Biologics, theranostics, and personalized medicine in drug delivery systems.

The progress in human disease treatment can be greatly advanced through the implementation of nanomedicine. This approach involves targeted and cell-specific therapy, controlled drug release, personalized dosage forms, wearable drug delivery, and companion diagnostics. By integrating cutting-edge technologies with drug delivery systems, greater precision can be achieved at the tissue and cellular levels through the use of stimuli-responsive nanoparticles, and the development of electrochemical sensor systems. This precision targeting - by virtue of nanotechnology - allows for therapy to be directed specifically to affected tissues while greatly reducing side effects on healthy tissues. As such, nanomedicine has the potential to transform the treatment of conditions such as cancer, genetic diseases, and chronic illnesses by facilitating precise and cell-specific drug delivery. Additionally, personalized dosage forms and wearable devices offer the ability to tailor treatment to the unique needs of each patient, thereby increasing therapeutic effectiveness and compliance. Companion diagnostics further enable efficient monitoring of treatment response, enabling customized adjustments to the treatment plan. The question of whether all the potential therapeutic approaches outlined here are viable alternatives to current treatments is also discussed. In general, the application of nanotechnology in the field of biomedicine may provide a strong alternative to existing treatments for several reasons. In this review, we aim to present evidence that, although in early stages, fully merging advanced technology with innovative drug delivery shows promise for successful implementation across various disease areas, including cancer and genetic or chronic diseases.

Aryl Hydrocarbon Receptor Agonism Antagonizes the Hypoxia-driven Inflammation in Cystic Fibrosis.

Hypoxia contributes to the exaggerated yet ineffective airway inflammation that fails to oppose infections in cystic fibrosis (CF). However, the potential for impairment of essential immune functions by HIF-1α (hypoxia-inducible factor 1α) inhibition demands a better comprehension of downstream hypoxia-dependent pathways that are amenable for manipulation. We assessed here whether hypoxia may interfere with the activity of AhR (aryl hydrocarbon receptor), a versatile environmental sensor highly expressed in the lungs, where it plays a homeostatic role. We used murine models of Aspergillus fumigatus infection in vivo and human cells in vitro to define the functional role of AhR in CF, evaluate the impact of hypoxia on AhR expression and activity, and assess whether AhR agonism may antagonize hypoxia-driven inflammation. We demonstrated that there is an important interferential cross-talk between the AhR and HIF-1α signaling pathways in murine and human CF, in that HIF-1α induction squelched the normal AhR response through an impaired formation of the AhR:ARNT (aryl hydrocarbon receptor nuclear translocator)/HIF-1ß heterodimer. However, functional studies and analysis of the AhR genetic variability in patients with CF proved that AhR agonism could prevent hypoxia-driven inflammation, restore immune homeostasis, and improve lung function. This study emphasizes the contribution of environmental factors, such as infections, in CF disease progression and suggests the exploitation of hypoxia:xenobiotic receptor cross-talk for antiinflammatory therapy in CF.

Biodrug Delivery Systems: Do mRNA Lipid Nanoparticles Come of Age?

As an appealing alternative to treat and prevent diseases ranging from cancer to COVID-19, mRNA has demonstrated significant clinical effects. Nanotechnology facilitates the successful implementation of the systemic delivery of mRNA for safe human consumption. In this manuscript, we provide an overview of current mRNA therapeutic applications and discuss key biological barriers to delivery and recent advances in the development of nonviral systems. The relevant challenges that LNPs face in achieving cost-effective and widespread clinical implementation when delivering mRNA are likewise discussed.

Low Efficacy of Genetic Tests for the Diagnosis of Primary Lymphedema Prompts Novel Insights into the Underlying Molecular Pathways.

Lymphedema is a chronic inflammatory disorder caused by ineffective fluid uptake by the lymphatic system, with effects mainly on the lower limbs. Lymphedema is either primary, when caused by genetic mutations, or secondary, when it follows injury, infection, or surgery. In this study, we aim to assess to what extent the current genetic tests detect genetic variants of lymphedema, and to identify the major molecular pathways that underlie this rather unknown disease. We recruited 147 individuals with a clinical diagnosis of primary lymphedema and used established genetic tests on their blood or saliva specimens. Only 11 of these were positive, while other probands were either negative (63) or inconclusive (73). The low efficacy of such tests calls for greater insight into the underlying mechanisms to increase accuracy. For this purpose, we built a molecular pathways diagram based on a literature analysis (OMIM, Kegg, PubMed, Scopus) of candidate and diagnostic genes. The PI3K/AKT and the RAS/MAPK pathways emerged as primary candidates responsible for lymphedema diagnosis, while the Rho/ROCK pathway appeared less critical. The results of this study suggest the most important pathways involved in the pathogenesis of lymphedema, and outline the most promising diagnostic and candidate genes to diagnose this disease.

