3. Cell Therapy Allogeneic SCT
• High dose chemotherapy/radiation usually does not
eradicate malignancy
– Higher relapse rate with identical twin or with T-cell
depletion
– Reduced relapse with GVHD
• Allogeneic GVL effect responsible for eradicating
residual disease.
5. Time(weeks)
SurvivalProbability
0 20 40 60 80 100 120
0.00.20.40.60.81.0
In remission, PB.blast=0
Active Disease, PB.blast=0
Active Disease, PB.blast>0
p<0.0001
Time(weeks)
Event-freeprobability
0 20 40 60 80 100 120
0.00.20.40.60.81.0
In remission, PB.blast=0
Active Disease, PB.blast=0
Active Disease, PB.blast>0
p<0.0001
Relapse is main cause of treatment failure
with Allogeneic HSCT for AML
6. Fundamental Problems with
HSCT
• Graft-vs.-malignancy which naturally
occurs post transplant is relatively weak
• Graft vs. Malignancy associated with
GVHD
• Relapse remains the major cause of
treatment failure
• Resistant infections can occur due to
post transplant immune deficiency
8. • T-cells that down regulate immune responses
termed regulatory T cells have been identified.
• CD4+CD25+FoxP3+
• Challenge to separate from Tconv
• Cord Blood vs. Peripheral Blood
• Can suppress GVHD
• Clinical Trials
• Natural T regs
• Inducible T regs
Regulatory T-Cells (Tregs)
9. Cord Blood Treg Expansion
and Activation
•Anti-CD3/antiCD28-coated beads.
•Supplemented with IL-2 300 IU/mL
Reduced incidence of grade II-IV aGVHD (43% vs 61%)
Brunstein et al Blood 2011
CD25 Selection Culture
10. Clinical outcomes of patients after nonmyeloablative umbilical cord blood transplantation who
received Treg ≥ 30 × 105/kg (dotted line; n = 18) and historical controls (solid line; n = 108).
Brunstein C G et al. Blood 2011;117:1061-1070
11. Questions with Tregs
• Production process, separation of Tregs
from Tconv
• Cell Dose
• Administration with calcineurin inhibitors
vs. sirolimus
• Impact on GVL effects?
12. Suicide Switch to Abrogate
GVHD
• Genetically modify T-cells to introduce
gene to induce apoptosis upon
treatment with an activating drug
• Herpes virus tyrosine kinase – activated
with ganciclovir
• Modified Caspace 9
19. Anti Viral T-cells
• Initial studies indicate feasibility and
suggest efficacy (CMV, EBV)
– Effective for EBV-LPD
• Rapid production techniques have been
developed
• Difficult to use in patients with GVHD-
must avoid high dose steroids
• Donor specific products
• Off the shelf 3rd party CTLs under study
21. Donor Lymphocyte Infusion
• Effective treatment for EBV-LPD,
relapsed CML, CLL, indolent NHL; less
effective for relapsed AML and ALL
• Planned DLI studied to enhance GVM
effects
• Frequently complicated by GVHD
– Related to cell dose, time post transplant
– Escalating cell dose
22. Targets for Graft-vs.-Malignancy
Broadly expressed minor
histocompatibility antigen (GVHD)
Lineage restricted
minor histocompatibility
antigen (G-vs-hematopoietic),
or Redirected CAR T-cells vs CD19
Aberrant overexpressed
normal cellular constituent
(Proteinase 3, WT1,
telomerase)
Allo-Specific Malignancy Specific
Idiotype, Fusion peptide of
translocation (bcr-abl)
23.
