3D printed clotrimazole intravaginal ring for the treatment of recurrent vaginal candidiasis

Vulvovaginal candidiasis is a vaginal infection caused by the fungal pathogen Candida albicans that, most commonly, affects women of reproductive age. Its first-line treatment consists in topical applications of conventional drug formulations (e.g., creams, gels, tablets) containing imidazole drugs. The treatment involves single or multiple daily applications and, in the case of recurrences, daily administration of oral antifungal drugs for up to one month. Intravaginal rings are flexible, biocompatible medical devices that, compared to conventional drug formulations, offer the possibility of a controlled vaginal drug delivery over a determined period with a single application, thus increasing patient compliance. Among innovative manufacturing techniques, in recent years, fused deposition modeling 3D printing has emerged in the pharmaceutical field to produce different therapeutics combining drugs and polymers. This technique allows to print objects layer by layer with many different thermoplastic materials after a computer-assisted design. Thermoplastic polyurethanes are flexible and biocompatible materials that can be efficiently employed for the manufacturing of drug release systems, already utilized to prepare vaginal devices. In this work, we produced a clotrimazole-loaded intravaginal ring by fused deposition modeling 3D printing combining the drug with thermoplastic polyurethane using hot melt extrusion. The rings were computer-designed and then printed with two different drug concentrations (i.e., 2% and 10% w/w). The intravaginal rings were first tested in an agar-diffusion test to evaluate their effectiveness against C. albicans; and the 10% loaded ring was selected for further studies. Drug release was evaluated in two different media (i.e., 50% ethanol and vaginal fluid simulant) showing a sustained release over a period of seven days. Next, in vitro time-kill analysis against C. albicans in simulated vaginal fluid was performed and displayed a complete growth inhibition after 5 days, compared to the control. These results suggest a potential application of these 3D printed intravaginal rings for the treatment of vulvovaginal candidiasis and for the long-time treatment of recurrences

An easy 3D printing approach to manufacture vertical diffusion cells for in vitro release and permeation studies

Vertical diffusion cells are commonly used in the pharmaceutical and cosmetic fields to study the release and permeation of active ingredients through synthetic or biological membranes. Nevertheless, the commercially available glass-based systems are expensive and need to be carefully handled due to their fragility. Fused deposition modeling 3D printing is an additive manufacturing technique that allows producing objects layer by layer using different thermoplastic materials. Among them, polypropylene is a robust, flexible, and chemically inert polymer that can resist to many organic solvents. In this work, we designed and printed a vertical diffusion cell following pharmacopeia requirements by using polypropylene in a fused deposition modeling 3D printer. To keep the system thermostated, the developed model fits in a heating block to avoid the use of water recirculating system. The vertical diffusion cells were leak-free and presented chemical resistance and no interaction with model molecules (i.e., caffeine, diclofenac sodium, and glycyrrhetinic acid). The 3D printed cells were compared to commercially available glass cells and then two different types of synthetic membranes (i.e., PDMS and Strat-M®) were used to evaluate the permeation of a caffeine hydrogel. The developed 3D printed testing system could represent an efficient alternative to the glass-based equipment

Poly(3-hydroxybutyrate): A potential biodegradable excipient for direct 3D printing of pharmaceuticals

During the past decades, 3D printing has revolutionised different areas of research. Despite the considerable progress achieved in 3D printing of pharmaceuticals, the limited choice of suitable materials remains a challenge to overcome. The growing search for sustainable excipients has led to an increasing interest in biopolymers. Poly(3-hydroxybutyrate) (PHB) is a biocompatible and biodegradable biopolymer obtained from bacteria that could be efficiently employed in the pharmaceutical field. Here we aimed to demonstrate its potential application as a thermoplastic material for personalised medicine through 3D printing. More specifically, we processed PHB by using direct powder extrusion, a one-step additive manufacturing technique. To assess and denote the feasibility and versatility of the process, a 3D square model was manufactured in different dimensions (sidexheight: 12x2 mm; 18x2 mm; 24x2 mm) and loaded with increasing percentages of a model drug (up to 30% w/w). The manufacturing process was influenced by the drug content, and indeed, an increase in the amount of the drug determined a reduction in the printing temperature, without affecting the other parameters (such as the layer height). The composition of the model squares was investigated using Fourier-transform infrared spectroscopy, the resulting spectra confirmed that the starting materials were successfully incorporated into the final formulations. The thermal behaviour of the printed systems was characterized by differential scanning calorimetry, and thermal gravimetric analysis. Moreover, the sustained drug release profile of the formulations was performed over 21 days and showed to be dependent on the dimensions of the printed object and on the amount of loaded drug. Indeed, the formulation with 30% w/w in the dimension 24x2 mm released the highest amount of drug. Hence, the results suggested that PHB and direct powder extrusion technique could be promising tools for the manufacturing of prolonged release and personalised drug delivery forms

reactive oxygen species responsive nanoplatforms as smart drug delivery systems for gastrointestinal tract targeting