Hazelnut Shells as Source of Active Ingredients: Extracts Preparation and Characterization.

Hazelnut shells represent a waste material (about 42% of the total biomass) deriving from hazelnut harvest. These are mainly used as a heating source; however, they represent an interesting source of polyphenols useful in health field. The impact on phenolic profile and concentrations of hazelnut shell extracts obtained by three extraction methods (maceration, ultrasonic bath, and high-power ultrasonic), as well as temperature, extraction time, and preventive maceration, was studied. The prepared extracts were characterized in terms of chemical composition, antioxidant and antimicrobial activities. Eighteen different phenolic compounds were identified and quantified by chemical analysis and gallic acid was the most abundant in all the extracts analyzed. Other relevant compounds were chlorogenic acid, protocatechuic acid and catechin. Preventive maceration had a positive effect on the extraction of different types of compounds regardless of the method performed. Application of the high-power ultrasonic method had different effects, either positive or negative, depending on the type of compound and extraction time. All the prepared extracts showed antioxidant activity especially those prepared by maceration, and many of them were able to inhibit the growth of both B. subtilis and B. cereus.

TIE1 as a Candidate Gene for Lymphatic Malformations with or without Lymphedema.

TIE1 is a cell surface protein expressed in endothelial cells. Involved in angiogenesis and lymphangiogenesis, including morphogenesis of lymphatic valves, TIE1 is important for lymphatic system functional integrity. The main purpose of this study was to identify different variants in the TIE1 gene that could be associated with lymphatic malformations or dysfunction and predisposition for lymphedema. In a cohort of 235 Italian lymphedema patients, who tested negative for variants in known lymphedema genes, we performed a further test for new candidate genes, including TIE1. Three probands carried different variants in TIE1. Two of these segregated with lymphedema or lymphatic dysfunction in familial cases. Variants in TIE1 could contribute to the onset of lymphedema. On the basis of our findings, we propose TIE1 as a candidate gene for comprehensive genetic testing of lymphedema.

Bioadhesive Polymeric Films Based on Red Onion Skins Extract for Wound Treatment: An Innovative and Eco-Friendly Formulation.

The onion non-edible outside layers represent a widely available waste material deriving from its processing and consumption. As onion is a vegetable showing many beneficial properties for human health, a study aiming to evaluate the use of extract deriving from the non-edible outside layers was planned. An eco-friendly extraction method was optimized using a hydroalcoholic solution as solvent. The obtained extract was deeply characterized by in vitro methods and then formulated in autoadhesive, biocompatible and pain-free hydrogel polymeric films. The extract, very soluble in water, showed antioxidant, radical scavenging, antibacterial and anti-inflammatory activities, suggesting a potential dermal application for wounds treatment. In vitro studies showed a sustained release of the extract from the hydrogel polymeric film suitable to reach concentrations necessary for both antibacterial and anti-inflammatory activities. Test performed on human keratinocytes showed that the formulation is safe suggesting that the projected formulation could be a valuable tool for wound treatment.

Increasing evidence of hereditary lymphedema caused by CELSR1 loss-of-function variants.

A whole exome sequencing approach was recently used to detect a CELSR1 truncating variant associated with lymphedema in a large pedigree. Since this first report, no other similar associations have been reported in the literature. Here, we present the genetic results of 95 probands tested using a next generation sequencing panel that covered all known lymphedema-associated genes, including CELSR1. Five out of 95 probands (5.3%) were found to carry novel loss-of-function variants in CELSR1. Family segregation studies were possible in four out of five probands and showed possible sex-specific differences: CELSR1 variants showed almost complete penetrance in females and were associated with early-onset lymphedema, whereas in males they showed incomplete penetrance and were associated with late onset of the condition. Since the percentage of lymphedema patients carrying CELSR1 variants is not negligible, we do not hesitate to recommend including this gene in routine genetic testing.

Tumor Targeting by Peptide-Decorated Gold Nanoparticles.