24. Shared Resources
Flow Cytometry and Cellular Imaging Facility, Genetically Engineered Mouse
Facility, Monoclonal Antibody Facility; Clinical Trials Support Resource
Antigen-Specific Immune Therapy for AML
P3
NE
Leukemia
PR1 peptide
PR1
PR1-CTL
Clinical trials with cord blood-derived
PR1-CTL are ongoing for transplant
recipients (AML, CML)
PR1-CTL are naturally enriched (0.1-0.4%) in fetal cord blood
AML
No AML
Molldrem et al
25. Redirect T-cell Specificity through the
Introduction of Chimeric Antigen Receptors
(CARs)
vL
vH
CH1
CL
Antibody
Fab
vH vL
Chimeric antigen receptor
TCR-complex
26. Production Methods
• Retroviral vectors
• Letiviral vectors
• Non viral systems, Sleeping Beauty
• Expansion using artificial APCs
28. 2nd and 3rd Generation
Chimeric Antigen Receptors
Propagation on Artificial APCs
Cooper et al
41BB
29. Chimeric Antigen Receptor T-cells
• Can target nonimmunogenic targets,
tissue/tumor specific antigens. Most
experience targeting CD19 for B-cell
lymphomas, CLL and ALL
• First, second and third generation constructs
including costimulatory molecules CD28,
CD137 enhance survival of the cells in vivo
and their proliferation
• Optimal design of CAR not established
– Affinity of antibody receptor, spacer, costimulatory
molecules, coexpressed receptors, homing
molecules
30. Clinical Trials of CAR T-cells
• lymphodepleting chemotherapy and
autologous CAR T-cells
• some complete remissions, eradicating
CD19+ cells (reported studies N=32;
CR-3 PR-10)
• Small number of HSCT patients treated
with autologous or allogeneic CAR+
cells
• Durable elimination of CD19+ normal B-
cells
31. Anti CD19 CAR T-cells for CLL
Porter DL et al. N Engl J Med 2011;365:725-733
32. Serum and Bone Marrow Cytokines before and after
Chimeric Antigen Receptor T-Cell Infusion.
Porter DL et al. N Engl J Med 2011;365:725-733
33. CAR Problem Areas
• Autologous vs. Allogeneic
• Survival, homing in vivo
• In vivo expansion needed for activity
• Toxicity, “cytokine storm” may occur,
particularly with CD137 containing
CARs- can produce respiratory failure
• Time/ expense in producing patient
specific products
• Complex, regulatory considerations
make multicenter studies difficult
36. NK Cells
• Component of innate immune system
• CD3- TCR-, CD16+, CD56+
• Mediates anti-tumor, anti-viral, BM rejection
• Activating and inhibitory receptors (KIR)
• Cytotoxicity governed by missing ligand hypothesis re:
inhibitory receptors
– Cw alleles that bind to KIR2DL1 have amino acid K at
position 80.
– Cw alleles that bind to KIR2DL2 or to KIR2DL3 have amino
acid N at position 80
– Bw4 or Bw6, KIR 3DL1 amino acids at positions 82-83
• Missing ligand model has “not” predicted
responses in most clinical trials
39. NK Cells- Clinical
• NK reactivity reported to reduce relapse
in AML following haploidentical
transplants
• Human studies infusing “selected” NK
cells (CD3-depleted +/- CD56 selected)
demonstrate safety, activity.
– Limited by low and variable frequency (5-
15%) in normal donors, cannot collect
more than 106/kg by apheresis
– NK cells already in PBPC, CB or BMT,
adding low doses from donor unlikely to
benefit
• Ex vivo expansion feasible, entering
human clinical studies
40. 4 Log expansion of NK cells using
mbIL21 APCs
Cryopreserve in
aliquots
41. IL-2 or IL-15
Haploidentical
Allo reactive
NK Cells
Busulfan
Fludarabine
Donor, Haploidentical
or Cord Blood
NK Cells
IL-2
Allo match
PBPC
Melphalan
Fludarabine
Haploidentical
Allo reactive
NK Cells
Haplo
BMT Cy-tacro-MMF
Flag-ida
43. Conclusions
• Adoptive cellular immunotherapy is a promising novel
treatment modality for treatment of cancer.
• Cellular immune therapy is a promising approach to
control alloreactivity to prevent GVHD. Tregs
successful to prevent GVHD in mice; improved
approaches needed to achieve similar benefit in man.
• Antigen specific CTLs and CAR T-cells can eradicate
experimental tumors. Preliminary human clinical trials
have been performed with autologous and allogeneic
cells, demonstrating activity and feasibility in
conjunction with HSCT.
44. Where do we go from here?
• Rapidly evolving technology; optimal
cellular designs and production
methods need to be determined.
• Need widely accepted products which
can be taken into larger scale phsae II
and III clinical trials.
• The needed multicenter “gene therapy”
trials will costly and complex to
administer
Notes de l'éditeur
Q6. retroviral vector issues.Adenoviral vector used for transient transfection situations Retroviral vector for stably transfected situations with low oncogenic risk (e.g. T cells)Lentivirus for stem cells (in development)
