Richard P Junghans, PhD, MD

Transcription

1 CANCER GENE THERAPY WITH DESIGNER T CELLS: TOXICITY VERSUS ACTIVITY Richard P Junghans, PhD, MD Director, Biotherapeutics Development Lab Associate Professor of Surgery and Medicine Boston University School of Medicine Chief, Division of Surgical Research Roger Williams Medical Center Providence, RI, USA No commercial relationships to disclose.

2 Anti- CEA Normal T cells Designer T cells IgTCR = Immunoglobulin-T cell receptor CIR = Chimeric immune receptor

3 Carcinoembryonic antigen (CEA) o Expression High on tumor, low on normal Topological sequestration o High clinical relevance: On colorectal, breast, pancreas, lung, others More than 100,000 deaths/yr for CEA+ tumors

4

5 CLINICAL TRIAL Clinical Data: 1 st Generation Phase I Study of Anti-CEA Designer T Cells in Adenocarcinoma ( 1st generation ) FDA BB IND 7301

6 TCR Anti-Cancer T Cell Gene Therapy Gene-Modified TCR

7 Interventions o Phlebotomy/Apheresis o Isolate patient s peripheral blood mononuclear cells (PBMC) o Activate/transduce with IgTCR o Expand in IL2 o Harvest cells equal to dose; Infuse o Monitor for Toxicity/Response

8 Clinical Summary o Number of doses administered (24) o Patients treated (7): 5 colorectal, 2 breast o Doses sizes administered 1 x 10 9, 3 x 10 9, 1 x 10 10, 3 x 10 10, 1 x cells

9 Pharmacokinetics Rapid Systemic Loss

10 CEA (NG/ML) Response: Proof of Principle T Cells CEA Increasing pain Pain resolved Day of Treatment BUT! Time-Limited in Duration

11 Pharmacodynamics o Immunogenicity: None. 0/7 with plasma reactivity against designer T cells o IL2 arm: Systemically active. Positive for NK cell expansion, T cells stable

12 Toxicity o Probably related or Definitely related No grade III toxicity, one grade IV toxicity (grade II fever >> grade IV SVT) No delayed grade III, IV toxicity Positive for low grade fevers, mild GI symptoms (<grade III) Transient hypoxemia (O 2 sat<90%) with high T cell dose

13 (-) IL 2 Toxicity Name Sex Age Diagnosis Dose Fever (24h) Adverse Events (Grade III-IV) Relatedness MS Dose 1 (6/10/98) Dose 2 (6/24/98) Dose 3 (7/8/98) F 75 Colon Ca 1x10 9 3x10 9 1x F Bilirubin GI Bleed * Death * Possibly (temporal only) # Possibly (temporal only) # RH Dose 1 (7/13/99) Dose 2 (7/27/98) Dose 3 (8/10/98) Dose 4 (8/26/98) M 55 Rectal Ca 1x10 9 3x10 9 1x x F 101 F <101 F <101 F Anemia Possibly (temporal only) # DB Dose 1 (7/15/98) Dose 2 (7/29/98) Dose 3 (8/12/98) Dose 4 (8/26/98) M 54 Colon Ca 1x10 9 3x10 9 1x x F Rigors F Bilirubin (9/23/98;28 days) Possibly (temporal only) # HF Dose 1 (8/11/98) Dose 2 (8/25/98) Dose 3 (9/14/98) Dose 4 (10/14/98) Dose 5 (2/1/99) M 75 Colon Ca 1x x x x x A Flutter* A Flutter Death * (2/3/99) Possibly (temporal only) Possibly (temporal only) # GT Dose 1 (6/8/99) M 69 Rectal Ca 1x F SVT/Hypotension* Rigors Probably (to fever) (+) IL 2 Name Sex Age Diagnosis Dose Fever (24H) Adverse Events (Grade III-IV) Relatedness JD Dose 1 (9/23/98) Dose 2 (10/5/98) Dose 3 (10/19/98) Dose 4 (11/2/98) F 39 Breast CA 1x10 9 3x10 9 1x x F 102 F EM Dose 1 (11/4/98) Dose 2 (11/16/98) Dose 3 (11/30/98) F 47 Breast CA 1x10 9 3x10 9 1x F Bilirubin Death* Possibly (temporal only ) # Possibly (temporal only) # * Event generating SAE report # Current progression