Cancer remains one of the most important challenges in biomedical sciences. Chemotherapeutic agents are very potent molecules that exhibit a significant level of toxicity in numerous tissues of the body, particularly in those characterized by high proliferative activity, such as the bone marrow. The scenario is even more complex in the case of the central nervous system, and in particular brain tumors, where the blood brain barrier limits the efficacy of drug therapies. Integrins, transmembrane proteins widely expressed in different types of cancer (glioblastoma, melanoma, and breast cancer), regulate the angiogenic process and play a pivotal role in tumor growth and invasion. Here, we report a nanotechnology strategy based on the use of AuNPs decorated with an arginine-glycine-aspartic acid-like peptide for the diagnosis and treatment of cancer. Two hours after administration in mice, the accumulation of the peptide-decorated NPs in the subcutaneous tumor was ∼4-fold higher than that of uncoated particles and ∼1.4-fold higher than that of PEGylated particles. Also, in the case of the intracranial tumor model, interesting results were obtained. Indeed, 2 h after administration, the amount of peptide-decorated particles in the brain was 1.5-fold that of undecorated particles and 5-fold that of PEGylated particles. In conclusion, this preliminary study demonstrates the high potential of this carrier developed for diagnostic and therapeutic applications.

Exploring the Nano Spray-Drying Technology as an Innovative Manufacturing Method for Solid Lipid Nanoparticle Dry Powders.

To take advantage of solid-state properties, the nano spray-drying (NSD) technique was investigated as an innovative one-step method to produce solid lipid nanoparticles (SLN) in the form of a dry powder starting from a lipid/leucine O/W emulsion. Compritol was chosen as wall-forming lipid. Rapamycin (Rp) was employed as a model drug to be loaded into SLN. Based on an initial screening, Lutrol F68 was chosen as surfactant and high-shear homogenization as an emulsification method. A two-level fractional factorial design and an extended factorial design were employed to determine critical factors and best preparation conditions. Compritol concentration, L-leucine/lipid ratio, and Lutrol F68 concentration resulted critical. Best conditions granted 51% yield, 3.2 µm L-leucine/SLN particle size, and a SLN population around 150 nm. All samples showed the presence of lipid aggregates. Material loss in the emulsification step was found responsible for SLN aggregation and low yield. The almost quantitative Rp loading increased SLN population span. Replacing compritol with cetyl palmitate produced aggregation of dry powders and SLN. Overall, NSD was found a fast method to produce SLN dry powders. More insightful assessment of the emulsification step and lipid property effects will be critical to the optimization of the NSD process. Hypotheses account for direct coupling of high-pressure homogenization with NSD for future successful development of this promising manufacturing method.

Dentifrice Based on Fluoride-Hydrotalcite Compounds: Characterization and Release Capacity Evaluation by Novel In Vitro Methods.

Anti-caries activity of fluoride ions is due to the protection against demineralization and the enhancement of remineralization of tooth enamel. Dentifrices available on the market contain sodium fluoride, sodium monofluorophosphate, stannous fluoride, and amine fluoride as source of these ions. A new compound working both as fluoride ion source and as abrasive was projected. Hybrids based on F- ions intercalated between the lamellae of hydrotalcite-like compounds (HTlc-F), namely MgAl-HTlc-F and ZnAl-HTlc-F, were prepared and characterized. Then, three different percentages (2, 3, and 4%) of both HTlc-F compounds were assayed. After the rheological characterization, the dentifrices containing 3 and 4% of MgAl-HTlc-F and ZnAl-HTlc-F, respectively, resulted to be the most suitable ones. Two novel in vitro methods, "rotary toothbrush method" and "manual brushing method," were developed and used in order to study the F- ions release from the prepared dentifrices. The obtained results showed that the dentifrice containing ZnAl-HTlc-F (4%) was the most effective in releasing fluoride ions. The "rotary toothbrush method" resulted to be the most suitable as the simulation of the brushing movements is standardizable and reproducible.

Towards Targeting the Aryl Hydrocarbon Receptor in Cystic Fibrosis.

Tryptophan (trp) metabolism is an important regulatory component of gut mucosal homeostasis and the microbiome. Metabolic pathways targeting the trp can lead to a myriad of metabolites, of both host and microbial origins, some of which act as endogenous low-affinity ligands for the aryl hydrocarbon receptor (AhR), a cytosolic, ligand-operated transcription factor that is involved in many biological processes, including development, cellular differentiation and proliferation, xenobiotic metabolism, and the immune response. Low-level activation of AhR by endogenous ligands is beneficial in the maintenance of immune health and intestinal homeostasis. We have defined a functional node whereby certain bacteria species contribute to host/microbial symbiosis and mucosal homeostasis. A microbial trp metabolic pathway leading to the production of indole-3-aldehyde (3-IAld) by lactobacilli provided epithelial protection while inducing antifungal resistance via the AhR/IL-22 axis. In this review, we highlight the role of AhR in inflammatory lung diseases and discuss the possible therapeutic use of AhR ligands in cystic fibrosis.

