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Cellular therapies in Acute Lymphocytic Leukemia
- 1. ELIZABETH J SHPALL, MD Houston, USA • Professor of Medicine and Chair of Cord Blood Transplant Program, UT MD Anderson Cancer Center • As a nationally recognized expert in stem cell transplantation, Medical Director of the GMP Cell Therapy Laboratory since 2002, and the Director of the Cord Blood Bank (CBB) at M. D. Anderson Cancer Center since 2004, Dr. Shpall's research covers a range of applications, among them: leukemia, lymphoma treatment, anti-tumor immunity enhancement, and tissue repair. Dr. Shpall’s influence is felt beyond the Texas Medical Center. She is the current vice-president of FACT Netcord, having chaired their Cord Blood Standards Committee since 1999, and she was the founding president and has been a board member of the Foundation Accreditation of Hematopoietic Cell Therapy since 1995.
- 2. Cellular Therapy for ALL with Chimeric Antigen Receptors Elizabeth J. Shpall MD •
- 3. Roadblocks to Successful Cellular Immunotherapy for ALL PROBLEM •Targeting •Expansion ex vivo •Proliferation in the host •Persistence •Effector:Target ratio – create sufficient effector cells for clinical success SOLUTION •CAR or TCR •GMP cell culture •?Young T cells •?Memory T cells •Ideally, there would be an efficacy signal, not just safety, in phase 1 Abbreviations: CAR, chimeric antigen receptor; GMP, good manufacturing practice; TCR, T-cell receptor
- 4. Gene Transfer of CARs Eshhar et al; PNAS 1993 Intracytoplasmic vscFv v Spacer Linker vLvLvLvH abTcR complex ge edzz Monoclonal antibody Monoclonal antibody T or NK Cell TumorTumor Antigen CARCAR CARs combine an antibody binding domain (scFv) that recognizes a desired tumor associated antigen with one or more T-cell receptor signaling endodomains. Forced expression of CAR on the T-cell surface leads to activation of T cells through CD3ζ CD28 and/or CD137.
- 5. Chimeric Antigen Receptors Possess Advantages Over Endogenous Receptors • Intact molecule recognized (including non-protein) • HLA- independent • Costimulation not needed CAR T Cells Endogenous T cells • Recognition of peptides processed through MHC • Need • costimulation Chimeric Antigen Receptors Possess
- 6. Redirecting the Specificity of T cells • Gene transfer technology is used to stably express CARs on T cells, conferring novel antigen specificity • Transduction: – Many retroviral – Some now using lentiviral – Non-viral Sleeping Beauty system • Temporary modification with RNA transfection • Suicide systems Viral or nonviral vector T cell CD19 Native TCR Tumor cell CTL019 cell Dead tumor cell Anti-CD19 CAR construct
- 7. Evolution of CARs with additional costimulatory domains Dotti et al. 2009 th this is hhh ext tt e x t Classic CARs Sports CARs Muscle CARs Sadelain, et al. Cancer Discov. 2013
- 8. CART19: Chimeric Antigen Receptor T cells against CD19
- 9. • Clinical scale up: no feeder cells required • Expansion >106 -fold • Repertoire preserved • T cells persist after infusion • Maintains function and homing of T cells • Induction of telomerase: minimizing replicative senescence Levine et al J Immunol 1997; Carroll et al Science 1997; Weng et al Imm Rev 1997; Humeau et al Mol Ther 2004 Properties of Anti-CD3/CD28 Costimulated Expansion System
