Preparation, Characterization, and Biological Evaluation of a Hydrophilic Peptide Loaded on PEG-PLGA Nanoparticles

The encapsulation of peptides and proteins in nanosystems has been extensively investigated for masking unfavorable biopharmaceutical properties, including short half-life and poor permeation through biological membranes. Therefore, the aim of this work was to encapsulate a small antimicrobial hydrophilic peptide (H-Ser-Pro-Trp-Thr-NH2, FS10) in PEG-PLGA (polyethylene glycol-poly lactic acid-co-glycolic acid) nanoparticles (Nps) and thereby overcome the common limitations of hydrophilic drugs, which because they facilitate water absorption suffer from rapid degradation. FS10 is structurally related to the well-known RNAIII inhibiting peptide (RIP) and inhibits S. aureus biofilm formation. Various parameters, including different method (double emulsion and nanoprecipitation), pH of the aqueous phase and polymeric composition, were investigated to load FS10 into PEG-PLGA nanoparticles. The combination of different strategies resulted in an encapsulation efficiency of around 25% for both the double emulsion and the nanoprecipitation method. It was found that the most influential parameters were the pH—which tailors the peptides charge—and the polymeric composition. FS10-PEG-PLGA nanoparticles, obtained under optimized parameters, showed size lower than 180 nm with zeta potential values ranging from −11 to −21 mV. In vitro release studies showed that the Nps had an initial burst release of 48–63%, followed by a continuous drug release up to 21 h, probably caused by the porous character of the Nps. Furthermore, transmission electron microscopy (TEM) analysis revealed particles with a spherical morphology and size of around 100 nm. Antimicrobial assay showed that the minimum inhibitory concentration (MIC) of the FS10-loaded Nps, against S. aureus strains, was lower (>128 µg/mL) than that of the free FS10 (>256 µg/mL). The main goal of this work was to develop polymeric drug delivery systems aiming at protecting the peptide from a fast degradation, thus improving its accumulation in the target site and increasing the drug-bacterial membrane interactions

Wound-Healing Promotion and Anti-Inflammatory Properties of Carvacrol Prodrugs/Hyaluronic Acid Formulations

Background. Wound healing (WH) is a complex process involving several stages, such as hemostasis, inflammation, re-epithelialization, and remodeling. Many factors can impair WH, and different pharmacological approaches were studied to date, but the increase in antibiotic resistance, invasiveness, treatment duration, and high cost, have often hampered the resolution of the wound. In this study, we investigated the possible application of water-soluble carvacrol prodrugs (WSCPs) and hyaluronic acid (HA) and their formulations (WSCPs/HA) to improve WH and regulate the inflammatory response. Materials and methods. Firstly, the cytotoxicity of 0.1, 1 and 10 µg/mL of HA, WSCPs and WSCPs/HA formulations were evaluated on HaCaT cells and THP-1 cell lines. The ability of WSCPs/HA formulations to modulate wound repair was evaluated in an in vitro model of WH, using HaCaT cells at 6, 18, and 24 h. The expression of WH mediators, after wound closure was determined by qRT-PCR. Following, we polarized THP-1 cells in M1/M2-like macrophages and tested the anti-inflammatory properties of WSCPs/HA formulations. After, we tested the in vitro WH model for the effects of conditioned medium (CM) from M1/M2-like cells cultured in the presence of WSCPs/HA. Results. Results showed that WSCPs/HA formulations were able to significantly raise the wound closure rate, compared to the single constituents, promoting a complete wound closure after 18 h for WSCP1/HA (10 µg/mL) and after 24 h for WSCP2/HA (10 µg/mL), modulating the MMPs, TGFβ, and COX-2 gene expression. The effects of CM derived from M1/M2 polarized cells cultured in the presence of WSCPs/HA determined WH regulation, with a better ability of the WSCP2/HA formulation to modulate the time-dependent expression of reparative and inflammatory mediators. Conclusion. Our data underline the possible application of WSCPs/HA formulations as bioactive agents for the regulation of the wound repair process by the modulation of inflammatory and remodeling phases, affecting the activity of immune cells

Healthcare Systems across Europe and the US: The Managed Entry Agreements Experience

This systematic study aims at analyzing the differences between the approach of the European healthcare systems to the pharmaceutical market and the American one. This paper highlights the opportunities and the limitations given by the application of managed entry agreements (MEAs) in European countries as opposed to the American market, which does not regulate pharmaceutical prices. Data were collected from the Organisation for Economic Co-operation and Development (OECD), the European Medicines Agency, and the national healthcare agencies of US and European countries. A literature review was undertaken in PubMed, Scopus, MEDLINE, and Google for a period ten years (2010–2019). The period 2020–2021 was considered to compare health expenditure before and after the SARS-CoV-2 pandemic. Scarce information from national agencies has been given in terms of MEAs related to the COVID-19 pandemic. The comparison between the United States approach and the European one shows the importance of a market access regulation to reduce the cost of therapies, increasing the efficiency of national healthcare systems and the advantages in terms of quality and accessibility to the final users: patients. Nevertheless, it seems that the golden age of MEAs for Europe was during the examined period. Except for Italy, countries will move to other forms of reimbursements to obtain higher benefits, reducing the costs of an inefficient implementation and outcomes in the medium term

