Novel ethosomal gel formulation for enhanced transdermal delivery of curcumin and cyclosporine: a preclinical approach to rheumatoid arthritis management

Abstract

Vesicular systems have demonstrated efficacy in the management of Rheumatoid Arthritis (RA). This study explores the synergistic effect of edge-activated ethosomal gel to enhance the transdermal delivery of Curcumin (CUR) and Cyclosporine (CYC). Ethosomal vesicles prepared via the ethanol injection method were incorporated into a gel, with the optimized formulation exhibiting an average particle size of 93.3 ± 1.17 nm and a zeta potential of −29.2 ± 0.17 mV. Ex vivo diffusion studies on porcine ear skin demonstrated 97.115 ± 0.40% CUR and 98.331 ± 1.08% CYC release over 18 hours, exhibiting Hixson-Crowell diffusion mechanisms. The steady-state flux and permeability coefficients were 0.095 µg/cm²/hr and 0.0095 cm/hr for CUR, and 0.0804 µg/cm²/hr and 0.01608 cm/hr for CYC respectively. In anti-inflammatory tests on lipopolysaccharide (LPS)-induced RAW 264.7 cells, the gel significantly increased IL-10 levels (p < 0.001), inhibited prostaglandin-E2, and reduced IL-6 and TNF-α levels (p < 0.001). Moreover, the ethosomal gel demonstrated nonirritating properties and exhibited significant reduction in arthritic symptoms in the Complete Freund’s Adjuvant induced 28-day rat model, surpassing the effects of marketed and conventional gel. These findings highlight the synergistic benefits of combining CUR and CYC in an ethosomal gel, offering a promising alternative for RA management. Future clinical investigations are warranted to validate its safety and efficacy in humans and facilitate potential therapeutic integration.

Introduction

Rheumatoid arthritis (RA) is a chronic, systemic inflammatory disease that predominantly affects synovial joints, leading to progressive disability, increased mortality rates, and significant socioeconomic burdens (Negi et al. Citation2024). Clinically, RA is characterized by symmetrical joint involvement manifested as hyperplasia, arthralgia, edema, erythema, and impaired mobility (Smolen et al. Citation2023). Systemic manifestations, including infection, glucocorticoid-induced osteoporosis (GIOP), and the involvement of extra-articular organs such as the lungs, heart, nervous system, and musculoskeletal system, further complicate disease progression (Wu et al. Citation2022). Women are disproportionately affected, being three times more likely to develop RA compared to men. Globally, RA affects an estimated 18 million individuals, as reported by the Global Burden of Disease (GBD) study in 2019 (Vos et al. Citation2020; Cai et al. Citation2023). In the United States, the prevalence ranges between 0.6–1.0%, whereas in India, it is approximately 0.75% (Malaviya et al. Citation1993; Xu and Wu Citation2021). Diagnosis typically occurs within three months to two years after symptom onset, with disease progression often resulting in irreversible joint deformities and significant impairment in daily functioning (Bullock et al. Citation2018). Notably, over 50% of RA patients in developed nations reportedly leave the workforce within a decade of diagnosis (Kirkeskov and Bray Citation2023).

The pathogenesis of RA involves a complex interplay of genetic, immunological, and inflammatory mechanisms (Siouti and Andreakos Citation2019). Genetic predisposition is linked to specific human leukocyte antigen (HLA) alleles, while the immune response targets self-antigens, leading to the production of autoantibodies such as rheumatoid factor (RF) and anti-cyclic citrullinated peptide (anti-CCP) (Van Delft and Huizinga Citation2020). These autoantibodies recognize citrullinated antigens, generated via peptidylarginine deaminase-mediated post-translational modifications, triggering inflammation. Infiltration of immune cells into the synovium and subsequent cytokine release activate inflammatory pathways, with key mediators such as tumor necrosis factor-α (TNF-α), interleukin-1 (IL-1), and interleukin-6 (IL-6) exacerbating synovial inflammation and systemic effects (Jang et al. Citation2022). Chronic inflammation results in pannus formation, a pathological structure comprising immune cells, fibroblasts, and blood vessels, which invades adjacent tissues and promotes cartilage degradation, bone erosion, and joint deformities. Concurrently, systemic inflammation contributes to complications such as cardiovascular disorders, pulmonary issues, and a generalized inflammatory response (Panagopoulos and Lambrou Citation2018; Nandakumar et al. Citation2023). Although disease-modifying antirheumatic drugs (DMARDs), nonsteroidal anti-inflammatory drugs (NSAIDs), steroids, and monoclonal antibodies effectively alleviate symptoms and slow disease progression in RA, these therapies are often inadequate, leaving a substantial unmet need for more effective treatments (Radu and Bungau Citation2021).

Cyclosporine (CYC), initially approved as an immunosuppressant to prevent organ rejection, has demonstrated efficacy in RA patients unresponsive to conventional treatments such as methotrexate and NSAIDs/DMARDs (Tapia et al. Citation2023). CYC suppresses immune activity by inhibiting T-cell activation, which is a critical mediator of the inflammatory response in RA (Liddicoat and Lavelle Citation2019). However, oral formulations of CYC, such as Sandimmune® Soft Gelatin Capsules and Oral Solution, exhibit erratic bioavailability (approximately 34%), attributed to incomplete gastrointestinal absorption and extensive first-pass metabolism (Schuetz et al. Citation2005; Ershad et al. Citation2023). Furthermore, CYC administration is often associated with nephrotoxicity, limiting its clinical utility (Steinmuller Citation1989; Wu et al. Citation2018). Interestingly, curcumin (CUR), a polyphenolic compound with potent anti-inflammatory and antioxidative properties, has shown potential in mitigating CYC-induced nephrotoxicity while alleviating RA-associated inflammation and pain (Huang et al. Citation2018; Kadhim et al. Citation2021).

