Transcript B Cell

SENSITIZATION:
Management Strategies
Khalid Al Meshari, MD, FACP
Director
Renal and Pancreas Transplant Program
King Faisal Specialist Hospital
and Research Centre
Layout
1. Definition
2. Scope of the problem
3. Detection techniques
4. Management Strategies, KFSH experience
5. Future
Layout
Definition
Sensitization
1. Sensitization is defined as the presence of
clinically relevant Abs in recipient sera
against donor’ antigens
2. Relevance is determined by the reactivity of
these Abs with donor’ lymphocytes
expressing these antigens (positive CXM)
Sensitization
1. methods:
NIH-CDC, AHG-CDC
Flow Cytometry
Solid phase
2. How much Ab:
Titer (1:2, 1:4, 1:8)
Fluorescence intensity (MFI)
Molecule of equivalent soluble fluorescence (MESF)
Clinical Relevance of the Detection of
HLA Antibody by Different Techniques
T-Cell (Class I)
B-Cell (Class II)
CDC (IgG)
Hyperacute
rejection
Flow (IgG)
Acute antibody
Acute antibody
‘mediated’ rejection ‘mediated’ rejection
Single antigen
beads
Relevance
unknown
*Hyperacute rejection – if very high titre only
Acute antibody
‘mediated’ rejection*
Relevance unknown
Immunological Memory:
A Barrier to Transplant
Decreased chance of
receiving kidney
Majority of those waiting
longer are women
35
1
30
0.8
25
0.6
%
0.4
20
15
10
0.2
0
5
0
No sensitization
Sensitized
Odds
No sensitization
Men
Sensitized
Women
Association of Preformed PRA with
Various Characteristics of Patients
Preformed PRA
Characteristics
Proportion
female
Proportion with
retransplant
Mean (SE)
pretransplant
blood
transfusions
None
(n=3001)
1-50%
(n=803)
>50%
(n=244)
P
1014 (34%)
361 (45%)
154 (53%)
<0.0001
159 (5%)
112 (14%)
97 (40%)
<0.0001
3.47 (0.15)
6.01 (0.43) 10.7 (1.12)
<0.0001
Opelz G. Lancet 2005;365:1570-76
TARGET ANTIGENS
HLA or HLA-related antigens
Major MHC antigens
MHC class I (HLA-A, B, C)
MHC class II (HLA-DR, DP, DQ)
Minor MHC antigens
MHC Class I polypeptide related sequences A (MICA) and B (MICB)
Non-HLA related antigens
Angiotensin II type 1 receptor
Endothelium/monocyte antigens
Perlecan
Collagen Type IV, and VI
Agrin
Vimentin
Myosin
ABO blood group antigens
Antigens Targeted by Anti-Donor
Antibodies
ABO
MHC
Antigen Class II
Polymorphism:
MHC
Class I
Minor
Tissue
HLA
Specific
Antigens Antigens
Limited
Very High
Very High
Limited
Varies
Vasc. Endothelium
+
+
+
+
+
Parenchyma
+
-
+
+
+
APC
+
+
+
+
-
T Cells
+
-
+
+
-
Antigens Targeted by Anti-Donor
Antibodies
ABO
MHC
Antigen Class II
Polymorphism
MHC
Class I
Limited
Very High
Very High
Vasc. Endothelium
+
+
+
Parenchyma
+
-
+
APC
+
+
+
T Cells
+
-
+
HLA Antibodies
MHC
Class II
Polymorphism:
MHC
Class I
Very High
Very High
Vascular
Endothelium
+
+
Parenchyma
-
+
APC
+
+
T Cells
-
+
Humoral Alloreactivity
Pre Transplant
Preformed Ab´s
historic
current
time
Humoral Alloreactivity
Post Transplant
De novo Ab´s
de novo
time
Humoral Alloreactivity
Pre Tx
Preformed Ab’s
Historic
Current
Post Tx
De novo Ab’s
Memory
Response
De novo
Time
But What Really is De Novo
De novo
Time
De-Novo Anti-HLA Antibody
may result from…
Pre-transplant antibody present at low levels
Recall memory response
Inadequate immunosuppression
Inflammation
Pathways to HLA Sensitization
•
•
•
•
•
•
•
•
Pregnancies
Previous transplant
Blood transfusions
Use of tissue allografts for vascular reconstruction
in congenital heart surgery
CMV infection
Bacterial infections
Vaccination
Left ventricular assist devices
Antigens Targeted by Anti-Donor
Antibodies
ABO
Antigen
Polymorphism
Limited
Vasc. Endothelium
+
Parenchyma
+
APC
