Structural and Functional Analysis of the D614G SARS-CoV-2 Spike Protein Variant.

The SARS-CoV-2 spike (S) protein variant D614G supplanted the ancestral virus worldwide, reaching near fixation in a matter of months. Here we show that D614G was more infectious than the ancestral form on human lung cells, colon cells, and on cells rendered permissive by ectopic expression of human ACE2 or of ACE2 orthologs from various mammals, including Chinese rufous horseshoe bat and Malayan pangolin. D614G did not alter S protein synthesis, processing, or incorporation into SARS-CoV-2 particles, but D614G affinity for ACE2 was reduced due to a faster dissociation rate. Assessment of the S protein trimer by cryo-electron microscopy showed that D614G disrupts an interprotomer contact and that the conformation is shifted toward an ACE2 binding-competent state, which is modeled to be on pathway for virion membrane fusion with target cells. Consistent with this more open conformation, neutralization potency of antibodies targeting the S protein receptor-binding domain was not attenuated.

IFIH1 (MDA5) is required for innate immune detection of intron-containing RNA expressed from the HIV-1 provirus.

Intron-containing RNA expressed from the HIV-1 provirus activates type 1 interferon in primary human blood cells, including CD4+ T cells, macrophages, and dendritic cells. To identify the innate immune receptor required for detection of intron-containing RNA expressed from the HIV-1 provirus, a loss-of-function screen was performed with short hairpin RNA-expressing lentivectors targeting twenty-one candidate genes in human monocyte-derived dendritic cells. Among the candidate genes tested, only knockdown of XPO1 (CRM1), IFIH1 (MDA5), or MAVS prevented activation of the interferon-stimulated gene ISG15. The importance of IFIH1 protein was demonstrated by rescue of the knockdown with nontargetable IFIH1 coding sequence. Inhibition of HIV-1-induced ISG15 by the IFIH1-specific Nipah virus V protein, and by IFIH1-transdominant 2-CARD domain-deletion or phosphomimetic point mutations, indicates that IFIH1 (MDA5) filament formation, dephosphorylation, and association with MAVS are all required for innate immune activation in response to HIV-1 transduction. Since both IFIH1 (MDA5) and DDX58 (RIG-I) signal via MAVS, the specificity of HIV-1 RNA detection by IFIH1 was demonstrated by the fact that DDX58 knockdown had no effect on activation. RNA-Seq showed that IFIH1 knockdown in dendritic cells globally disrupted the induction of IFN-stimulated genes by HIV-1. Finally, specific enrichment of unspliced HIV-1 RNA by IFIH1 (MDA5), over two orders of magnitude, was revealed by formaldehyde cross-linking immunoprecipitation (f-CLIP). These results demonstrate that IFIH1 is the innate immune receptor for intron-containing RNA from the HIV-1 provirus and that IFIH1 potentially contributes to chronic inflammation in people living with HIV-1, even in the presence of effective antiretroviral therapy.

From Preformulative Design to In Vivo Tests: A Complex Path of Requisites and Studies for Nanoparticle Ocular Application. Part 1: Design, Characterization, and Preliminary In Vitro Studies.

Ocular pathologies are widely diffused worldwide, and their effective treatment, combined with a high patient compliance, is sometimes challenging to achieve due to the barriers of the eye; in this context, the use of nanoparticles for topical ophthalmic application could represent a successful strategy. Aiming to develop nanoplatforms with potential clinical applications, great attention has to be paid to their features, in relation to the route of administration and to the pharmacopoeial requirements. This review (part 1) thus embraces the preliminary steps of nanoparticle development and characterization. At the beginning, the main barriers of the eye and the different administration routes are resumed, followed by a general description of the advantages of the employment of nanoparticles for ocular topical administration. Subsequently, the preformulative steps are discussed, deepening the choice of raw materials and determining the quantitative composition. Then, a detailed report of the physicochemical and technological characterization of nanoparticles is presented, analyzing the most relevant tests that should be performed on nanoparticles to verify their properties and the requisites (both mandatory and suggested) demanded by regulatory agencies. In conclusion, some preliminary noncellular in vitro evaluation methods are described. Studies from in vitro cellular assays to in vivo tests will be discussed in a separate (part 2) review paper. Hence, this overview aims to offer a comprehensive tool to guide researchers in the choice of the most relevant studies to develop a nanoplatform for ophthalmic drug administration.

