Off-the-Shelf Cellular Immunotherapy with Cord Blood-Derived Natural Killer Cells Loaded with AFM13 for CD30+ Malignancies

Transcription

1 Off-the-Shelf Cellular Immunotherapy with Cord Blood-Derived Natural Killer Cells Loaded with AFM13 for CD30+ Malignancies Yago Nieto, MD PhD Department of Stem Cell Transplantation and Cellular Therapy The University of Texas MD Anderson Cancer Center

2 COI Disclosures Research funding from Otsuka, Celgene and Novartis

3 Cornerstones of this Clinical Project Strengths of AFM13 as a bispecific NK cell engager Expanded CB-derived NK cells for immunotherapy Strong preclinical data Important clinical niche

4 Hodgkin s Lymphoma Node structure effaced by malignant cells (Reed Sternberg) within a reactive mixed cellular infiltrate (tumor microenvironment) Reed Sternberg cells 1% tumor mass tumor microenvironment 99% tumor mass Immunosuppressive crosstalk between cytokines secreted by Reed Sternberg cells and receptors on cells in the tumor microenvironment

5 NK effector function is impaired in Hodgkin Lymphoma patients E:T 5:1 (L428 cell line) Autologous NK cells from patients with HL have deficiency in target cell killing Reiners K. R. et al. Molecular Therapy. 2013

6 We hypothesized that we can infuse massive numbers of expanded cord blood-derived functional NK cells whose specificity is redirected against CD30+ tumors by preloading those cells with AFM13 prior to adoptive infusion Cord blood NK cells (CB- NK cells) AFM13 loaded CB-NK cells

7 Allogeneic off-the-shelf umbilical cord blood-derived NK cells Advantages: Readily available Can easily be expanded to a large scale (around 1,000-fold expansion in 14 days) Feasibility and safety of therapy have been shown by our group in clinical trials in myeloma and non-hodgkin lymphoma, where > 50 patients have been treated to date with no toxicity and promising clinical activity (PLoS One, 2013, Leukemia 2015, Br J Haematol 2017)

8 Step 1- Cord Blood-Derived NK cell Expansion

9 Step 2- Loading of NK cells with AFM13 AFM13 1 μg/ml 48h Karpas-299 Expression of activation receptors on AFM13 loaded NK cells NKG2A NKG2D NKG2C NKp30 NKp44 NKp46 DNAM-1 CD69 CD25 CD57 Pan-KIR TIM-3 Trail CD62L T-bet Eomes CD94 2B4 CB-NK cells CB-NK cells + AFM13 Receptors % CB-NK cells + Karpas CB-NK cells + AFM13 + Karpas % Positive CD56+ CD3- % Positive CD56+ CD3-100 % Positive CD56+ CD CD69 CD69 **** **** CD25 CD25 **** **** **** TRAIL TRAIL **** **** CB-NK cells Unloaded CB-NK cells + Loaded Loaded + Karpas CB-NK cells + Unloaded + Karpas L Nassif, K Rezvani, ASH 2018 CB-NK cells +

10 AFM13 loaded, but not unloaded, NK cells kill CD30+ target cell (Karpas-299) AFM13: 100ug/ml AFM13 loaded NK cell Unloaded NK cell n=3 % Positive CD56+ CD3- % Positive CD56+ CD CD107a * * * * * * CD107a INFg INFg TNFa TNFa AFM13 loaded NK cell+ Karpas CB-NK cells + AFM ug/ml vs Karpas CB-NK cells + AFM ug/ml vs Karpas CB-NK Unloaded vs Karpas NK cell+ Karpas CB-NK vs Karpas L Nassif, K Rezvani, ASH 2018

