Dietary phytochemicals are particularly attractive for chemoprevention and are able to modulate several signal transduction pathways linked with cancer. Olive oil, a major component of the Mediterranean diet, is an abundant source of phenolic compounds. Olive oil production is associated with the generation of a waste material, termed 'olive mill wastewater' (OMWW) that have been reported to contain water-soluble polyphenols. Prostate cancer (PCa) is considered as an ideal cancer type for chemopreventive approaches, due to its wide incidence but relatively long latency period and progression time. Here, we investigated activities associated with potential preventive properties of a polyphenol-rich olive mill wastewater extract, OMWW (A009), on three in vitro models of PCa. A009 was able to inhibit PCa cell proliferation, adhesion, migration, and invasion. Molecularly, we found that A009 targeted NF-κB and reduced pro-angiogenic growth factor, VEGF, CXCL8, and CXCL12 production. IL-6/STAT3 axis was also regulated by the extract. A009 shows promising properties, and purified hydroxytyrosol (HyT), the major polyphenol component of A009, was also active but not always as effective as A009. Finally, our results support the idea of repositioning a food waste-derived material for nutraceutical employment, with environmental and industrial cost management benefits.

Epidemiological studies showed that diet can play a relevant role in reducing the risk of developing colon cancer (CC) and lower rate of CC insurgence has been observed amongst populations living within the Mediterranean basin. Olive oil, a major component of the Mediterranean diet, is an abundant source of phenolic compounds. Olive oil production is associated with the generation of waste material, termed ‘olive mill wastewaters’ (OMWW), that have been reported to be enriched in polyphenols as well. Given the beneficial activity of polyphenols on human health, we investigated whether the use of different batches of purified extracts from OMWW, termed A009, might be effective in exerting chemopreventive activities in vitro and in vivo, on CC cell lines. Cell proliferation and survival were evaluated on A009 treated cells by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, while the induction of apoptosis was assessed by flow cytometry. Further, functional studies to investigate the ability of A009 to interfere with CC cell line adhesion, migration, sprouting and invasion were performed. Finally, the ability of A009 to interfere with CT-26 CRC tumour cell growth was assessed in vivo. Purified hydroxytyrosol, the major component in the A009 extracts, was used as a control. A009 inhibited cell proliferation, migration, invasion, adhesion and sprouting of CC cells along with the release of pro-angiogenic and pro- inflammatory cytokines (VEGF, IL-8) similar to hydroxytyrosol alone. In vivo, A009 inhibited CT-26 tumour growth in a significant manner over that of hydroxytyrosol alone. Our results show that A009 extracts exert promising chemopreventive properties, suggesting that different polyphenols act synergistically, improving their single component effects in CC cell lines. Finally, our results support the idea of repositioning a waste derived material for nutraceutical employment, with environmental and industrial cost management benefits. Citation Format: Barbara Bassani, Teresa Rossi, Daniela De Stefano, Daniele Pizzichini, Paola Corradino, Antonino Bruno, Douglas M. Noonan, Adriana Albini. Chemopreventive activities of a polyphenol rich purified extract from olive oil processing on colon cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5272. doi:10.1158/1538-7445.AM2017-5272

