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. 2023 Jan;6(1):2200199.
doi: 10.1002/adtp.202200199. Epub 2022 Oct 11.

Emulsion-induced polymersomes taming tetrodotoxin for prolonged duration local anesthesia

Xiaosi Li  1 , Qi Li  1 , Shenghan Song  2 , Amy O Stevens  2 , Zach Broemmel  1 , Yi He  2   3 , Ursula Wesselmann  4 , Tony Yaksh  5 , Chao Zhao  1   6   7
Affiliations

Emulsion-induced polymersomes taming tetrodotoxin for prolonged duration local anesthesia

Xiaosi Li et al. Adv Ther (Weinh). 2023 Jan.

Abstract

Injectable local anesthetics that can provide a continuous nerve block approximating the duration of a pain state would be a life-changing solution for patients experiencing post-operative pain or chronic pain. Tetrodotoxin (TTX) is a site 1 sodium channel blocker that is extremely potent compared to clinically used local anesthetics. Challengingly, TTX doses are limited by its associated systemic toxicity, thus shortening the achievable duration of nerve blocks. Here, we explore emulsion-induced polymersomes (EIP) as a drug delivery system to safely use TTX for local anesthesia. By emulsifying hyperbranched polyglycerol-poly (propylene glycol)-hyperbranched polyglycerol (HPG-PPG-HPG) in TTX aqueous solution, HPG-PPG-HPG self-assembled into micrometer-sized polymersomes within seconds. The formed polymersomes have microscopically visible internal aqueous pockets that encapsulate TTX with an encapsulation efficiency of up to 94%. Moreover, the polymersomes are structurally stable, enabling sustained TTX release. In vivo, the freshly prepared EIP/TTX formulation can be directly injected and increased the tolerated dose of TTX in Sprague-Dawley rats to 11.5 μg without causing any TTX-related systemic toxicity. In the presence of the chemical penetration enhancer (CPE) sodium octyl sulfate (SOS), a single perineural injection of EIP/TTX/SOS formulation produced a reliable sciatic nerve block for 22 days with minimal local toxicity.

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Figures

Figure 1.
Figure 1.
Synthesis and characterization of HPG-PPG-HPG copolymer. A) Synthesis route for HPG-PPG-HPG. B) 1H NMR spectra of HPGL-PPG4000-HPGL. C) MALDI-ToF MS spectra of HPGL-PPG4000-HPGL. D) A comparison of the intensity in MS spectrometry between HPGL-PPG4000-HPGL and pure PPG4000.
Figure 2.
Figure 2.
Characterization of particles fabricated after emulsifying HPG-PPG-HPG in the water using a vortex mixer at the speed of 3000 rpm for 20 seconds. A) Optical microscope images of HPG-PPG-HPG particles. Red arrow: aqueous pockets. Scale bars: 20 μm. B) A comparison of the percentage of space occupied by the particles in the emulsion between HPG-PPG-HPG and PPG4000. The emulsions were left at room temperature for 192 hours after the vortex procedure. Data are means ± SD, n = 3.
Figure 3.
Figure 3.
The average binding energy of six unique PPG polymers.
Figure 4.
Figure 4.
Characterization of drug encapsulation and release characteristics from polymersomes fabricated by the vortex procedure. A) A comparison of the encapsulation efficiency of SF in different polymersomes. B) Effect of the HPGL-PPG4000-HPGL concentration on the encapsulation efficiency of SF. C) Confocal fluorescence microscopy images of polymersomes. Green: SF. Red: Dil. Scale bars: 20 μm. D) A comparison of the cumulative release of SF eluted from different polymersomes in PBS at 37 °C. D-F) A comparison of the cumulative release of SF eluted from different polymersomes in PBS at 37 °C. G) Mathematical modeling of SF release from polymersomes. H) A comparison of the encapsulation efficiency of TTX in polymersomes. I) A comparison of the cumulative release of TTX eluted from different polymersomes in PBS at 37 °C. Data are means ± SD, n = 3.
Figure 4.
Figure 4.
Characterization of drug encapsulation and release characteristics from polymersomes fabricated by the vortex procedure. A) A comparison of the encapsulation efficiency of SF in different polymersomes. B) Effect of the HPGL-PPG4000-HPGL concentration on the encapsulation efficiency of SF. C) Confocal fluorescence microscopy images of polymersomes. Green: SF. Red: Dil. Scale bars: 20 μm. D) A comparison of the cumulative release of SF eluted from different polymersomes in PBS at 37 °C. D-F) A comparison of the cumulative release of SF eluted from different polymersomes in PBS at 37 °C. G) Mathematical modeling of SF release from polymersomes. H) A comparison of the encapsulation efficiency of TTX in polymersomes. I) A comparison of the cumulative release of TTX eluted from different polymersomes in PBS at 37 °C. Data are means ± SD, n = 3.
Figure 5.
Figure 5.
Neurobehavioral assessment of rats that received a single sciatic nerve injection of 0.3 mL PBS containing 60 mg HPG-PPG-HPG polymer and TTX and/or SOS. Ipsilateral and contralateral paw thermal latency to thermal stimulation. n = 4. Data presented are the mean ± SD of the respective treatment groups.
Figure 6
Figure 6
Tissue reaction to formulations 4, 14, and 28 days after injection, including representative photographs of the site of injection upon dissection, representative hematoxylin–eosin stained sections of muscles and adjacent loose connective tissue, and toluidine blue stained sections of nerve. Data are representative of 2–5 animals in each group. Scale bars, 200 μm (H&E), 100 μm (toluidine blue)
Scheme 1.
Scheme 1.
Schematic illustration of emulsion-induced polymersomes encapsulation of hydrophilic small molecules.
See this image and copyright information in PMC

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