Mesenchymal stromal cells loaded with Paclitaxel (PacliMES) a potential new therapeutic approach on mesothelioma.

Malignant pleural mesothelioma is an asbestos-related tumor originating in mesothelial cells of the pleura that poorly responds to chemotherapeutic approaches. Adult mesenchymal stromal cells derived either from bone marrow or from adipose tissue may be considered a good model for cell-based therapy, a treatment which has experienced significant interest in recent years. The present study confirms that Paclitaxel is effective on mesothelioma cell proliferation in 2D and 3D in vitro cultures, and that 80,000 mesenchymal stromal cells loaded with Paclitaxel inhibit tumor growth at a higher extent than Paclitaxel alone. An in vivo approach to treat in situ mesothelioma xenografts using a minimal amount of 106 mesenchymal stromal cells loaded with Paclitaxel showed the same efficacy of a systemic administration of 10 mg/kg of Paclitaxel. These data strongly support drug delivery system by mesenchymal stromal cells as a useful approach against many solid tumors. We look with interest at the favourable opinion recently expressed by the Italian Drug Agency on the procedure for the preparation of mesenchymal stromal cells loaded with Paclitaxel in large-scale bioreactor systems and their storage until clinical use. This new Advanced Medicinal Therapy Product, already approved for a Phase I clinical trial on mesothelioma patients, could pave the way for mesenchymal stromal cells use as drug delivery system on other solid tumors for adjuvant therapy associated with surgery and radiotherapy.

Photo-Inactivation of Staphylococcus aureus by Diaryl-Porphyrins.

Photodynamic Antimicrobial Chemotherapy (PACT) has received great attention in recent years since it is an effective and promising modality for the treatment of human oral and skin infections with the advantage of bypassing pathogens' resistance to antimicrobials. Moreover, PACT applications demonstrated a certain activity in the inhibition and eradication of biofilms, overcoming the well-known tolerance of sessile communities to antimicrobial agents. In this study, 13 diaryl-porphyrins (mono-, di-cationic, and non-ionic) P1-P13 were investigated for their potential as photosensitizer anti-Staphylococcus aureus. The efficacy of the diaryl-porphyrins was evaluated through photo-inactivation tests. Crystal-violet staining combined with viable count techniques were aimed at assaying their anti-biofilm activity. Among the tested compounds, the neutral photosensitizer P4 was better than the cationic ones, irrespective of their corresponding binding rates. In particular, P4 was active in inhibiting the biofilm formation and in impairing the viability of the adherent and planktonic populations of a 24 h old biofilm. The inhibitory activity was also efficient against a methicillin resistant S. aureus strain. In conclusion, the diaryl-porphyrin family represents a reservoir of promising compounds for photodynamic applications against the pathogen S. aureus and in preventing the formation of biofilms that cause many infections to become chronic.

Searching for antimicrobial photosensitizers among a panel of BODIPYs.

In recent years, antimicrobial Photodynamic Therapy (aPDT) gained increasing attention for its potential to inhibit the growth and spread of microorganisms, both as free-living cells and/or embedded in biofilm communities. In this scenario, compounds belonging to the family of boron-dipyrromethenes (BODIPYs) represent a very promising class of photosensitizers for applications in antimicrobial field. In this study, twelve non-ionic and three cationic BODIPYs were assayed for the inactivation of Staphylococcus aureus, Pseudomonas aeruginosa and Candida albicans. As expected, S. aureus showed to be very sensitive to BODIPYs and mild conditions were sufficient to reach good rates of photoinactivation with both neutral and monocationic ones. Surprisingly, one neutral compound (named B9 in this study) resulted the best BODIPY to photoinactivate P. aeruginosa PAO1. The photoinactivation of C. albicans was reached with both neutral and mono-cationic BODIPYs. Furthermore, biofilms of the three model microorganisms were challenged with BODIPYs in light-based antimicrobial technique. S. aureus biofilms were successfully inhibited with milder conditions than those applied to P. aeruginosa and C. albicans. Notably, it was possible to eradicate 24-h-old biofilms of both S. aureus and P. aeruginosa. In conclusion, this study supports the potential of neutral BODIPYs as pan-antimicrobial PSs.

Bacterial pigments: A colorful palette reservoir for biotechnological applications.

Synthetic derivatives are currently used instead of pigments in many applicative fields, from food to feed, from pharmaceutical to diagnostic, from agronomy to industry. Progress in organic chemistry allowed to obtain rather cheap compounds covering the whole color spectrum. However, several concerns arise from this chemical approach, as it is mainly based on nonrenewable resources such as fossil oil, and the toxicity or carcinogenic properties of products and/or precursors may be harmful for personnel involved in the productive processes. In this scenario, microorganisms and their pigments represent a colorful world to discover and reconsider. Each living bacterial strain may be a source of secondary metabolites with peculiar functions. The aim of this review is to link the physiological role of bacterial pigments with their potential use in different biotechnological fields. This enormous potential supports the big challenge for the development of strategies useful to identify, produce, and purify the right pigment for the desired application. At the end of this ideal journey through the world of bacterial pigments, the attention will be focused on melanin compounds, whose production relies upon different techniques ranging from natural producers, heterologous hosts, or isolated enzymes. In a green workflow, the microorganisms represent the starting and final point of pigment production.