Nuclear Factor-κB (NF-κB) is a transcription factor regulating several genes involved in important physiological and pathological processes. NF-κB has been found constitutively activated in many inflammatory/immune diseases. In addition, a positive correlation between persistent activation of NF-κB and tumor promotion has been demonstrated. Since the IKK (IκB kinase) activation is an indispensable component of all pro-inflammatory signaling pathways leading to NF-κB activation, considerable efforts have been done in order to develop novel anti-inflammatory therapeutics targeting IKK. Association of the IKK complex relies on critical interactions between the C-terminus NBD (NEMO binding domain) of the catalytic subunits IKKα and IKKβ, and the regulatory subunit NEMO (NF-κB Essential Modulator). Thus, this IKK/NEMO interacting region provides an attractive target to prevent the IKK complex formation and NF-κB activation. In this regard, we have identified non-peptide small molecule disruptors of IKKβ/NEMO complex through a structure-based virtual screening (SBVS) of the NCI chemical library. Phenothiazine 22 and its close analogues (22.2, 22.4 and 22.10) were able to reduce nitrite production and iNOS mRNA expression in J774 murine macrophages stimulated with LPS for 24h. These effects were associated with a reduced NF-κB/DNA binding activity as well as a decreased expression of phosphorylated IKKβ, IκBα and NF-κB/p65 in these cells. These observations suggest that compound 22 and its three structural analogues by inhibiting IKKβ/NEMO association mediate the blockage of NF-κB signaling pathway and may prove effective in treatment of diseases in which the IKK/NF-κB pathway is dysregulated.

Traumatic brain injury (TBI) initiates a neuroinflammatory cascade that contributes to neuronal damage and behavioral impairment. In the present study, we performed a widely used model of TBI to determine the neuroprotective propriety of palmitoylethanolamide (PEA) and the antioxidant effect of a flavonoid luteolin (Lut), given as a co-ultramicronized compound Co-ultraPEALut. We demonstrated that the treatment with Co-ultraPEALut resulted in a significant improvement of motor and cognitive recovery after controlled cortical impact, as well as markedly reducing lesion volumes. Moreover, our results revealed the ability of Co-ultraPEALut to reduce brain trauma through modulation of nuclear factor-κB activation. In addition, treatment with Co-ultraPEALut significantly enhanced the post-TBI expression of the neuroprotective neurotrophins glial cell line-derived neurotrophic factor compared with vehicle. Co-ultraPEALut at the dose of 1 mg/kg also modulated apoptosis, the release of cytokine and reactive oxygen species, the activation of chymase, tryptase, and nitrotyrosine, and inhibited autophagy. Thus, our data demonstrated that Co-ultraPEALut at a lower dose compared with PEA alone can exert neuroprotective effects and the combination of both could improve their ability to counteract the neurodegeneration and neuroinflammation induced by TBI.

Olive oil, a major feature of the Mediterranean diet, is an abundant source of phenolic compounds. Olive oil production is associated with the generation of waste material, termed ‘olive mill wastewater’ (OMWW), that has been reported to be enriched in soluble polyphenols. Given the known beneficial activity of polyphenols, we investigated whether the use of purified extracts from OMWW, termed A009, rich in hydroxytyrosol, might have anticancer activities on colon cancer (CC) cell lines in vitro and in vivo and could represent a chemopreventive preparation for CC. A009 from different batches inhibited proliferation, migration, invasion, adhesion, sprouting of CC cells and release of angiogenic, pro-inflammatory cytokines (VEGF, IL-8). Our data demonstrate that a novel purified, polyphenol enriched extract, obtained from food industry waste material, with similar activity than purified hydroxytyrosol but easier to produce in large quantities and with an environment-sensitive approach, has potential cancer chemopreventive properties for colon cancer cells.

The increasing use of carbon nanotubes (CNTs) in several industrial applications raises concerns on their potential toxicity due to factors such as tissue penetrance, small dimensions, and biopersistence. Using an in vivo model for CNT environmental exposure, mimicking CNT exposition at the workplace, we previously found that CNTs rapidly enter and disseminate in the organism, initially accumulating in the lungs and brain and later reaching the liver and kidneys via the bloodstream in CD1 mice. Here, we monitored and traced the accumulation of single-walled CNTs (SWCNTs), administered systemically in mice, in different organs and the subsequent biological responses. Using the novel in vivo model, MITO-Luc bioluminescence reporter mice, we found that SWCNTs induce systemic cell proliferation, indicating a dynamic response of cells of both bone marrow and the immune system. We then examined metabolic (water/food consumption and dejections), functional (serum enzymes), and morphological (organs and tissues) alterations in CD1 mice treated with SWCNTs, using metabolic cages, performing serum analyses, and applying histological, immunohistochemical, and ultrastructural (transmission electron microscopy) methods. We observed a transient accumulation of SWCNTs in the lungs, spleen, and kidneys of CD1 mice exposed to SWCNTs. A dose- and time-dependent accumulation was found in the liver, associated with increases in levels of aspartate aminotransferase, alanine aminotransferase and bilirubinemia, which are metabolic markers associated with liver damage. Our data suggest that hepatic accumulation of SWCNTs associated with liver damage results in an M1 macrophage-driven inflammation.

