(M0930-03-13) Development and Characterization of Primaquine and Chloroquine Solid Dispersion Loaded Dissolving Microneedles for the Improved Treatment of Malaria

Purpose: Affecting over 240 million people worldwide each year, malaria is an infectious disease which accounts for 881,000 deaths annually, with nine out of ten deaths occurring in sub-Saharan Africa^1,2. This mosquito-borne disease arises when plasmodium parasites are injected into the human systemic circulation during a mosquito bite^3,4. It is known that P. falciparum is the deadliest species of plasmodium to infect humans. Besides the cardinal symptoms of malaria, which include fever, muscle ache and lethargy, patients infected with P. falciparum also experience persistent vomiting upon hospital admission^5,6. This eliminates the use of the conventional oral drug delivery route, thus necessitating an intravenous infusion and, in some instances, intramuscular injection. Nevertheless, these routes require skilled health-care-workers, which is particularly challenging in low- and middle-income countries⁶. In light of this limitation, we have engineered and developed microneedles (MNs) loaded with two efficacious antimalarial drugs for the treatment of malaria. It is hoped that these MNs offer a suitable alternative treatment strategy which can be painlessly self-administered by patients in low- and middle-income countries.

Methods: Solid dispersions (SD) of primaquine (PQ) and chloroquine (CQ) in Soluplus® were prepared individually via lyophilization. The powder of SD-PQ and SD-CQ then were prepared as the first layer of dissolving MAPs by mixing the powder with deionized water (ratio of powder and water 1:2). The aqueous mixtures were then cast into the silicone moulds (600 pyramidal needles on a 0.76 cm² area with 750 μm needle height), as illustrated in Figure 1A. All MN formulations were visualized using a digital microscope and scanning electron microscope (SEM). A Texture Analyzer was used to evaluate the mechanical properties of the MAPs when subjected to a compressive force of 32 N, analogous to manual thumb pressure⁷. The insertion capability of the MAPs into the skin simulant Parafilm® M⁷ and ex vivo full-thickness neonatal porcine skin were evaluated in situ using optical coherence tomography (OCT). Dermatokinetic studies were conducted using modified Franz diffusion cells in order to quantify the amount of drug deposited into the skin as well as the amount of drug that permeated through the neonatal porcine skin. The efficacy of the formulation was also evaluated over a period of 7 days using female mice (C57BL/6JUnib, 6-7 weeks old, 20 ± 3g) infected with Plasmodium berghei.

Results: The formulations of PQ-MNs and CQ-MNs were successfully able to form homogeneous MN arrays, as presented in Figure 1(D-E). The application force of 32N/array reduced needles height by less than 10% in all cases (Figure 1B). For the insertion studies, both formulations were able to penetrate down into the dermis to a depth of 430 µm, which was confirmed by OCT analysis (Figure 1 H-K). The drug content for PQ-MN was determined to be 2.8 mg/patch, while that of CQ-MN was found to be 3.4 mg/patch (Figure 1 C). Dermatokinetic studies revealed that the delivery efficiencies of PQ-MN and CQ-MN found in the skin deposition study were 23% and 61%, respectively (Figure 2A-B). Over the course of 24 h, we observed that both formulations resulted in an increase in drug delivery into and across the skin with time. After 24 h, most of the drug had been delivered across the skin It was also found that, after 24 h, 96.00 ± 3.29 μg and 260.43 ± 67.22 μg of the drug were still deposited in the skin for PQ and CQ respectively. The presence of the drug in the skin, even after MNs removal may serve as a potential depot for sustained delivery of PQ and CQ via MNs for the treatment of malaria. Efficacy studies in a murine model for malaria revealed that mice that received a treatment combination of PQ-MNs and CQ-MNs demonstrated a 99.96% reduction in parasitemia after 7 days post-infection (Figure 2 (E)).

Conclusion: The present results describe the successful development and characterization of dissolving MNs which are loaded with solid dispersions of PQ and CQ in the tips of the needles. Furthermore, dermatokinetic studies showed that, after 24 h, most of the drug had been delivered transdermally across the skin. However, it was also found that some of the drugs still remain deposited in the skin, which may serve as an intradermal depot for the controlled and sustained release of both PQ and CQ during the treatment of malaria. Additionally, efficacy studies in a murine model for malaria showed that mice treated with PQ-MNs and CQ MNs was able to reduce in parasitemia up to 99.96% in the treated animals. Overall, the current work highlighted that the MNs engineered in the current work may provide a minimally invasive, needle-free drug delivery strategy for the treatment of malaria in low-resource settings.

References: 1. World Health Organization. World Malaria Report 2021.; 2021.
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4. Milner DA, Jr. Malaria Pathogenesis. Cold Spring Harbor Perspectives in Medicine. 2018;8(1). doi:10.1101/CSHPERSPECT.A025569
5. Bria YP, Yeh CH, Bedingfield S. Significant symptoms and nonsymptom-related factors for malaria diagnosis in endemic regions of Indonesia. International Journal of Infectious Diseases. 2021;103:194-200. doi:10.1016/J.IJID.2020.11.177
6. Eisenhut M, Omari AA, Group CID. Intrarectal quinine versus intravenous or intramuscular quinine for treating Plasmodium falciparum malaria. The Cochrane Database of Systematic Reviews. 2009;2009(1). doi:10.1002/14651858.CD004009.PUB3
7. Larran ̃eta E, Moore J, Vicente-Perez EM, Gonzalez-Vazquez P, Lutton R, Woolfson AD, Donnelly RF, A proposed model membrane and test method for microneedle insertion studies, International Journal of Pharmaceutics. 472 (2014) 65–73. doi:10.1016/j.ijpharm.2014.05.042.


Figure 1. (A) The drug-loaded tips were cast from an aqueous blend of drug solid dispersion and deionized water. (B) Needle tip reduction after compression (Means + SD, n = 15). (C) Drug contents in the MNs (Means + SD, n = 4). Digital image of MN array loaded with (D) primaquine and (E) chloroquine. SEM images of (F) PQ-MNs and (G) CQ-MNs. OCT photos of the skin insertion of (H) PQ-MNs and (I) CQ-MNs. Parafilm M® insertion of (J) PQ-MNs and (K) CQ-MNs.


Figure 2. Drug distribution at different times after application of (A) CQ-MNs and (B) PQ-MNs (Means + SD, n = 4). (C) Peripheral blood parasitemia curves as a function of time to day 7 post-infection in groups of mice infected with murine parasite P. berghei. Control group: MN Blank without drug. Treated group: PQ-MN, CQ-MN and combination of PQ-MN and CQ-MN applied application (Mean + SD, n = 4).