Biological activity In vitro, absorption, BBB penetration and tolerability of nanoformulation of BT44, a RET agonist with disease-modifying potential for the treatment of neurodegeneration

27 September 2022, Version 2
This content is a preprint and has not undergone peer review at the time of posting.

Abstract

BT44 is a novel, second generation glial cell line-derived neurotropic factor (GDNF) mimetic, with improved biological activity and a lead compound for the treatment of neurodegenerative disorders. Like many other small molecules, it suffers from intrinsic poor aqueous solubility, posing significant hurdles at various levels for its preclinical development and clinical translation. Herein, we report a poly(2-oxazoline)s (POx) based BT44 micellar nanoformulation with ultra-high drug loading capacity of 47 wt.%. The BT44 nanoformulation was comprehensively characterized by 1H-NMR spectroscopy, differential scanning calorimetry (DSC), powder X-ray diffraction (XRD), dynamic light scattering (DLS) and cryo-transmission/scanning electron microscopy (cryo-TEM/SEM). The DSC, XRD and redispersion studies collectively confirmed that the BT44 formulation can be stored as a lyophilized powder and can be redispersed upon need. The DLS suggested that the redispersed formulation is suitable for parenteral administration (Dh ≈ 70 nm). The cryo-TEM measurements showed the presence of worm-like structures both in plain polymer and BT44 formulation. The BT44 formulation retained biological activity in immortalized cells and in cultured dopamine neurons. The micellar nanoformulation of BT44 exhibited improved absorption (after subcutaneous injection) and blood-brain barrier (BBB) penetration and no acute toxic effects in mice were observed. In conclusion, herein, we have developed an ultra-high BT44 loaded aqueous injectable nanoformulation, which can be used to pave way for its preclinical and clinical development for the management of neurodegenerative disorders.

Keywords

Neurodegeneration
receptor tyrosine kinase agonist
blood brain barrier
solubility enhancement
poly(2-oxazoline)
nanomedicine

Supplementary materials

Title
Description
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Title
Supporting information for BT44 Micellar formulation
Description
Synthesis of 2-n-pentyl-2-oxazoline monomer and triblock copolymers, DSC thermogram of three POx based triblock copolymer, groups contributing towards HSPs determination, quantification of BT44 by HPLC, visual appearance of BT44 formulation, 1H-NMR spectra of polymers and formulation, DSC thermogram of lyophilized formulation.
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