14 Patient GT o 69M, rectal ca, baseline RBBB Febrile 103.8F (39.9C) within 6 hrs SVT, responsive to IV beta blockers blood pressure, 20 mm drop asymptomatic o blood cultures neg, cell cultures neg o expect it: >>immune response indicators recommend as pharmacodynamic measure o avoid predisposing cardiac conditions

15 Temperature Heart Rate GT Vital Signs Temperature Heart Rate :18 13:00 13:40 14:34 14:50 15:20 17:34 22:34 0:10 5:00 11:00 18:00 21:00 22:00 15:00 22:00 24:00:00 2:00 Time (Hour)

16 CEA (NG/ML) T Cells CEA Increasing pain Pain resolved Day of Treatment

17 GVHD in bowel Apoptotic bodies at base of crypts

18 GT biopsy: no apoptotic bodies Conclude: cytokine syndrome

19 CEA targeting by TCR Parkhurst et al, ASGCT 2010 o Cloned murine TCR for HLA-A2 presented CEA peptide o TCR gene transfer (not CAR ~ 1 st gen), 3 subjects Lymphodepletion 2-4 x 10^8 cells 1-20% engraftment o Toxicity against bowel, all had grade 2/3 diarrhea T cell infiltration epithelial destruction o Some tumor partial response noted o However, concluded not safe to target CEA What is the difference???

20 MHC class I on enterocytes Topology MHC I: Primarily basal expression

21 CEA on enterocytes Antigen sequestration: CEA confined to microvilli

22 Lesson on Ag selection o What are normal tissues expressing? o What is topology of expression? Epithelial cancers Antigens Expression CEA Lumenal Class I Basal Her2 Basal CA IX Basal

23 Liver, etc Infuse Causes of Activity Loss? Tissues T T * Blood T #1 #2 T * Cancer

24 New Directions: Hypotheses o Tumor eradication would follow if EITHER #1. T cells persisted systemically [QUANTITY] ( bypass co-stimulation ) OR #2. T cells expanded intratumorally [QUALITY] ( provide co-stimulation )

25 Hypothesis 2: 2 nd Generation Provide co-stimulation Incorporate Signal 2 into designer T cells

26 Immunology 101

27 T Cell Activation Antigen Presenting Cell MHC B7 TCR CD T Cell o o o Gene expression - Cytokines (IL-2, 4, IFN-, etc) - Surface molecules (CD25, CD40L, etc) Cytotoxicity Proliferation

28 Designer T Cells First Generation o IgTCR chimeric immunoglobulin T cell receptor Tumor Cell Antigen Ig TCR 1 Modified T Cell CD28 MIPCEA Advantage: IgTCR provides Signal 1: adequate T cell cytotoxicity. Disadvantage: Lacking Signal 2, undergoes Activation-Induced Cell Death (AICD) after killing target cells. [HYPOTHESIS]

29 Signals o Signal 1: Activated: T cells kill tumor >> and die by AICD Resting: anergy o Signal 1+2: Activated: T cells kill tumor >> and proliferate Resting: activation, see above

30 1 st and 2 nd Gen Constructs IgTCR Ig28TCR ( Tandem ) VL VH VH VL VL VH VH VL Signal 1 Signal 1+2

31 2nd Gen T Cell Tumor-Induced Proliferation Expansion on MIP101 Expansion on MIPCEA 1.0E+07 CEA(-) 1.0E+07 CEA(+) 8.0E E E E+06 UnTd IgTCR Tandem 6.0E E+06 UnTd IgTCR Tandem 2.0E E E+00 Day 0 Day 3 Day 7 0.0E+00 Day 0 Day 3 Day 7 Day Day Emtage et al. Clin Cancer Res 2008

32 Adjuvant model: Superior in vivo Tumor Suppression by 2nd Gen T Cells Tumor size (mm ) 2 Tumor size (mm ) 300 MIP MIPCEA 250 CEA(-) 250 CEA(+) Untransduced IgTCR Tandem 200 2Untransduced IgTCR 150 Tandem Day post tumor cell injection Day post tumor cell injection

