Preclinical and clinical phase I studies of a new recombinant Filgrastim (BK0023) in comparison with Neupogen®

Abstract

Background: Filgrastim or methionyl-granulocyte colony-stimulating factor (Met-G-CSF), is a recombinant therapeutic protein widely used to treat severe neutropenia caused by myelosuppressive drugs in patients with nonmyeloid malignancies. In addition to its role in the regulation of granulopoiesis, treatment with G-CSF is considered the standard approach to mobilize CD34 positive (CD34+) mononuclear cells for reconstituting hemopoietic ability for bone marrow transplantation. An intended biosimilar filgrastim (coded BK0023) was produced in GMP conditions by E.coli fermentation according to an original recombinant process and showed physico-chemical properties and purity profile similar to Neupogen®, a commercial preparation of filgrastim. The aim of the present study was to demonstrate the comparability of BK0023 to Neupogen® in terms of both in vitro biological activities and in vivo toxicology, pharmacokinetics and pharmacodynamics.

Methods: Cell proliferation and radioligand binding assays were conducted in NFS-60 cells to compare the biological activity and functional interaction with the G-CSF receptor in vitro, while preclinical in vivo studies, including pharmacokinetics and pharmacodynamics after repeated dose were performed in normal and neutropenic rats. A phase I study was carried out in healthy male volunteers treated by multiple-dose subcutaneous administration of BK0023 and Neupogen® to evaluate their pharmacodynamic effects as well as their pharmacokinetic and safety profile and to demonstrate their pharmacodynamic equivalence and pharmacokinetic bioequivalence.

Results: The results reported in this work demonstrate that BK0023 is comparable in terms of biological activity, efficacy and safety to Neupogen®.

Conclusions: BK0023 has the same pharmacokinetic profile, efficacy and safety as the reference commercial filgrastim Neupogen® and therefore could be further developed to become a convenient option to treat neutropenia in oncological patients.

Trial registration: ClinicalTrials.gov NCT01933971.

Figure 1

Concentration-dependent inhibition of [¹²⁵I]-G-CSF binding by BK0023 (black circle) and Neupogen^® (open circle) in NFS-60 cells expressing G-CSF receptor. Data are reported as percent of specific binding at each concentration of displacer and are the means ± S.E.M. of four separate experiments. Statistical analysis by Student’s t-test did not show any significant difference between BK0023 and Neupogen^®.

Figure 2

Mean neutrophile counts (× 10⁹/l) obtained in groups of 10 normal rats treated for 4 days by subcutaneous injections of 10, 30 and 100 μg/kg/day of BK0023 and Neupogen^® are respectively shown in left, central and right panels where full circles, squares and triangles are respectively referred to control rat, BK0023 and Neupogen. No significant difference was found between BK0023 and Neupogen treated groups analysed by Student’s test. Significant differences between treated and control groups analyzed by Dunnet’s test were found at P < 0.05 (*) or P < 0.01 (**).

Figure 3

Mean neutrophile counts (× 10⁹/l) ± SD obtained in groups of 10 neutropenic rats treated for 4 days by subcutaneous injections of 10, 30 and 100 μg/kg/day of BK0023 and Neupogen^® are respectively shown in left, central and right panels where full circles, squares and triangles are respectively referred to control rats, BK0023 and Neupogen. No significant difference was found between BK0023 and Neupogen treated groups analysed by Student’s test. Significant differences between treated and control groups analysed by Dunnet’s test were found at P < 0.05 (*) or P < 0.01 (**).

Figure 4

Mean neutrophil counts (× 10 ³ /μl) ± SD measured in human healthy volonteers treated for 7 days by subcutaneous injections of BK0023 (triangles) and Neupogen^® (circles) at dosage of 2.5 μg/kg/day (upper panel) and 5.0 μg/kg/day (central panel) and for 5 days by subcutaneous injections of BK0023 and Neupogen^® at dosage of 10.0 μg/kg/day (lower panel).

Figure 5

Plasma level profiles of G-CSF (pg/ml) measured by immunoenzymatic assay at day 1 (solid lines) and at the last day of treatment (dashed lines) for BK0023 2.5 μg/kg/day (A); Neupogen 2.5 μg/kg/day (B); BK0023 5 μg/kg/day (C); Neupogen 5 μg/kg/day (D); BK0023 10 μg/kg/day (E); Neupogen 10 μg/kg/day (F).