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Background: A great number of therapeutic limitations, such as chemoresistance, high dosage, and long treatments, are still present in cancer therapy, and are often followed by side effects such as infections, which represent the primary cause of death among patients. Methods: We report pH- and enzymatic-responsive hybrid clustered nanoparticles (HC-NPs), composed of a PCL polymeric core loaded with an anticancer drug, such as Imatinib Mesylate (IM), and coated with biodegradable multilayers embedded with antibacterial and anticancer baby-ship silver NPs, as well as a monoclonal antibody for specific targeting of cancer cells conjugated on the surface. Results: The HC-NPs presented an onion-like structure that serially responded to endogenous stimuli. After internalization into targeted cancer cells, the clustered nanoparticles were able to break up, thanks to intracellular proteases which degraded the biodegradable multilayers and allowed the release of the baby-ship NPs and the IM loaded within the pH-sensible polymer present inside the mothership core. In vitro studies validated the efficiency of HC-NPs in human chronic leukemic cells. This cellular model allowed us to demonstrate specificity and molecular targeting sensitivity, achieved by using a combinatorial approach inside a single nano-platform, instead of free administrations. The combinatory effect of chemotherapic drug and AgNPs in one single nanosystem showed an improved cell death efficacy. In addition, HC-NPs showed a good antibacterial capacity on Gram-negative and Gram-positive bacteria. Conclusions: This study shows an important combinatorial anticancer and antimicrobial effect in vitro.

Recently, the discovery of natural compounds capable of modulating nervous system function has revealed new perspectives for a healthier brain. Here, we investigated the effects of oleic acid (OA) and hydroxytyrosol (HTyr), two important extra virgin olive oil compounds, on lipid synthesis in C6 glioma cells. OA and HTyr inhibited both de novo fatty acid and cholesterol syntheses without affecting cell viability. The inhibitory effect of the individual compounds was more pronounced if OA and HTyr were administered in combination. A reduction of polar lipid biosynthesis was also detected, while triglyceride synthesis was marginally affected. To clarify the lipid-lowering mechanism of these compounds, their effects on the activity of key enzymes of fatty acid biosynthesis (acetyl-CoA carboxylase-ACC and fatty acid synthase-FAS) and cholesterologenesis (3-hydroxy-3-methylglutaryl-CoA reductase-HMGCR) were investigated in situ by using digitonin-permeabilized C6 cells. ACC and HMGCR activities were especially reduced after 4 h of 25 μM OA and HTyr treatment. No change in FAS activity was observed. Inhibition of ACC and HMGCR activities is corroborated by the decrease of their mRNA abundance and protein level. Our results indicate a direct and rapid downregulatory effect of the two olive oil compounds on lipid synthesis in C6 cells.

In this study we have applied an integrated system biology approach to characterize the metabolic landscape of Streptomyces ambofaciens and to identify a list of potential metabolic engineering targets for the overproduction of the secondary metabolites in this microorganism. We focused on an often overlooked growth period (i.e., post-first rapid growth phase) and, by integrating constraint-based metabolic modeling with time resolved RNA-seq data, we depicted the main effects of changes in gene expression on the overall metabolic reprogramming occurring in S. ambofaciens. Moreover, through metabolic modeling, we unraveled a set of candidate overexpression gene targets hypothetically leading to spiramycin overproduction. Model predictions were experimentally validated by genetic manipulation of the recently described ethylmalonyl-CoA metabolic node, providing evidence that spiramycin productivity may be increased by enhancing the carbon flow through this pathway. The goal was achieved by over-expressing the ccr paralog srm4 in an ad hoc engineered plasmid. This work embeds the first metabolic reconstruction of S. ambofaciens and the successful experimental validation of model predictions and demonstrates the validity and the importance of in silico modeling tools for the overproduction of molecules with a biotechnological interest. Finally, the proposed metabolic reconstruction, which includes manually refined pathways for several secondary metabolites with antimicrobial activity, represents a solid platform for the future exploitation of S. ambofaciens biotechnological potential.