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502 Discovery of HX017, a novel NKG2A/CD94 mAb, for potential cancer immunotherapy
  1. Hang Ke1,
  2. Faming Zhang2,
  3. Jialing Li1,
  4. Dawei Chen3,
  5. Feiyu Peng1,
  6. Cen Chen4 and
  7. Henry Li2
  1. 1Hanx Biopharmaceuticals, Inc., Wuhan, HuBei, China
  2. 2Hanx Biopharmaceuticals, Inc., Oceanside, CA, USA
  3. 3Innomodels, Ltd., Beijing, China
  4. 4Crown Bioscience Inc., San Diego, CA, USA
  • Journal for ImmunoTherapy of Cancer (JITC) preprint. The copyright holder for this preprint are the authors/funders, who have granted JITC permission to display the preprint. All rights reserved. No reuse allowed without permission.


Background NKG2A (Natural Killer Group 2A), an inhibitory immune checkpoint, forms a heterodimer receptor with CD94. NKG2A/CD94 is often expressed on subsets of natural killer (NK) cells, CD8+ tumor-infiltrating T (CD8+ TIL) cells and γδ T cells, etc. Its ligand HLA-E, a non-classical MHC-I molecule, is frequently up-regulated in many solid tumors or hematological malignancies, protecting from NKG2A-expressing immune cell mediated killing. Several NKG2A/CD94 blocking mAb (e.g. Monalizumab) are being tested in clinic, demonstrating preliminary antitumor activity in certain solid tumors including head and neck carcinoma (HNSCC) and non-small cell lung carcinoma (NSCLC).

Methods A humanized anti-NKG2A/CD94 mAb HX017 was recently constructed from mouse hybridoma screening. SPR/ELISA and flow cytometry were used to assay the binding of HX017 to human NKG2A/CD94 proteins; The blockade activity was investigated by flow cytometry as HX017 competitive binding to NKG2A/CD94-expressing 293T cells in the presence of hHLA-E-peptide tetramer (ligand blocking) or biotin-labelled Monalizumab (epitope binning); HX017-mediated NK activations were determined by 1) CD107α-expression using NK/CD4+ autoblast co-culture from different donors and 2) NKG2A/CD94+ Jurkat reporter cell (luciferase) when co-cultured with the APC expressing HLA-E/β2m; K562 target cell killing by IL-15 treated NKL cells (NKG2A/CD94+) in the presence of HLA-B7 signaling peptide was determined (CytoTox-Glo™). Another luciferase reporter assay using NKG2A/CD94+ PD-1+ Jurkat cell and APC expressing HLA-E/β2m/PD-L1 was assessed to investigate combination with anti-PD-1 in vitro. In vivo tumor growth inhibition by HX017 were evaluated using two types of humanized models.

Results HX017 was shown to have high binding affinity to NKG2A/CD94 (EC50 ~0.22 nM) and ligand-blocking (IC50 ~0.63 nM). HX017 efficiently triggered NK activation and enhanced NK-cytotoxicity to target cells in vitro, alone or in combination with anti-PD1 mAb. HX017 showed antitumor activity, as a single agent or, in particular, in combination with PD1 mAb, in two types of humanized mice: 1) MC38-hHLA-E/hPD-L1 HuGEMM model where hPD-1/hPD-L1/hCD94/hNKG2A were knocked-in, and A431 tumor in huPBMC reconstituted NSG-IL-15 mice model. Finally, HX017 has good stability.

Conclusions HX017 could potentially be developed as a promising candidate cancer immunotherapy, particularly in combination with other ICIs.

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