- 10. Sleeping Beauty system to genetically modify T cells to target CD19 via CAR Transposon (Donor) sequences flanked by inverted repeats are integrated into genome Transposase (Helper) expression is transient Plasterk RH, Cell 74(5):781, 1993 Ivics Z, et al, Cell 91 (4):501, 1997 Nucleus Transposase Transposon CAR Cytoplasm Laurence Cooper Laboratory
- 11. Schematic of T-cell manufacture Methods available at: Huls MH, et al. Clinical application of Sleeping Beauty and artificial antigen presenting cells to genetically modify T cells from peripheral and umbilical cord blood. J Vis Exp. 2013 Feb 1;(72):e50070. SB system with Perry Hackett aAPC with Carl June Irradiated aAPC Derived from K-562 cells and modified to co-express CD19, CD86, CD137L, membrane-bound IL-15 (and CD64)
- 12. Authors Target Disease N Positive Outcomes City of Hope 2010 Follicular lymphoma (FL) 2 -- NCI 2010 FL 1 1 PR BCM 2011 DLBCL, FL 6 -- U. Penn 2011 CLL 3 2 CR 1 PR Memorial SK 2011 CLL, ALL 9 1 PR, 1 cCR CARs Targeting CD19 Initial Results: 2010, 2011
- 13. Author CD19+ Disease N infused Vector/ Activation Endo- domain Chemo CAR+ cell Response Persist Key Pts Cruz BCM 152 Post Allo 9 Retrovirus EBV/CMV/A deno CD28/ CD3z None Donor T 2/6 DLI bulk PR->PD 3/3 adult 100% CR 9 wks Novel CAR Bispecific Trivirus Davila MMSK 69 ALL 13 Adults Retrovirus CD3x28 b CD28/ CD3z Lympho depleting Auto-pbpc 106 /Kg 10/12 MRD- 6 revers CS 83% CR NR Potent bridge to SCT Grupp Penn 67 ALL 16 Peds 4 Adults Lentivirus CD3x28 b 41BB/ CD3z Lymph- depleting Auto-PB 11 Post Allo 107 -108 /Kg 14/17 82% CR CS in responders 1-15+ Mos respon Potent No GVH with hx Kalos Penn 163 CLL -24 ALL-14ped 3 adults Lentivirus CD3x28 b 41BB/ CD3z Lymph- depleting Auto-PB 5CR 7PR 50% CR 8/17 47% Max5% All resp and SD Deep seq CRs Lee NCI, Peds 614 CD19+ ALL 7 NHL1 Retrovirus CD3x28 b. CD28/ CD3z Flu-Cy Auto-PB Pre/Post Allo 4/4 ALL CR5/7 71% All pts 1-38% 55days Active Fresh Infusion ASH 2013 CAR Clinical Abstracts
- 14. • Disease Target: CD19+ B ALL • Subjects: 16 adults • Vector: Retrovirus • Manufacturing: CD3/28 Beads • Signaling Endodomains: CD28 and CD3 zeta • Pre T Cell Prep: Cyclophosphamide • Overall 14 CR (88%) – higher than salvage chemo (44%) • CAR T cells associated with CRS Chimeric Antigen Receptor (CAR)Memorial Sloan Kettering Experience (Davila et al 2014)
- 15. U. Penn Experience (Maude et al 2014) • Disease Target: CD19+ B ALL • Subjects: 25 between 5-22, and 5 older adults • Vector: Lentivirus • Manufacturing: CD3/28 beads • Signaling Endodomains: 41BB and CD3 zeta • Pre T Cell Prep: None, Cy/VP, Flu/Cy, Etoposide/Cytarabine, Cyclophosphamide, CVAD-B, CVAD-A, or clofarabine • 27 CR (90%), 6 month EFS 67%, overall survival 78% • T cells detectable in blood, bone marrow, and CSF of patients who responded • All patients had CRS (mild to moderate in 22/30, severe in 8) University of Pennsylvania Experience (Maude et al 2014)
- 16. Modified T cells detectable at high levels by flow cytometry with longer peristence Grupp, et al. NEJM 2013