Therapeutic strategies aimed at correcting the defect of the cystic fibrosis transmembrane conductance regulator (CFTR) (“CFTR-repair”) constitute a new avenue towards the treatment of patients affected by Cystic Fibrosis (CF), the most common lethal monogenic disease in Caucasians. CF is caused by mutations in the CFTR gene, which codes for a 1480 amino-acid protein normally functioning as a chloride channel at apical membrane from epithelial cells (1). In the CFTR gene, more than 1900 mutations, most of which are disease-relevant, have been identified and then categorized in six different classes according to their functional impact (2). The phenotypic consequences of such genotypes comprise insufficiency of the exocrine pancreas, increased electrolytes in sweat, male infertility and—most prevalent—a debilitating and eventually lethal inflammation/infection-triggered respiratory dysfunction. Mutation-specific approaches are focusing on the identification of small molecules capable of correcting the deficient subcellular trafficking of the CFTR protein (“correctors”: agents that ensure the expression of the mutated protein at the apical plasma membrane) or defective gating (“potentiators”: agents that reinstate the channel function of mutated CFTR proteins that are orthotopically expressed).

Cystic fibrosis (CF) is a lethal monogenic disease with mortality and morbidity mostly associated to lung destruction (O'Sullivan and Freedman, 2009O'Sullivan B.P. Freedman S.D. Cystic fibrosis.Lancet. 2009; 373: 1891-1904Summary Full Text Full Text PDF PubMed Scopus (1017) Google Scholar). This, in turn, is strictly dependent on recurrent pulmonary infections, with bacteria growing also in biofilms, leading to an unsatisfactory effect of antibiotics (Ratjen and Döring, 2003Ratjen F. Döring G. Cystic fibrosis.Lancet. 2003; 361: 681-689Summary Full Text Full Text PDF PubMed Scopus (891) Google Scholar). The CF registry of patients reports an improvement of both patient survival and quality of life, due to a global care of several aspects of the disease. Yet, it is an unmet need, hence, the urgency of new antibiotics. Recently, Vertex Ph. has introduced in therapy a new potentiator (a channel gating activator), namely ivacaftor (Kalydeco), whose effects, are beneficial, to date, for patients exhibiting the G551D mutation (Ramsey et al., 2011Ramsey B.W. Davies J. McElvaney N.G. et al.A CFTR potentiator in patients with cystic fibrosis and the G551D mutationVX08-770-102 study group.N. Engl. J. Med. 2011; 365: 1663-1672Crossref PubMed Scopus (1609) Google Scholar). Furthermore, the same company has introduced a combination of lumacaftor (a corrector of defective CFTR folding/cellular processing, Orkambi) and ivacaftor (Kalydeco) for patients exhibiting F508del-CFTR, the most common mutation (Wainwright et al., 2015Wainwright C.E. Elborn J.S. Ramsey B.W. et al.Lumacaftor–ivacaftor in patients with cystic fibrosis homozygous for Phe508del CFTR.N. Engl. J. Med. 2015; 373: 220-231Crossref PubMed Scopus (963) Google Scholar). The effects of this approach have been discussed in several papers (see an extensive comment in (Maiuri et al., 2015Maiuri L. De Stefano D. Raia V. et al.The holy grail of cystic fibrosis research: pharmacological repair of the F508del-CFTR mutation.Ann. Transl. Med. 2015; 3: S24PubMed Google Scholar). Interestingly, a recent paper reported the effects of Cysteamine in combination with epigallocatechin gallate, in restoring CFTR function and expression at plasma membrane in nasal brushing from CF patients, via rescue of defective autophagy (De Stefano et al., 2014De Stefano D. Villella V.R. Esposito S. et al.Restoration of CFTR function in patients with cystic fibrosis carrying the F508del-CFTR mutation.Autophagy. 