Immune Reconstitution and Immune Responses after Haematopoietic Cell Transplantation

1
Immune Reconstitutionand Immune Responses
afterHaematopoietic Cell Transplantation
2
Bone marrow
Precursor pool size
Thymus
Thymic activity
Peripheral pool
Naïve repertoire
Homeostatic proliferation
Antigen exposure
Death
Memory repertoire
Homeostatic proliferation Antigen exposure
3
Flow Cytometry
CD4
CD8
Memory
Naive
Naive
Memory
CD27
CD27
CD45RO
CD45RO
Effector
Effector
4
TRECs TCR rearrangement excision circles
5
Factors Influencing the Rate of Immune
Reconstitution

• Conditioning
• Age - both of donor and recipient
• Stem cell source
• T cell depletion
• Infection
• Acute or chronic GvHD

6
Campath-1H conditioning reduces early T cell
numbers
CD4
CD8
7
SCID Patient 18 Years Post-Transplant Increasing
clinical immunodeficiency but normal T cell
numbers
V? 1
V? 2
V? 3
V? 4
V? 5.1
V? 5.2
No TRECs have been detected in this patient
8
Thymic Function after Hematopoietic Stem-cell
Transplantation for the Treatment of Severe
Combined Immunodeficiency(Patel et al (2000) N
Engl J Med 3421325-32)
9
TREC Levels in Immunodeficiency Syndromes
30000
20000
TREC per µg DNA
10000
0
Normal
DiGeorge
Con
Non-Con
SCID
10
CD3
Months post-transplant
CD4
CD8
5000
4000
1000
1000
T-cells per ?l
T-cells per ?l
100
100
10
10
Months post-transplant
Months post-transplant
11
Jamieson et al Immunity 10569-575 (1999)
Peripheral T cells
Thymic output
Age
12
CD4
CD8
1000
1000
100
100
Absolute TREC count (TREC per ?l blood)
10
10
1
1
0.1
0.1
6
12
9
3
6
12
9
3
2000
2000
1000
1000
Naïve T-cells per ?l blood
100
100
10
10
6
12
9
3
6
12
9
3
Months post-HCT
6-30 yrs
31-53 yrs
13
Memory CD4
Effector CD4
1000
10000
1000
T-cells per ?l blood
100
T-cells per ?l blood
100
10
10
6
12
3
9
6
12
3
9
Non-naïve CD8
CD85728-
5000
10000
1000
1000
T-cells per ?l blood
T-cells per ?l blood
100
100
10
10
6
12
3
9
6
12
3
9
Months post-HCT
6-30 yrs
31-53 yrs
14
Flow Cytometry
CD4
CD8
Memory
Naive
Naive
Memory
CD27
CD27
CD45RO
CD45RO
Effector
Effector
15
Naïve, memory and effector cells in the CD4
compartment (3 months post transplant)
16
21.3
6.7
89.9
NAÏVE
43.5
17.6
59.9
MEMORY
82.3
71.1
13.2
EFFECTOR
87.7
81.3
7.6
27-/45RO-
TNF?
IFN?
CCR7
17
Aberrant T cell subsets persist for over 2years
Patient 47
Control
CD4
59
21
10
10
CD8
CD27
CD45RO
18
Normal healthy control
CD45RA
CD27
CD45RO
CD45RO
Time post-HSCT
3 months
6 months
9 months
1 year
Patient 1
CD45RA
Patient 12
CD45RO
Patient 1
Patient 12
CD27
CD45RO
19
CD27 and CD27- subsets of CD4CD45RA T cells
show opposite correlations with TREC
0.05gtpgt0.01 0.01gtpgt0.001
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SUMMARY (Children)

• TREC levels rise with time post-transplant and
  correlate with naïve T-cell numbers
• (CD4 r0.88, Plt0.0001 CD8 r0.96, Plt0.0001)
• Memory and effector cells do not increase in
  parallel
• Thymus dependent pathways rapidly restore
  T-cell numbers in children (lt1 year)
• Peripheral expansion of graft derived memory and
  effector cells plays only a minor role in T-cell
  reconstitution in children

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SUMMARY (Adults)

