PowerPoint Presentation - Hematopoietic Stem Cells as Vehicles for the Treatment of Human Disease

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Allogeneic transplantation for Adults with
Severe Sickle Cell Disease
John F. Tisdale, MD Senior Investigator Molecular
and Clinical Hematology Branch
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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

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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

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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.

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Application to sickle cell disease?

• NIH experience overall (ngt100)
• 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

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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
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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
• 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)

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Why Rapamycin??
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Rapamycin but not Cyclosporine Maintains
Chimerism in the Absence of Long-term
Immunosuppression










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Sickle Hemoglobin is Replaced by Donor Hemoglobin
in Chimeric Homozygote Mice
Powell, J, Fitzhugh, C. et al., Transplantation,
80(11)1541-5, 2005
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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
• gt 2 hospital admissions per year for pain crises
  (VOC)
• previous acute chest syndromes (ACS)
• red cell alloimmunization
• osteonecrosis of multiple joints

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Screening and Accrual of Patients
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Characteristics of 10 Patients Undergoing
Nonmyeloablative Hematopoietic Stem-Cell
Transplantation (HSCT)
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Conditioning regimen
Sirolimus taper if full donor chimerism achieved
in the absence of GVHD
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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

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Donor chimerism after transplantation
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Improvement in hemolysis parameters after
transplantation
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Normalization of hemoglobin levels after
transplantation
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Replacement by donor derived red cells allows
tapering of narcotic analgesics
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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
  gt50
• 5 patients off immunosuppression with stable
  mixed chimerism
• Longer follow-up and further accrual necessary
• Barriers remain for widespread application

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Barriers to therapeutic gene delivery using
HSCsHSC 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.

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Barriers to therapeutic gene delivery using
HSCsHSC source/MUD bone marrow
Patient 1 2 3 4 5 6 7 8 9 10
BMDW Potential 6/6 allele match Potential 10/10 allele match 0 0 0 0 0 0 1 1 2 2 4 4 6 6 9 8 32 28 42 36
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 0 0 0 0 0 0 0 0 0 0 0 0 1 1 1 1 1 1 1 1 2 2 2 1 4 3 0 1 4 4 1 50 5 5 0 1 13 5 0 1

• Seven of 10 with potential 6/6 donor

Using Haplogic, only 1/7 with gt1 chance of
having a 6/6 donor
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Barriers to therapeutic gene delivery using
HSCsHSC source/Cord blood
gt4/6 HLA match gt5/6 HLA match 6/6 HLA match of pts matched 9 of 10 6 of 10 1 of 10 TNC gt 1.5 x 107/kg of pts with matched UCB ( with ABO matched) 9 (9) 5 (2) 0 (0) TNC gt 2.5 x107/kg of pts with matched UCB ( with ABO matched) 8 (7) 2 (2) 0 (0)
gt4/6 HLA match gt5/6 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 gt1.5 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 Median UCB with TNC gt2.5 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
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Barriers to therapeutic gene delivery using
HSCsHaploidentical 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.
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Post-Transplant Cy and Sirolimus are Synergistic
Will Sirolimus Prevent Post Transplant Cy effect
on Engraftment?
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Conditioning regimen
Escalating dose post transplant Cy
TBI 200 cGy
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2
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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
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Hematopoietic stem cells as vehicles for
therapeutic gene delivery Future efforts for
human application
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Crew

• Tisdale lab
• Pat Weitzel
• Naoya Uchida
• Courtney Fitzhugh
• O.J. Phang
• Kareem Washington
• Matt Hsieh
• Department of Transfusion Medicine
• Charley Carter
• Susan Leitman
• Dave Stoncek

• Roger Kurlander
• Elizabeth Kang
• Jonathan Powell
• Terri Wakefield
• Beth Link
• Karen Kendrick
• Griffin Rodgers