- 17. NCI Experience (Kochenderfer et al 2014) • Disease Target: CD19+ B cell malignancies (DLBCL, indolent lymphomas, CLL) • Subjects: 15 adults • Vector: Retrovirus • Manufacturing: 10-Day Culture • Signaling Endodomains: CD28 and CD3 zeta • Pre T Cell Prep: Cyclophosphamide and fludarabine • 8 CR, 4 PR, 1 SD, 2 not evaluable • Acute toxicities (fever, hypotension, delirium, neurotoxicity) resolved within 3 weeks; 1 patient died 16 days post infusion • 1st successful treatment of DLBCL with anti CD19 CAR T cells NCI Experience (Kochenderfer et al 2014)
- 18. Pediatrics NCI Experience (Lee et al 2014) • Disease Target: CD19+ ALL or Non-Hodgkin’s lymphoma • Subjects: 21 children and young adults (1-30 years of age) • Vector: Retrovirus • Manufacturing: Bead Stimulated 11 Day Culture • Signaling Endodomains: CD28 and CD3 zeta • Pre T Cell Prep: Cyclophosphamide and fludarabine • 12 CR (MRD Neg), 1 CR, 1 Cri, 3 SD, 4 PD • All toxicities fully reversible Peds NCI Experience (Lee et al 2014)
- 19. MD Anderson Kebriaei et al. 2014: Lymphodepletion and Sleeping Beauty Autologous CAR+ T Cells for Active Disease Age Dx Stage CAR-T source Dose Level (%) CAR expression Ds. Status post CAR Toxicity 33 ALL Refractory relapse MSD 106 /m2 94.0 Progressed, 1 mo. None 25 ALL s/p haploHCT,Refractory Haplo 106 /m2 95.0 Progressed, 6 mo. None 53 NHL Recurrent MSD 106 /m2 58.3 Cru, 6 mo. None 21* ALL s/p alloHCT, Rel 3 MSD 107 /m2 96.8 Progressed, 9 mo. None 31* ALL s/p alloHCT, Rel 2 MSD 107 /m2 93.4 CR, 1 mo. VOD/ GVHD 23* ALL s/p alloHCT, Rel 2 MSD 5x107 /m2 96.5 Progressed, 1 mo. None 58 ALL s/p alloHCT, CR2, MRDpos alloSelf 5x108 /m2 76.7 CR, 6 mo. None 52 NHL s/p autoHCT, refractory Self 5x108 /m2 89.7 Progressed, 1 mo. None 29 ALL s/p alloHCT, CR3, MRDpos alloSelf 5x108 /m2 90.1 CR, 3 mo. GVHD 76 CLL Refractory relapse Self 5x108 /m2 31.5 Stable, 12 mo. None 63 CLL Refractory relapse Self 5x108 /m2 57.3 Progressed, 1 mo. None 76 CLL Refractory relapse Self 5x108 /m2 32.9 Progressed, 2 mo. None 31 ALL s/p alloHCT, refractory CNS alloSelf 5x109 /m2 88.9 CNS progressed, 1 mo. None 63 CLL s/p alloHCT, refractory alloSelf 5x109 /m2 92.2 Too early None N=14 pts treated, 5 with disease regression med. 6 mo follow-up
- 20. • Extraordinarily high ferritin levels after highly active CAR infusion suggest MAS/HLH/CRS • 16,000 to 415,000 in Penn studies • Mild coagulopathy, transaminitis • Elevated D-dimer and low fibrinogen • High fevers, hypotension • Moderate marrow hematophagocytosis Macrophage Activation Syndrome (MAS) Hemophagocytic Lymphohistiocytosis (HLH) Cytokine Release Syndrom (CRS) “Storm”
- 21. Cytokine Release Syndrome (CRS) • Can be significant, but highly variable across patients and studies • May or may not scale with dose, may scale with disease burden • ? Different with different diseases • Different in the era of highly active CAR cell therapy • IL-6 is an important driver of toxicity • Entirely reversible with cytokine blockade – IL-6R blocking agent tocilizumab
- 22. Tocilizumab (Actemra) • IL-6 receptor antagonist • Blocks IL-6 mediated effects • Indicated in: – juvenile idiopathic arthritis (JIA) – Rheumatoid arthritis (RA) – In Japan, indication for Castleman’s Disease • Typically given monthly • Rare side effects of transaminitis and neutropenia • Penn uses 8 mg/kg