Artificial apolipoprotein corona enables nanoparticle brain targeting.

Many potential therapeutic compounds for brain diseases fail to reach their molecular targets due to the impermeability of the blood-brain barrier, limiting their clinical development. Nanotechnology-based approaches might improve compounds pharmacokinetics by enhancing binding to the cerebrovascular endothelium and translocation into the brain. Adsorption of apolipoprotein E4 onto polysorbate 80-stabilized nanoparticles to produce a protein corona allows the specific targeting of cerebrovascular endothelium. This strategy increased nanoparticle translocation into brain parenchyma, and improved brain nanoparticle accumulation 3-fold compared to undecorated particles (119.8 vs 40.5 picomoles). Apolipoprotein decorated nanoparticles have high clinical translational potential and may improve the development of nanotechnology-based medicine for a variety of neurological diseases.

The long and winding road to inhaled TB therapy: not only the bug's fault.

Not all of the issues impacting the success of tuberculosis (TB) treatment arise from pathogen-related disease characteristics. Nowadays, there is an increasing awareness that antibiotic treatment is not the only answer to the TB problem, promoting the search for alternative administration strategies and host-directed therapies. Among all the administration routes, being the lungs the main TB focus, inhalation is conceptually a logical solution to enhance treatment effectiveness and compliance. Nevertheless, research efforts and funding are almost entirely conveyed to conventional approaches. This review will critically evaluate the reasons constraining research in this field, providing some future perspectives. The most recent advances in inhalation approaches for TB will be discussed, either at the preclinical or clinical phase, illustrating the risk of failure and chances of success.

Development and characterization of an emulgel based on a snail slime useful for dermatological applications.

Snail slime is an interesting material for effective dermatological use (e.g. wounds). Its properties are stricly connected to the origin. In this paper a snail slime, deriving from the species Helix aspersa Muller and obtained from a company, was deeply characterized and then properly formulated. The slime, obtained by Donatella Veroni method, was firstly submitted to NMR analysis in order to evaluate the chemical composition. The main molecules found are glycolate and allantoin, well known for their activities in wound healing promotion. In vitro experiments performed on keratinocytes, revealed the snail slime ability to promote cellular well-being. Moreover, the microbiological analysis showed high activity against many strains involved in wounds infections such as gram+ (e.g. S. aureus, S. pyogenes), gram- (e.g. P. aeruginosa, E. coli) and the yeast C. albicans. The effect on skin elasticity was evaluated as well by the instrument Cutometer® dualMPA580. The snail slime was then formulated as hydrophilic gel, using a combination of corn starch and sodium hyaluronate as polymers, then used as external water phase of an O/W emulgel. The formulation is physically stable and easily spreadable and demonstrated antimicrobial activity as observed for slime alone, suggesting its suitability to be used for wound treatment.

The immune and microbial homeostasis determines the Candida-mast cells cross-talk in celiac disease.

Celiac disease (CD) is an autoimmune enteropathy resulting from an interaction between diet, genome, and immunity. Although many patients respond to a gluten-free diet, in a substantive number of individuals, the intestinal injury persists. Thus, other factors might amplify the ongoing inflammation. Candida albicans is a commensal fungus that is well adapted to the intestinal life. However, specific conditions increase Candida pathogenicity. The hypothesis that Candida may be a trigger in CD has been proposed after the observation of similarity between a fungal wall component and two CD-related gliadin T-cell epitopes. However, despite being implicated in intestinal disorders, Candida may also protect against immune pathologies highlighting a more intriguing role in the gut. Herein, we postulated that a state of chronic inflammation associated with microbial dysbiosis and leaky gut are favorable conditions that promote C. albicans pathogenicity eventually contributing to CD pathology via a mast cells (MC)-IL-9 axis. However, the restoration of immune and microbial homeostasis promotes a beneficial C. albicans-MC cross-talk favoring the attenuation of CD pathology to alleviate CD pathology and symptoms.

A microbially produced AhR ligand promotes a Tph1-driven tolerogenic program in multiple sclerosis.