Effect of Ferulic Acid Loaded in Nanoparticle on Tissue Transglutaminase Expression Levels in Human Glioblastoma Cell Line.

Glioblastoma (GBM) is one of the most aggressive cancers, characterized by a decrease in antioxidant levels. Evidence has demonstrated that ferulic acid (FA), a natural antioxidant particularly abundant in vegetables and fruits, could be a promising candidate for GBM treatment. Since FA shows a high instability that compromises its therapeutic application, it has been encapsulated into Nanostructured Lipid Carriers (NLCs) to improve its bioavailability in the brain. It has been demonstrated that tissue transglutaminase (TG2) is a multi-functional protein implicated in many physiological and pathological processes, including cancer. TG2 is also involved in GBM correlated with metastasis formation and drug resistance. Therefore, the evaluation of TG2 expression levels and its cellular localization are important to assess the anti-cancer effect of FA against GBM cancer. Our results have demonstrated that treatment with free FA and FA-NLCs in the U87-MG cancer cell line differently modified TG2 localization and expression levels. In the cells treated with free FA, TG2 appeared expressed both in the cytosol and in the nucleus, while the treatment with FA-NLCs showed that the protein is exclusively localized in the cytosol, exerting its pro-apoptotic effect. Therefore, our data suggest that FA loaded in NLCs could represent a promising natural agent for supplementing the current anti-cancer drugs used for the treatment of GBM.

Human Anti-HIV-1 gp120 Monoclonal Antibodies with Neutralizing Activity Cloned from Humanized Mice Infected with HIV-1.

Broadly neutralizing, anti-HIV-1 gp120 mAbs have been isolated from infected individuals, and there is considerable interest in developing these reagents for Ab-based immunoprophylaxis and treatment. As a means to identify potentially new anti-HIV Abs, we exploited humanized NOD-scid IL2rγnull mice systemically infected with HIV-1 to generate a wide variety of Ag-specific human mAbs. The Abs were encoded by a diverse range of variable gene families and Ig classes, including IgA, and several showed significant levels of somatic mutation. Moreover, the isolated Abs not only bound target Ags with similar affinity as broadly neutralizing Abs, they also demonstrated neutralizing ability against multiple HIV-1 clades. The use of humanized mice will allow us to use our knowledge of HIV-1 gp120 structure and function, and the immune response targeting this protein, to generate native human prophylactic Abs to reduce the infection and spread of HIV-1.

Nanotechnologies for intranasal drug delivery: an update of literature.

Scientific research has focused its attention on finding an alternative route to systemic oral and parenteral administration, to overcome their usual drawbacks, such as hepatic first-pass which decreases drug bioavailability after oral administration, off-target effects, low patient compliance and low speed of onset of the pharmacological action in first-aid cases. Innovative drug delivery systems (DDS), mainly based on polymer and lipid biocompatible materials, have given a great prompt in this direction in the last years. The intranasal (IN) route of administration is a valid non-invasive alternative. It is highly suitable for self-administration, the drug quickly reaches the bloodstream, largely avoiding the first pass effect, and can also reach directly the brain bypassing BBB. Association of IN route with DDS can thus become a winning strategy for the controlled delivery of drugs, especially when a very quick effect is desired or needed. This review aims at analyzing the scientific literature regarding IN-DDS and their different ways of administration (systemic, topical, pulmonary, nose-to-brain). In particular, attention was devoted to polymer- and lipid-based micro- and nanocarriers, being the topic of most published articles in the last decade, but the whole plethora of colloidal DDS investigated in recent years for IN administration was presented.

Coating Lacticaseibacillus rhamnosus GG in Alginate Systems: an Emerging Strategy Towards Improved Viability in Orange Juice.

Fruit juices are successfully proposed as suitable probiotic vehicles, but researchers' efforts should be developed to avoid effects of bacteria overgrowing on sensory and nutritional cues of final products and to preserve viability of probiotic bacteria during storage. In the present study, encapsulation of Lacticaseibacillus rhamnosus GG strain in alginate systems was performed through ionotropic gelation technology. The alginate systems were optimized by using Box-Behnken Design to investigate the influence of three independent variables at three different levels: particle mean size and polydispersity index. The optimized probiotic-loaded alginate particles were added to orange juice samples. The viability of the probiotic strain, both as free and microencapsulated, was evaluated in orange juice stored at 5°C for 35 days. Morphology and size of probiotic-loaded alginate particles were found suitable for incorporation into juice. TEM analysis revealed that unloaded systems were clustered as nanoparticles (CL_NP), while the loaded sample appeared as a coated system (Coated_LGG). Microbiological evaluation revealed that the encapsulation assured the survival of Coated_LGG, with a reduction of less than 1-unit log in cellular density after 35 days of refrigerated storage in orange juice. Results indicated that the encapsulated bacteria did not affect the macroscopic properties neither the microbiological characteristic of orange juice; thus, it can be proposed as functional food.