Antimicrobial Activity of Nanoconjugated Glycopeptide Antibiotics and Their Effect on Staphylococcus aureus Biofilm.

In the era of antimicrobial resistance, the use of nanoconjugated antibiotics is regarded as a promising approach for preventing and fighting infections caused by resistant bacteria, including those exacerbated by the formation of difficult-to-treat bacterial biofilms. Thanks to their biocompatibility and magnetic properties, iron oxide nanoparticles (IONPs) are particularly attractive as antibiotic carriers for the targeting therapy. IONPs can direct conjugated antibiotics to infection sites by the use of an external magnet, facilitating tissue penetration and disturbing biofilm formation. As a consequence of antibiotic localization, a decrease in its administration dosage might be possible, reducing the side effects to non-targeted organs and the risk of antibiotic resistance spread in the commensal microbiota. Here, we prepared nanoformulations of the 'last-resort' glycopeptides teicoplanin and vancomycin by conjugating them to IONPs via surface functionalization with (3-aminopropyl) triethoxysilane (APTES). These superparamagnetic NP-TEICO and NP-VANCO were chemically stable and NP-TEICO (better than NP-VANCO) conserved the typical spectrum of antimicrobial activity of glycopeptide antibiotics, being effective against a panel of staphylococci and enterococci, including clinical isolates and resistant strains. By a combination of different methodological approaches, we proved that NP-TEICO and, although to a lesser extent, NP-VANCO were effective in reducing biofilm formation by three methicillin-sensitive or resistant Staphylococcus aureus strains. Moreover, when attracted and concentrated by the action of an external magnet, NP-TEICO exerted a localized inhibitory effect on S. aureus biofilm formation at low antibiotic concentration. Finally, we proved that the conjugation of glycopeptide antibiotics to IONPs reduced their intrinsic cytotoxicity toward a human cell line.

Photoinactivation of Pseudomonas aeruginosa Biofilm by Dicationic Diaryl-Porphyrin.

In recent years, antimicrobial photodynamic therapy (aPDT) has received increasing attention as a promising tool aimed at both treating microbial infections and sanitizing environments. Since biofilm formation on biological and inert surfaces makes difficult the eradication of bacterial communities, further studies are needed to investigate such tricky issue. In this work, a panel of 13 diaryl-porphyrins (neutral, mono- and di-cationic) was taken in consideration to photoinactivate Pseudomonas aeruginosa. Among cationic photosensitizers (PSs) able to efficiently bind cells, in this study two dicationic showed to be intrinsically toxic and were ruled out by further investigations. In particular, the dicationic porphyrin (P11) that was not toxic, showed a better photoinactivation rate than monocationic in suspended cells. Furthermore, it was very efficient in inhibiting the biofilms produced by the model microorganism Pseudomonas aeruginosa PAO1 and by clinical strains derived from urinary tract infection and cystic fibrosis patients. Since P. aeruginosa represents a target very difficult to inactivate, this study confirms the potential of dicationic diaryl-porphyrins as photo-activated antimicrobials in different applicative fields, from clinical to environmental ones.

Effect of blue light at 410 and 455 nm on Pseudomonas aeruginosa biofilm.

Pseudomonas aeruginosa is an opportunistic pathogen resistant to many antibiotics, able to form biofilm and causes serious nosocomial infections. Among anti-Pseudomonas light-based approaches, the recent antimicrobial Blue Light (aBL) treatment seems very promising. The aim of this study was to evaluate the efficiency of blue light in inhibiting and/or eradicating P. aeruginosa biofilm. Light at 410 nm has been identified as successful in inhibiting biofilm formation not only of the model strain PAO1, but also of CAUTI (catheter-associated urinary tract infection) isolates characterized by their ability to form biofilm. Results of this work on 410 nm light also demonstrated that: i) at the lowest tested radiant exposure (75 J cm-2) prevents matrix formation; ii) higher radiant exposures (225 and 450 J cm-2) light impairs the cellular components of biofilm, adherent and planktonic ones; iii) light eradicates with a good rate young and older biofilms in a light dose dependent manner; iv) it is also efficient in inactivating catalase A, a virulence factor playing an important role in pathogenic mechanisms. Light at 455 nm, even if at a lower extent than 410 nm, showed a certain anti-Pseudomonas activity. Furthermore, light at 410 nm caused detrimental effects on enzyme activity of ß-galactosidase and catalase A, and changes on plasmid DNA conformation and ortho-nitrophenyl-ß-D-galactopyranoside structure. This study supports the potential of blue light for anti-infective and disinfection applications.

Magnetic Nanoconjugated Teicoplanin: A Novel Tool for Bacterial Infection Site Targeting.

