Long acting injectables for therapeutic proteins

Biotherapeutic development presents a myriad of challenges in relation to delivery, in particular for protein therapeutics. Protein delivery is complicated due to hydrophilicity, size, rate of degradation in vivo, low permeation through biological barriers, pH and temperature sensitivity, as well as the need to conserve its quaternary structure to retain function. To preserve therapeutic levels in vivo, proteins require frequent administration due to their short half-lives. Formulation strategies combining proteins with lipid carriers for parenteral administration show potential for improving bioavailability, while preserving protein activity and bypassing the mucosal barriers of the body. Encapsulating protein in long acting injectable delivery systems can improve therapeutic indices by prolonging and controlling protein release and reducing the need for repeat interventions.

Highlights

• Bulk and dispersed lipid cubic phases (LCP) can be injected parenterally

• Bulk and dispersed LCP can incorporate large proteins

• Size and surface properties of proteins incorporated impact on release kinetics

• Enzyme activity can last 3 times longer in LCPs compared to free protein in solution

Two lyotropic crystal forming lipids, monoolein and phytantriol, have been formulated to produce lipidic cubic phases and assessed for their use as long acting protein eluting injectables. Three soluble proteins, cytochrome c, glyceraldehyde-3-phosphate dehydrogenase and aldehyde dehydrogenase and one membrane protein, cytochrome c oxidase, were incorporated into bulk cubic phase formulations of each lipid system to comparatively assess protein release kinetics. The activity of the soluble proteins was measured upon release from a phytantriol bulk cubic phase and phytantriol cubosomes, produced using a liquid precursor method.

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Materials

Commercial grade monoolein (1-(9Z-octadecenoyl)-rac-glycerol) (9.9 MAG) was received and used without further purification from Jena Biosciences. Phytantriol (3,7,11,15-tetramethyl-1,2,3-hexadecanetriol) was received and used without further purification from TCI (Tokyo Chemical Industry). Tween 80, propylene glycol, chloroform, cytochrome c from equine heart ≥ 95% purity, phosphate buffered saline (PBS) tablets, 2,2(-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) tablets, β-nicotinamide adenine dinucleotide (NAD⁺), hexanal 98% and GAPDH Activity assay kit were received and used without further purification from Sigma-Aldrich. Hydrogen peroxide (30% w/v) was received and used without further purification from Mallinckrodt Chemicals. Water used throughout was purified using a Milli-Q Water System (Millipore Corporation, Bedford, MA) consisting of a carbon filter cartridge, two ion-exchange filter cartridges and an organic removal cartridge.

Sally Ryan, Kim Shortall, Michele Dully, Ahmed Djehedar, David Murray, James Butler, John Neilan, Tewfik Soulimane, Sarah P. Hudson,
Long acting injectables for therapeutic proteins,
Colloids and Surfaces B: Biointerfaces, 2022, 112644, ISSN 0927-7765,
https://doi.org/10.1016/j.colsurfb.2022.112644.