+
T Cells
+
The ABO Blood Group Antigens
NAG
Precursor
oligosaccharide
Gal
H
NAG
Gal
H antigen
Gal
A
B
NAG
A antigen
Gal NAGA
NAG
Fuc
Gal NAGA
Fuc
B antigen
Blood group
(phenotype)
Genotypes
Antigens
Antibodies to ABO
in serum
A
AA, AO
A
Anti-B
B
BB, BO
B
Anti-A
AB
AB
A and B
None
O
OO
H
Anti-A and anti-B
Complement-Fixing Antibodies
The structure of IgM
Antigen
Idiotype
Epitope
Fab
(variable region)
C1q-binding region
J Chain
C3b- and C4bbinding region
CH2
CH2
CH3
CH3
FC
Fc receptor types I, II, and III
Antibodies involved in Pre-transplant
Humoral Allo-immunity
Antigenic
target
Type of
Antigen
Antibody
Sensitizing events
MHC
Protein
IgG or mix of IgG
and IgM
Previous transplant,
pregnancy, or
transfusion
ABO
Carbohydrate
IgM or mix of IgM
and IgG
“Natural” (crossreaction with gut
bacterial flora)
Nature of the Alloantibody
Response
-HLA
Isoagg
Previous
exposure
T-cell
help
Repertoire
Clonality
YES
YES
Extensive
Polyclonal
Limited
Polyclonal
Naturally No autoproduced reactive
T-cell?
Layout
Scope of the problem
Defining the Scope of HLA
Sensitization
• 34% of USA waiting list is sensitized
• Frequency of highly sensitized patients (14%) is
increasing
– Increasing number waiting for regraft
– More sensitive techniques for detecting Ab
• Some highly sensitized patients will never be
transplanted
• Many sensitized patients have living donors who
are eliminated because of a positive XM
Patients on the UNOS Wait-List 31 JAN ‘05
vs. Patients Receiving Deceased Donor
Grafts Dec ’99 to NOV ‘04
Current PRA%
0 – 19%
20 – 79%
80 – 100%
Waiting #
44,889
7,934
5,754
77%
13%
10%
%
23%
Transplanted
#
34,036
3,613
1,882
%
86%
9%
5%
14%
OPTN 18 FEB 2005
Waiting Time (days)
Waiting Times for Sensitized
Renal Candidates
3000
0-9
10-79
80+
2500
2000
1500
1000
500
0
97-98
www.optn.org July 2006
99-00
Years Listed
01-02
<half transplanted
Reasons Patients Removed From UNOS Wait-list
Dec 1999 to Nov 2004
Current PRA%
0-19%
20-79%
80-100%
D D Transplant
47% (33,998)
42% (3,611)
37% (1,879)
Living Donor Transplant
20% (14,853)
12% (1,002)
5% ( 276)
Condition Deteriorated
4% ( 3,065)
7% ( 561)
8% ( 409)
Died
17% (12,290)
27% (2,292)
37% (1,906)
6% (4,216)
1% ( 401)
2% (1,221)
3% (2,302)
<1% ( 243)
7% ( 595)
<1% ( 61)
3% ( 235)
2% ( 141)
<1% ( 24)
7% ( 383)
1% ( 55)
3% ( 147)
1% ( 55)
<1% ( 15)
Other
• Not clarified
• Medically unsuitable
• Refused Transplant
• Tx’d Kid-Pan or Panc
• Condition Improved
OPTN 25 Feb 2005
Defining Scope of ABO
Incompatibility
• Based on blood group frequencies in the USA there is a
35% chance that any 2 individuals will be ABO
incompatible
• Blood type incompatible recipients have natural occurring
antibodies against carbohydrate epitopes on disparate
blood group molecules which can result in hyperacute
rejection
• Up to 1/3 of potential live donors are excluded on the
basis of ABO incompatibility
• About 1500 patients/yr in the US present with a willing but
ABO incompatible live donor
Kidney Median Waiting Times
by ABO
2500
97-98
99-00
01-02
2000
1500
1000
500
0
O
A
B
OPTN/UNOS Data as of 5-14-06
AB
Immunologic Barriers
• Sensitization to HLA
–
–
–
–
Increases waiting time
Associated with reduced graft survival
Affects most, if not all transplanted organs
Greater impact on women
• ABO
– Increases waiting time
– Affects differ among ethnic groups because of
differences in ABO distribution
– ABOi Tx can experience hyperacute rejection
Problems with Sensitized Patients
• Hyperacute Rejection
• Primary Nonfunction
• Delayed Function
• Accelerated Humoral Rejection
