Ultrasound-mediated topical delivery of econazole nitrate with potential for treating Raynaud's phenomenon
Graphical abstract
Introduction
Topical and transdermal drug delivery possess several advantages over conventional and parenteral administration, such as prevention of drug metabolism by the gastrointestinal (GI) lumen/hepatic first- pass (Prausnitz and Langer, 2008), reduction of GI side-effects resulting from prolonged exposure to certain drugs (Polat et al., 2010), non-invasiveness with no pain or discomfort associated with needles and better patient adherence (Alsaab et al., 2016a, Han and Das, 2015, Schoellhammer et al., 2014). However, a drug molecule has to overcome the skin barrier properties in order to enter the systemic circulation. The skin acts as a major rate-limiting factor to topical and transdermal drug permeation (Alsaab et al., 2016b). The stratum corneum is the outermost layer of the skin, which is ~20 µm thick. It is made up of hexagonal, highly ordered, compact cells called corneocytes that are embedded in the lipid bilayer matrix. This peculiar structure of the stratum corneum acts as a diffusional barrier (Forslind and Lindberg, 2004). Thus, the actual drug concentration that permeates the skin is less. Several physical and chemical methods to overcome the skin barrier properties have been employed to enhance the drug permeation. These include ablation, microneedles, iontophoresis, electrophoresis, sonophoresis or ultrasound – assisted transdermal delivery and the use of chemical penetration enhancers (Alexander et al., 2012, Azagury et al., 2014, Oberli et al., 2014, Park et al., 2014).
Ultrasound has been widely investigated as a non-invasive method to enhance the transdermal permeation of drugs. This technique was first used to deliver hydrocortisone across the skin for treating digital polyarthritis in 1954 (Fellinger and Schmid, 1954). In 2004, SonoPrep® (Sontra Medical) was approved by FDA for the transdermal delivery of local anesthesia (Alkilani et al., 2015, Watkinson, 2012). Ultrasound-assisted drug delivery has been tested for its ability to enhance localized permeation of drugs such as anti-inflammatory agents (dexamethasone, cortisol), local anesthetics (lidocaine and prilocaine) and drugs used in orthopedic diagnosis (Daftardar et al., 2019, Katz et al., 2004) Recently, many macromolecules such as protein and peptides have also been successfully administered using ultrasound-assisted transdermal delivery (Luis et al., 2007). Ultrasound is defined as a longitudinal sound wave, i.e., mechanical energy transferred by oscillating neighboring particles from one point to the other. Ultrasonic acoustic waves have frequency above 20 kHz and is categorized into three frequencies: low-frequency ultrasound (20–100 kHz), therapeutic ultrasound (0.7–3 MHz) and high-frequency ultrasound (>3 MHz). Studies have reported that low-frequency ultrasound is more effective in enhancing the transdermal drug permeation than high-frequency ultrasound (Daftardar et al., 2019, Mitragotri, 2006). The extent to which the ultrasound waves can penetrate into the skin is inversely proportional to the ultrasound frequency (Machet and Boucaud, 2002). The exact mechanism by which ultrasound enhances drug permeation is still unclear, although there have been several reports on the mechanisms of ultrasound namely, acoustic and thermal streaming and inertial cavitation (Azagury et al., 2014). Cavitation is characterized by bubble formation due to pressure changes induced in the coupling medium by ultrasound (Tang et al., 2002, Tezel et al., 2002). The formed bubbles cause disruption of the stratum corneum lipids, thereby increasing the skin permeabilization.
