (+) FCXM P<0.001

HLA Ab, Specificity, Titer and FCXMHLA Ab, Specificity, Titer and FCXM

HLA AbHLA Ab

DSDS Non-DSNon-DS TiterTiter

FCXMFCXM

(+)(+) (-)(-)

12 mo.GraftSurvival

12 mo.GraftSurvival

4444 ++ ++ ≤16≤16 (-)(-) 91%91%2.2.

NN

44/300 = 15% of total recipients

(+) DSA , but a (-) FCXM

44/54 ( 81.5% ) DSA (+) recips were XM (-)

44/300 = 15% of total recipients

(+) DSA , but a (-) FCXM

44/54 ( 81.5% ) DSA (+) recips were XM (-)

18 A-A; 11 women; 6 Hispanic (25/44 = 57%)18 A-A; 11 women; 6 Hispanic (25/44 = 57%)

HLA Ab, Specificity, Titer and FCXMHLA Ab, Specificity, Titer and FCXM

HLA AbHLA Ab

DSDS Non-DSNon-DS TiterTiter

FCXMFCXM

(+)(+) (-)(-)

12 mo.GraftSurvival

12 mo.GraftSurvival

2323 00 ++ ≥256≥256 (+)(+) 87%87%5.5.

3030 00 ++ ≤16≤16 (+)(+) 87%87%7.7.

2525 00 00 -- (+)(+) 88%88%9.9.

NN

78/300 = 26% of total recipients

(-) DSA but a (+) FCXM

78/231 ( 38% ) DSA (-) recips were XM (+)

78/300 = 26% of total recipients

(-) DSA but a (+) FCXM

78/231 ( 38% ) DSA (-) recips were XM (+)

ConclusionsConclusionsConclusionsConclusionsConclusionsConclusionsConclusionsConclusions

1. High titer DS-HLA Ab and a (+) FCXM are bad!

2. The presence of (low titer) DS-HLA Ab and a (-) FCXM is not a contraindication to transplant (vs a virtual crossmatch).

3. Absence of DS-HLA Ab does not guarantee a (-) FCXM.

1. High titer DS-HLA Ab and a (+) FCXM are bad!

2. The presence of (low titer) DS-HLA Ab and a (-) FCXM is not a contraindication to transplant (vs a virtual crossmatch).

3. Absence of DS-HLA Ab does not guarantee a (-) FCXM.

4. The binding of IgG to donor targets results in poor clinical outcome.

5. Negative FCXMs result in excellent clinical outcome (whether DS, non-DS HLA or non-HLA Abs are present).

4. The binding of IgG to donor targets results in poor clinical outcome.

5. Negative FCXMs result in excellent clinical outcome (whether DS, non-DS HLA or non-HLA Abs are present).

ATC 2007 Abstract: Am J Trans, 2007

Transplantation of DSA+/FCXM-Kidney Recipients

1. We previously reported the 85% 1-year graft survival for 26 DSA(+)/FCXM(-) renal allograft recipients treated with Basiliximab, Thymo, CNI, Steroids.

2. From January, 2008 we prospectively treated 33 DSA(+)/FCXM(-) renal allograft recipients with:

Thymoglobulin induction (7 – 14 days) 5 – 10 plasmapheresis followed by Rituxan/IVIgCyclosporine, MMF, Steroids

3. There were 15 male / 18 female recipients; 11 African-American, 4 Hispanic, 11 Caucasian, 2 Asian; 7 - 1o, 7 re-transplant recipients.

• In order to transplant these patients DSA is not listed for calculated PRA.

• Patients receive no points for PRA, but are crossmatched for every donor.

• If cytotoxic and flow crossmatches are negative, DSA protocol is initiated.

All recipients were ABO compatible and transplanted following negative AHG and FCXMs.

Sera tested included pre-transplant (days 0-7) highest historic PRA, and an intermediate serum of 3-6 months pre-transplant.

All but 1 recipient had positive DSA in pre-transplant and other prior sera.

1 recipient had positive DSA in a historic serum, but not pre-transplant.