– Not easily controlled by drugs
– May cause lasting damage
• Acute Cellular Rejection
• Chronic Rejection
Correlation of Immediate Graft
Failures with Crossmatch Test
80% loss if XM+
15 loss if XM180
Patients
150
42% loss if Ab+
2% loss if Ab-
4
27
120
90
164
60
34
30
48
36
18
4
24
23
9
0
0
Preformed antibodies
No preformed
antibodies
Random donor lymphocytes
6
+XM
-XM
Donor lymphocytes
Patel, Terasaki. NEJM 280: 1969
1-year graft survival analysis of kidney transplants
from cadaver donors in relation to PRA
Opelz G, Lancet 2005 ;365:1570-6
Opelz G. Lancet 2005;365:1570-76
10-Year follow-up of kidney grafts
from HLA-identical sibling donors
Death censored functional graft survival of
HLA-identical sibling transplants
KFSH DD List
60
58
Listed patients
HSP
50
40
37
32
30
20
23
17 53%
10
0
40%
16 43%
5
2
O
A
B
AB
40%
KFSH DD List
150
Listed patients
HSP
132
125
100
75
50
25
0
58
44%
Layout
Detection techniques
Antibody
Mediated Rejection
Endothelium
PMN
Anti-HLA Ab
C1
Fc Receptor
Mediated Binding
C’ Activation
ADCC
CDC
C4
C4a + C4b
Donor HLA
MAC
Platelet microthrombi
C4d
Evolution of Detection Techniques
• 1999
–
–
–
–
T-AHG-CDC screening
No solid phase assays
Many important DSA not detected
AMR histology not well characterized
2009
Transplant Programs have seen a revolution in technology
•
•
•
•
•
HLA typing
HLA Ab Screen
HLA Ab Specificity
Donor Specific HLA Ab
C4d staining
→ Molecular (low to high (allele) resolution)
→ Solid Phase (increase sensitivity)
→ Solid Phase (increase in resolution)
→ Flow Crossmatch (increase sensitivity)
→ More accurate Diagnosis of AMR
Some issues to consider next:
• Needs to standardize HLA Ab quantitation
• Further studies needed to define relative risks of low levels of donor specific
antibodies (e.g. detected by solid phase only)
• Further studies needed to validate significance of HLA Cw, DP, DQ
antibodies, and non-HLA Ab (e.g. MICA, MICB)
Stages of Humoral Alloreactivity
Banff Conference 2001/2003
Humoral Rejection Conference, NIH, April 2003
I. Latent Humoral Response
Circulating
Antibody
II. Silent Humoral Rejection
Circulating
C4d
Antibody deposition
Accommodation?
Pre-rejection?
Circulating
C4d
III. Subclinical Humoral Rejection Antibody deposition
Tissue
Injury
Circulating
C4d
Antibody deposition
Tissue
Injury
IV. Humoral Rejection
Graft
Dysfunction
Takemoto SK, et al. Am J Transplant. 2004; 4: 1033-1041 (with permission from E. Akalin)
Layout
Management Strategies
Management Strategies
Matching Strategies
• Regional or national sharing
of crossmatch trays
• Mandatory sharing of
0-antigen mismatched
kidneys
• Acceptable mismatch /
virtual crossmatch / HLA
matchmaker
• Paired kidney exchange
Desensitization
Diagnostics
• Flow or Luminex® PRA
• PD flow crossmatch
• HLASA by solid phase assay
• C4d staining
Therapeutics
• Apheresis
• IVIG
• Rituximab
• Thymoglobuline®
Acceptable Mismatch
Recognition that matching is not for everyone.
85% of DD Txs are mismatched.
Focus on appropriate mismatching rather than
looking for a perfect HLA “match”.
Shift emphasis to antibody evaluation for the
identification of “acceptable mismatches”.
Requires detailed evaluation of the patient’s HLA
antibody specificities.
Exposure to IPA and NIMA During
Pregnancy Can Lead to Either
Immunization or Tolerization
Immunization
Adult exposure to IPA
Maternal site
NIMA/IMA
Tolerance
Fetal exposure to NIMA
Possible mechanisms
Chimerism
Privileged site: modulation of APC
Soluble HLA
Immune deviation
Regulatory T cells: HLA-DR sharing?