Nanoconjugated antibiotics can be regarded as next-generation drugs as they possess remarkable potential to overcome multidrug resistance in pathogenic bacteria. Iron oxide nanoparticles (IONPs) have been extensively used in the biomedical field because of their biocompatibility and magnetic properties. More recently, IONPs have been investigated as potential nanocarriers for antibiotics to be magnetically directed to/recovered from infection sites. Here, we conjugated the "last-resort" glycopeptide antibiotic teicoplanin to IONPs after surface functionalization with (3-aminopropyl) triethoxysilane (APTES). Classical microbiological methods and fluorescence and electron microscopy analysis were used to compare antimicrobial activity and surface interactions of naked IONPs, amino-functionalized NPs (NP-APTES), and nanoconjugated teicoplanin (NP-TEICO) with non-conjugated teicoplanin. As bacterial models, differently resistant strains of three Gram-positive bacteria (Staphylococcus aureus, Enterococcus faecalis, and Bacillus subtilis) and a Gram-negative representative (Escherichia coli) were used. The results indicated that teicoplanin conjugation conferred a valuable and prolonged antimicrobial activity to IONPs toward Gram-positive bacteria. No antimicrobial activity was detected using NP-TEICO toward the Gram-negative E. coli. Although IONPs and NP-APTES showed only insignificant antimicrobial activity in comparison to NP-TEICO, our data indicate that they might establish diverse interaction patterns at bacterial surfaces. Sensitivity of bacteria to NPs varied according to the surface provided by the bacteria and it was species specific. In addition, conjugation of teicoplanin improved the cytocompatibility of IONPs toward two human cell lines. Finally, NP-TEICO inhibited the formation of S. aureus biofilm, conserving the activity of non-conjugated teicoplanin versus planktonic cells and improving it toward adherent cells.

Catalase A is involved in the response to photooxidative stress in Pseudomonas aeruginosa.

BACKGROUND: Pseudomonas aeruginosa is the etiological agent of systemic and skin infections that are often difficult to treat. Photodynamic therapy (PDT) and, more recently, phototherapy (PT), are emerging among antimicrobial treatments to be combined with antibiotics. Visible light, either alone or combined with a photosensitizer (PS), elicits photooxidative stress that induces microbial death. The response of bacteria to phototherapy seems to involve the antioxidant machinery. This study relies on the effects of detoxifying catalase A (KatA) in response to PDT and PT-induced photooxidative stress. METHODS: The photo- and photodynamic inactivation experiments have been targeted at P. aeruginosa PAO1 and its isogenic derivative katA- mutant. The microorganisms were irradiated by a wide-spectrum halogen-tungsten lamp or light-emitting diodes (LEDs). Two photosensitizers, Tetrakis-(1-methyl-4-pyridyl)-21H, 23porphine, tetra-p-tosylate (TMPyP) porphyrin and Toluidine Blue O (TBO), were applied as part of the photodynamic approach. RESULTS: P. aeruginosa katA- mutant was more sensitive than wild-type strain PAO1 to wide-spectrum light and blue LED (464 nm) treatments. The complementation of KatA, in katA- mutant, restored the light response of wild-type PAO1. Upon TBO treatment and irradiation by visible light (halogen lamp or LED), the sensitivity of katA- mutant was significant higher (p = 0.028 and p = 0.045, respectively) than that of the PAO1 strain. CONCLUSIONS: This study provides the first description of KatA in the response to photooxidative stress induced by photo- and photodynamic therapy.

Response to photo-oxidative stress of Pseudomonas aeruginosa PAO1 mutants impaired in different functions.

Clinicians often have to deal with infections that are difficult to control because they are caused by superbugs resistant to many antibiotics. Alternatives to antibiotic treatment include antimicrobial photodynamic therapy (aPDT). The photodynamic process causes bacterial death, inducing oxidative stress through the photoactivation of photosensitizer molecules in the presence of oxygen. No PDT-resistant bacteria have been selected to date, thus the response to photo-oxidative stress in non-phototrophic bacteria needs further investigation. The opportunistic pathogen Pseudomonas aeruginosa, in particular, has been shown to be more tolerant to PDT than other micro-organisms. In order to find any genetic determinants involved in PDT-tolerance, a panel of transposon mutants of P. aeruginosa PAO1 involved in the quorum sensing signalling system and membrane cytoplasmic transport were photoinactivated as part of this study. Two pseudomonas quinolone signalling (PQS) knock-out mutants, pqsH- and pqsC-, were as PDT-sensitive as the PAO1 wild-type strains. Two PQS hyperproducer variants, pqsA- and rsaL-, were shown to be more tolerant to photo-oxidative stress than the wild-type strain. In the pqsA- mutant, the hyperpigmentation due to the presence of phenazines could protect cells against PDT stress, while in rsaL- no pigmentation was detectable. Furthermore, a mutant impaired in an ATP-binding cassette transport involved in maintaining the asymmetry of the outer membrane was significantly more tolerant to photo-oxidative stress than the wild-type strain. These observations support the involvement of quorum sensing and the importance of the bacterial cell envelope when dealing with photo-oxidative stress induced by photodynamic treatment.