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Permeability-enhancing effects of three laurate-disaccharide monoesters across isolated rat intestinal mucosae

none8sìLaurate (C12)-sucrose esters are established intestinal epithelial permeation enhancers (PEs) with potential for use in oral delivery. Most studies have examined blends of ester rather than specific monoesters, with little variation on the sugar moiety. To investigate the influence of varying the sugar moiety on monoester performance, we compared three monoesters: C12-sucrose, C12-lactose, and C12-trehalose. The assays were: critical micellar concentration (CMC) in Krebs-Henseleit buffer, MTS and lactate dehydrogenase assays in Caco-2 cells, transepithelial electrical resistance (TEER) and apparent permeability coefficient (Papp) of [14C] mannitol across isolated rat intestinal mucosae, and tissue histology. For CMC, the rank order was C12-trehalose (0.21 mM) < C12-sucrose (0.34 mM) < C12-lactose (0.43 mM). Exposure to Caco-2 cells for 120 min produced TC50 values in the MTS assay from 0.1 to 0.4 mM. Each ester produced a concentration-dependent decrease in TEER across rat mucosae with 80% reduction seen with 8 mM in 5 min, but C12-trehalose was less potent. C12-sucrose and C12-lactose increased the Papp of [14C] mannitol across mucosae with similar potency and efficacy, whereas C12-trehalose was not as potent or efficacious, even though it still increased flux. In the presence of the three esters, gross intestinal histology was unaffected except at 8 mM for C12-sucrose and C12-lactose. In conclusion, the three esters enhanced permeability likely via tight junction modulation in rat intestinal tissue. C12-trehalose was not quite as efficacious, but neither did it damage tissue to the same extent. All three can be considered as potential PEs to be included in oral formulations.openMcCartney, Fiona; Perinelli, Diego R; Tiboni, Mattia; Cavanagh, Robert; Lucarini, Simone; Filippo Palmieri, Giovanni; Casettari, Luca; Brayden, David JMccartney, Fiona; Perinelli, Diego R; Tiboni, Mattia; Cavanagh, Robert; Lucarini, Simone; Filippo Palmieri, Giovanni; Casettari, Luca; Brayden, David

Peptide-guided resiquimod-loaded lignin nanoparticles convert tumor-associated macrophages from M2 to M1 phenotype for enhanced chemotherapy

Nanomedicines represent innovative and promising alternative technologies to improve the therapeutic effects of different drugs for cancer ablation. Targeting M2-like tumor-associated macrophages (TAMs) has emerged as a favorable therapeutic approach to fight against cancer through the modulation of the tumor microenvironment. However, the immunomodulatory molecules used for this purpose present side effects upon systemic administration, which limits their clinical translation. Here, the biocompatible lignin polymer is used to prepare lignin nanoparticles (LNPs) that carry a dual agonist of the toll-like receptors TLR7/8 (resiquimod, R848). These LNPs are targeted to the CD206-positive M2-like TAMs using the “mUNO” peptide, in order to revert their pro-tumor phenotype into anti-tumor M1-like macrophages in the tumor microenvironment of an aggressive triple-negative in vivo model of breast cancer. Overall, we show that targeting the resiquimod (R848)-loaded LNPs to the M2-like macrophages, using very low doses of R848, induces a profound shift in the immune cells in the tumor microenvironment towards an anti-tumor immune state, by increasing the representation of M1-like macrophages, cytotoxic T cells, and activated dendritic cells. This effect consequently enhances the anticancer effect of the vinblastine (Vin) when co-administered with R848-loaded LNPs to the M2-like macrophages, using very low doses of R848, induces a profound shift in the immune cells in the tumor microenvironment towards an anti-tumor immune state, by increasing the representation of M1-like macrophages, cytotoxic T cells, and activated dendritic cells. This effect consequently enhances the anticancer effect of the vinblastine (Vin) when co-administered with R848-loaded LNPs. Statement of significance Lignin-based nanoparticles (LNPs) were successfully developed to target a potent TLR7/8 agonist (R848) of the tumor microenvironment (TME). This was achieved by targeting the mannose receptor (CD206) on the tumor supportive (M2-like) macrophages with the "mUNO" peptide, to reprogram them into an antitumor (M1-like) phenotype for enhanced chemotherapy. LNPs modified the biodistribution of the R848, and enhanced its accumulation and efficacy in shifting the immunological profile of the cells in the TME, which was not achieved by systemic administration of free R848. Moreover, a reduction in the tumor volumes was observed at lower equivalent doses of R848 compared with other studies. Therefore, the co-administration of R848@LNPs is a promising chemotherapeutic application in aggressive tumors, such as the triple-negative breast cancer. (c) 2020 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.Peer reviewe