CUR, derived from the rhizomes of Curcuma longa, has demonstrated efficacy in reducing oxidative stress and chronic inflammation, making it a promising candidate for RA therapy. CUR modulates TNF-α expression to attenuate cartilage breakdown and inflammation (Sivani et al. Citation2022; Kunnumakkara et al. Citation2023). However, its therapeutic potential is hampered by low solubility, chemical instability, and limited bioavailability due to rapid hepatic metabolism into water-soluble conjugates like curcumin-glucuronide and curcumin-sulfonate (Sohn et al. Citation2021; El-Saadony et al. Citation2023). Addressing these limitations, this study explores a transdermal delivery system for CUR in combination with CYC, bypassing first-pass metabolism. Despite their potential, poor aqueous solubility and low permeability hinder the transdermal delivery of both CYC and CUR (El Hosary et al. Citation2024; Elhabal et al. Citation2024). While previous studies have explored the transdermal delivery of curcumin (CUR) and cyclosporine (CYC) individually using carriers such as liposomes (Pierre and dos Santos Miranda Costa Citation2011), solid lipid nanoparticles (Liu et al. Citation2020), and nanostructured lipid carriers (Khan et al. Citation2023), many of these systems do not comply with the Food and Drug Administration’s (FDA) Inactive Ingredient Database (IIG) limits. This is primarily due to the incorporation of high concentrations of surfactants, which may result in skin irritation or systemic toxicity. Moreover, several reported formulations do not account for the stability of CYC during processing, often involving harsh solvents or aggressive manufacturing conditions that could degrade the active compounds.

In our previously published study, a nanoemulsion-based gel system was developed for the co-delivery of CUR and CYC for the management of RA (Gharat et al., Citation2024). While the nanoemulsion approach improved drug solubilization and provided enhanced permeation via nanosized emulsion droplets, the current study explores an alternative vesicular strategy using ethosomes. Ethosomal gels offer a unique advantage due to their high ethanol content, which not only fluidizes the stratum corneum lipids but also enhances vesicle deformability, facilitating deeper transdermal penetration. This distinct mechanism positions ethosomes as a promising carrier for the effective delivery of both CUR and CYC through the skin, addressing the limitations associated with conventional transdermal systems. Ethosomes, phospholipid vesicles enriched with ethanol, offer a promising solution by enhancing solubility and permeability. Their unique structure enables deeper skin penetration, improved drug delivery, and increased therapeutic efficacy. Furthermore, smaller particle sizes enhance absorption while reducing aggregation and coalescence. Soya lecithin and cholesterol are widely utilized for ethosomal formulation due to their ability to stabilize lipid bilayers and enhance vesicular flexibility for effective skin penetration. Soya lecithin, rich in phosphatidylcholine, forms bilayers encapsulating drugs, while cholesterol maintains optimal bilayer fluidity and structural stability (Jadhav et al. Citation2024). Edge-activated ethosomes, a specialized form of ethosomes that incorporate edge activators such as Tween 80, Span 80, and sodium cholate, further enhance deformability and skin penetration. These surfactants disrupt phospholipid packing, increasing vesicle flexibility and preventing aggregation, thereby ensuring a stable and uniform formulation (Kumar Sarwa et al. Citation2015).

The objective of the present study is to develop and evaluate a transdermal delivery system for the co-delivery of CUR and CYC using ethosomal vesicles, with the aim of overcoming the limitations associated with previously reported carriers. The proposed system emphasizes the use of biocompatible excipients, minimal surfactants, and mild processing techniques to enhance drug solubility, permeability, and therapeutic efficacy. Specifically, edge-activated ethosomes encapsulating CUR and CYC were developed via the ethanol injection method and incorporated into a gel using Carbopol® Ultrez 10 NF. The final ethosomal gel was subjected to a series of evaluations including cell line studies, in vitro and ex vivo permeation analyses, and preclinical assessments using a Complete Freund’s Adjuvant (CFA)-induced arthritic rat model to establish its potential for rheumatoid arthritis management.

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Materials

Curcumin (99%) was purchased from Otto chemie Pvt. Ltd, Mumbai, India. Cyclosporine was provided as a gift sample by Concord Biotech Ltd, India. Tween 80 was procured from S.D. Fine chemicals Ltd., Mumbai, India. Soya lecithin (97%) and Cholesterol (99%) were purchased from Otto chemie. Pvt. Ltd, Mumbai, India. Absolute ethanol (99.9%), Polyvinyl alcohol (Molecular weight- 85,000 Da to 124,000 Da), Potassium dihydrogen phosphate (purified), and Disodium hydrogen phosphate (LR Grade) were obtained from S.D. Fine-Chem Limited, India. Carbopol® Ultrez 10 NF was generously provided by Lubrizol India Pvt Ltd, Mumbai, India. The RAW 264.7 cells were obtained from National Center for Cell Science, Pune, India. Enzyme-linked immunoassay (ELISA) kits for mouse interleukin-10 (IL-10), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) were purchased from Ray Biotech Labs, Norcross, Georgia. All excipients used in the formulation are ‘Generally Recognized as Safe’ listed.

Gharat, S., Momin, M., Panchal, U., & Omri, A. (2025). Novel ethosomal gel formulation for enhanced transdermal delivery of curcumin and cyclosporine: a preclinical approach to rheumatoid arthritis management. Drug Delivery, 32(1). https://doi.org/10.1080/10717544.2025.2512620