Anti-HLA Ab towards paternal
HLA molecules
Non-Inherited Maternal HLA Antigens
are Often “Acceptable” Mismatches
NIMA
NIPA
Antibodies
46
72
No antibodies
43
6
p<0.001
Donor Selection on Basis of
Acceptable Mismatches
Patient:
A24,31; B27,51; DR4,- and 100% PRA
AM:
A25, A26, B44
Possible kidney donors:
A25, A31; B27, B51; DR4
A26, A31; B27, B51; DR4
A24, A25; B27, B51; DR4
A24, A26; B27, B51; DR4
A24, A31; B44, B51; DR4
A24, A31; B27, B44; DR4
A25, A31; B44, B51; DR4
A26, A31; B44, B51; DR4
A25, A31; B27, B44; DR4
A26, A31; B27, B44; DR4
A24, A25; B44, B51; DR4
A24, A26; B44, B51; DR4
A25, A31; B27, B44; DR4 etc.
Current Approaches to Determine
Acceptable Mismatches
• Cross-matches with a patient specific panel.
(Leiden)
• Use of single antigen expressing cell lines (SALs)
(Leiden)
• Consider non-inherited maternal HLA antigens
• Single antigen beads
• HLA Matchmaker program
HLA Matchmaker
• The HLA Type of the antibody producer
determines what structural components of an
immunizing HLA antigen that will be seen as nonself
• Donor HLA-A, B mismatches are defined by
triplets of amino acid residues (epitopes) on
alloantibody-accessible sites of HLA molecules
HLA Matching on the
Triplet Level
Donor mm B18
Patient
B7
Immune system of the recipient recognizes:
A single HLA mismatch or 11 triplet mismatches
HLA Matching on the
Triplet Level
Donor mm
B18
Patient B7
B52
A33
Immune system of the recipient recognizes:
No triplet mismatches !
Allocation of Cadaver Kidneys to
Highly Sensitized Patients
Class et al. Transplantation 2004;78:190-193
40
35
30
25
20
AM
15
ETKAS
10
5
0
<6
>6
>12
>24
>36
Waiting time in months
AM: allocation through the Acceptable Mismatch Program (n=57)
KAS: Kidney Allocation System
ET: allocation through the standard Eurotransplant system (n=11)
Graft Survival
Class et al. Transplantation 2004;78:190-193
100
80
60
AM
40
<5% PRA
5-85% PRA
20
>85% PRA
0
12
24
Post Tx [m]
Comparison of graft survival between acceptably mismatched patients (n=112) and
nonsensitized (5% panel reactive antibody (PRA), sensitized (6%-84% PRA), and HS (85% PRA)
recipients whose transplants were arranged through the ET-KAS (14.328). GS, graft survival.
Virtual Crossmatch
Virtual Crossmatching:
• Determining the presence of donorspecific HLA antibodies (DSA) without
performing a crossmatch, by comparing
the HLA type of the donor with the HLA
antibody specificities of the recipient.
Emory Algorithm for Evaluating the Sensitized Patient
AJT Oct. 2006
Depictions of Kidney Paired Donations
Conventional Kidney Paired Donation
Donor
Recipient
Blood
Type A
Blood
Type B
Intended Pairs
Blood Type Incompatible
Blood
Type B
Blood
Type A
Unconventional Kidney Paired Donation
Donor
Recipient
Blood
Type A
Blood
Type O
Blood
Type O
Blood
Type A
Positive Cross-match
Matched Pairing
Domino Paired Donation
Donor 1: NDD
incompatible
Donor 2
Recipient 1
Recipient 2
1st eligible recipient
From UNOS match run
Montgomery et al. Lancet. 2006; 368: 419
NEAD Chains
Donor 1: NDD
incompatible
Donor 2
Recipient 1
incompatible
Donor X
Recipient X
A Nonsimultaneous,extended,Altruistic-Donor Chain
Rees M, etal. New Eng J Med 2009;360 (11):1096-1101
1
2
3
4
5
6
7
8
9
10
Transplant Date:
JUL 2007
JUL 2007
SEP 2007
SEP 2007
FEB 2008
FEB 2008
FEB 2008
FEB 2008
MAR 2008
MAR 2008
Recipient’s State:
AZ
OH
OH
OH
MD
MD
MD
NC
MD
OH
O
O
A
A
B
A
A
A
A
A
O
A
A
B
A
A
A
AB
A
AB
52%
0%
23%
0%
82%
78%
64%
3%
100%
46%
Cauc
Cauc
Cauc
Cauc
Cauc
Hispanic
Cauc
Cauc
Cauc
AA
Brother
Sister
Husband
Wife
Daughter
Father
Wife
Husband
Friend
Friend
Brother
Brother
Mother
Daughter
Recipient’s Sex and
ABO type:
Donor’s Sex and
ABO type:
O
Recipient’s PRA:
Recipient’s Ethnicity:
Recipient-to-donor
Relationship:
Wife
Husband
The initiating donor was an unpaired altruistic donor from Michigan.