2014; 10: 2053-2074Crossref PubMed Scopus (120) Google Scholar). Noteworthy, cysteamine has received an Orphan Drug Designation in CF also for its mucolytic properties. Such an effect is extremely attractive, being thick mucus an important issue for CF patients. Charrier et al. described the antimicrobial, antibiofilm and mucoactive properties of cysteamine, either alone or in combination with recommended antibiotics, a novel mucoactive antimicrobial and antibiofilm agent for the treatment of CF (Charrier et al., 2014Charrier C. Rodger C. Robertson J. et al.Cysteamine (Lynovex®), a novel mucoactive antimicrobial & antibiofilm agent for the treatment of cystic fibrosis.Orphanet J. Rare Dis. 2014; 9: 189https://doi.org/10.1186/s13023-014-0189-2Crossref PubMed Scopus (52) Google Scholar). The authors subsequently reported in the study recently published and herein discussed, that patients were invited to provide their sputum, the most being infected with P. aeruginosa (Devereux et al., 2015Devereux G. Fraser-Pitt D. Robertson J. et al.Cysteamine as a future intervention in cystic fibrosis against current and emerging pathogens: a patient-based ex vivo study confirming its antimicrobial and mucoactive potential in sputum.EBioMedicine. 2015; 2: 1507-1512Summary Full Text Full Text PDF PubMed Scopus (27) Google Scholar). The results of the study prove that daily incubation of sputum with cysteamine significantly decreased the sputum bacterial load and decreased mucus viscosity. The major limitation regarding this study is that no patient has been directly administered. Although cysteamine exhibits also a mucolytic activity that might help in reaching the infected pulmonary site, such an issue must be addressed. Moreover, no characterization of the bacterial type has been provided, meaning that it is also difficult to establish a proper mechanism of action. Finally, the patient mutation type should be taken into account. The study, however, has a great appeal. In fact, cysteamine is an already FDA approved drug for another rare disease, cystinosis, and, thus, in therapy in children. Moreover, side effects have been extensively documented and cysteamine is a safe drug for cystinosis patients although it is well known that pharmacokinetic profile and, thus, tolerability, might be different in CF patients. Moreover, cysteamine is endogenously present, although at very low levels, as a consequence of coenzyme A metabolism (Besouw et al., 2013Besouw M. Masereeuw R. van den Heuvel L. et al.Cysteamine: an old drug with new Potential.Drug Discov. Today. 2013; 18: 785-792Crossref PubMed Scopus (131) Google Scholar). The herein cited study, per se, offers the opportunity to address a new mechanism of action for cysteamine and to have a novel drug exhibiting multiple mechanisms of action, being cysteamine an antibiotic, mucolytic and capable of functionally rescuing misfolded CFTR at plasma membrane. These results have been developed from independent research groups and allow to prospectively be positive on the opportunity that cysteamine will be also authorized for CF therapy. In conclusion, there is a strong hope that future studies, carried on with CF patients, will clarify the antibiotic effects of cysteamine, possibly identifying also a dose regimen and, if the case, the bacterial strains that can be treated. Cysteamine, therefore, represents an old drug with several new targets for CF therapy. The authors declare no conflicts of interest. DDS and MCM are supported by E-Rare-2 and Fondazione Italiana per la Fibrosi cistica (FFC 4#2015). Cysteamine as a Future Intervention in Cystic Fibrosis Against Current and Emerging Pathogens: A Patient-based ex vivo Study Confirming its Antimicrobial and Mucoactive Potential in SputumCysteamine has recently been shown to have in vitro properties potentially therapeutically beneficial in cystic fibrosis (CF). In this study we investigated the antimicrobial and mucolytic activity of cysteamine against the complex biologic matrix of CF sputum. Full-Text PDF Open Access