33 Phase Ia/Ib Trial of 2 nd Generation Anti-CEA Designer T Cells in Adenocarcinoma FDA BB IND 10791

34 Hypotheses o 2nd gen Signal 1+2 designer T cells will escape AICD and proliferate intratumorally o Expanding designer T cells in tumor will maintain anti-tumor activity until tumor elimination

35 -IL2 Pt # Cohort 1x10 9 1x x10 11 #1 #2 #3 #4 #5 #6 I II X X X T Cell Dose, Number of Cells X X X Phase Ia Safety Study Design #7 #8 #9 III X X X Any CEA+ cancer Funded by FDA Orphan Products Division

36 Summary 2 nd Generation CEA o 5 patients treated 1 st cohort completed 2 nd cohort in process o Safety, no SAEs (no new risks with 2 nd gen) o Responses one minor (brain and lung) one SD 12+ months o Continue in escalation o Need to assess value of IL2 supplementation

37 Hypothesis #1 T Cell Engraftment Auto-Transplant: Engraft designer T cells via lympho-expansive capacities of the body after lympho-depletion treatments (Bypass co-stimulation)

38

39 NMA Melanoma TILs Tumor Harvest Melanoma CD8+ TIL X Non-myeloablative (NMA) Conditioning Hematologic Recovery 6/13 major responses Tumor Response Dudley et al Science 2002;298:850

40 Prostate Cancer T Cell Harvest CIR+ Ex vivo gene therapy Anti-PSMA designer T cells Non-myeloablative (NMA) Conditioning +IL2 low dose (outpatient) Hematologic Recovery CIR+ CD3+ Tumor Response

41 Prostate Specific Membrane Antigen (PSMA) o Surface membrane glycoprotein 100,000 Daltons o Unrelated to PSA o Normal prostate epithelium and vasculature (reportedly on type II astrocytes?) o Elevated expression in metastatic lesions and hormone refractory disease o High clinical relevance: 28,000 deaths per year from PSMA+ prostate tumors o Antibody (3D8) from G. Murphy and A. Boynton

42 PSMA on prostate cancer

43 Normal tissue PSMA Tasch et al. Crit Rev Immunol 2001 Prostatic epithelium and vasculature Dispensable white gray And brain??!! Kinoshita et al. W J Surg 2006 Sacha et al. Neuroscience 2007

44 Thought: If DTC toxic against brain, then best to have a format that allows T cell suppression and escape T cell quantity: Can be controlled with infusion Engraftment with vast numbers hard to control But: No 1 st gen infused will be therapeutically successful T cell quality (potency): After remove steroid suppression: 1 st gen DTC won t reactivate 2 nd gen will Conclude: 1 st gen safe with engraftment (2 nd gen safe with infusion)

45 Phase I Study of Autologous Transplantation of Anti-PSMA Designer T Cells after NMA Conditioning in Prostate Cancer BB IND 12084

46 Treatment Schema Designer T cells modify T cells microbiologic testing NMA T cell infusion T cell collection G-CSF PSC collection chemotherapy start IL end IL2-21* -20* -16* Study Day Biopsy* CTX 60 mg/kg d-7, d-6 Fludarabine 25 mg/m2 d-5 to d-1

47 Phase I Study Enrollment Plan o T Cell Dose, Number of Cells o Pt # Cohort o #1 X o #2 I X o #3 X o #4 X o #5 X o #6 X o #7 X o #8 II X o #9 X o #10 X o #11 X o #12 X o (Bx) o #13 X o #14 III X o #15 X o #16 X o #17 X o #18 X omonitoring Safety Designer T cell persistence/expansion in blood In tumor Tumor response Funded in part by US Army

48 Counts per ul Counts per ul Peripheral Blood Recovery o Chemo d-8 to d-2 o T cells d0 o IL2 start d0-d28 o ANC=0 x5-8 d Recovery ANC> Pt #1 ANC and ALC over Time T cells days ANC ALC T cells days AC ALC

49 IL2 (pg/ml) IL2 via Continuous Infusion IL2 of JN IU/ml prescreen preinfusion post- 0 hr post 1 hr post 3 hr post 8 hr post 24 hr post 48 hr post-day 5 post-day 7 post-day 14 post-day 21 post-day 28 75,000 IU/kg/d ~ 3 MIU/m2/d

