Double-layer dissolving microneedles for delivery of mesenchymal stem cell Secretome: Formulation, characterisation and skin irritation study.

Regenerative therapy based on stem cells have been developed, focusing on either stem cell or secretome delivery. Most marketed cellular and gene therapy products are available as injectable dosage forms, leading to several limitations requiring alternative routes, such as the intradermal route. Microneedles, capable of penetratingthe stratum corneumbarrier, offer a potential alternative for intradermal delivery. This present study aimed to develop double-layer dissolving microneedles (DMN) for the delivery of freeze-dried mesenchymal stem cell secretome. DMNs were fabricated using a two-step casting method and composed of two polymer combinations: poly(vinyl pyrrolidone) (PVP) with poly(vinyl alcohol) (PVA) or PVP with sodium hyaluronate (SH). The manufactured DMNs underwent assessments for morphology, mechanical strength, in skin dissolution, protein content, in vitro permeation, in vivo skin irritation, and physical stability. Based on evaluations of morphology and mechanical strength, two formulas (F5 and F12) met acceptance criteria. Evaluation of protein content revealed that F12 (PVP-SH combination) had a higher protein content than F5 (PVP-PVA combination), 99.02 ± 3.24 µg and 78.36 ± 3.75 µg respectively. In vitro permeation studies showed that F5 delivered secretome protein by 100.84 ± 0.88%, while F12 delivered 99.63 ± 9.21% in 24 h. After four days of observation onSprague-Dawleyrat's skin, no signs of irritation, such as oedema and redness, was observed after applying both formulations. The safety of using PVP-PVA and PVP-SH combinations as excipients for DMN secretome delivery has been confirmed, promising significant advancements in biotherapeutic development in the future.

Development of ropivacaine hydrochloride-loaded dissolving microneedles as a local anesthetic agent: A proof-of-concept.

Ropivacaine hydrochloride (RPL) is a local anesthetic agent that has been widely used for the treatment of pain during or after surgery. However, this drug is only available in parenteral dosage form and may contribute to the infiltration of RPL into the plasma, causing some undesirable side effects. Intradermal delivery of RPL using dissolving microneedles may become a promising strategy to deliver such drugs into the skin. This research aimed to develop RPL-loaded dissolving microneedles (DMN-RPLs) as a proof of the concept of intradermal delivery of a local anesthetic. The DMN-RPLs were fabricated using either centrifugation or air-pressurized chamber methods. Several polymers, such as poly(vinyl pyrrolidone) (PVP), poly(vinyl alcohol) (PVA), and sodium hyaluronate (SH), were utilized for manufacturing the DMN-RPLs. The prepared DMN-RPLs were assessed for their thermal properties, chemical bonds, mechanical strength, insertion ability, skin-dissolution study, and drug content. Furthermore, in-skin deposition and dermatokinetic studies were also performed. The results showed that F9 (30 % w/w PVP-4 % w/w SH) and F10 (30 % w/w PVP-5 % w/w PVA) containing 5 % w/w of RPL were the most promising formulations, as shown by their needle height reduction (<10 %) and insertion depth (∼400 µm). Both formulations were also able to deliver more than 60 % of the RPL contained in the DMNs into the epidermis, dermis, and receiver compartment. This study, for the first time, has provided a proof concept to deliver RPL as a local anesthetic using DMNs and the intradermal route, aiming to minimize pain and discomfort during administration and improve the patient's experience.

Quercetin loaded polymeric dissolving microarray patches: fabrication, characterisation and evaluation.

Quercetin, a natural compound, shows promising potential in wound healing by reducing fibrosis, limiting scar formation, and boosting fibroblast proliferation. However, its effectiveness is hindered by poor solubility, resulting in low bioavailability and necessitating high doses for therapeutic efficacy. This study presents a novel approach, fabricating quercetin-loaded microarray patches (MAPs) using widely employed solubility enhancement strategies. Fabricated MAPs exhibited favourable mechanical strength and could be inserted into excised porcine skin to a depth of 650 µm. Furthermore, formulations containing Soluplus® significantly increased the drug loading capacity, achieving up to 2.5 mg per patch and complete dissolution within an hour of application on excised porcine skin. In vitro studies on full-thickness neonatal porcine skin demonstrated that Soluplus®-enhanced MAPs effectively delivered quercetin across various skin layers, achieving a delivery efficiency exceeding 80% over 24 h. Additionally, these prototype MAPs displayed anti-inflammatory properties and demonstrated biocompatibility with human keratinocyte skin cells. Therefore, quercetin-loaded MAPs employing Soluplus® as a solubility enhancer present a promising alternative strategy for wound healing and anti-inflammatory therapy applications.

