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Pharmacokinetics, Pharmacodynamics and Antiviral Efficacy of the MEK Inhibitor Zapnometinib in Animal Models and in Humans



doi: 10.3389/fphar.2022.893635.


eCollection 2022.

Affiliations

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Julia Koch-Heier et al.


Front Pharmacol.


.

Abstract

The mitogen-activated protein kinase (MEK) inhibitor zapnometinib is in development to treat acute viral infections like COVID-19 and influenza. While the antiviral efficacy of zapnometinib is well documented, further data on target engagement/pharmacodynamics (PD) and pharmacokinetics (PK) are needed. Here, we report zapnometinib PK and PD parameters in mice, hamsters, dogs, and healthy human volunteers. Mice received 25 mg/kg/day zapnometinib (12.5 mg/kg p. o. twice daily, 8 h interval). Syrian hamsters received 30 mg/kg (15 mg/kg twice daily) or 60 mg/kg/day once daily. Beagle dogs were administered 300 mg/kg/day, and healthy human volunteers were administered 100, 300, 600 and 900 mg zapnometinib (once daily p. o.). Regardless of species or formulation, zapnometinib maximum plasma concentration (Cmax) was reached between 2-4 h after administration with an elimination half-life of 4-5 h in dogs, 8 h in mice or hamsters and 19 h in human subjects. Doses were sufficient to cause up to 80% MEK inhibition. Across all species approximately 10 μg/ml zapnometinib was appropriate to inhibit 50% of peripheral blood mononuclear cells (PBMC) MEK activity. In mice, a 50%-80% reduction of MEK activity was sufficient to reduce influenza virus titer in the lungs by more than 90%. In general, while >50% MEK inhibition was reached in vivo at most doses, 80% inhibition in PBMCs required significantly higher doses and appeared to be the practical maximal level obtained in vivo. However, the period of reduced phosphorylated extracellular-signal regulated kinase (pERK), a measure of MEK inhibition, was maintained even after elimination of zapnometinib from plasma, suggesting a sustained effect on MEK consistent with regulatory effects or a slow off-rate. These data suggest a target plasma Cmax of at least 10 μg/ml zapnometinib in further clinical studies.


Keywords:

MEK inhibitor; WES; antiviral therapy; pharmacodynamic; pharmacokinetic; zapnometinib.

Conflict of interest statement

OP, CW and SC are shareholder of Atriva Therapeutics GmbH. OP and SC are consultants for Atriva Therapeutics GmbH. CW, JK-H, AS, LW, JV, and YF are employees of Atriva Therapeutics GmbH. MB is managing director of Synovo GmbH.

Figures



FIGURE 1

MEK Inhibition, Plasma Concentration and Reduction of Virus Titer in Influenza Virus Infected and Zapnometinib Treated Mice. Female C57BL/6 mice were infected with influenza A virus strain H1N1pdm09 and treated twice with 25 mg/kg zapnometinib by oral gavage 1 h prior and 7 h post infection. Blood was collected at different time points (t = 0–24 h) for pharmacokinetic and pharmacodynamic analyses. (A) Association between zapnometinib plasma concentration (left y-axis, red line) and MEK inhibition (right y-axis, blue line). Data presented as mean ± SD (B,C) Antiviral efficacy of zapnometinib against H1N1pdm09 in mice. (B) Virus titers in the lung of zapnometinib treated mice (n = 12) compared to DMSO treated control (n = 3) were given as log10 ffu/ml. Unpaired t test with Welch’s correction was used to test for statistical significance of the difference between control and zapnometinib treated group (***p < 0.001). (C) Virus titer in % reduction of zapnometinib treated mice (n = 12) compared to DMSO treated control (n = 3) Unpaired t-test with Welch’s correction was test for statistical significance of the difference between the two groups (***p < 0.001).