Multiple sclerosis is a debilitating autoimmune disease, characterized by chronic inflammation of the central nervous system. While the significance of the gut microbiome on multiple sclerosis pathogenesis is established, the underlining mechanisms are unknown. We found that serum levels of the microbial postbiotic tryptophan metabolite indole-3-carboxaldehyde (3-IAld) inversely correlated with disease duration in multiple sclerosis patients. Much like the host-derived tryptophan derivative L-Kynurenine, 3-IAld would bind and activate the Aryl hydrocarbon Receptor (AhR), which, in turn, controls endogenous tryptophan catabolic pathways. As a result, in peripheral lymph nodes, microbial 3-IAld, affected mast-cell tryptophan metabolism, forcing mast cells to produce serotonin via Tph1. We thus propose a protective role for AhR-mast-cell activation driven by the microbiome, whereby natural metabolites or postbiotics will have a physiological role in immune homeostasis and may act as therapeutic targets in autoimmune diseases.

Development of spray-dried N-acetylcysteine dry powder for inhalation.

N-acetylcysteine (NAC) has both antioxidant and immunomodulatory activities and has been used as adjuvant therapy in several viral infections. Recently, NAC attracted attention for its possible role in reducing the affinity of the spike protein receptor binding domain to angiotensin-converting enzyme (ACE2) receptors. Since only NAC solutions are available for inhalation, the purpose of the work was to develop a NAC dry powder for inhalation using mannitol or leucine as excipient. The powder was successfully produced using co-spray-drying with leucine. ATR-FTIR analyses evidenced spectral variations ascribed to the formation of specific interactions between NAC and leucine. This effect on the NAC environment was not evident for NAC-mannitol powders, but mannitol was in a different polymorphic form compared to the supplied material. Both the feedstock concentration and the leucine content have an impact on the powder aerodynamic features. In particular, to maximize the respirable fraction, it is preferable to produce the powder starting from a 0.5 % w/v feedstock solution using 33 to 50 % w/w leucine content. The NAC-leucine powder was stable for ten months maintaining NAC content of 50 % (w/w) and about 200 µg of NAC was able to deposit on a transwell insert, useful for future in vitro studies.

Turning Microbial AhR Agonists into Therapeutic Agents via Drug Delivery Systems.

Developing therapeutics for inflammatory diseases is challenging due to physiological mucosal barriers, systemic side effects, and the local microbiota. In the search for novel methods to overcome some of these problems, drug delivery systems that improve tissue-targeted drug delivery and modulate the microbiota are highly desirable. Microbial metabolites are known to regulate immune responses, an observation that has resulted in important conceptual advances in areas such as metabolite pharmacology and metabolite therapeutics. Indeed, the doctrine of "one molecule, one target, one disease" that has dominated the pharmaceutical industry in the 20th century is being replaced by developing therapeutics which simultaneously manipulate multiple targets through novel formulation approaches, including the multitarget-directed ligands. Thus, metabolites may not only represent biomarkers for disease development, but also, being causally linked to human diseases, an unexploited source of therapeutics. We have shown the successful exploitation of this approach: by deciphering how signaling molecules, such as the microbial metabolite, indole-3-aldehyde, and the repurposed drug anakinra, interact with the aryl hydrocarbon receptor may pave the way for novel therapeutics in inflammatory human diseases, for the realization of which drug delivery platforms are instrumental.

Ketorolac Loaded Poly(lactic-co-glycolic acid) Coating of AZ31 in the Treatment of Bone Fracture Pain.

Biodegradable metal alloys may be successfully used to support bone repair, avoiding second surgery commonly needed when inert metal alloys are used. Combining a biodegradable metal alloy with a suitable pain relief agent could improve patient quality of life. AZ31 alloy was coated using a poly(lactic-co-glycolic) acid (PLGA) polymer loaded with ketorolac tromethamine using the solvent casting method. The ketorolac release profile from the polymeric film and the coated AZ31 samples, the PLGA mass loss of polymeric film, and the cytotoxicity of the optimized coated alloy were assessed. The coated sample showed a ketorolac release that was prolonged for two weeks, which was slower than that of just the polymeric film, in simulated body fluid. PLGA mass loss was complete after a 45-day immersion in simulated body fluid. The PLGA coating was able to lower AZ31 and ketorolac tromethamine cytotoxicity observed in human osteoblasts. PLGA coating also prevents AZ31 cytotoxicity, which was identified in human fibroblasts. Therefore, PLGA was able to control ketorolac release and protect AZ31 from premature corrosion. These characteristics allow us to hypothesize that the use of ketorolac tromethamine-loaded PLGA coating on AZ31 in the management of bone fractures can favor osteosynthesis and relief pain.