The recipient of Transplant 6 required desensitization to HLA DSA by T and B cell flow cytometry.
3 The recipient of Transplant 9 required desensitization to blood group (AHG titer of 1:8)
1
2
Daughter Mother
Mother Daughter
What is Desensitization?
• Lower baseline DSA to below threshold of renal
endothelial cell injury at time of transplant
• Prevent memory response
• Maintain low DSA level posttransplant
– Detect and treat response early
Desensitization as a Solution
• Goal of desensitization is to reduce relevant antibodies to
a level where a previously “incompatible” transplant can
be performed safely and successfully.
• Requirements
– Accurate and thorough assessment of a patient’s
immunologic risk
– Timely and careful monitoring both pre- and post-transplant
– Open and timely communication between clinical and
laboratory staffs
Desensitization
Therapeutics
Diagnostics
• Flow or Luminex® PRA
• Apheresis:
• PD flow crossmatch
• IVIG
• SA by solid phase assay
• Rituximab
• C4d staining
• Thymoglobuline®
Specialized,
dedicated Staff
Transplant Team
Transplant Surgeon
Challenges to the Clinicians:
Develop new therapeutic
strategies to improve outcomes
Transplant
Physician
Clinical
Laboratory
Challenges to the Labs:
Researcher
Challenges to the Researchers:
Provide insights into the
mechanisms underlying
allograft injury
Provide diagnostics tools to
predict and monitor for allograft
injury
Provide environment for
translation of basic science into
clinical practice
Risk Assessment
• Sensitization History
– Transplants: Ag exposure / immunologic response
– Pregnancies: determine paternal HLA
– Recent or large numbers of transfusions
• Current Ab: specificity and titer
– Specificity: DSA or not
– Titer: correlates best with risk and treatment needed
• Immunologic Profile of Current Tx
– Number of mismatches
– Child → mother, husband → wife
– Nature of mismatch: repeat, highly immunogenic
Zachary AA and Hart JM in Handbook of Human Immunology, 1997
Zachary et al., in Manual of Clinical Laboratory Immunology, 6th Edition, 2003
Desensitization
High-dose IVIG
Jordan SC, et al. Am J Transplant 2006; 6:459-66.
Low-dose IVIG and Pheresis
Zachary AA, et al. Hum Immunol 2005; 66:364-70.
Rituximab
Pescovitz MD, et al. Am J Transplnt 2005; 5(suppl11):324
Rituximab / High-dose IVIG
Ashley A, et al. NEJM 2008;359:242-251
Controlling B &T Memory
T and B – cell memory occurs together
HLA A2
Pregnancy associated sensitization
Anti-HLA A2 Ab
Transplantation (1996) 62:672
B
Transfusion associated sensitization
IFNg
Transplantation (1990) 45:987
Th
APC
IL-2
CTL
Antibody and T-cell mediated rejection occur together
Ab mediated rejection: T2 + T3 Tubulitis 50% (12/24) of cases
Transplantation (1996) 61:1586
Fas
Granzyme B
Perforin
Strategy
Use new solid phase technologies to fully define level of risk then
proceed to modulate the immune system prior to and around the
time of transplant
•Requires approaches specific to T & B-cell memory
T-cell
B-cell
Plasma
cell
HLA Ab
• IVIG
-
+
-
++
• Pheresis
-
-
-
++
• Rituximab
-
+++
-
-
+++
++
+
-
• Thymoglobulin
M. Zand, Transplantation 2005
Activity of rATG, Alemtuzumab,
and Rituximab in Debulking the
B-Cell Mass
Naïve
Activated
Memory
Pre-Plasma
Cell
Plasma Cell
CD19
+++
+++
+++
+/-
-
CD20
++
++
++
+/-
-
CD27
-
+/-
+
+++
+++
CD38
-
+
+/-
++
+++
CD40
++
++
++
++
-
CD52
+
+
-
-
-
CD80/86
-
++
++
+++
-
CD95
+
+
+
++
+
HLA-DR
+
++
++
++
-
sIgD
++
++
+/-
-
-
-
+/-
++
++
+
sIgM/G
rTAG
Rituximab
Alemtuzumab
Anti B-Cell Activity
Induced apoptosis in
Resting B-cell
Activated
B-cell
Plasma
cell
Rabbit Anti-thymocyte
Globulin
Yes
Yes
Yes
Rituximab (anti-CD20)
No
Yes
No
Sirolimus
No
Yes
No
Mycophenolate
Mofetil
Inhibits B cell proliferation only
Cyclosporine
No direct activity on resting or activated B
cells
The Limitations of
Immunomodulation (desensitization)
• May not be available to all patients
– Center expertise
– High Titers
– Deceased donors more difficult
• Short term outcomes may be worse
– Higher rejection than the general transplant population
• Long term outcomes are not known
• Burden of immunosuppression
Limitations of Anti-CD20
Treatment
• CD20 expression:
– Low on pre-B cells, high in normal B cells, progressive
loss in plasma B cells
– Anti-CD20 depletes mature and memory B cells but not
plasma cells nor pre-existing Ab titers
• Vascular access to B cell compartment:
– Blood > lymph nodes/spleen > peritoneal cavity
– Follicular > MZ > GC
• Survival factors in the B cell microenvironment:
– BAFF/ BLyS B cell survival factors
Which Option is Best for an
Individual Incompatible Pair
Defining the Immunologic Phenotype
How difficult will they be to match?