11 AFM13 loaded and washed vs AFM13 loaded and unwashed NK cells have similar level of cytotoxicity against Karpas % Specific Lysis of Karpas **** 1 hour *** ** AFM13 binding with CB-NK cells n=4 Wash twice with PBS (loaded and washed) OR Loaded (but not washed) AFM13 loaded and washed NK cells AFM13 loaded NK cells CB-NK cells loaded by AFM13 (washed before co-c CB-NK cells with AFM13 (direactly added, no wash AFM13 Concentration (ug/ml) 0 E:T 10:1 L Nassif, K Rezvani, ASH 2018

12 CD56 Retention of AFM13 on NK cell surface for up to 7 days after labelling By Flow Cytometry 0h 24h 48h 72h 7 days Unloaded 0.32% 0.02% 0.66% 0.53% 0.9% AFM13 Loaded 41.3% 36.2% 61.7% 76.1% 58.8% By ImageStream AFM13 0h Bright Field CD56 CD16 AFM13 Overlay 24h 48h L Nassif, K Rezvani, ASH 2018

13 Retention of AFM13 on NK cell surface enhances cytotoxicity against CD30-positive Karpas-299, but not CD30-negative Daudi cells Percent (%) specific lysis of Karpas at 10:1 (E:T) ratio Percent (%) specific lysis of Daudi at 5:1 (E:T) ratio AFM13: 100ug/ml n=4 n=3 AFM13 loaded washed NK cells AFM13 loaded NK cells Unloaded NK cells AFM13 loaded NK cells Unloaded NK cells

14 AFM13 loaded NK cells improve survival of mice engrafted with CD30+ lymphoma (Karpas-299) Week NSG Null mice, female, 12-week-old * Day -1 Irradiation 300 cgy * Karpas 299-ffluc Karpas 299-ffuc+ NK cells Karpas 299-ffuc+ NK cells loaded by AFM13 Day 0 Day 7 Karpas-luc Karpas 299-ffuc+ AFM13 Weekly 0.1x x10 6 BLI and Flow cytometry NK cells Overall survival Karpas 299-ffluc Karpas 299-ffluc + unloaded NK cells Karpas 299-ffluc + AFM13 loaded NK cells Karpas 299-ffluc + AFM13 (no NK cells) Percent survival days L Nassif, K Rezvani, ASH 2018

15 Day AFM13-loaded NK cells decrease tumor growth in vivo Radiance Radiance [p/s/cm²/sr] [p/s/cm²/sr] Radiance [p/s/cm²/sr] Week 4 5 Week Karpas 299-ffluc Karpas 299-ffluc + unloaded NK cells Karpas 299-ffluc + AFM13 loaded NK cells Karpas 299-ffluc + AFM13 (no NK cells) Karpas Karpas + Unloaded NK Karpas + AFM13 loaded NK Radiance [p/s/cm²/sr] NS Karpas only Groups 0.0 *** *** NS Groups Karpas + Unloaded NK Karpas cells only Karpas + AFM13 loaded Karpas NK cells + Unloaded NK L Nassif, cells K Rezvani, ASH 2018

16 CD56 AFM13-loaded NK cells successfully home to sites of disease Gated on human CD45 positive cells (on day 18 after NK cell infusion) Liver Spleen Bone Marrow Blood CD3 L Nassif, K Rezvani, ASH 2018

17 Role of New CD30-targeting and Immunological therapies in the Treatment of Hodgkin Frontline (ABVD, BV-AVD) Primary refractory CR 1 st salvage (chemo ± BV) CR HDC + auto-sct No CR CR/PR 2 nd salvage (BV) 3 rd salvage (anti-pd-1) ± xrt Relapse Salvage (BV, anti-pd-1) ± Maintenance BV CR Surveillance PR Allo-SCT

