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THU0110 Affinity and potency of the anti-NKG2A MAB NNC141-0100: Implications for mabel and dosing in the first-in-man trial in rheumatoid arthritis
  1. L. Alifrangis1,
  2. P. André2,
  3. V. Pascal3,
  4. E. Bonnet2,
  5. L. Radzikowski4,
  6. M.B. Petersen1,
  7. M. Bléry2
  1. 1Non-Clinical Development, Diabetes Research Unit, Novo Nordisk A/S, Måløv, Denmark
  2. 2Innate Pharma, Marseille, France
  3. 3Translational Immunology, Biopharmaceutical Research Unit
  4. 4Dev. Bioanalysis, Diabetes Research Unit, Novo Nordisk A/S, Måløv, Denmark

Abstract

Background The anti-NKG2A monoclonal antibody (mAb), NNC141-0100, may potentially reduce inflammation in Rheumatoid Arthritis (RA) and other immune disorders via a novel mechanism of action. NNC141-0100 has been proposed to promote the cytotoxicity of natural killer (NK) cells towards pro-inflammatory cells expressing HLA-E. NNC141-0100 blocks the inhibitory interaction between HLA-E and the heterodimer CD94/NKG2A receptor on the NK-cells by binding to the NKG2A subunit. Given the novelty of the mechanism of action involving the immune system, the starting dose in the First-in-Man trial should be supported by determination of the Minimum Anticipated Biological Effect Level (MABEL)1.

Objectives To characterise the affinity and potency of NNC141-0100 in human in vitro systems designed to facilitate the translation to humans. The results should be used to calculate the MABEL and set the starting dose following implementation in a PK/PD model.

Methods The affinity (Kd) and maximum receptor binding capacity was assessed in whole blood incubated with increasing concentrations of NNC141-0100. The number of free and bound CD94/NKG2A receptors on relevant peripheral blood leucocytes subsets was assessed by flow cytometry. For comparison, Kd was assessed by surface plasmon resonance (Biacore®) and on isolated peripheral blood mononuclear cells (PBMC). The potency for the biological effect (EC50) was tested in an in vitro system using cell types resembling the in vivo situation. Cytokine-stimulated human PBMC (effector cells) were incubated with autologous CD4+ T-cells (target cells), and increasing concentrations of NNC141-0100. The cytotoxic potential of the effector cells was assessed by flow cytometry using the degranulation marker CD107, known as a sensitive biomarker preceding cytotoxicity. Both healthy subjects and Rheumatoid Arthritis (RA) subjects provided blood samples. In the PK/PD model, the Kd and EC50 values provided the basis for simulating receptor occupancy and CD107 profiles versus time and dose levels in humans.

Results NNC141-0100 was bound with a similar, high affinity to the human CD94/NKG2A receptor in all test systems. The EC50 in the CD107 assay was approximately 100-fold higher than the Kd. This difference may be due to competitive interaction from HLA-E on the target cells. Similar CD107 potencies were observed for healthy and RA subjects. By means of PK/PD simulations, the i.v. MABEL dose in humans was calculated to give a predicted CD107 response of 10% at Cmax. The corresponding receptor occupancy at Cmax was 90%. Due to the high occupancy and the uncertainty of the in vitro-in vivo correlation for cytotoxicity and potentiating the immune system, a safety factor was applied to the MABEL to determine the starting dose.

Conclusions Given the observed difference between affinity and potency, the MABEL dose for the First-In-Man trial in RA patients had a high predicted CD94/NKG2A receptor occupancy of 90%, while the expected cytotoxicity as measured by the degranulation marker CD107 was much lower (<1%).

  1. EMA guideline: EMEA/CHMP/SWP/294648/2007

Disclosure of Interest L. Alifrangis Shareholder of: Novo Nordisk A/S, Employee of: Novo Nordisk A/S, P. André Shareholder of: Innate Pharma, Employee of: Innate Pharma, V. Pascal Shareholder of: Novo Nordisk A/S, Employee of: Novo Nordisk A/S, E. Bonnet Shareholder of: Innate Pharma, Employee of: Innate Pharma, L. Radzikowski Shareholder of: Novo Nordisk A/S, Employee of: Novo Nordisk A/S, M. Petersen Shareholder of: Novo Nordisk A/S, Employee of: Novo Nordisk A/S, M. Bléry Shareholder of: Innate Pharma, Employee of: Innate Pharma

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