Raynaud’s phenomenon (RP) is characterized by reversible vasospasm of the digits induced by cold exposure or emotional stress (Wigley, 2002). The exact cause of RP is undetermined and the current treatment modalities are largely unsatisfactory and associated with significant side effects. Transient Receptor Potential Melastatin 8 (TRPM8) is one of the TRP proteins, which is a calcium permeable, non-selective cation transporter protein that is directly activated by cold temperatures below 26˚C. TRPM8 has been convincingly demonstrated to be the main molecular transducer responsible for the sensitivity to innocuous cold (Bautista et al., 2007, Dhaka et al., 2007). Recently, the role for TRPM8 in mediating attacks of RP has been identified. We have earlier reported the role of TRPM8 in mediating attacks of RP and the antagonizing properties of econazole nitrate (EN) (Bahl et al., 2019, Kahaleh et al., 2017). Ensuring the delivery of EN to the vasculature beneath the stratum corneum and epidermal layers is important for achieving a topical therapy for RP. Econazole nitrate is available as a water-miscible cream (1%) and is only used as a topical agent for the treatment of superficial skin infections such as tinea, athlete’s foot, ringworm and yeast infections. Moreover, local reactions such as irritation, erythema, burning, stinging and rash have been reported in patients. Physical phase separation of the cream is also reported due to the drug salting out effect (Verma and Pathak, 2012). More than 90% of the topically applied drug remains on the skin due to its low solubility and thus resulting in poor skin permeation (Firooz et al., 2015).
In an attempt to design a topical therapy for RP and minimize the undesirable effects of the marketed EN cream, our lab has previously developed and characterized four formulations containing 3% EN: F1_solution, F2_HPMC dispersion, F3_VersaBase cream® and F4_Lipoderm® Activemax™ Cream for ensuring the penetration of econazole through the skin to the dermal vasculature, where cold sensing receptors are expressed by microvascular endothelial cells. These formulations demonstrated an increase in the drug permeability compared to the marketed cream (Bahl et al., 2019). The use of newer technologies such as ultrasound can actively drive drug molecules through the transdermal barrier, may reduce time to onset and allow formulation to be highly useful in the management of RP.
In this study, the effect of low-frequency ultrasound on the skin permeability of EN from the previously developed formulations (F2_HPMC dispersion and F4_Lipoderm® Activemax™ Cream) was evaluated across the pig ear skin. For effective treatment of RP, it is imperative that the drug penetrates the skin rapidly providing prompt therapeutic action. We hypothesize that the use of ultrasound would enhance the permeation of EN across the skin. Optimization of the critical ultrasound parameters such as the horn distance, application time and amplitude were performed. The influence of formulations and ultrasound exposure on the skin stratum corneum was determined based on the spectral shifts of the CH stretching vibrations using attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy. Further, the skin toxicity assessment of formulations and ultrasound exposure was investigated.
Section snippets
Materials
Econazole nitrate with 99.53% purity was purchased from Alfa Aesar, (Ward Hill, MA), Distilled deionized water was used for the preparation of buffers. High Performance Liquid Chromatography (HPLC) grade solvents such as methanol (Lot 165479) and ammonium dihydrogen phthalate (ADP) (Lot 120935) were provided from Fisher Scientific (Pittsburgh, PA). The ingredients used for the preparation of phosphate buffer pH 6.8 as coupling medium and receptor fluid consisted of potassium phosphate monobasic
Results and discussion
In our previous study, we have successfully designed and characterized four different formulations with 3% EN (F1_solution, F2_HPMC dispersion, F3_Versabase® cream and F4_Lipoderm® Activemax™ Cream). Percutaneous absorption data showed better permeation of EN from all formulations compared to the marketed EN cream (1%). Among all formulations, F2_HPMC dispersion and F4_Lipoderm® Activemax™ cream were found to be the best (Bahl et al., 2019). As a logical extension of our previous work, this
Conclusion
In the present study, we have evaluated the ultrasound-assisted transdermal delivery of EN from two topical formulations. The ultrasound parameters such as amplitude, application time and distance of the horn were optimized and used for the in vitro percutaneous absorption study across the porcine ear skin. A constant frequency (20 kHz) ultrasound application with 40% amplitude, 0.5 cm distance between ultrasound horn and skin surface for 2 min were considered optimum. The percutaneous
Author contributions
Conceptualization, Data curation: S.H.S.B, B.K, N.A; Formal analysis: S.D, D.B; Funding acquisition: S.H.S.B, B.K, N.A; Methodology: S.D, D.B; Supervision: S.H.S.B, B.K, N.A; Writing - original draft: S.D, D.B; Writing - review & editing: S.H.S.B, B.K, N.A.
Declaration of Competing Interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
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