Chronic inflammation, a condition frequently associated with several pathologies, is characterized by angiogenic and fibrogenic responses that may account for the development of granulomatous tissue. We previously demonstrated that the chymase, rat mast cell protease-5 (rMCP-5), exhibits pro-inflammatory and pro-angiogenic properties in a model of chronic inflammation sustained by mast cells (MCs), granuloma induced by the subcutaneous carrageenan-soaked sponge implant in rat. In this study, we investigated the effects of palmitoylethanolamide (PEA), an anti-inflammatory and analgesic endogenous compound, on rMCP-5 mRNA expression and Microphtalmia-associated Transcription Factor (MITF) activation in the same model of chronic inflammation. The levels of rMCP-5 mRNA were detected using semi-quantitative RT-PCR; the protein expression of chymase and extracellular signal-regulated kinases (ERK) were analyzed by western blot; MITF/DNA binding activity and MITF phosphorylation were assessed by electrophoretic mobility shift assay (EMSA) and immunoprecipitation, respectively. The administration of PEA (200, 400 and 800 µg/ml) significantly decreased rMCP-5 mRNA and chymase protein expression induced by λ-carrageenan. These effects were associated with a significant decrease of MITF/DNA binding activity and phosphorylated MITF as well as phosphorylated ERK levels. In conclusion, our results, showing the ability of PEA to inhibit MITF activation and chymase expression in granulomatous tissue, may yield new insights into the understanding of the signaling pathways leading to MITF activation controlled by PEA.

Aim of this work was to investigate the influence of hyaluronic acid (HA) molecular weight on the thermogelation and biocompatibility of its blends with methyl cellulose in view of a possible application in drug delivery and/or wound healing. We found out that it was possible to obtain MC/HA blends showing a rheological behavior typical of a viscous solution at 20 °C and of a weak gel at 37 °C only when blending MC with low molecular weight HA. Moreover, the blends containing low molecular weight HA did not affect human foreskin fetal fibroblasts viability, proliferation and migration. On the contrary, the cell incubation with high molecular weight HA resulted in a marked and significant reduction of cell viability, compared to control cells. Finally, the optimized blends, in terms of rheological properties and biocompatibility, proved to be able to control and prolong bovine serum albumin release by a combined mechanism of platform dissolution and drug diffusion.

Trans-resveratrol, a polyphenol extracted from Vitis vinifera, has different beneficial effects following its administration on the skin. Here the potential use of binary systems to enhance in vitro and in vivo activity of trans-resveratrol was investigated. Thus the aqueous solubility of trans-resveratrol was investigated in the presence of growing concentrations of polyethylene glycol (PEG) or β-cyclodextrin (βCD) as solubilizing excipients. Then, the solid dispersion of trans-resveratrol with PEG or inclusion complexes trans-resveratrol/βCD were prepared and characterised by different methods. Cytotoxicity and inhibition of reactive oxygen species (ROS) following H2O2 challenge in the presence of trans-resveratrol, alone or associated to the excipients, was evaluated on human keratinocyte HaCaT cell line. Both the trans-resveratrol-containing binary systems induced significant reduction of H2O2-induced ROS production, especially in the case of βCD that was selected for the following phase of the study. Thus, the effect of a cream containing trans-resveratrol, alone or associated to βCD, on different skin parameters such as corneometry, colorimetry and elastometry, was evaluated on human volunteers. All patients showed a visible improvement of clinical conditions with a remarkable decrease of aging signs, but this effect was higher of the hemi face treated with the βCD-containing formulation versus formulation containing trans-resveratrol alone.