How difficult will they be to desensitize?
How Difficult Will They Be to
Desensitize? (+) XM
Major Factors
DSA titer (depth ) and multiplicity (breadth)
Repeat mismatches
Sensitized from previous transplant(s)
How Difficult Will They Be to
Desensitize? ABOi
Major Factors
Iso-hemoagglutinin Titer
Donor Type:
Difficulty: A1 >B >A2
Matching Donor/Recipient Pair to
Transplant Modality
KPD
Desensitization
Easy-to-match pair
• O donor
• Low PRA
Difficult-to-match pair
• AB donor
• Broad sensitization
Difficult-to-desensitize
• High titer DSA
• High immunologic risk
Easy-to-desensitize
• Low titer DSA
• Low immunologic risk
Highly sensitized
No live donor available
Desensitization
High dose IVIg
Wait for (-) XM
DD kidney
Live donor available
Desensitization
PP/IVIg or IVIg
Tx
Kidney Paired
donation
Desensitization
with KPD
Tx
The Cost
Segev,etal JAMA,2005;293:1883-1890
Per-Patient Analysis of Transplant Costs and Desensitization While
Awaiting Donation From the Deceased Donor List vs Kidney Paired
Donation (KPD)
Years on Waitlist*
Cost of
Dialysis†
Cost of
Desensitizationŧ
Cost of KPD§
Highly sensitized
6.73
$485,038
$233,717
$204,738
Blood type O
4.85
$374,605
$201,544
$172,565
Blood type A
2.97
$264,172
$169,372
$140,393
Blood type B
5.48
$411,309
$212,237
$183,258
Blood type AB
1.63
$185,453
$146,439
$117,460
Recipient type
* Average years awaiting deceased donor donation are based on UNOS data.
† Average per patient cost of dialysis calculated according to Medicare payments of $58,758/year for
dialysis, for the number of years the average patient awaits a donation from the deceased donor waitlist,
plus $89,508 for the cost of transplantation and the first year of postoperative care.
Ŧ Average per patient cost of desensitization calculated according to Medicare payments of $28,979 for
desensitization. $89,508 for transplantation and the first year of post operative care, and $17,118/year of
immunosuppression for the number of years the patient would have awaited a donation from the
deceased donor waitlist.
§ Average per patient cost of a patient successfully matched through a KPD calculated according to
Medicare payments of $89,508 for the cost of transplantation and the first year of post operative care, and
$17,118/year of immunosuppression for the number of years the patient would have awaite donation from
the deceased donor waitlist.