Curcumin Containing PEGylated Solid Lipid Nanoparticles for Systemic Administration: A Preliminary Study.

Curcumin (CUR) has a wide range of pharmacological properties, including anti-inflammatory and antioxidant activities, and it can be considered a good candidate for the potential treatment of central nervous system (CNS) pathologies, although its use in clinical practice is compromised due to its high lipophilicity. Solid lipid nanoparticles (SLNs) are well-known nanocarriers representing a consolidated approach for the delivery of lipophilic compounds, but their systemic use is limited due their short half-life. The formulation of stealth SLNs (pSLNs) could be a valid strategy to overcome this limit. Curcumin-loaded-pSLNs were prepared by the solvent evaporation method. Formulation was characterized for their mean size, zeta potential, size distribution, and morphology. Drug antioxidant activity was evaluated by Oxygen Radical Absorbance Capacity (ORAC) assay. Finally, the obtained formulations were analyzed in terms of long-term stability. Curcumin-loaded-pSLNs showed good technological parameters with a mean particle size below 200 nm, as confirmed by TEM images, and a zeta potential value around -30 mV, predicting good long-term stability. Differential Scanning Calorimetry (DSC) analysis confirmed that PEG micelles interacted with the SLN surface; this suggests the location of the PEG on the pSLN surface. Therefore, these preliminary studies suggest that the produced formulation could be regarded as a promising carrier for the systemic administration.

W/O/W Microemulsions for Nasal Delivery of Hydrophilic Compounds: A Preliminary Study.

The administration of hydrophilic therapeutics has always been a great challenge because of their low bioavailability after administration. For this purpose, W/O/W microemulsion resulted to be a potential successful strategy for the delivery of hydrophilic compounds, interesting for the nasal mucosal therapy. Herein, an optimized biphasic W/O microemulsion was designed, through a preliminary screening, and it was inverted in a triphasic W/O/W microemulsion, intended for the nasal administration. In order to enhance the mucosal retention, surface modification of the biphasic W/O microemulsion was performed adding didodecyldimethylammonium bromide, and then converting the system into a cationic triphasic W/O/W microemulsion. The developed samples were characterized in terms of droplet size, polydispersity, zeta potential, pH and osmolality. The physical long-term stability was analyzed storing samples at accelerated conditions (40 ± 2 °C and 75 ± 5 % RH) for 6 months in a constant climate chamber, following ICH guidelines Q1A (R2). In order to verify the potential retention on the nasal mucosa, the two triphasic systems were analyzed in terms of mucoadhesive properties, measuring the in vitro interaction with mucin over time. Furthermore, fluorescein sodium salt was selected as a model hydrophilic drug to be encapsulated into the inner core of the two triphasic W/O/W microemulsions, and its release was analyzed compared to the free probe solution. The cytocompatibility of the two platforms was assessed on two cell lines, human fibroblasts HFF1 and Calu-3 cell lines, chosen as pre-clinical models for nasal and bronchial/tracheal airway epithelium.

Nose-to-Brain Drug Delivery and Physico-Chemical Properties of Nanosystems: Analysis and Correlation Studies of Data from Scientific Literature.