Microneedle-Mediated Transdermal Delivery of Genetic Materials, Stem Cells, and Secretome: An Update and Progression.

Medical practitioners commonly use oral and parenteral dosage forms to administer drugs to patients. However, these forms have certain drawbacks, particularly concerning patients' comfort and compliance. Transdermal drug delivery presents a promising solution to address these issues. Nevertheless, the stratum corneum, as the outermost skin layer, can impede drug permeation, especially for macromolecules, genetic materials, stem cells, and secretome. Microneedles, a dosage form for transdermal delivery, offer an alternative approach, particularly for biopharmaceutical products. In this review, the authors will examine the latest research on microneedle formulations designed to deliver genetic materials, stem cells, and their derivatives. Numerous studies have explored different types of microneedles and evaluated their ability to deliver these products using preclinical models. Some of these investigations have compared microneedles with conventional dosage forms, demonstrating their significant potential for advancing the development of biotherapeutics in the future.

Fabrication of dissolving microneedles for transdermal delivery of protein and peptide drugs: polymer materials and solvent casting micromoulding method.

Proteins and peptides are rapidly developing pharmaceutical products and are expected to continue growing in the future. However, due to their nature, their delivery is often limited to injection, with drawbacks such as pain and needle waste. To overcome these limitations, microneedles technology is developed to deliver protein and peptide drugs through the skin. One type of microneedles, known as dissolving microneedles, has been extensively studied for delivering various proteins and peptides, including ovalbumin, insulin, bovine serum albumin, polymyxin B, vancomycin, and bevacizumab. This article discusses polymer materials used for fabricating dissolving microneedles, which are poly(vinylpyrrolidone), hyaluronic acid, poly(vinyl alcohol), carboxymethylcellulose, GantrezTM, as well as other biopolymers like pullulan and ulvan. The paper is focused solely on solvent casting micromoulding method for fabricating dissolving microneedles containing proteins and peptides, which will be divided into one-step and two-step casting micromoulding. Additionally, future considerations in the market plan for dissolving microneedles are discussed in this article.

Combination of Dissolving Microneedles with Nanosuspension and Co-Grinding for Transdermal Delivery of Ketoprofen.

Ketoprofen is an anti-inflammatory agent that may cause gastric irritation if administered orally. Dissolving microneedles (DMN) can be a promising strategy to overcome this issue. However, ketoprofen has a low solubility; therefore, it is essential to enhance its solubility using certain methods, namely nanosuspension (NS) and co-grinding (CG). This research aimed to formulate DMN containing ketoprofen-loaded NS and CG. Ketoprofen NS was formulated with poly(vinyl alcohol) (PVA) at concentrations of 0.5%, 1%, and 2%. CG was prepared by grinding ketoprofen with PVA or poly(vinyl pyrrolidone) (PVP) at different drug-polymer ratios. The manufactured ketoprofen-loaded NS and CG were evaluated in terms of their dissolution profile. The most promising formulation from each system was then formulated into microneedles (MNs). The fabricated MNs were assessed in terms of their physical and chemical properties. An in vitro permeation study using Franz diffusion cells was also carried out. The most promising MN-NS and MN-CG formulations were F4-MN-NS (PVA 5%-PVP 10%), F5-MN-NS (PVA 5%-PVP 15%), F8-MN-CG (PVA 5%-PVP 15%), and F11-MN-CG (PVA 7.5%-PVP 15%), respectively. The cumulative amounts of drug permeated after 24 h for F5-MN-NS and F11-MN-CG were 3.88 ± 0.46 µg and 8.73 ± 1.40 µg, respectively. In conclusion, the combination of DMN with nanosuspension or a co-grinding system may be a promising strategy for delivering ketoprofen transdermally.

Enhancing Intradermal Delivery of Lidocaine by Dissolving Microneedles: Comparison between Hyaluronic Acid and Poly(Vinyl Pyrrolidone) Backbone Polymers.