FIGURE 2


FIGURE 2

Pharmacokinetics and pharmacodynamics of zapnometinib in syrian hamsters. Syrian hamsters were treated with either a single dose of 60 mg/kg or twice daily with 15 mg/kg zapnometinib (n = 5 animals per group). The first treatment was given at t = 0 h and the second at 12 h post the first treatment (only for 15 mg/kg). Blood was collected at pre-dose, and 0.5, 1, 2, 4, 8, 12, and 24 h after the first treatment and analyzed for substance and ERK phosphorylation levels. Since no data were collected between 12 and 24 h the theoretical course of plasma concentration was calculated by adding the values from the first 12 h to the respective time points >12 h (A,B) MEK inhibition in PBMCs compared to pre-dose at (A) Cmax (t = 4 h; n
(60 mg/kg) = 4; n
(15 mg/kg) = 3) or (B) 24 h post treatment (n
(60 mg/kg) = 4; n
(15 mg/kg) = 3). Data are presented as mean ± SD. (C,D) Plasma concentration (red line, left y-axis) of (C) 60 mg/kg and (D) 15 mg/kg of zapnometinib correlated with MEK inhibition (blue line, right y-axis). Dashed red line between 12 and 24 h shows extrapolated course of the plasma concentration and dashed blue line indicates that no values between 12 and 24 h were analyzed for MEK inhibition. Data are presented as mean ± SD (n = 5).


FIGURE 3


FIGURE 3

Pharmacokinetic Profiles and Pharmacodynamics of Different Zapnometinib Formulations in Beagle Dogs. Beagle dogs were treated with different formulations of zapnometinib. Blood was taken at pre-dose, 0.5, 1, 2, 4, 6, 9, 12, 15, and 24 h after treatment different timepoints and analyzed for plasma concentration and MEK inhibition. (A,B) PBMC samples of dogs (A) at the time of the highest plasma concentration Cmax (t = 2 or 4 h; n
(formulation 1) = 9; n
(formulation 2) = 7; n
(formulation 3) = 10; n
(formulation 4) = 8; n
(formulation 5) = 10) or (B) 24 h post treatment (n
(formulation 1) = 8; n
(formulation 2) = 10; n
(formulation 3) = 10; n
(formulation 4) = 10; n
(formulation 5) = 10) were analyzed with WesTM for MEK inhibition levels and compared to pre-dose. Data are presented as mean ± SD. (C–G) Plasma concentration (red line, left y-axis) was plotted with MEK inhibition (blue line, right y-axis). Data are presented as mean ± SD (n = 10). (H) Relationship of zapnometinib concentration and target suppression (reduction of ERK phosphorylation) until reaching Cmax is shown as a non-linear regression of ERK phosphorylation. The individual points represent individual values of ERK phosphorylation in dogs treated with zapnometinib.


FIGURE 4


FIGURE 4

Pharmacokinetic and pharmacodynamic assessment of zapnometinib in humans. Healthy volunteers were randomized to four cohorts for a single ascending dose in a phase 1 clinical trial and were treated with either 100, 300, 600 or 900 mg zapnometinib. Blood was taken 0.5, 1, 2, 4, 8, 12, and 24 h later and analyzed for plasma concentration and MEK inhibition. Additionally, blood was taken at 36, 48, 72, and 96 h after treatment for the MAD part. (A) Plasma concentrations following a single dose of 100, 300, 600 and 900 mg zapnometinib. Data are presented as mean ± SD (n = 8). (B) Mean MEK inhibition at Cmax compared to pre-dose analyzed by ERK phosphorylation with WesTM in PBMCs at the highest zapnometinib plasma concentration (Cmax,t = 2–4 h; n
(100 mg) = 7; n
(300 mg) = 7; n
(600 mg) = 6; n
(900 mg) = 5) and (C) 24 h post treatment (n
(100 mg) = 8; n
(300 mg) = 8; n
(600 mg) = 3; n
(900 mg) = 3). (D)Human whole blood was treated with different concentration of zapnometinib (100 μg/ml to 0 μg/ml) for 1 h at 37°C and 5% CO2. PBMCs were isolated and analysed for MEK inhibition. Determination of IC50 value by plotting %ERK phosphorylation against log10 of zapnometinib concentration, using nonlinear regression fit in GraphPad Prism. Data are presented as mean ± SD (n = 3 blood donors).

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