Apoptosis, a form of programmed cell death, is a critical defence mechanism against the formation and progression of cancer and acts by eliminating potentially deleterious cells without causing such adverse effects, as inflammatory response and ensuing scar formation. Therefore, targeting apoptotic pathways becomes an intriguing strategy for the development of chemotherapeutic agents. In last decades, marine natural products, such as sesterterpenoids, have played an important role in the discovery and development of new drugs. Interestingly, many of these compounds have a strong potential as anticancer drugs by inhibiting cell proliferation and/or inducing cell death. In the present study, we investigated the effects of scalaradial and cacospongionolide, two sesterterpenoids from Cacospongia scalaris and Fasciospongia cavernosa marine sponges, on the apoptotic signalling pathway in three different human tumoral cells. Results were obtained by using DNA fragmentation, comet and viability assays, quantification of the mitochondrial transmembrane potential and Western blot. The T47D (human breast carcinoma), A431 (human epidermoid carcinoma), HeLa (human cervix carcinoma) and HCT116 (human colon carcinoma) cells were incubated for 24 h with scalaradial or cacospongionolide. Treatment of T47D cells with scalaradial or cacospongionolide for 24 h brought about a significant increase in DNA migration as well as fragmentation. Moreover, incubation of HCT116 and HeLa cells with scalaradial or cacospongionolide for 24 h caused an increased expression of pro-apoptotic proteins. Furthermore, scalaradial or cacospongionolide, added to HCT116 and HeLa cells overnight, induced a significant and concentration-dependent loss of mitochondrial transmembrane potential, an early apoptosis signalling event. These effects paralleled with those achieved with p50 and p65, NF-κB subunits, nuclear level. In conclusion, scalaradial and cacospongionolide, by determining human cancer cell apoptosis, may represent new promising compounds to inhibit cancer cell proliferation.

The purpose of this special issue is to point the attention to the RNA functions, which in the last decades emerged as extremely important. Particularly, physiopathological roles of microRNA, short hairpin RNA, and small interfering RNA have been reported, in turn leading to the opportunity to consider RNAs as both novel therapeutic targets and tools. The contributions of this special issue cover a variety of subjects and reflect the complexity of the field. In particular, RNA is a difficult molecule, due to its rapid degradation. Several strategies have been described to overcome these issues, in order to render the use of RNA possible. M. Gaglione et al. reported chemical modifications of siRNAs containing terminal amide linkages by introducing hydroxyethylglycine PNA (hegPNA) moieties at 5′, at 3′ positions, and on both terminals and demonstrated that some of these modifications are compatible with the RNAi machinery, as they markedly increased the resistance to serum-derived nucleases. Indeed, Z. Deng et al. reported the role of PNPase as a major enzyme of sRNAs degradation. I. Scognamiglio et al. describe the development and effects of stable nucleic acid lipid vesicles (SNALPs) encapsulating miR-34a to treat multiple myeloma. In this study, the authors show that it is possible to overcome the targeted delivery of SNALPs as well as miRNA encapsulation efficiency drawbacks by conjugating SNALPs with the transferrin and chemically modifying the miRNA with a 2′-O-methylation. The formulation was effective in reducing tumor growth. The miRNAs are the subject of another contribution. A. Hsu et al. demonstrated that an alteration of miRNA levels may be used as diagnostic markers of myocardial infarction. The differentially expressed miRNAs were evaluated in a separate cohort of 62 subjects to show that serum miR-486-3p and miR-150-3p were upregulated while miR-126-3p, 14 miR-26a-5p, and miR-191-5p were significantly downregulated suggesting that serum miRNAs may be used as potential diagnostic biomarkers. F. Amato et al. reported the effectiveness of an inhibitor of miR-5093p for the regulation of cystic fibrosis-related gene expression. Primary miRNA (pri-miRNA) is the subject of M. B. Mowa and colleagues study. The authors investigated the utility of the murine transthyretin receptor (MTTR) promoter for expression of artificial anti-HBV primary miRNA (pri-miRNA) sequences to develop their use in liver-specific transcription regulatory elements. They demonstrated that the expression of anti-HBV pri-miR mimics from MTTR promoter is well suited to countering HBV replication and development of HD Ads through attenuation of their immunostimulatory effects. The present issue constitutes an important update in a constantly developing field. The efforts to carry on these studies will, possibly, generate new therapeutic opportunities in the near future. Maria Chiara Maiuri Daniela De Stefano Ammad Ahmad Farooqi