Background: In the last few decades, nose-to-brain delivery has been investigated as an alternative route to deliver molecules to the Central Nervous System (CNS), bypassing the Blood-Brain Barrier. The use of nanotechnological carriers to promote drug transfer via this route has been widely explored. The exact mechanisms of transport remain unclear because different pathways (systemic or axonal) may be involved. Despite the large number of studies in this field, various aspects still need to be addressed. For example, what physicochemical properties should a suitable carrier possess in order to achieve this goal? To determine the correlation between carrier features (eg, particle size and surface charge) and drug targeting efficiency percentage (DTE%) and direct transport percentage (DTP%), correlation studies were performed using machine learning. Methods: Detailed analysis of the literature from 2010 to 2021 was performed on Pubmed in order to build "NANOSE" database. Regression analyses have been applied to exploit machine-learning technology. Results: A total of 64 research articles were considered for building the NANOSE database (102 formulations). Particle-based formulations were characterized by an average size between 150-200 nm and presented a negative zeta potential (ZP) from -10 to -25 mV. The most general-purpose model for the regression of DTP/DTE values is represented by Decision Tree regression, followed by K-Nearest Neighbors Regressor (KNeighbor regression). Conclusion: A literature review revealed that nose-to-brain delivery has been widely investigated in neurodegenerative diseases. Correlation studies between the physicochemical properties of nanosystems (mean size and ZP) and DTE/DTP parameters suggest that ZP may be more significant than particle size for DTP/DTE predictability.

In vitro and in vivo studies of ocular topically administered NLC for the treatment of uveal melanoma.

Uveal melanoma is one of the most common and aggressive intraocular malignancies, and, due to its great capability of metastasize, it constitutes the most incident intraocular tumor in adults. However, to date there is no effective treatment since achieving the inner ocular tissues still constitutes one of the greatest challenges in actual medicine, because of the complex structure and barriers. Uncoated and PEGylated nanostructured lipid carriers were developed to achieve physico-chemical properties (mean particle size, homogeneity, zeta potential, pH and osmolality) compatible for the ophthalmic administration of (S)-(-)-MRJF22, a new custom-synthetized prodrug for the potential treatment of uveal melanoma. The colloidal physical stability was investigated at different temperatures by Turbiscan® Ageing Station. Morphology analysis and mucoadhesive studies highlighted the presence of small particles suitable to be topically administered on the ocular surface. In vitro release studies performed using Franz diffusion cells demonstrated that the systems were able to provide a slow and prolonged prodrug release. In vitro cytotoxicity test on Human Corneal Epithelium and Human Uveal Melanoma cell lines and Hen's egg-chorioallantoic membrane test showed a dose-dependent cytotoxic effect of the free prodrug on corneal cells, whose cytocompatibility improved when encapsulated into nanoparticles, as also confirmed by in vivo studies on New Zealand albino rabbits. Antiangiogenic capability and preventive anti-inflammatory properties were also investigated on embryonated eggs and rabbits, respectively. Furthermore, preliminary in vivo biodistribution images of fluorescent nanoparticles after topical instillation in rabbits' eyes, suggested their ability to reach the posterior segment of the eye, as a promising strategy for the treatment of choroidal uveal melanoma.

High dickkopf-1 levels in sera and leukocytes from children with 21-hydroxylase deficiency on chronic glucocorticoid treatment.

Children with 21-hydroxylase deficiency (21-OHD) need chronic glucocorticoid (cGC) therapy to replace congenital deficit of cortisol synthesis, and this therapy is the most frequent and severe form of drug-induced osteoporosis. In this study, we enrolled 18 patients (9 females) and 18 sex- and age-matched controls. We found in 21-OHD patients high serum and leukocyte levels of dickkopf-1 (DKK1), a secreted antagonist of the Wnt/ß-catenin signaling pathway known to be a key regulator of bone mass. In particular, we demonstrated by flow cytometry, confocal microscopy, and real-time PCR that monocytes, T lymphocytes, and neutrophils from patients expressed high levels of DKK1, which may be related to the cGC therapy. In fact, we showed that dexamethasone treatment markedly induced the expression of DKK1 in a dose- and time-dependent manner in leukocytes. The serum from patients containing elevated levels of DKK1 can directly inhibit in vitro osteoblast differentiation and receptor activator of NF-κB ligand (RANKL) expression. We also found a correlation between both DKK1 and RANKL or COOH-terminal telopeptides of type I collagen (CTX) serum levels in 21-OHD patients on cGC treatment. Our data indicated that DKK1, produced by leukocytes, may contribute to the alteration of bone remodeling in 21-OHD patients on cGC treatment.

Cellular mechanisms of multiple myeloma bone disease.