Lidocaine hydrochloride (LiH), an amide-type local anesthetic agent, is commonly used in dermatological procedures. LiH is categorized as a BCS (biopharmaceutics classification system) class III group, which has high solubility and poor permeability. It should be noted that, in this context, LiH is intended as a local anesthetic, so the level of LiH in systemic circulation should be minimized to avoid toxicity and unwanted side effects such as hypotension and bradycardia. This study aimed to formulate and evaluate LiH-loaded dissolving microneedles (DMNs) with different polymer bases. Moreover, an in vitro permeation study using Franz diffusion cells and in vivo study were also performed. LiH-loaded DMNs were prepared using polymer groups of poly(vinyl pyrrolidone) (PVP-K30) and hyaluronic acid (HA). DMNs were created using the micro-molding method with centrifugation. The formulations selected based on the evaluation were F3 (HA 10%) and F5 (PVP-K30 25%). Based on the in vitro permeation study, the amount of drug permeated and deposited in the skin at F3 (HA 10%) was 247.1 ± 41.85 and 98.35 ± 12.86 µg, respectively. On the other hand, the amount of drug permeated and deposited in the skin at F5 (PVP-K30 25%) was 277.7 ± 55.88 and 59.46 ± 9.25 µg, respectively. Our in vivo drug-permeation study showed that only one rat from the PVP-K30 polymer group-with a concentration of 150.32 ng/mL-was detected on rat plasma. Therefore, LiH can be formulated into a DMN and can be deposited in the skin with a safe concentration of the drug permeating into systemic circulation.

Enhancement strategies for transdermal drug delivery systems: current trends and applications.

Transdermal drug delivery systems have become an intriguing research topic in pharmaceutical technology area and one of the most frequently developed pharmaceutical products in global market. The use of these systems can overcome associated drawbacks of other delivery routes, such as oral and parenteral. The authors will review current trends, and future applications of transdermal technologies, with specific focus on providing a comprehensive understanding of transdermal drug delivery systems and enhancement strategies. This article will initially discuss each transdermal enhancement method used in the development of first-generation transdermal products. These methods include drug/vehicle interactions, vesicles and particles, stratum corneum modification, energy-driven methods and stratum corneum bypassing techniques. Through suitable design and implementation of active stratum corneum bypassing methods, notably microneedle technology, transdermal delivery systems have been shown to deliver both low and high molecular weight drugs. Microneedle technology platforms have proven themselves to be more versatile than other transdermal systems with opportunities for intradermal delivery of drugs/biotherapeutics and therapeutic drug monitoring. These have shown that microneedles have been a prospective strategy for improving transdermal delivery systems.

Dissolving microneedle patches loaded with amphotericin B microparticles for localised and sustained intradermal delivery: Potential for enhanced treatment of cutaneous fungal infections.

Fungal infections affect millions of people globally and are often unreceptive to conventional topical or oral preparations because of low drug bioavailability at the infection site, lack of sustained therapeutic effect, and the development of drug resistance. Amphotericin B (AmB) is one of the most potent antifungal agents. It is increasingly important since fungal co-infections associated with COVID-19 are frequently reported. AmB is only administered via injections (IV) and restricted to life-threatening infections due to its nephrotoxicity and administration-related side effects. In this work, we introduce, for the first time, dissolving microneedle patches (DMP) loaded with micronised particles of AmB to achieve localised and long-acting intradermal delivery of AmB for treatment of cutaneous fungal infections. AmB was pulverised with poly (vinyl alcohol) and poly (vinyl pyrrolidone) to form micronised particles-loaded gels, which were then cast into DMP moulds to form the tips. The mean particle size of AmB in AmB DMP tips after pulverisation was 1.67 ± 0.01 µm. This is an easy way to fabricate and load microparticles into DMP, as few steps are required, and no organic solvents are needed. AmB had no covalent chemical interaction with the excipients, but the crystallinity of AmB was reduced in the tips. AmB was completely released from the tips within 4 days in vitro. AmB DMP presented inhibition of Candida albicans (CA) and the killing rate of AmB DMP against CA biofilm inside porcine skin reached 100% within 24 h. AmB DMP were able to pierce excised neonatal porcine skin at an insertion depth of 301.34 ± 46.86 µm. Ex vivo dermatokinetic and drug deposition studies showed that AmB was mainly deposited in the dermis. An in vivo dermatokinetic study revealed that the area under curve (AUC0-inf) values of AmB DMP and IV (Fungizone® bolus injection 1 mg/kg) groups were 8823.0 d∙µg/g and 33.4 d∙µg/g, respectively (264-fold higher). AmB remained at high levels (219.07 ± 102.81 µg/g or more) in the skin until 7 days after the application of AmB DMP. Pharmacokinetic and biodistribution studies showed that AmB concentration in plasma, kidney, liver, and spleen in the AmB DMP group was significantly lower than that in the IV group. Accordingly, this system addressed the systemic side effects of intravenous injection of AmB and localised the drug inside the skin for a week. This work establishes a novel, easy and effective method for long-acting and localised intradermal drug delivery.

Natural Deep Eutectic Solvents (NADES): Phytochemical Extraction Performance Enhancer for Pharmaceutical and Nutraceutical Product Development.