In this study, a sterile and biocompatible chitosan (CHI) gel for wound healing applications was formulated. CHI powder was treated in autoclave (ttCHI) to prepare sterile formulations. The heat treatment modified the CHI molecular weight, as evidenced by GPC analysis, and its physical–chemical features. Differential scanning calorimetry studies indicated that the macromolecules, before and after thermal treatment, differ in the strength of water-polymer interaction leading to different viscoelastic and flow properties. Thermally treated CHI exhibited the following effects: (i) increased the proliferation and migration of human foreskin foetal fibroblasts at 24 h; (ii) accelerated wound healing (measured as area of lesion) at 3 and 10 days in an in vivo model of pressure ulcers. These effects were linked to the increase of the hydroxyproline and haemoglobin content as well as Wnt protein expression. Moreover, we found a reduction of myeloperoxidase activity and TNF-α mRNA expression. These observations suggest the potential of this novel CHI gel in wound healing and other therapeutic applications.

Restoration of BECN1/Beclin 1-dependent autophagy and depletion of SQSTM1/p62 by genetic manipulation or autophagy-stimulatory proteostasis regulators, such as cystamine, have positive effects on mouse models of human cystic fibrosis (CF). These measures rescue the functional expression of the most frequent pathogenic CFTR mutant, F508del, at the respiratory epithelial surface and reduce lung inflammation in Cftr(F508del) homozygous mice. Cysteamine, the reduced form of cystamine, is an FDA-approved drug. Here, we report that oral treatment with cysteamine greatly reduces the mortality rate and improves the phenotype of newborn mice bearing the F508del-CFTR mutation. Cysteamine was also able to increase the plasma membrane expression of the F508del-CFTR protein in nasal epithelial cells from F508del homozygous CF patients, and these effects persisted for 24h after cysteamine withdrawal. Importantly, this cysteamine effect after washout was further sustained by the sequential administration of epigallocatechin gallate (EGCG), a green tea flavonoid, both in vivo, in mice, and in vitro, in primary epithelial cells from CF patients. In a pilot clinical trial involving 10 F508del-CFTR homozygous CF patients, the combination of cysteamine and EGCG restored BECN1, reduced SQSTM1 levels and improved CFTR function from nasal epithelial cells in vivo, correlating with a decrease of chloride concentrations in sweat, as well as with a reduction of the abundance of TNF/TNF-alpha (tumor necrosis factor) and CXCL8 (chemokine [C-X-C motif] ligand 8) transcripts in nasal brushing and TNF and CXCL8 protein levels in the sputum. Altogether, these results suggest that optimal schedules of cysteamine plus EGCG might be used for the treatment of CF caused by the F508del-CFTR mutation.

The trans-resveratrol (t-res), a non-flavonoid polyphenol extracted from different plants, has recently earned interest for application on the skin for different applications. In this work, the potential of nanocarriers, namely transfersomes and ethanol-containing vesicles, to deliver t-res into/through the skin was investigated. Thus, transfersomes with different surfactants, namely polysorbate 80 (Tw80), sodium cholate (SC) and sodium deossicholate (SDC) and ethanol-containing vesicles with different lipid composition, namely soy phosphatidylcholine (SPC) and cholesterol (chol), encapsulating t-res were prepared and characterized. The nanocarriers had a mean diameter ranging between 83 and 116nm with a high t-res encapsulation efficiency (≥70%). Moreover, cytotoxicity as well as the inhibition of production of reactive oxygen species (ROS) and lipid peroxidation, following incubation of H2O2-stimulated human keratinocyte (HaCaT) with t-res, as free or encapsulated into the nanocarriers, were investigated. Only blank nanocarriers containing Tw80 or ethanol were cytotoxic and led to increase of ROS, but this effect was not observed when using nanocarriers encapsulating t-res. Finally, permeation studies on porcine skin carried out on Franz diffusion cells, showed that only ethanol-containing vesicles based SPC were able to promote t-res permeation through the skin.