50 Engraftment 1.1% 2.5% Blood sample Day +14 #1 CIR+ CD3 61% #2 CIR+ 7.3% Dose CD3+

51 PSA PSA PSA Response 3.50 T cells Conditioning d-8 to d-2 T cells infused d0 Low dose IL2 d0 to d chemo Days T cells chemo Days

52 Summary Prostate o Five patients treated Safely administered; toxicity due to conditioning Neutropenic fevers, hospitalization for antibiotics Malaise, fatigue (IL2) 50 and 70% reduction in PSA in two subjects o More doses to follow in escalation 10^10 10^11 o Limitations??? 1st generation: signal 1 only, After resting, do not reactivate 2 nd generation in preclinical o Conclude: PSMA safe for DTC targeting

53 Considerations for Antigen Targeting o The target Normal tissues that express Essential/Dispensible? Topology of expression (epithelial) Unknowability is central to safety testing design Safety with Ab targeting no assurance for T cells o The method of targeting Potency: Co-stimulation, can reactivate after suppression Quantity: Infuse vs engraft Infusion allows control of patient exposures (DLI)

54 T cells homing in on target THE END

55 Table 1. Features of designer T cell versions On Contact with Antigen. CIR Activated T cells Resting T cells 1st gen Signal 1 killing, AICD anergy 2nd gen Signal 1+2 superactivation, killing, activation, killing, cytokines, proliferation cytokines, proliferation

56 Table 2. Features of T cell administration methods Conditioning Risk? In blood In tumor Cost* Infuse None No Transient Transient $5-10K Engraft Chemo, XRT Toxic Stable Sustained $60-100K *Clinical, non-manufacturing costs of patient treatment and monitoring. (Manufacturing ~$15K for 10^11 dose; less for lower doses.)

57 [ QUALITY ] Table 3. Options matrix for designer T cell Strategies (2x2) [ QUANTITY ] Administration method Infuse Engraft Designer 1st gen 1 2 T cell version 2nd gen is untested state Strategy 1 = 1 st gen infused Strategy 2 = 1 st gen engrafted Strategy 3 = 2 nd gen infused Strategy 4 = 2 nd gen engrafted

58 Table 4. Likely toxicity response of auto-reactive DTC to immune suppression on different Strategies On steroids Off steroids Comment 1 st gen only 1 st gen; 0 1 Suppress Suppressed don t reactivate 0 2 Suppress Suppressed don t reactivate 2 nd gen 2 nd gen reactivate; 0 3 Suppress Suppressed small numbers* 0 4 Suppress Autoimmune very large numbers *Known from BMT that dosage of allo-reactive T cells in donor lymphocyte infusions (DLI) critical to safety. Low dose controlled endogenous mechanisms (Treg, etc). High dose uncontrolled except with max suppress, lethal.

59 Safe paths of Strategy Escalation for exposures to designer T cells 1. Avoid maximum exposure for first-inhuman DTC tests for Ag (Strategy 4) 2. Once a Strategy safely tested for Ag, other Strategies safe for that Ag 3. But NOT all paths advisable (---) 0 is untested state Strategy 1 = 1 st gen infused Strategy 2 = 1 st gen engrafted Strategy 3 = 2 nd gen infused Strategy 4 = 2 nd gen engrafted

60 Optimal paths of Strategy Escalation for exposures to designer T cells All paths to 2 nd gen engrafted pass through 2 nd gen infused 1. Lower hazard 2. Lower cost 3. Better science: Test hypothesis of more signals to create effective immune response (without engraftment) NB: No 2 nd gen DTC has yet been tested by infusion to full doses with appropriate cytokine support; hypothesis of sufficiency OR deficiency still untested. 0 is untested state Strategy 1 = 1 st gen infused Strategy 2 = 1 st gen engrafted Strategy 3 = 2 nd gen infused Strategy 4 = 2 nd gen engrafted

61 Considerations for Antigen Targeting o The target Normal tissues that express Essential/Dispensible? Topology of expression (epithelial) Unknowability central to safety testing design Safety with Ab targeting no assurance for T cell o The method of targeting Co-stimulation, can reactivate after suppression Infuse vs engraft Infusion allows control of patient exposures (DLI)

62 T cells homing in on target THE END

63