KFSH & RC Experience
Desensitization
Measures of Success
• Prevention of hyper-acute AMR
• Minimization and reversal of antibody mediated rejection
• Induction of a state of relative donor specific unresponsiveness
or a state of accommodation
• patient and graft survival comparable to sensitized CXM
negative recipients
• Prevention of neoplasia
• Prevention and/or minimization of opportunistic infections
Diagnostic Algorithm
KFSH&RC
Sensitization present
Clinical relevance to donor’s HLA
Strength of reaction to donor’s HLA
Diagnostic Algorithm
KFSH&RC
Sensitization present
• LAB screen® mixed
• LAB screen ® class I & II
Clinical relevance to donor’s HLA
• LAB screen ® single antigen class I & II
• Molecular typing of donor’s HLA ( low
resolution and occasionally high
resolution)
Strength of reaction to donor’s HLA
• SFI, MESF
• PD FCXM (T and/or B IgG)
• CDC / CD AHG CXM (T and/or B IgG)
Management Approach
KFSH&RC Protocol
Induction of desensitization
High risk:
• Positive CDC/AHG CDC CXM
• Positive PD FCXM
• Positive DSA
Low risk:
• Negative CDC/AHG CDC CXM
• Positive PD FCXM
• Positive DSA
• Antibody depleting:
IA
• Antibody modulation:
High dose IVIG
• Anti-B cell xx
Rituximab®
• Antibody modulation:
High dose IVIG
• Anti-B cell xx
Rituximab®
Pre-conditioning Regimen
High Risk Patients
High
dose
IVIG
(2.0gm/
kg)
Daily Immunoadsorption : 3.5 plasma volume/ treatment
5 – 10 treatments
Rituximab
Single dose of
500mg IV
MMF 500 mg BID
-14
-7
-1
0
1
Pre-conditioning Regimen
Moderate Risk
HD
IVIG
(2.0gm/
kg)
Rituximab
Single dose of
500mg IV
-14
-2
0
Pre-conditioning Regimen
Low Risk
IVIG
(1.0gm/
kg)
-1
0
Measures To Control Activated T & B
Lymphocytes during the
Pre-adaptation Phase
Steroids (Standard Dose)
Tacrolimus 0.15 – 0.3 mg/kg BID
Target level: 10-15 ng /dl
MMF 1 gm BID
Thymoglobulin
1.5 mg/kg/day
0
1
2
Weeks Post Engraftment
3
4
Monitoring
Post-Transplant:
Intense monitoring
1 month
0
• High index of clinical suspicion for AMR
• Class I or class II SA Luminex®
• Renal allograft biopsy with C4d Staining
Monitoring
Post-Transplant:
Routine Monitoring
3
*
6
♣*
12
♣*
24
♣*
Months
*Protocol biopsy
♣*Protocol biopsy and SA Luminex®
36
♣
Outcomes of Desensitization
Protocols at Different Centers
Long-Term Outcomes of Desensitization Protocols for HS ESRD Patients
Trans.
Program
Patient #
Protocol
Used
Patient
Survival
(3-5 Yrs)
Graft
Survival
(3-5 Yrs)
AR Rates
Mean SCr
(3-5 Yrs)
Mayo Clinic
#94
HD IVIG
PE/IVIG LD
97% at
5 Yrs
80% at
5 Yrs.
35%
1.6  0.6
mg/dl
JHU
#90
PE + CMV-Ig
95% at
3 Yrs.
80.9% at
3 Yrs.
62%
1.2  0.3
mg/dl
CSMC
#96
HD IVIG
97% at
5 Yrs.
87% at
5 Yrs.
36%
1.5  0.4
mg/dl
KFSH
#85
Anti CD20Ab
HD IVIG
IA or PE/IVIG
LD
98% at
5 Yrs
94% at
5Yrs
21% AMR*
13% ACR*
0.9 ± 0.4
mg/ dl
Adapted from Jordan and Pescovitz Clin J Am Soc Nephrol 1:421-432, 2006
*Both clinical and subclinical rejections
Pre-conditioning Regimen
ABO Incompatible Tx
KFSH&RC Protocol
Plasma exchange (PE): 1.5 plasma volume qod
4 – 10 treatments each followed by small dose IVIG
Rituximab
Single dose of
500mg IV
PE
followed
by high
dose IVIG
MMF 500 mg BID
-14
-7
0
1
2
3
4
Recent Published Reports of ABOIncompatible Kidney Transplantation
Reference
Patients
1-Yr graft
survival (%)
Comment
Schneulle and van der Woude
(1998)
108
NA
Meta-analysis of A2 donors
Nelson et al. (1998)
46
94
10-yr experience with A2
and A2B donors
Alkhunaizi et al. (1999)
15
93
A2 donors with
plasmapheresis
Sorensen et al. (2001)
15
93
A2 donors without
plasmapheresis
Gloor et al. (2003)
10
80
Included high titer anti-A
recipients
Shimura et al. (2000)
67
81
A1 and B donors; 8 yr graft
survival, 66%
Gloor et al. (2003)
8
100
Splenectomy and
plasmapheresis
Almeshari et al (2009)
7
100
Plasmapheresis / high dose
IVIG/Rituximab
A2 and A2B donors
Non-A2 donors
a
Two year graft survival.