The inflammatory process plays a crucial role in the onset and progression of several lung pathologies, including cystic fibrosis (CF), and the involvement of NF-κB is widely recognized. The specific inhibition of NF-κB by decoy oligonucleotides delivered within the lung may be beneficial, although rationally designed systems are needed to optimize their pharmacological response. Prompted by this need, we have developed and tested in vivo an inhalable dry powder for the prolonged delivery of a decoy oligodeoxynucleotide to NF-κB (dec-ODN), consisting of large porous particles (LPPs) based on poly(lactic-co-glycolic) acid. First, LPPs containing dec-ODN (dec-ODN LPPs) were engineered to meet the aerodynamic criteria crucial for pulmonary delivery, to gain an effective loading of dec-ODN, to sustain its release, and to preserve its structural integrity in lung lining fluids. We then investigated the effects of dec-ODN LPPs in a rat model of lung inflammation induced by the intratracheal aerosolization of LPS from Pseudomonas aeruginosa. The results show that a single intratracheal insufflation of dec-ODN LPPs reduced the bronchoalveolar neutrophil infiltration induced by LPS for up to 72 hours, whereas naked dec-ODN was able to inhibit it only at 6 hours. The persistent inhibition of neutrophil infiltrate was associated with reduced NF-κB/DNA binding activity, as well as reduced IL-6, IL-8, and mucin-2 mRNA expression in lung homogenates. We consider it noteworthy that the developed LPPs, preventing the accumulation of neutrophils and NF-κB-related gene expression, may provide a new therapeutic option for the local treatment of inflammation associated with lung disease.

A specific and promising approach to limit inflammation and mucin iperproduction in chronic lung diseases relies on specific inhibition of nuclear Factor-κB (NF-κB) by a decoy oligonucleotide (dec-ODN). To fulfill the requirements dictated by translation of dec-ODN therapy in humans, inhalable dry powders were designed on a rational basis to provide drug protection, sustained release and to optimize pharmacological response. To this end, large porous particles (LPP) for dec-ODN delivery made of a sustained release biomaterial (poly(lactic-co-glycolic) acid, PLGA) and an "adjuvant" hydrophilic polymer (polyethylenimine, PEI) were developed and their effects on LPS-stimulated human airway epithelial cells evaluated. The composite PLGA/PEI particles containing dec-ODN (i.e., LPPPEI) were successfully engineered for widespread deposition in the lung and prolonged release of intact dec-ODN in vitro. LPPPEI caused a prolonged inhibition of IL-8 and MUC2 expression in CF human bronchial epithelial cells and human epithelial pulmonary NCI-H292 cells, respectively, as compared to naked dec-ODN. Nonetheless, as compared to previously developed LPP, the presence of PEI was essential to construct a dec-ODN delivery system able to act in mucoepidermoid lung epithelial cells. In perspective, engineering LPP with PEI may become a key factor for tuning carrier properties, controlling lung inflammation and mucin production which, in turn, can foster in vivo translation of dec-ODN therapy.

In the last decade, the nanotechnology advancement has developed a plethora of novel and intriguing nanomaterial application in many sectors, including research and medicine. However, many risks have been highlighted in their use, particularly related to their unexpected toxicity in vitro and in vivo experimental models. This paper proposes an overview concerning the cell death modalities induced by the major nanomaterials.