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Drug Delivery

Drug Delivery - Poster Session C

Poster J4 - Lithium Carbonate Encapsulation in a Liposomal Delivery Vehicle for Bipolar Medication Usage During Pregnancy

Friday, October 25, 2024
10:00 AM – 11:00 AM EST
Location: Exhibit Hall E, F & G

Introduction:  Bipolar disorder is defined as a mental illness which can lead to sudden mood shifts[1]. Lithium carbonate (Li2CO3) is a medication used for bipolar disorder shown to offset symptoms, prevent suicide, and improve mental wellbeing over time[2]. The exact neurobiological and chemical pathways of lithium treatment are still unknown; however, it is well understood that the lithium ions interfere with the metabolism of phosphatidylinositol[7]. Use of mood stabilizers, such as lithium, during pregnancy and postpartum have been shown to reduce relapse[2]; however, continuation also risks fetal malformations[3]. Lithium carbonate has been shown to cross the placental barrier between mother and fetus, which can result in various complications including cardiac malformations, Epstein Syndrome, or miscarriage[3]. For breastfeeding women, lithium ions can enter the breast milk due to the contents being highly dependent on blood and plasma composition[4]. There is a need to develop a controlled delivery vehicle for lithium treatment, and liposomes are one potential avenue. Liposomes are a simple nanoparticle which features 3 common components: Dipalmitoyl phosphatidylcholine (DOPC), cholesterol, and polyethylene glycol (PEG). In this work, liposomes were chosen due to their high loading efficiency and extensive use in previous drug delivery systems[5]. This study aims to test the viability of using liposomes to load and deliver lithium carbonate.

Materials and

Methods:  Four liposome formulations and multilamellar variations were formulated to contain varying molar ratios of DOPC, PEG, and cholesterol (Figure 1A). Formulation 1 was based on a previous liposome study using lithium[6], and the additional formulations were iterated to investigate differences in the PEG and cholesterol content. A thin film hydration method followed by extrusion was used to develop the liposomes. Briefly, lipids stored in chloroform were dried under nitrogen gas and vacuumed to create a thin film. The thin lipid film was hydrated using nuclease-free water containing 60.7 mM of lithium carbonate solution. A set of 5 freeze-thaw-vortex cycles were conducted to establish multilamellar vesicles. Then, extrusion through a 100 nm polycarbonate membrane was performed. Finally, Amicon ultra centrifugal filters (100 kDa molecular weight cut-off) were used to separate unencapsulated lithium carbonate from the liposomes, and the flowthrough was collected for analysis. Physicochemical characterization of the extruded liposomes and multilamellar liposomes was performed using a Zetasizer to measure size, polydispersity index (PDI), and zeta potential. Encapsulation of lithium carbonate within the liposomes was measured using a conductivity meter. A standard curve was established with known lithium concentrations. Empty liposomes of formulations 1-3 were measured as a control to assess the lipid membrane effects on the conductivity readings. To assess rupturing the liposomes, Triton X-100 was added to the liposomes to destabilize the membrane and release any loaded lithium.

Results, Conclusions, and Discussions: Multilamellar and extruded liposomes resulted in uniform size distributions for all conditions tested (Figure 1). All extruded formulations resulted in particles with smaller diameters and PDIs compared to multilamellar liposomes. After liposome characterization, conductivity was assessed to measure lithium loading. Using the conductivity meter, an initial standard curve with lithium carbonate (3.125 mg/L – 400 mg/L) resulted in an R2 value of 0.99. At a lithium carbonate concentration of 3.125 mg/L, the conductivity was determined to be 7-8 µS. Due to the dilutions necessary for the samples, this resulted in a need to produce a lower limit of detection. Next, the standard curve was produced with lithium carbonate at a lower concentration of 0.78125 mg/L, which resulted in a limit of detection of ~2.9 µS. The multilamellar liposome formulation 2 resulted in a conductivity reading ranging between 9.44 µS and 9.75 µS, which provides a proof-of-concept measurement of lithium loading within the conductivity standard curve . Using the multilamellar liposome formulation 2, a test to disrupt the liposomes using Triton X-100 was performed. Before the addition of Triton X-100, conductivity was measured at 9.76 µS, and after Triton, it was 12.64 µS, which indicates release of encapsulated lithium carbonate. Finally, an increase in conductivity was observed when measuring the lithium loaded liposomes kinetically over multiple days, which will be further investigated in future work.
In this work, multilamellar and extruded liposomes were developed and characterized. These different liposome structures were assessed for their ability to load lithium carbonate. Conductivity measurements were optimized for lithium carbonate using this measurement technique. The multilamellar formulation 2 (DOPC:PEG:Chol at 62.5:5.0:32.5 mol%) resulted in a measurable conductivity within the standard curve produced, and results with Triton X-100 indicate initial promise of lithium carbonate loading. Further, increases in conductivity was observed over time, which motivated future assessment of the lithium carbonate encapsulation stability. In the future, increasing the concentration of lithium carbonate may provide additional insights into the loading and stability. Ultimately, this initial work provides a foundation for future testing to establish a lithium-loaded particle to improve controlled delivery for use during pregnancy and lactation.

Acknowledgements (Optional):  [1] D. K. Hall-Flavin, “Bipolar treatment: Are bipolar I and bipolar II treated differently?,” 2018.
[2] S. A. Barroilhet and S. N. Ghaemi, “When and how to use lithium,” Acta Psychiatrica Scandinavica, vol. 142, pp. 161–172, June 2020.
[3] E. M. P. Poels, H. H. Bijma, M. Galbally, and V. Bergink, “Lithium during pregnancy and after delivery: a review,” International Journal of Bipolar Disorders, vol. 6, p. 26, Dec. 2018.
[4] T. W. Hale and K. Krutsch, Hale’s Medication and Mothers’ Milk. Springer Publishing Company, LLC, twentieth ed., 2023.
[5] Luka Šturm, & Nataša Poklar Ulrih. (2021). Basic Methods for Preparation of Liposomes and Studying Their Interactions with Different Compounds, with the Emphasis on Polyphenols. International Journal of Molecular Sciences, 22(12), 6547–6547. https://doi.org/10.3390/ijms22126547
[6] Hosseini, Y., Seyed Ebrahim Alavi, Azim Akbarzadeh, & Amir Heidarinasab. (2015). Improving lithium carbonate therapeutics by pegylated liposomal technology: an in vivo study. Comparative Clinical Pathology, 25(1), 211–218. https://doi.org/10.1007/s00580-015-2172-2
[7] A. Czarnywojtek, M. Zgorzalewicz-Stachowiak, B. Czarnocka, N. Sawicka-Gutaj, P. Gut, I. KrelaKazmierczak, and M. Ruchala, “Effect of lithium carbonate on the function of the thyroid gland:´ mechanism of action and clinical implications,” Journal of physiology and pharmacology: an official journal of the Polish Physiological Society, vol. 71, Apr. 2020.