Layout
Future
Development of B Lineage
Immune Response and Memory
Plasmablast/
short-lived plasma cell
TH
germinal
center
Memory
B cell
antigen
TH
Naïve
B cells
TH
Activated
B cells
Long-lived
plasma cell
short-lived
plasma cell
Development of B Cells in the
Germinal Center
T helper
B cell
cell
Antigen
Cytokines
Proliferation
Somatic hypermutatino of
IgV genes – selection of B
cells with high affinity for
antigen
Follicular
dendritic
cell
presents
antigen to
B cell
Death of nonselected cells
Plasma
cell
Memory
B cell
Somatic hypermutation, affinity maturation and memory B cell formation
Acquisition of Diversity, Specificity
and Functional Capacity
• Ig Gene re-arrangement
• Allelic exclusion
• Somatic hypermutation
• Affinity maturation
• Isotype switching
BLyS (BAFF) / APRIL System
Ligands
Receptors
BLyS
BAFF-R
BCMA
APRIL Heterotrimer
TACI Proteoglycans
• Increased B-cell survival
• Co-stimulation of B-cell
proliferation
• Ig class switch recombination
• Enhanced APC function
• Germinal centre formation
• Regulation of B-cell tolerance
• Sequester APRIL at cell
surface to improve TACI
and/or BCMA signaling?
• Mediator plasma cell
trafficking
Dillon et al. Nature Reviews Drug Discovery 5, 235 (2006)
Copyright @ 2006 Nature Publishing Group. Nature Reviews / Drug Discovery
Synergistic Depletion of Memory B Cells
and Plasma Cells with Anti-CD20 and
BLyS (BAFF)/APRIL Blockade
• Maintenance of long-lived plasma cells and
serological memory despite mature and
memory B cell depletion during CD20
immunotherapy in mice.
Di Lillo et al. J Immunol. 2008; 180: 361
• The Dependence of Plasma cells and
Independence of Memory B Cells on BAFF and
APRIL.
O’Conner at al. J Immunol. 2008; 180(6): 3655.
B Cell Development
Shortlived
plasma
cell
Long-lived
Antibody removal strategies
plasma cell
Plasmablast
Short-lived
plasma cell
Clonal
expansion
Memory
cell
Space-directed strategies
Activated
Follicular
B cell
Fetal liver and
bone marrow Tolerance strategies
CLP
Pro-B
IgM
μ
μ
Large
Pre-B
Small
Pre-B
IgM
IgD
Targeted cell depletion strategies
Clonal
expansion
Immature Activated
(T1/T2)
Marginal
Zone B
(spleen)
Plasma
Plasmablast cell
Novel Monoclonal Antibodies and
Fusion Receptor Proteins That Deplete
and/or Block Activation of B Cells
Belimumab
Atacicept
BR3Fc
Anti-CD40
BCR
BAFF
(BLyS)
BR3
Epratuzumab
syk
CD19
Ras
AME-133
PKC
Ocrelizumab
Ofatumumab
BCMA
TACI
CD22
p13K
APRIL
Activation/Survival
B Cell
Atacicept
Long-lived plasma cell
138+
20-
DSA
Bone marrow, secondary
lymphoid organs
Baseline anti-donor antibody titer variable
person to person, variable from time to time
The Effect of Desensitization Protocols
on Human Splenic B Cell Populations in Vivo
Ramos, et al. AJT 2007; 7: 402 – 407.
The Ubiquitin-Proteasome Pathway
Elimination of Cells
Bortezomib (Velcade TM, Pyz-Phe-borolLeu)
O
N
N
Pyrazinoic acid
N
H
H
N
Indication
Treatment of myeloma
OH
B
OH
O
phenylalanine
leucine
boric acid
Side effects
GI 87%
Neutropenia 20%
Thrombocytopenia 35%
Neuropathy 36%
Proteasome Inhibition Causes Apoptosis of
Human Plasma cells and is an Effective
Therapy for AMR and ACR
Bortezomib
(Velcade®)
Everly MJ,Transplantation
2008;86 (12):1754-1761
Perry DK,AJT
2009;9 (12):201-209
Proteasome Inhibitors
Role of Proteasomes
• T & B cell activation
• Cell cycle control
• Cell adhesion and migration
• Apoptosis of cells of T, B, and monocyte
lineage
• Antigen presentation
Eculizumab : Anti C5
Eculizimab
C9
C5a
C5 C7
C8
C5
C5 convertase
C5b
C5 C5b
C5 C5b
Poly-C9
C5 C5b
C7
C7 C8
C7 C8
Plasma membrane
MAC
Abbas AK, et al, Cellular and Molecular Immunology, 4th edn, 2000.
Strategies for Future
• Avoid antibodies
• Remove antibodies mechanically
• Deplete or suppress B cells by targeting developmentally
regulated cell surface molecules (CD20, CD19)
• Impair B cell proliferation by targeting space-dependent
cell receptors (TACl, BMCR)
• Eliminate or suppress mature antigen-specific antibodyproducing plasma cells (Bortezomib)
• B cell tolerance/combined B/T cell tolerance
• Encourage graft accommodation