Multiple myeloma (MM) is a hematologic malignancy of differentiated plasma cells that accumulates and proliferates in the bone marrow. MM patients often develop bone disease that results in severe bone pain, osteolytic lesions, and pathologic fractures. These skeletal complications have not only a negative impact on quality of life but also a possible effect in overall survival. MM osteolytic bone lesions arise from the altered bone remodeling due to both increased osteoclast activation and decreased osteoblast differentiation. A dysregulated production of numerous cytokines that can contribute to the uncoupling of bone cell activity is well documented in the bone marrow microenvironment of MM patients. These molecules are produced not only by malignant plasma cells, that directly contribute to MM bone disease, but also by bone, immune, and stromal cells interacting with each other in the bone microenvironment. This review focuses on the current knowledge of MM bone disease biology, with particular regard on the role of bone and immune cells in producing cytokines critical for malignant plasma cell proliferation as well as in osteolysis development. Therefore, the understanding of MM pathogenesis could be useful to the discovery of novel agents that will be able to both restore bone remodelling and reduce tumor burden.

IFIH1 (MDA5) is required for innate immune detection of intron-containing RNA expressed from the HIV-1 provirus.

Antiretroviral therapy (ART) suppresses HIV-1 viremia and prevents progression to AIDS. Nonetheless, chronic inflammation is a common problem for people living with HIV-1 on ART. One possible cause of inflammation is ongoing transcription from HIV-1 proviruses, whether or not the sequences are competent for replication. Previous work has shown that intron-containing RNA expressed from the HIV-1 provirus in primary human blood cells, including CD4+ T cells, macrophages, and dendritic cells, activates type 1 interferon. This activation required HIV-1 rev and was blocked by the XPO1 (CRM1)-inhibitor leptomycin. To identify the innate immune receptor required for detection of intron-containing RNA expressed from the HIV-1 provirus, a loss-of-function screen was performed with shRNA-expressing lentivectors targeting twenty-one candidate genes in human monocyte derived dendritic cells. Among the candidate genes tested, only knockdown of XPO1 (CRM1), IFIH1 (MDA5), or MAVS prevented activation of the IFN-stimulated gene ISG15. The importance of IFIH1 protein was demonstrated by rescue of the knockdown with non-targetable IFIH1 coding sequence. Inhibition of HIV-1-induced ISG15 by the IFIH1-specific Nipah virus V protein, and by IFIH1-transdominant inhibitory CARD-deletion or phosphomimetic point mutations, indicates that IFIH1 filament formation, dephosphorylation, and association with MAVS, are all required for innate immune activation in response to HIV-1 transduction. Since both IFIH1 and DDX58 (RIG-I) signal via MAVS, the specificity of HIV-1 RNA detection by IFIH1 was demonstrated by the fact that DDX58 knockdown had no effect on activation. RNA-Seq showed that IFIH1-knockdown in dendritic cells globally disrupted the induction of IFN-stimulated genes. Finally, specific enrichment of unspliced HIV-1 RNA by IFIH1 was revealed by formaldehyde crosslinking immunoprecipitation (f-CLIP). These results demonstrate that IFIH1 is required for innate immune activation by intron-containing RNA from the HIV-1 provirus, and potentially contributes to chronic inflammation in people living with HIV-1.

The Therapeutic Potential of Novel Carnosine Formulations: Perspectives for Drug Development.

Carnosine (beta-alanyl-L-histidine) is an endogenous dipeptide synthesized via the activity of the ATP-dependent enzyme carnosine synthetase 1 and can be found at a very high concentration in tissues with a high metabolic rate, including muscles (up to 20 mM) and brain (up to 5 mM). Because of its well-demonstrated multimodal pharmacodynamic profile, which includes anti-aggregant, antioxidant, and anti-inflammatory activities, as well as its ability to modulate the energy metabolism status in immune cells, this dipeptide has been investigated in numerous experimental models of diseases, including Alzheimer's disease, and at a clinical level. The main limit for the therapeutic use of carnosine is related to its rapid hydrolysis exerted by carnosinases, especially at the plasma level, reason why the development of new strategies, including the chemical modification of carnosine or its vehiculation into innovative drug delivery systems (DDS), aiming at increasing its bioavailability and/or at facilitating the site-specific transport to different tissues, is of utmost importance. In the present review, after a description of carnosine structure, biological activities, administration routes, and metabolism, we focused on different DDS, including vesicular systems and metallic nanoparticles, as well as on possible chemical derivatization strategies related to carnosine. In particular, a basic description of the DDS employed or the derivatization/conjugation applied to obtain carnosine formulations, followed by the possible mechanism of action, is given. To the best of our knowledge, this is the first review that includes all the new formulations of carnosine (DDS and derivatives), allowing a decrease or complete prevention of the hydrolysis of this dipeptide exerted by carnosinases, the simultaneous blood-brain barrier crossing, the maintenance or enhancement of carnosine biological activity, and the site-specific transport to different tissues, which then offers perspectives for the development of new drugs.

