Standard of care for cancer is commonly a combination of surgery with radiotherapy or chemoradiotherapy. However, in some advanced cancer patients this approach might still remaininefficient and may cause many side effects, including severe complications and even death. Oncolytic viruses exhibit different anti-cancer mechanisms compared with conventional therapies, allowing the possibility for improved effect in cancer therapy. Chemotherapeutics combined with oncolytic viruses exhibit stronger cytotoxic responses and oncolysis. Here, we have investigated the systemic delivery of the oncolytic adenovirus and paclitaxel encapsulated in extracellular vesicles (EV) formulation that, in vitro, significantly increased the transduction ratio and the infectious titer when compared with the virus and paclitaxel alone. We demonstrated that the obtained EV formulation reduced the in vivo tumor growth in animal xenograft model of human lung cancer. Indeed, we found that combined treatment of oncolytic adenovirus and paclitaxel encapsulated in EV has enhanced anticancer effects both in vitro and in vivo in lung cancer models. Transcriptomic comparison carried out on the explanted xenografts from the different treatment groups revealed that only 5.3% of the differentially expressed genes were overlapping indicating that a de novo genetic program is triggered by the presence of the encapsulated paclitaxel: this novel genetic program might be responsible of the observed enhanced antitumor effect. Our work provides a promising approach combining anticancer drugs and viral therapies by intravenous EV delivery as a strategy for the lung cancer treatment.

Extracellular vesicles (EVs) are naturally occurring cargo delivery vesicles that have recently received considerable attention for their roles in intercellular communication in many physiological and pathological processes, including tumourigenesis. EVs generated by different tissues demonstrated specific homing: in particular, cancer-derived EVs showed a selective tropism for the tumor tissue from which the vesicles originated. For this property, EVs have been proposed as drug delivery tools for anti-cancer therapies, although the limited knowledge about their in vivo tropism hinders their therapeutic applications. The current study aimed to characterize the targeting properties of cancer-derived EVs in vitro and their biodistribution in vivo, by using an imaging approach.Methods: EVs were generated from: i) murine lung (LL/2) and colon (MC-38) cancer lines, ii) human lung cancer cell line (A549) and iii) human liver biopsy samples from healthy individuals. EVs were loaded with fluorescent dyes alone or in combination with a biopharmaceutical agent, the oncolytic adenovirus (OV), characterized for charge and size and tested for their activity in cancer cell lines. Finally, optical imaging was extensively applied to study in vivo and ex vivo the biodistribution of EVs originated from different sources in different mouse models of cancer, including xenograft, syngeneic graft and the MMTV-NeuT genetically modified animal.Results: We initially demonstrated that even loading EVs even with a large biopharmaceutical oncolytic viruses (OVs) did not significantly change their charge and dimension properties, while increasing their anti-neoplastic activity compared to the virus or EVs alone. Interestingly, this activity was observed even if the EVs derived from lung cancer were applied to colon carcinoma cell lines and vice versa, suggesting that the EV uptake occurred in vitro without any specificity for the cancer cells from which the vesicles originated. When administered i.v (intravenously) to the mouse models of cancer, the tumour-derived EVs, but not the EVs derived from a healthy tissue, demonstrated a selective accumulation of the fluorescence at the tumour site 24 h after injection; adding OVs to the formulation did not change the tumour-specific tropism of the EVs also in vivo. Most interestingly, the in vivo experiments confirmed the in vitro observation of the generalized tropism of tumour-derived EVs for any neoplastic tissue, independent of the tumour type or even the species originating the vesicles.Conclusions: Taken together, our in vitro and in vivo data demonstrate for the first time a heterologous, cross-species tumour-tropism for cancer-derived EVs. This finding challenges our current view on the homing properties of EVs and opens new avenues for the selective delivery of diagnostic/therapeutic agents to solid tumours.

Background Nonsteroidal anti-inflammatory drugs (NSAIDs) have been proposed as chemopreventive agents for many tumours; however, the mechanism responsible for their anti-neoplastic activity remains elusive and the side effects due to cyclooxygenase (COX) inhibition prevent this clinical application. Methods Molecular biology, in silico, cellular and in vivo tools, including innovative in vivo imaging and classical biochemical assays, were applied to identify and characterise the COX-independent anti-cancer mechanism of NSAIDs. Results Here, we show that tumour-protective functions of NSAIDs and exisulind (a sulindac metabolite lacking anti-inflammatory activity) occur through a COX-independent mechanism. We demonstrate these NSAIDs counteract carcinogen-induced proliferation by inhibiting the sirtuin 1 (SIRT1) deacetylase activity, augmenting acetylation and activity of the tumour suppressor p53 and increasing the expression of the antiproliferative gene p21. These properties are shared by all NSAIDs except for ketoprofen lacking anti-cancer properties. The clinical interest of the mechanism identified is underlined by our finding that p53 is activated in mastectomy patients undergoing intraoperative ketorolac, a treatment associated with decreased relapse risk and increased survival. Conclusion Our study, for the first-time, links NSAID chemopreventive activity with direct SIRT1 inhibition and activation of the p53/p21 anti-oncogenic pathway, suggesting a novel strategy for the design of tumour-protective drugs.