18 Unmet Needs of Current Therapeutic Armamentarium 20% of patients fail frontline therapy Up to 50% of patients who become candidates for HDC and autologous SCT eventually relapse Results poor in patients who are not in PET-negative remission Allogeneic SCT has substantial toxicity and only works for patients in response Anti-CD30 brentuximab vedotin and anti-pd-1 have substantial activity in R/R HL but most responses are not durable Brentuximab vedotin: 75% ORR, 34% CR Only around 1/3 of CR are durable (9% of all patients) Most pts relapse within 1-2 years Unclear what the activity of brentuximab will be in patients already exposed to this drug in 1 st line Anti-PD-1 mabs (nivolumab and pembrolizumab): 70% ORR, 15% CR Most patients relapse within 2 years Median DOR: 12 months for PRs and 20 months for CRs

19 Cord Blood NK cells Loaded with AFM13 as Adoptive Immunotherapy for Refractory/Relapsed CD30+ Malignancies MD Anderson Cancer Center; Affimed CB-derived NK cells NK NK NK NK NK NK NK Expansion of NK cells Cyclophosphami de 300 mg/m 2 Fludarabine 30 mg/m2 NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK NK Day -15 Banked umbilical cord blood Day -14 Pre-activated with cytokines Wash and co-culture with feeder cells Day -7 Restimulate with feeder cells Day -4 Day -3 Day -2 Day 0 Loaded NK cells with AFM13 Cell Infusion Day +7 Day +14 Day +21 AFM13 Injection AFM13 Injection AFM13 Injection

20 Treatment Plan Day -15 Begin CB NK production incubated with AFM13-4 Fludarabine 30 mg/m 2 IV / cyclophosphamide 300 mg/m 2 IV -3 Fludarabine 30 mg/m 2 IV / cyclophosphamide 300 mg/m 2 IV -2 Fludarabine 30 mg/m 2 IV / cyclophosphamide 300 mg/m 2 IV 0 AFM13 + CB NK infusion +7 AFM AFM AFM13 Day +28: Restaging if not CR repeat treatment

21 CB NK cell Dose Escalation Cohort NK cell dose AFM13 1 1x10 6 per Kg Preloaded 2 1x10 7 per Kg Preloaded 3 1x10 8 per Kg Preloaded 4 RP2D Not preloaded Concurrent IV 3+3 escalation Target N = 24 patients

22 Relapsed/refractory classical Hodgkin, ALCL, PTCL or B-NHL that are CD30+ by IHC at 1% HL and ALCL patients must also be refractory or intolerant to brentuximab vedotin Age years INCLUSION CRITERIA ECOG performance status 2 Adequate renal function, as defined by estimated serum creatinine clearance 50 ml/min and/or serum creatinine 1.8 mg/dl Adequate hepatic function (SGOT and/or SGPT 3 x ULN; bilirubin and ALP 2 x ULN Adequate pulmonary function (FEV1, FVC and DLCOc 50% Adequate cardiac function with left ventricular ejection fraction 40% Negative Beta HCG in woman with child-bearing potential EXCLUSION CRITERIA Grade 3 non-hematologic toxicity from prior therapy that has not resolved to grade 1 Active hepatitis B, either active carrier (HBsAg +) or viremic (HBV DNA 10,000 copies/ml, or 2,000 IU/mL) Active infection requiring parenteral antibiotics HIV infection Current participation or past participation within 4 weeks in a study of an investigational agent or device Active central nervous system (CNS) involvement Life expectancy 6 months

23 Study Endpoints Primary Endpoint To establish the safety and recommended dose of CB NK cells Secondary Endpoints: To assess the overall response rate (complete and partial response rates) To evaluate overall survival (OS) time To quantify persistence of infused donor CB AFM-NK cells in the recipient To conduct comprehensive immune reconstitution studies

24 Conclusions We have developed a novel premixed product, comprised of expanded CB-NK cells loaded with AFM13 to redirect their specificity against CD30+ malignancies In vivo studies confirm the antitumor activity of AFM13-NK cells Our encouraging in vitro and in vivo data observed in this study, provide the rationale for the planned clinical trial testing the strategy of off-the-shelf adoptive immunotherapy with massive numbers of AFM13-loaded CB-NK cells in patients with relapsed/refractory CD30+ malignancies