• TREC levels rise with time post-transplant and
  correlate with naïve T-cells
• BUT TREC levels and naïve T-cell numbers are
  10 fold lower than children
• Memory and effector T-cells remain the dominant
  cell type over the first year
• peripheral expansion is important in adults to
  restore normal T-cell numbers
• Thymic dependent pathways are less able to
  contribute to T-cell reconstitution in adults

22
1000
1000
CD4
CD8
B
A
100
100
10
10
Absolute TREC count (TREC per ?l blood)
1
1
0.1
0.1
6
12
9
3
6
12
9
3
1000
1000
C
D
100
100
Naive T cells per ?l bloo)
10
10
1
1
0.1
0.1
6
12
9
3
6
12
9
3
No GvHD
GvHD
23
Patient 1
Patient 9
Patient 17
3 months
6 months
9 months
12 months
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Effect of GvHD on thymic output

• Inflammatory reaction in the thymus?
• Immunosuppression - methotrexate, Cyclosporin A,
  steroids?
• Or does failure of thymic output (of regulatory
  cells) cause GvHD?

25
New T cell production is inhibited by Steroids
in the absence of GvHD
200
6
4
Naïve T-cells per ul blood
TREC T-cells per ul blood
100
2
29 year old Autoimmune cytopaeniaTCD MisMUD No
GvHD
0
0
6
12
1500
1000
Memory and effector T-cells per ul blood
500
0
12
6
Months post-HSCT
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To Optimise Post Transplant Reconstitution

• Conditioning allows stable stem cell engraftment
• If possible, reactivation of the thymus would
  benefit adult patients
• Prevention of GvHD or alternative (non-steroid)
  therapies will improve reconstitution

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Immune reconstitution and immunotherapy
Does patient respond?
Yes
No treatment required
No
Does patient have an adequate T cell repertoire?
Yes
No
Vaccination
Adoptive Immunotherapy
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What immune responses do we want in the
transplant recipient?
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Potential Targets for Immunotherapy

• Viral antigens
• CMV, EBV
• Alloantigens and minor HAg
• HA1, HY
• Tumour associated antigens
• Leukaemic antigens, eg PR1
• Idiotopes
• other tumour associated antigens, eg melanoma

30
CMV specific T cells in CMV PCR- patients
all data CD3 gated
31
Healthy / post SCT
Post transplant, patients have significantly
higher frequencies of CMV specific T cells
than healthy controls.
Post stem cell transplantation, patients tend to
be lymphopenic,which could account for the
higher frequency.
However the absolute numberof CMV specific T
cells is alsoincreased.
32
Patient 4 - Disease AML - Male, 54 years
old - Patient/Donor CMV / - HLA identical
sibling PBSC
33
Patient 3 - Disease AML - Male, 32 years
old - Patient/Donor CMV / - HLA non
identical unrelated Bone Marrow
34
Patient 1 - Disease AML - Male, 39 years
old - Donor/Patient CMV -/ - HLA identical
sibling PBSC
35
Tet cells / CMV PCR status
The number of CMV specific T cells correlates
with the detection of viral replication by PCR.
All data sets are statistically different.
This shows that the number of CMV specific T
cells can be used in the clinic as an indicator
of the level of immunity to CMV.
36
Monitoring (3)

• CD8 counts measured by TRUECOUNT analysis
• Absolute IPS response calculated by
  combination with tetramer analysis

37
Monitoring (4)
-no subsequent reactivations are seen in this
B35 individual -the specific T cell count
remains around 14x106/l

• HLA-A0201 restricted CTLs confer protection
  against CMV reactivation above a threshold level
  of 107-2x107/l
• (Meijer, E 2003, Chen et al. Cytotherapy 2002
  4(1)41)

38
Monitoring (5)
Example Patient CMS, HLA-B3501, CMV/- Tx
type Allo BM DOB 24/9/68 (age at Tx
35) HLA Id Sib Diagnosis AML M4 Engraftment
d17 GvHDgrade I/II since d21 Conditioning
TBI
39
Monitoring (6)
- no CMV reactivations - nevertheless high
number of tetramer staining cells at d46
Prednisolone d62 - d91?Cyclosporin from d42
40
Monitoring (7)