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From the past decade, extracellular vesicles (EVs) have attracted considerable attention as tools for the selective delivery of anti-neoplastic drugs to cancer tissues. Compared to other nanoparticles, EVs display interesting unique features including immune compatibility, low toxicity and the ability to encapsulate a large variety of small-and macro-molecules. However, in virtually all studies, investigations on EVs have been focused on fully transformed cancers: the possibility to apply EV technology also to early-stage tumors has never been explored. Methods: Herein, we studied the ability of cancer-derived EVs to recognize and deliver their cargo also to incipient cancers. To this purpose, EV biodistribution was studied in MMTV-NeuT genetically modified mice during early mammary transformation, in fully developed breast tumors and in the normal gland of wild type syngeneic mice. EVs were loaded with indocyanine green (ICG), a near-infrared (NIR) dye together with oncolytic viruses and i.v. injected in mice. The nanoparticle biodistribution was assayed by in vivo and ex vivo optical imaging (detecting the ICG) and semiquantitative real-time PCR (measuring the adenoviral genome) in different tissues. Results: Our results demonstrate the ability of cancer-derived EVs to recognize early-stage neoplastic tissues opening the possibility to selectively deliver theranostics also for tumor prevention. Conclusions: Taken together our study demonstrates the ability of EVs to recognize and deliver diagnostic and therapeutic agents not only to fully transformed tissues but also to early stage tumors. These findings pave the way for the synthesis of "universal" EVs-based formulation for targeted cancer therapy.

Dysfunction of the complex cerebral networks underlying wakefulness and awareness is responsible for Disorders of Consciousness (DoC). Traumatic Brain Injury (TBI) is a common cause of DoC, and it is responsible for a multi-dimensional pathological cascade that affects the proper functioning of the brainstem and brain consciousness pathways. Iron (Fe), Zinc (Zn), and Copper (Cu) have a role in the neurophysiology of both the ascending reticular activating system, a multi-neurotransmitter network located in the brainstem that is crucial for consciousness, and several brain regions. We aimed to summarize the role of these essential metals in TBI and its possible link with consciousness alterations. We found that TBI alters many neuronal molecular mechanisms involving essential metals, causing neurodegeneration, neural apoptosis, synaptic dysfunction, oxidative stress, and inflammation. This final pattern resembles that described for Alzheimer’s disease (AD) and other neurological and psychiatric diseases. Furthermore, we found that amantadine, zolpidem, and transcranial direct current stimulation (tDCS)—the most used treatments for DoC recovery—seem to have an effect on essential metals-related pathways and that Zn might be a promising new therapeutic approach. This review summarizes the neurophysiology of essential metals in the brain structures of consciousness and focuses on the mechanisms underlying their imbalance following TBI, suggesting their possible role in DoC. The scenario supports further studies aimed at getting a deeper insight into metals’ role in DoC, in order to evaluate metal-based drugs, such as metal complexes and metal chelating agents, as potential therapeutic options.

Computed tomography (CT) is a diagnostic medical imaging modality commonly used to detect disease and injury. Contrast agents containing iodine, such as iohexol, are frequently used in CT examinations to more clearly differentiate anatomic structures and to detect and characterize abnormalities, including tumors. However, these contrast agents do not have a specific tropism for cancer cells, so the ability to detect tumors is severely limited by the degree of vascularization of the tumor itself. Identifying delivery systems allowing enrichment of contrast agents at the tumor site would increase the sensitivity of detection of tumors and metastases, potentially in organs that are normally inaccessible to contrast agents, such as the CNS. Recent work from our laboratory has identified cancer patient-derived extracellular vesicles (PDEVs) as effective delivery vehicles for targeting diagnostic drugs to patients’ tumors. Based on this premise, we explored the possibility of introducing iohexol into PDEVs for targeted delivery to neoplastic tissue. Here, we provide preclinical proof-of-principle for the tumor-targeting ability of iohexol-loaded PDEVs, which resulted in an impressive accumulation of the contrast agent selectively into the neoplastic tissue, significantly improving the ability of the contrast agent to delineate tumor boundaries.