Novel strategy for immunomodulation: Dissolving microneedle array encapsulating thymopentin fabricated by modified two-step molding technology

Highlights

• TP5-DMNA was developed using a two-step molding technology for immunomodulation.

• BSA was used as co-material to fabricate TP5-DMNA for higher mechanical strength.

• The TP5-DMNA had equivalent immunomodulation efficiency to intravenous injection.

• The TP5-DMNA can be self-administrated with minimal pain and good compliance.

Abstract

Thymopentin (TP5) is commonly used in the treatment for autoimmune diseases, with a short plasma half-life (30 s) and a long treatment period (7 days to 6 months). It is usually administrated by syringe injection, resulting in compromised patient compliance. Dissolving microneedle array (DMNA) offers a superior approach for transdermal delivery of biological macromolecules, as it allows painless penetration through the stratum corneum and generates minimal biohazardous waste after dissolving in the skin. Despite recent advances in DMNA as a novel approach for transdermal drug delivery, problem of insufficient mechanical strength remains to be solved. In this study, TP5-loaded DMNA (TP5-DMNA) was uniquely developed using a modified two-step molding technology. The higher mechanical strength was furnished by employing bovine serum albumin (BSA) as a co-material to fabricate the needles. The obtained TP5-DMNA containing BSA displayed better skin penetration and higher drug loading efficiency than that without BSA. The in vivo pharmacodynamics study demonstrated that TP5-DMNA had comparative effect on immunomodulation to intravenous injection of TP5, in terms of ameliorating the CD4+/CD8+ ratio, SOD activity and MDA value to the basal level. Only mild irritation was observed at the site of administration. These results suggest that the novel TP5-DMNA utilizing BSA provides an alternative approach for convenient and safe transdermal delivery of TP5, which is a promising administration strategy for future clinical application.

Introduction

Currently, there emerged an increasing interest in the immunotherapy with proteins and polypeptides, due to their promising therapeutic efficiency. Hypodermic injection is the most common way for the delivery of these therapeutic agents, because of their unstable structure and high molecular weight [1]. However, frequent injection can lead to pain, infection and biohazardous waste, which compromise the patient compliance.

Thymopentin (TP5), a synthetic water-soluble pentapeptide (Arg-Lys-Asp-Val-Tyr), is an immunomodulating agent consisting of the 32nd to 36th residues of the 49 amino acid thymopoietin [2]. TP5 shows promise in promoting the differentiation of thymocytes and affecting the function of mature T-cells. Clinically, TP5 is used for treating autoimmune diseases [3] such as cancer, infections, acquired immunodeficiency syndrome [4], rheumatoid arthritis [5], [6], as well as primary and secondary immune deficiency. The commercially available product of TP5 is in the form of lyophilized powder for intramuscular or intravenous (i.v.) injection. And the patients have to receive frequent injections due to the long treatment period (7 days to 6 months) but very short half-life of TP5 in plasma (about 30 s) [7], which greatly reduces the patient compliance. Ideal formulation of TP5 should be readily administrated without pain, maximizing the patients’ compliance and improving therapeutic effect. Previous studies about TP5 were mainly focused on the development of sustained-release delivery systems for TP5 by hypodermic injection, such as microsphere, nanoparticle, liposome, and hydrogel, without considering pain and potential burst influence [8], [9], [10].

Recently, microneedles have been proposed as an alternative administration route for immunotherapeutic proteins and polypeptides. Microneedles (<1 mm) can create temporary conduits in an invasion-minimized manner [11], [12]. Being directly and readily utilized in skin without pain, microneedle arrays can be self-administrated with good patient compliance [13], [14]. Dissolving microneedle array (DMNA) fabricated from biocompatible and biodegradable polymers can gradually dissolve inside the skin without generating any biohazardous waste [15], [16]. Moreover, the application of DMNA can effectively prolong the shelf-life of proteins/polypeptides, and lower the cost of cold chain processes compared with the conventional solution counterparts [17]. Therefore, DMNA can be used as potential vehicles for transdermal delivery of TP5 as a novel treatment for immunodeficiency syndrome [18], [19].

Various methods has been used to fabricate DMNA, including micromolding, high-temperature molding [20], [21], UV photo-polymerization curing [22] and aqueous solution casting [23]. The problems involved in these methods are complicated steps, and potential impairment to the stability of biomacromolecular drugs caused by the radiation, polymerizing reagent or elevated temperature utilized in these processes [13], [24]. To overcome these problems, a two-step molding technology (Fig. 1) was utilized for fabricating DMNA. However, drug concentration gradient between the needles and the base causing the drug diffusion from the needle to the base results in lower drug-encapsulation efficiency in the needle. In our previous study, the evaporation of base solvent was found to be a key factor determining needle loading proportion (NDP), and improved NDP (90%) was achieved by replacing slow-evaporating water with highly volatile ethanol as the base solvent [25]. However, the mechanical strength of the obtained microneedles was still insufficient for porcine skin penetration.

The aim of this study was to deliver TP5 via novel transdermal route using DMNA (TP5-DMNA) with robust mechanical strength. To achieve higher mechanical strength for easier and better skin penetration of needle, the preparation of DMNA was optimised by employing BSA as a co-material. The mechanical strength of drug-free DMNA was evaluated, and the insertion capability of the TP5-DMNA in porcine cadaver skin was investigated. The immunomodulatory efficacy of this novel delivery system was confirmed via pharmacodynamic study in the induced immunosuppressed Sprague-Dawley (SD) rat model. A histological study of the administration site was also carried out to estimate the biocompatibility of TP5-DMNA.