Natural products from plants were extracted and widely studied for their activities against many disease conditions. The selection of the extracting solvent is crucial to develop selective and effective methods for the extraction and isolation of target compounds in the plant matrices. Pharmacological properties of plant extracts and their bioactive principles are related to their excellent solubility, stability, and bioavailability when administered by different routes. This review aims to critically analyze natural deep eutectic solvents (NADES) as green solvents in their application to improve the extraction performance of plant metabolites in terms of their extractability besides the stability, bioactivity, solubility, and bioavailability. Herein, the opportunities for NADES to be used in pharmaceutical formulations development including plant metabolites-based nutraceuticals are discussed.

Versatility of hydrogel-forming microneedles in in vitro transdermal delivery of tuberculosis drugs.

Current therapy of tuberculosis (TB) has several limitations, such as risk of liver injury and intestinal dysbiosis due to frequent oral administration of antibiotics. Transdermal administration could be used to improve antibiotic delivery for treatment of Mycobacterium tuberculosis infection. Therefore, we developed a novel approach, using hydrogel-forming microneedle (MN) arrays to transdermally deliver TB drugs, namely rifampicin, isoniazid, pyrazinamide and ethambutol, which have different physicochemical properties. These drugs were individually prepared into three types of drug reservoirs, including lyophilised tablets, directly compressed tablets and poly(ethylene glycol) tablets. Formulations of each drug reservoir type were optimised to achieve a rapidly dissolving tablet, and further integrated with hydrogel-forming MN arrays for in vitro permeation studies. Three types of hydrogel formulation were manufactured using different type of polymers and crosslinking processes. These MN arrays were then evaluated in terms of swelling ability, morphology and physical properties. Results of solute diffusion studies showed that drug permeation across the swollen hydrogel membrane was affected mostly by physiochemical properties and functional groups of each drug. In the in vitro studies, the amount of permeated drug through the hydrogel-forming MN arrays across the dermatomed neonatal porcine skin was affected by the drug solubility and reservoir design. The highest permeation of rifampicin (3.64 mg) and ethambutol (46.99 mg) were achieved using MN arrays combined with the poly(ethylene glycol) tablets and directly compressed tablet, respectively. For isoniazid and pyrazinamide, the highest drug permeation was attained using lyophilised reservoir with the amount of drug delivered approximately 58.45 mg and 20.08 mg, respectively. These equate to transdermal delivery of approximately 75% (rifampicin), 79% (isoniazid), 20% (pyrazinamide) and 47% (ethambutol) of the drugs loaded into the reservoirs on average. Importantly, the results of this work have demonstrated the versatility of hydrogel formulations to deliver a TB drug regime using MN arrays. Accordingly, this is a promising approach to deliver high dose of TB drugs.

Development, Evaluation, and Pharmacokinetic Assessment of Polymeric Microarray Patches for Transdermal Delivery of Vancomycin Hydrochloride.

Methicillin-resistant Staphylococcus aureus (MRSA) can cause harmful and potentially deadly infections. Vancomycin remains the first-line antibiotic treatment for MRSA-derived infections. Nevertheless, as a peptide drug, it is poorly absorbed when administered orally because of its high molecular weight and low permeability in the gastrointestinal tract and is therefore administered intravenously for the treatment of systemic diseases. In order to circumvent some of the many drawbacks associated with intravenous injection, other routes of drug delivery should be investigated. One of the strategies which has been employed to enhance transdermal drug delivery is based on microarray patches (MAPs). This work, for the first time, describes successful transdermal delivery of vancomycin hydrochloride (VCL) using dissolving MAPs (DMAPs) and hydrogel-forming MAPs (HFMAPs). VCL was formulated into DMAPs and reservoirs [film dosage forms, lyophilized wafers, and compressed tablets (CSTs)] using excipients such as poly(vinyl pyrrolidone), poly(vinyl alcohol), sodium hyaluronate, d-sorbitol, and glycerol. In this study, HFMAPs were manufactured using aqueous blends containing poly(methylvinyl ether-co-maleic acid) cross-linked by esterification with poly(ethylene glycol). The VCL-loaded CSTs (60% w/w VCL) were the most promising reservoirs to be integrated with HFMAPs based on the physicochemical evaluations performed. Both HFMAPs and DMAPs successfully delivered VCL in ex vivo studies with the percentage of drug that permeated across the neonatal porcine skin recorded at 46.39 ± 8.04 and 7.99 ± 0.98%, respectively. In in vivo studies, the area under the plasma concentration time curve from time zero to infinity (AUC0-∞) values of 162.04 ± 61.84 and 61.01 ± 28.50 µg h/mL were achieved following the application of HFMAPs and DMAPs, respectively. In comparison, the AUC0-∞ of HFMAPs was significantly greater than that of the oral administration control group, which showed an AUC0-∞ of 30.50 ± 9.18 µg h/mL (p < 0.05). This work demonstrates that transdermal delivery of VCL is feasible using DMAPs and HFMAPs and could prove effective in the treatment of infectious diseases caused by MRSA, such as skin and soft tissue infections, lymphatic-related infections, and neonatal sepsis.