Healthcare Systems across Europe and the US: The Managed Entry Agreements Experience

This systematic study aims at analyzing the differences between the approach of the European healthcare systems to the pharmaceutical market and the American one. This paper highlights the opportunities and the limitations given by the application of managed entry agreements (MEAs) in European countries as opposed to the American market, which does not regulate pharmaceutical prices. Data were collected from the Organisation for Economic Co-operation and Development (OECD), the European Medicines Agency, and the national healthcare agencies of US and European countries. A literature review was undertaken in PubMed, Scopus, MEDLINE, and Google for a period ten years (2010–2019). The period 2020–2021 was considered to compare health expenditure before and after the SARS-CoV-2 pandemic. Scarce information from national agencies has been given in terms of MEAs related to the COVID-19 pandemic. The comparison between the United States approach and the European one shows the importance of a market access regulation to reduce the cost of therapies, increasing the efficiency of national healthcare systems and the advantages in terms of quality and accessibility to the final users: patients. Nevertheless, it seems that the golden age of MEAs for Europe was during the examined period. Except for Italy, countries will move to other forms of reimbursements to obtain higher benefits, reducing the costs of an inefficient implementation and outcomes in the medium term

In Vitro Wound-Healing Properties of Water-Soluble Terpenoids Loaded on Halloysite Clay

This research was funded by the Italian Ministry of Education, University and Research (University "G. d'Annunzio" of Chieti-Pescara), grant number FAR 2020.Recently, mineral healing clays have gained much attention for wound-dressing applications. Here, we selected halloysite (HAL) clay as a biocompatible, non-toxic material that is useful as a drug delivery system to enhance the healing properties of water-soluble terpenoids 1-3 (T1-3). Terpenoids-loaded HAL clay (TH1-3) was prepared and characterized by adsorption equilibrium studies, X-ray powder diffraction (XRPD), thermogravimetric analysis (TGA), differential scanning calorimetry (DSC), Fourier-transform infrared (FTIR) spectroscopy, and release studies. The results reveal that T1-3 were adsorbed at the HAL surface with good efficiency. The prevalent mechanism of drug retention is due to the adsorption via electrostatic interactions between the cationic groups of the T1-3 and the HAL’s external surface. Release studies demonstrated that T3 was released in a higher percentage (>60%) compared to T1-2 ( 50%). Additionally, TH1-3 were assessed for their antimicrobial activity and capability to promote the re-epithelialization of scratched HaCat monolayers, through the time-kill test and the wound-healing assays, respectively. The results reveal that all the tested formulations were able to reduce the microbial growth after 1 h of incubation and that they ensured complete wound closure after 48 h. Furthermore, at the concentration of 1 g/mL, TH3 exhibited 45% wound closure at 24 h, compared to TH1 (27%) and TH2 (30%), proving to be the best candidate in making the tissue-repair process easier and faster.Italian Ministry of Education, University and Research (University "G. d'Annunzio" of Chieti-Pescara) FAR 202

A novel point mutation within the juxtamembrane domain of the gene in acute myeloid leukemia

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Gene expression profile of protein kinases reveals a distinctive signature in chronic lymphocytic leukemia and in vitro experiments support a role of second generation protein kinase inhibitors

To investigate the role of protein kinases (PKs) in chronic lymphocytic leukemia (CLL), we performed gene expression profile on 505 PK genes. Comparison between CLL with acute lymphocytic leukemia (ALL) patients highlighted an homogeneous up-modulation of several PKs in CLL, 16 also overexpressed in two additional CLL cohorts. Q-PCR analysis confirmed these findings. No differences were observed in the main prognostic subclasses, indicating that PK overexpression is specific of the disease itself. Tests in vitro showed that Dasatinib partially reduced CLL cells viability, mostly in IGHV germline patients. These findings suggest that treatment with second generation tyrosine kinase (TK) inhibitors may represent an attractive therapeutic strategy for CLL patients. © 2009 Elsevier Ltd

Minimal residual disease monitoring in chronic lymphocytic leukaemia patients. A comparative analysis of flow cytometry and ASO IgH RQ-PCR.

Minimal residual disease (MRD) is becoming increasingly important in chronic lymphocytic leukaemia (CLL) as treatment strategies are progressively improving. The primary objective of this study was to compare the applicability of three different flow cytometric approaches: basic 4-colour analysis, European Research Initiative in CLL (ERIC) consensus method and 8-colour analysis. Secondly, we investigated the sensitivity and specificity of flow cytometry (FC) compared to molecular analyses for MRD detection. A total of 462 CLL samples were evaluated by basic FC; in 143, ERIC consensus method was also performed and all three FC methodologies were applied in a subgroup of 10 cases. No discordance in defining MRD-positive/negative samples was observed between the FC methods; within positive samples, the ERIC consensus method and 8-colour analysis showed the most accurate results. MRD was analysed by FC and polymerase chain reaction (PCR) in 243 cases: concordant results were obtained in 199/243 samples (81·9%); 42/243 were FC-/PCR+. Overall, the sensitivity and specificity of FC compared to PCR was 96·5% and 77·2%, respectively. Both FC and PCR proved suitable for the detection of MRD and prediction of progression-free survival, which was significantly reduced in MRD-positive patients, regardless of the methodology. These results offer the rationale for a strategy to monitor MRD in CLL patients