Prunus spinosa Extract Loaded in Biomimetic Nanoparticles Evokes In Vitro Anti-Inflammatory and Wound Healing Activities

none14sìPrunus spinosa fruits (PSF) contain different phenolic compounds showing antioxidant and anti-inflammatory activities. Innovative drug delivery systems such as biomimetic nanoparticles could improve the activity of PSF extract by promoting (i) the protection of payload into the lipidic bilayer, (ii) increased accumulation to the diseased tissue due to specific targeting properties, (iii) improved biocompatibility, (iv) low toxicity and increased bioavailability. Using membrane proteins extracted from human monocyte cell line THP-1 cells and a mixture of phospholipids, we formulated two types of PSF-extract-loaded biomimetic vesicles differing from each other for the presence of either 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) or 1,2-dioleoyl-sn-glycero-3-phospho-(1'-rac-glycerol) (DOPG). The biological activity of free extract (PSF), compared to both types of extract-loaded vesicles (PSF-DOPCs and PSF-DOPGs) and empty vesicles (DOPCs and DOPGs), was evaluated in vitro on HUVEC cells. PSF-DOPCs showed preferential incorporation of the extract. When enriched into the nanovesicles, the extract showed a significantly increased anti-inflammatory activity, and a pronounced wound-healing effect (with PSF-DOPCs more efficient than PSF-DOPG) compared to free PSF. This innovative drug delivery system, combining nutraceutical active ingredients into a biomimetic formulation, represents a possible adjuvant therapy for the treatment of wound healing. This nanoplatform could be useful for the encapsulation/enrichment of other nutraceutical products with short stability and low bioavailability.openTiboni, Mattia; Coppari, Sofia; Casettari, Luca; Guescini, Michele; Colomba, Mariastella; Fraternale, Daniele; Gorassini, Andrea; Verardo, Giancarlo; Ramakrishna, Seeram; Guidi, Loretta; Di Giacomo, Barbara; Mari, Michele; Molinaro, Roberto; Albertini, Maria CristinaTiboni, Mattia; Coppari, Sofia; Casettari, Luca; Guescini, Michele; Colomba, Mariastella; Fraternale, Daniele; Gorassini, Andrea; Verardo, Giancarlo; Ramakrishna, Seeram; Guidi, Loretta; Di Giacomo, Barbara; Mari, Michele; Molinaro, Roberto; Albertini, Maria Cristin

Prunus spinosa Extract Loaded in Biomimetic Nanoparticles Evokes In Vitro Anti-Inflammatory and Wound Healing Activities

Prunus spinosa fruits (PSF) contain different phenolic compounds showing antioxidant and anti-inflammatory activities. Innovative drug delivery systems such as biomimetic nanoparticles could improve the activity of PSF extract by promoting (i) the protection of payload into the lipidic bilayer, (ii) increased accumulation to the diseased tissue due to specific targeting properties, (iii) improved biocompatibility, (iv) low toxicity and increased bioavailability. Using membrane proteins extracted from human monocyte cell line THP-1 cells and a mixture of phospholipids, we formulated two types of PSF-extract-loaded biomimetic vesicles differing from each other for the presence of either 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) or 1,2-dioleoyl-sn-glycero-3-phospho-(1&prime;-rac-glycerol) (DOPG). The biological activity of free extract (PSF), compared to both types of extract-loaded vesicles (PSF-DOPCs and PSF-DOPGs) and empty vesicles (DOPCs and DOPGs), was evaluated in vitro on HUVEC cells. PSF-DOPCs showed preferential incorporation of the extract. When enriched into the nanovesicles, the extract showed a significantly increased anti-inflammatory activity, and a pronounced wound-healing effect (with PSF-DOPCs more efficient than PSF-DOPGs) compared to free PSF. This innovative drug delivery system, combining nutraceutical active ingredients into a biomimetic formulation, represents a possible adjuvant therapy for the treatment of wound healing. This nanoplatform could be useful for the encapsulation/enrichment of other nutraceutical products with short stability and low bioavailability

Microfluidics for nanomedicines manufacturing: An affordable and low-cost 3D printing approach

During the last decade, an innovative lab on a chip technology known as microfluidics became popular in the pharmaceutical field to produce nanomedicines in a scalable way. Nevertheless, the predominant barriers for new microfluidics users are access to expensive equipment and device fabrication expertise. 3D printing technology promises to be an enabling new field that helps to overcome these drawbacks expanding the realm of microfluidics. Among 3D printing techniques, fused deposition modeling allows the production of devices with relatively inexpensive materials and printers. In this work, we developed two different microfluidic chips designed to obtain a passive micromixing by a "zigzag" bas-relief and by the presence of "split and recombine" channels. Computational fluid dynamics studies improved the evaluation of the mixing potential. A fused deposition modeling 3D printer was used to print the developed devices with polypropylene as manufacturing material. Then, two different model nanocarriers (i.e., polymeric nanoparticles and liposomes), loading cannabidiol as model drug, were formulated evaluating the influence of manufacturing parameters on the final nanocarrier characteristics with a design of experiments approach (2-level full factorial design). Both the chips showed an effective production of nanocarriers with tunable characteristics and with an efficient drug loading. These polypropylene-based microfluidic chips could represent an affordable and low-cost alternative to common microfluidic devices for the effective manufacturing of nanomedicines (both polymer- and lipid-based) after appropriate tuning of manufacturing parameters