A sensitive HPLC-UV method for quantifying vancomycin in biological matrices: Application to pharmacokinetic and biodistribution studies in rat plasma, skin and lymph nodes.

Vancomycin (VCN) is an antibiotic used in the treatment of methicillin-resistant Staphylococcus aureus (MRSA)-derived infections. As it has a relatively narrow therapeutic window, it is imperative to develop a sensitive and reliable analytical method for drug monitoring and pharmacokinetic purposes. In the present study, quick sample preparations and a sensitive high-performance liquid chromatography method using UV detection (HPLC-UV) have been developed and validated. The analytical method for detection and quantification of VCN in rat plasma, skin and lymph node samples was validated based on the Food and Drug Administration (FDA) and European Medicine Agency (EMEA) bioanalytical method validation guidelines. The optimised plasma sample preparation involved a simple protein precipitation step, with extraction recovery of 100.3 ±â€¯0.92 %. VCN in all biological matrices was analysed in a HPLC-UV system (215 nm) using a Cortecs® C18 column (4.6 × 150 mm, 2.7 µm particle size) fitted with a guard cartridge set at 20 °C. Reverse phase HPLC under gradient conditions, with mobile phase consisting of 20 mM phosphate buffer containing 0.5 % v/v of triethylamine and a mixture of methanol - acetonitrile (70:30, v/v), and runtime of 12 min/sample was employed. The calibration standards used for plasma ranged between 0.1-50 µg/ml, while in the skin and lymph node matrices, standards ranged between 0.05-50 µg/ml with correlation coefficients (R2) of ≥ 0.9995 for all matrices. The method was selective, sensitive, accurate and precise for detecting and quantifying VCN in the biological matrices used. The validated method was successfully utilised in the detection of VCN in a pharmacokinetic and organ biodistribution study carried out in rats following oral and IV bolus administration of the drug. This validated bioanalytical method offers a wide range of potential applications in clinical therapeutic drug monitoring, pharmacokinetics and toxicology.

Solid lipid nanoparticle-based dissolving microneedles: A promising intradermal lymph targeting drug delivery system with potential for enhanced treatment of lymphatic filariasis.

Conventional oral therapy of lymphatic filariasis drugs is only effective to kill microfilariae in the bloodstream, but is often ineffective to kill adult filarial (macrofilariae) in the complex anatomy of the lymphatic system. The encapsulation of drugs into lipid-based nanoparticles with sizes of <100nm, and administration intradermally, could be used to enhance lymphatic uptake. Therefore, we developed an innovative approach, using solid lipid nanoparticles (SLNs) and dissolving microneedles (MNs) to deliver antifilariasis drugs, namely doxycycline, diethylcarbamazine and albendazole, intradermally. The SLNs were prepared from Geleol® and Tween®80 as a lipid matrix and stabilizer, respectively. The formulations were optimized using a central composite design, producing SLNs with sizes of <100nm. Drug release was sustained over 48h from SLNs, compared to pure drugs. The SLNs were then incorporated into a polymeric hydrogel which was casted to form SLNs-loaded MNs. SLNs-loaded MNs exhibited sufficient mechanical and insertion properties. Importantly, dermatokinetic studies showed that>40% of drugs were retained in the dermis of excised neonatal porcine skin up to 24h post-MN application, indicating the high possibility of the SLNs to be taken by the lymphatic system. In in vivo studies, the maximal lymph concentrations of the three drugs in rat, achieved following intradermal delivery, ranged between 4- and 7-fold higher than that recorded after oral administration. Additionally, compared to oral administration, despite the lower plasma Cmax and organ-distribution, the AUC and relative bioavailability of the three drugs in rat plasma was also higher using our delivery approach. Accordingly, this delivery approach could maximize the drugs concentrations in the lymph system without essentially increasing their plasma concentrations. This could potentially deliver the drugs efficiently to the bloodstream, where the microfilariae reside, while also targeting drug to the lymph nodes, where filarial nematodes reside in infected patients, leading to an effective therapy for lymphatic filariasis.