Transplantation of DSA+/FCXM-Kidney Recipients

Results: post-transplant (2 – 37 months, mean 16.2):97% graft survival (32/33)100% patient survivalMean SCr of 1.5 mg/dL (n=32)1 Non-immunological graft loss at 21 monthsSCr of 8.7 mg/dL (n=1)

10 early rejections: 2 ACR, 8 AbMR (all C4d+ ) at 7 – 21 days post-transplant

2 delayed rejections at 8 months (ACR) and 12 months (mixed) post-transplant

Transplantation of DSA+/FCXM-Kidney Recipients

Pre-transplant Immune Studies:

1. 14 recipients with HLA Class I DSAs; titers from 1:2 – 1:128

DSA-specific MFIs of 1,275 – 7,213 (median MFI 2,386)

2. 19 recipients with HLA Class II DSAs; titers from 1:1 – 1:128

DSA-specific MFIs of 1,390 – 24,317 (median MFI 5,796)

The pre-transplant class of HLA Ab, DSA, DSA titers or

DSA MFI were not predictive of rejection or graft loss

(median MFI 3,408).

Post-Tx PRA and DSA Follow-up

N=33 (Follow-up of 2 to 30 mos. post-Tx)

5 - No DSA

10 - recips w/ class I DSA titers (1:1 - 1:64)

15 - recips w/ class II DSA titers (1:64 - 1:128)

2 year old African-American male, living-related transplant from mother. Managed with excellent renal function on cyclosporine / prednisone until auto accident led to loss of graft at age 17.

On wait-list 4 years with 95% PRA Class I and II.

Donor Antigen Specificities removed from UNOS listing. Within 1 week transplanted (10/15/08) with DSA(+), crossmatch(-) deceased donor.

Plasmapheresis x 1 week, thymoglobulin, CsA, MPA.

No rejection, creatinine 1.8 mg (2/8/11).

Conclusions

These results suggest that patients with surrogate HLA antigen-bead identified Ab may be successfully transplanted as long as the donor-specific FCXMs are negative and the patients receive aggressive immunosuppression.

These patients should not be excluded from transplantation (because of virtual crossmatch considerations) and may not need pre-transplant desensitization.

Graft Survival vs. CrossmatchGraft Survival vs. Crossmatch

Graft Survival vs. CrossmatchGraft Survival vs. Crossmatch

To Treat or Not to Treat ?

AbMR

To Treat or Not to Treat ?

Identifying Clinically Relevant Ab

1. During a clinical event or by biopsy

2. During a stable clinical course

3. Methods identifying the presence and specificity

of Abs do not characterize the function of the Ab.

4. The binding of an Ab in a XM is a functional assay

which may reflect the amount of Ab, the Ab

avidity for the target epitope or other factors.

Thoughts

The binding of patient Ab to Ag-coated beads is not a binding to donor Ag, but a third party, surrogate Ag.The fact that this might not correlate to crossmatch activity is not surprising. All Abs are not bad!We do not understand everything we see clinically. We have problems, but since they are identified we can solve them and take advantage of the new technology.

Thoughts

The binding of patient Ab to Ag-coated beads is not a binding to donor Ag, but a third party, surrogate Ag.The fact that this might not correlate to crossmatch activity is not surprising. All Abs are not bad!We do not understand everything we see clinically. We have problems, but since they are identified we can solve them and take advantage of the new technology.

Non-HLA AbsNon-HLA Abs

HLA Ab (Class I / Class II)HLA Ab (Class I / Class II)

Relevant ConsiderationsRelevant Considerations

(+) Donor-specific XM(+) Donor-specific XM

IgG AbIgG Ab

To identify the presence of clinically relevant Abs in recipient sera vs donor antigens(HLA, Non-HLA)

Determine antibody specificity (HLA, Non-HLA)

Determine antibody titer

XM by most sensitive method (FCXM)

Knowledge of the patient’s antibody status can help us to understand the crossmatch results

To identify the presence of clinically relevant Abs in recipient sera vs donor antigens(HLA, Non-HLA)

Determine antibody specificity (HLA, Non-HLA)

Determine antibody titer

XM by most sensitive method (FCXM)

Knowledge of the patient’s antibody status can help us to understand the crossmatch results

Role of the HLA Laboratory:Role of the HLA Laboratory: