Allogeneic transplantation for Adults
with Severe Sickle Cell Disease
John F. Tisdale, MD
Senior Investigator
Molecular and Clinical Hematology Branch
Hematopoietic stem cells as vehicles for
therapeutic gene delivery
Allogeneic stem cell transplantation
–Transplantation using allogeneic
stem cells from a normal donor
•HLA-matched sibling
Autologous stem cell gene transfer
–Transplantation using autologous stem
cells which have been corrected by
transfer of a normal or therapeutic gene
•Retroviral vectors
Hematopoietic stem cells as vehicles for
therapeutic gene delivery
Allogeneic stem cell transplantation
Myeloablative transplantation curative in
children with sickle cell disease
•Stable mixed chimerism sufficient
•13/50 surviving patients 11-99% donor
chimerism
(Walters et al., BBMT, 7, 665, 2001)
•None experienced recurrent painful
crises or other related events
•Toxic conditioning and GVHD limit
application to children
•Engraftment without ablation
Nonmyeloablative conditioning sufficient for
reliable allogeneic PBSC engraftment
• Cytoxan/fludarabine based immune ablative
conditioning piloted in patients with metastatic
cancer
– Childs, R.W., et al., JCO, 17, 2044, 1999.
– Childs, R., et al., NEJM, 343: 750-758, 2000.
• Extended to high-risk patients ineligible for
conventional myeloablative conditioning
– Kang, E.M., et al., Blood, 99, 698-701, 2002.
– Kang, E.M., et al., J Hematother and Stem Cell Res, 11, 809-816, 2002.
Application to sickle cell disease?
• NIH experience overall (n>100)
– Engraftment through donor
alloimmune response
– GVHD common
• T cell alloreactivity not
necessary in nonmalignant
disorders
– Treatment related mortality 21%
• GVHD principal cause
• Prohibitive in nonmalignant
disorders
Cumulative non-ablative BMT experience in
sickle cell disease
Chakrabarti S et al, BBMT 2004
A non-ablative protocol for adults with severe sickle
cell disease is needed
A Murine Model of Nonmyeloablative Stem
Cell Transplantation for the Treatment of
Sickle Cell Disease
•Develop regimen that:
– Promotes tolerance without need for
long term immunosuppression
– Allow for stable mixed chimerism
•F1-Hybrid donor mice
6 Days
G-CSF
(200 ug/kg)
– Myeloid-flow cytometry
– Erythroid-Hb electrophoresis
•Donors mobilized with G-CSF
•Mobilized cells collected day 6
•Recipient mice conditioned with
300 cGy and a 30d course of either
• Cyclosporine (CSA)
• Rapamycin (RAPA)
Harvest mobilized
stem cells
F1-Hybrid
C57Bl6 (Kb) X BalbC(Kd)
Recipient
C57Bl6 (Kb)
100x106
cells
Day -1
Week
Day 0
(300 cGy)
RAPA (3mg/kg)
or CSA (20mg/kg)
Why Rapamycin??
IL-2
CSA
TcR-CD3
RAPA
CD28
Effector Function
Proliferation
Anergy
Induction of tolerance
Rapamycin but not Cyclosporine Maintains
Chimerism in the Absence of Long-term
Immunosuppression
100
CSA
Rapa
% Donor
80
60
40
20
0
0
8
16
Weeks
24
32
Sickle Hemoglobin is Replaced by Donor
Hemoglobin in Chimeric Homozygote Mice
Powell, J, Fitzhugh, C. et al., Transplantation, 80(11):1541-5, 2005
Nonmyeloablative Allogeneic PBSC
Transplantation for Adults with Severe
Congenital Anemias
Eligibility: Adults with Hb SS, SC, or Sb0-thal
Severe end-organ damage
– stroke or abnormal CNS vessel
– TRV ≥2.5 m/s
– renal damage
• Or modifiable complication(s), not ameliorated by
hydroxyurea
– > 2 hospital admissions per year for pain crises
(VOC)
– previous acute chest syndromes (ACS)
– red cell alloimmunization
– osteonecrosis of multiple joints
Screening and Accrual of Patients
Characteristics of 10 Patients Undergoing Nonmyeloablative
Hematopoietic Stem-Cell Transplantation (HSCT)
Conditioning regimen
Sirolimus taper if full donor chimerism achieved in the absence of GVHD
Transplant course
• All patients tolerated conditioning
without serious adverse events
– No need for nutritional support
– No acute or chronic GVHD
– No sickle cell anemia related events
• Nine of 10 with stable engraftment
Donor chimerism after transplantation
Improvement in hemolysis parameters after
transplantation
Normalization of hemoglobin levels after
transplantation
Replacement by donor derived red cells allows
tapering of narcotic analgesics
IV morphine equivalent (mg)
2000
1600
1200
800
400
0
0
4
8
12
16
Weeks
20
24
28
Interim conclusions
• Allogeneic PBSC transplantation after low dose TBI,
campath, rapamycin conditioning and resulting mixed
hematopoietic chimerism sufficient to revert the sickle
phenotype
– Accrual currently at 23 patients, 20 without SCD
• Low toxicity allows application in adults with severe
disease
• ‘Split’ or mixed chimerism and absence of acute or
chronic GvHD suggests operational tolerance
– Protocol amended for weaning of rapamycin at CD3 >50%
• 5 patients off immunosuppression with stable mixed chimerism
• Longer follow-up and further accrual necessary
• Barriers remain for widespread application
Barriers to therapeutic gene delivery using HSCs:
HSC source
“The regimen use by Hsieh et al. is clearly an important development, but its
applicability is still limited by the small number of available HLA-matched siblings.”
Barriers to therapeutic gene delivery using HSCs:
HSC source/MUD bone marrow
Patient 1
2
3
4
5
6
7
8
9
10
BMDW
Potential 6/6 allele match
0
Potential 10/10 allele match 0
0
0
0
0
1
1
2
2
4
4
6
6
9
8
32
28
42
36
0
0
0
0
0
0
0
0
0
1
1
1
1
1
1
2
2
2
4
3
0
4
4
1
5
5
0
13
5
0
0%
0%
0%
1%
1%
1%
1%
50% 1%
NMDP (duplicated in BMDW
report)
Potential 6/6 allele match
Potential 10/10 allele match
Potential 10/10 allele match
of African descent
HapLogic: highest
probability of being 6/6
allele match
Seven of 10 with potential 6/6 donor
Using Haplogic, only 1/7 with >1% chance of having a 6/6 donor
1%
Barriers to therapeutic gene delivery using HSCs:
HSC source/Cord blood
>4/6 HLA match
>5/6 HLA match
6/6 HLA match
9 of 10
6 of 10
1 of 10
TNC > 1.5 x 107/kg
# of pts with matched UCB
(# with ABO matched)
9 (9)
5 (2)
0 (0)
>4/6 HLA match
>5/6 HLA match
6/6 HLA match
Median # of units
available per pt
(range)
103.5 (20-875)
2.5 (1-114)
0 (0)
Median UCB with TNC >1.5 x 107/kg
(# with ABO matched)
[TNC range x107/kg]
29 (8) [1.5-7.04]
0.5 (0) [1.51-4.39]
0 (0) [0]
# of pts matched
TNC > 2.5 x107/kg
# of pts with matched UCB
(# with ABO matched)
8 (7)
2 (2)
0 (0)
Median UCB with TNC >2.5 x107/kg
(# with ABO matched)
[TNC range x107/kg]
3 (1) [2.51-7.04]
0 (0) [2.55-4.39]
0 (0) [0]
Nine of 10 have at least one 4/6 cord blood match identified
Higher degree of matching, higher cell dose, and ABO compatibility limits applicability
Barriers to therapeutic gene delivery using HSCs:
Haploidentical grafts?
• Haploidentical donors
– Most accessible
– Large cell doses feasible
– Repeat collections feasible
• Immunologic barrier greater
– Higher degree of immunosuppression
• Post-graft cyclophosphamide
– Reduce graft rejection/GvHD
– Targets proliferating lymphocytes
– Early success in ongoing clinical trials
Luznik L et al. Blood, 2001. 98(12): 3456-3464.
Will Sirolimus Prevent
Transplant
Cy effect
Post-Transplant
Cy and Post
Sirolimus
are Synergistic
on Engraftment?
Sirolimus (Sir, 3mg/kg IP) days -1 or +4 for 30 days
200cGy TBI, bone marrow cells infused on day 0
Cyclophosphamide (Cy, 200mg/kg IP) day +2
Conditioning regimen
Escalating dose
post transplant Cy
TBI
200 cGy
+1
+2
+3
Cohort
Cy dose
Day Post Tx
Accrual ceiling 32/cohort
1
0
NA
3/3 rejected
2
50 mg/kg
+3
3/4 engrafted (no GvHD)
3
50 mg/kg
+3 and +4
NA
Hematopoietic stem cells as vehicles for therapeutic
gene delivery: Future efforts for human application
Allogeneic stem cell transplantation
Validate results with continued accrual
(Trial plan for 25 subjects)
Expand to multicenter trial design
(Facilitate recruitment)
Determine engraftment level sufficient to revert phenotype
(Compare marrow progenitor chimerism with peripheral blood)
Characterize immunologic recovery
(Mechanistic studies of tolerance induction)
Tolerance for alternative donor transplantation
(Haploidentical donors)
Crew
•
•
Tisdale lab
– Pat Weitzel
– Naoya Uchida
– Courtney Fitzhugh
– O.J. Phang
– Kareem Washington
– Matt Hsieh
•
Roger Kurlander
•
Elizabeth Kang
•
Jonathan Powell
Department of Transfusion Medicine
– Charley Carter
– Susan Leitman
– Dave Stoncek
•
•
•
•
Terri Wakefield
Beth Link
Karen Kendrick
Griffin Rodgers