Osteogenic differentiation of dental follicle stem cells.

BACKGROUND: Stem cells are defined as clonogenic cells capable of self-renewal and multi-lineage differentiation. A population of these cells has been identified in human Dental Follicle (DF). Dental Follicle Stem Cells (DFSCs) were found in pediatric unerupted wisdom teeth and have been shown to differentiate, under particular conditions, into various cell types of the mesenchymal tissues. AIM: The aim of this study was to investigate if cells isolated from DF show stem features, differentiate toward osteoblastic phenotype and express osteoblastic markers. METHODS: We studied the immunophenotype of DFSCs by flow cytometric analysis, the osteoblastic markers of differentiated DFSCs were assayed by histochemical methods and real-time PCR. RESULTS: We demonstrated that DFSCs expressed a heterogeneous assortment of makers associated with stemness. Moreover DFSCs differentiated into osteoblast-like cells, producing mineralized matrix nodules and expressed the typical osteoblastic markers, Alkaline Phosphatase (ALP) and Collagen I (Coll I). CONCLUSION: This study suggests that DFSCs may provide a cell source for tissue engineering of bone.

Nanoencapsulation Strategies for Active Compounds Delivery.

Nanoencapsulation strategies, including the possibility to deliver natural compounds, synthetic molecules, or other actives (viruses) for the treatment of different human diseases, represent a hot topic of great interest [...].

Drug Nanocrystals: Focus on Brain Delivery from Therapeutic to Diagnostic Applications.

The development of new drugs is often hindered by low solubility in water, a problem common to nearly 90% of natural and/or synthetic molecules in the discovery pipeline. Nanocrystalline drug technology involves the reduction in the bulk particle size down to the nanosize range, thus modifying its physico-chemical properties with beneficial effects on drug bioavailability. Nanocrystals (NCs) are carrier-free drug particles surrounded by a stabilizer and suspended in an aqueous medium. Due to high drug loading, NCs maintain a potent therapeutic concentration to produce desirable pharmacological action, particularly useful in the treatment of central nervous system (CNS) diseases. In addition to the therapeutic purpose, NC technology can be applied for diagnostic scope. This review aims to provide an overview of NC application by different administration routes, especially focusing on brain targeting, and with a particular attention to therapeutic and diagnostic fields. NC therapeutic applications are analyzed for the most common CNS pathologies (i.e., Parkinson's disease, psychosis, Alzheimer's disease, etc.). Recently, a growing interest has emerged from the use of colloidal fluorescent NCs for brain diagnostics. Therefore, the use of NCs in the imaging of brain vessels and tumor cells is also discussed. Finally, the clinical effectiveness of NCs is leading to an increasing number of FDA-approved products, among which the NCs approved for neurological disorders have increased.

Soluplus® polymeric nanomicelles improve solubility of BCS-class II drugs.

The issue of poor aqueous solubility is often a great hitch in the development of liquid dosage forms for those drugs that the Biopharmaceutics Classification System (BCS) includes in classes II and IV. Among the possible technological solutions, inclusion of the drug molecule within polymeric micelles, and particularly nanomicelles, has been proposed in the last years as a valid strategy. Our attention has been recently attracted by Soluplus®, an amphiphilic polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer able to form small and stable nanomicelles. The aim of this study was to characterize Soluplus® nanomicelles to enhance the apparent solubility of three model APIs, categorized in BCS class II: ibuprofen (IBU), idebenone (IDE), and miconazole (MIC). Drug-loaded Soluplus® micelles with a mean size around 60-70 nm were prepared by two methods (direct dissolution or film hydration method). The prepared nanosystems were characterized in terms of mean particle size and Zeta potential, physical stability, drug solubility, and in vitro drug release. The solubility of the tested APIs was shown to increase linearly with the concentration of graft copolymer. Soluplus® can be easily submitted to membrane filtration (0.2 µm PES or PTFE membranes), showing the potential to be sterilized by this method. Freeze-drying enabled to obtain powder materials that, upon reconstitution with water, maintained the initial micelle size. Finally, viscosity studies indicated that these nanomicelles have potential applications where a bioadhesive material is advantageous, such as in topical ocular administration.