Abstract 51 Immunodepletion of MDSC by AMV564, a Novel Tetravalent Bispecific CD33/CD3 T-Cell Engager, Restores Immune Homeostasis in MDS In Vitro Pingyan Cheng 1 , Erika A. Eksioglu 1 , Xianghong Chen 1 , Max Wei 1 , Jeanmarie Guenot 2 , Judith A. Fox 2 , Alan List 1 and Sheng Wei 1 . 1 Immunology and Malignant Hematology Program, H. Lee Moffitt Cancer Center & Research Institute, Tampa, FL; 2 Amphivena Therapeutics, San Francisco, CA
Disclosures • J. Guenot: Amphivena Therapeutics, Inc: Employment • J.A. Fox: Amphivena Therapeutics, Inc: Consultancy • Sheng Wei : scientific advisor for Amphivena Therapeutics, Inc • Alan List : medical & scientific advisor for Amphivena Therapeutics, Inc 2
MDSC Are Key Innate Immune Effectors in MDS Pathogenesis • Immature CD33 + myeloid cells with distinct function & phenotype ‐ Induce tumor immune tolerance and regulatory T-cell expansion ‐ Suppress autologous hematopoiesis ‐ Suppress T-cell proliferation and IFN- γ production ‐ S100A9 is an autocrine driver of MDSC expansion and paracrine inducer of pyroptosis in autologous hematopoietic progenitors Tumor Microenvironment ↑ROS ↑RNS ↑NO T Cell ↓L -arginine ↓L -cysteine MDSC Altered TCR-function ↓Proliferation ↑Apoptosis ↓IFN - γ production T reg expansion 3 IFN- γ , interferon-gamma; RNS, reactive nitrogen species; ROS, reactive oxygen species; TCR, T cell receptor
MDSC Express High Density CD33 & are Increased in MDS Bone Marrows % BM Lin - /CD33 Hi MDSC MDSC CD33 Surface Density P <0.0005 P <0.0001 • MDSCs are distinguished phenotypically by CD33 high and HLA-DR − Lin − • Genetically distinct and not derived from the mutant clone 4 Chen X et al. J Clin Invest. 2013; 123:4595-4611.
MDSC are Key Effectors of Progenitor Cell Death & Ineffective Hematopoiesis Granzyme B Polarization Cell Death MDSC Depletion & Add-Back P <0.001 5 Chen X et al. J Clin Invest. 2013; 123:4595-4611.
AMV564 is a Highly Potent CD33xCD3 T- Cell Engager Targeting CD33 Hi Cells • AMV564 is a bispecific, AMV564 bivalent, 2X2 T-cell engager ‐ Composed of human antibody variable fragments (scFv) ‐ Two recognition sites for both CD33 & CD3 with strong avidity ‐ Results in T-cell directed lysis of CD33 myeloid cells 6
Methods MDSC: • percent reduction AMV564 ( ± α - T cells: Primary BM • Percent reduction mononuclear PD1) or • isotype IgG Proliferation cells (BMMNC) (BrdU/CellTracker TM ) from15 MDS (control) 5-7 days • Function (IFN- γ) patients in vitro Stem cells : • DNA damage (gH2AX) • CFC BrdU, bromodeoxyuridine; gH2AX, phosphorylated histone 2AX; IFN- γ , interferon-gamma, CFC, 7 colony-forming capacity; AMV564 50ng/ml, α -PD1 10 ug/ml
AMV564 Treatment Depletes CD33 Hi MDSC in BMMNC from MDS Patients IgG (control) AMV564 Media P ≤ 0.001 IgG AMV564 8 BMMNC, bone marrow mononuclear cells, 7 day incubation
AMV564 Treatment Activates CD4 + T-cells in MDS BMMNC Control Media AMV564 IFN-g CD4 Brdu CD4 9
AMV564 Treatment Activates CD8 + T-cells in MDS BMMNC Samples Control Media AMV564 IFN-g CD8 Brdu CD8 10
Dose-dependent Depletion of MDSC by AMV564 Induces Proportionate T-Cell Activation CD4 + T cells C D 4 + T c e lls 6 0 Ig G (C o n tro l) A M V 564 0.1 µg /m l P ro p o rtio n o f C e lls A M V 564 0.2 µg /m l 4 0 A M V 564 0.5 µg /m l M D S C 2 0 MDSC 5 0 4 0 0 3 0 P ro p o rtio n P ro life ra tio n F u n c tio n P ro p o r tio n o f C e lls (B rd U ) (IF N -g a m m a ) 2 0 C D 8 + T c e lls CD8 + T cells 1 0 8 0 3 P ro p o rtio n o f C e lls 6 0 2 4 0 1 0 2 0 P ro p o rtio n 0 P ro p o rtio n P ro life ra tio n F u n c tio n (B rd U ) (IF N -g a m m a ) 11
AMV564 Depletion of MDSC Improves CFC & Decreases DNA Damage CFU-GEMM Formation DNA Damage (gH2AX) 35 P=0.018 30 Colonies (1x105 MDS BM 25 4.98 20 IgG (control) cells) 15 10 5 0 IgG AMV546 P ≤ 0.01 [n=6] 1.35 AMV564 gH2AX IgG AMV564 CD34 12 gH2AX, phosphorylated histone 2AX
AMV564 Depletion of MDSC Enhances CD8 T-cell Response to PD-1 Blockade α -PD1 AMV564 + α -PD1 IgG AMV564 BrdU IFN- γ 13 BrdU, bromodeoxyuridine; IFN- γ , interferon-gamma; PD-1, programmed cell death protein 1
AMV564 Depletion of MDSC Enhances CD4 T-cell Response to PD-1 Blockade α -PD1 AMV564 + α -PD1 IgG AMV564 BrdU IFN- γ 14 BrdU, bromodeoxyuridine; IFN- γ , interferon-gamma; PD-1, programmed cell death protein 1
In Vivo Proof of Principal Case from AMV564 Phase I AML Trial • Medical History ‐ 85 years/male ‐ Secondary AML (AML with MDS-related changes) ‐ Complex karyotype ‐ Resistant to HMA therapy • Baseline characteristics ‐ Low disease burden (5% blasts at baseline) ‐ Low blood counts ▪ Neutrophils: 0.5 x 10 9 /L (500/ μ L) ▪ Hemoglobin: 8.3 g/dL (transfusion-dependent) ‐ High inflammatory state ▪ CRP: 93 mg/L 15 CRP, C-reactive protein; FIH, first-in-human; HMA, hypomethylating agent
Patient 004: Blood count changes from baseline 16
Conclusions • AMV564 effectively depletes CD33 Hi MDSCs in a dose- dependent fashion • AMV564 restores immune homeostasis ‐ proliferation of CD4 + and CD8 + T-cells more than doubled with AMV564 treatment ‐ IFN- γ secretion markedly increased in AMV564 -treated cells • Suppression of MDSCs by AMV564 reduced DNA damage in CD34 + cells and improved colony-forming capacity • AMV564 depletion of MDSC enhances CD4/CD8 T-cell response to PD-1 blockade and warrants investigation in lower risk MDS 17
Acknowledgements Wei Lab • John F. DiPersio • Pingyan Cheng • Peter Westervelt • Erika Eksioglu • Michael Rettig • Xianghong Chen • Alexis Burnette Collaborators • Eric J. Feldman • Alan List • Tae H. Han 18
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