- 23. Blinatumomab (CD3-CB19 BITE) causes HLH/MAS as well, reversible w/ tocilizumab Elevated cytokines •IL-10 (5338 pg/mL) •IL-6 (681 pg/mL) •INF-γ (192 pg/mL) •IL-2R (4872 pg/mL) David Teachey, Stephan Grupp. Blood, 2013 Blinatumomab (CD3/CD19 Bite) causes HLH/MAS as well, reversible w/ tocilizumab
- 24. Neurotoxicity issues • Seen in several CD19 trials: – NCI – CHOP/U. Penn – CD19-CAR-T and Blinatumomab • Self-limited, generally untreated, fully resolves • ? Related to CRS • Not prevented by tocilizumab
- 25. • CD19-directed CAR T cells • Adverse events related to cytokine release syndrome occur --- but most are manageable • Partnerships with industry bring technology closer to being part of clinical care • In July 2014, FDA designated CTL019 CAR T cells with “breakthrough therapy” status • Improvements in CAR therapies underway CAR Therapy: Summary Recent Trials
- 26. • Persistence: Correlates with Outcome • Vector • sCFv or Endodomain Construction • Bulk of Disease • Cell Type Modified (auto vs allo) CAR Therapy: Outstanding Issues
- 27. CAR Therapy: Outstanding Issues • Ex Vivo Expansion • CD3x28 Beads • OKT3/IL2 • Antigen Presenting Cells • Cytokines • Duration of Ex Vivo Culture • Pre-Infusion Therapy: Lymphopenia? Treg Decrease? • No Therapy • Lymphodepleting Chemotherapy • Stem Cell Transplant CAR Therapy: Oustanding Issues
- 28. CAR Therapy: Outstanding Issues • Homing • CNS • Other Sanctuarial Sites • On Target Toxicity: B Cell Aplasia, Cytokine Storm Syndrome • Tocilizumab • Conditional Ablation: Caspase, HSV-TK CAR Therapy: Oustanding Issues
- 29. • Consolidate patients with MRD What is the Potential for CAR Therapy in ALL? • Reinduce remission • Produce MRD(-) state prior to SCT (Bridge to SCT) • Multicenter trial/s in pediatric ALL are enrolling Potential Roles for CAR Therapy in ALL
- 30. Acknowledgements Richard Champlin Katy Rezvani Ian McNiece Laurence Cooper Marcos de Lima Amin Alousi Simrit Parmar Chitra Hosing Nina Shah Borje Andersson Michael Andreeff Simon Robinson Steven Kornblau Yago Nieto Hong Yang Roy Jones Mark Munsell Xiaoying Liu Martin Korbling Paolo Anderlini Qaiser Bashir Jeffrey Molldrem Patrick Zweidler-McKay Michael Thomas Partow Kebriaei Sue Armitage Tara Sadeghi Laura Worth Demetri Petropoulos Indreshpal Kaur Mark Munsell Sairah Ahmed Muzaffar Qazilbash Qing Ma Issa Khouri Jared Burks Lisa St John Beatriz Martin-Antonio Gabriela Rondon Doyle Bosque Lori Griffin Kai Cao Rima Saliba Peter Thall Betul Oran Funding NCI R01- CA061508 NCI PO1- CA148600 CPRIT RP100469 CPRIT RP 100430 CLL Global Research Fund HRSA 234200737 Funding NCI R01- CA061508 NCI PO1- CA148600 CPRIT RP100469 CPRIT RP 100430 CLL Global Research Fund HRSA 234200737 Catherine Bollard Michael Keating William Wierda
- 31. Redirected by gene transfer * Ex Vivo Approaches for Adoptive T Cell Therapy
Editor's Notes
- Genetically modify T-cells to express an antigen recognition domain of a specific antibody, in this case CD-19. Traditionally did not lead to in vivo expansion. The newest generation of CAR T-cells were enhanced by adding a co-stimulatory domain. In our model the co-stimulatory domain is 4-1BB (CD137) CARs are not all the same. The co-stimulatory domains were murine but now humanized to try to further minimize rejection.