Paolo Dambruoso

Mesenchymal stem cells as delivery vehicle of porphyrin loaded nanoparticles: Effective photoinduced in vitro killing of osteosarcoma

Mesenchymal stem cells (MSC) have the unique ability to home and engraft in tumor stroma. These features render them potentially a very useful tool as targeted delivery vehicles which can deliver therapeutic drugs to the tumor stroma. In the present study, we investigate whether fluorescent core-shell PMMA nanoparticles (FNPs) post-loaded with a photosensitizer, namely meso-tetrakis (4-sulfonatophenyl) porphyrin (TPPS) and uploaded by MSC could trigger osteosarcoma (OS) cell death in vitro upon specific photoactivation. In co-culture studies we demonstrate using laser confocal microscopy and time lapse imaging, that only after laser irradiation MSC loaded with photosensitizer-coated fluorescent NPs (TPPS@FNPs) undergo cell death and release reactive oxygen species (ROS) which are sufficient to trigger cell death of all OS cells in the culture. These results encourage further studies aimed at proving the efficacy of this novel tri-component system for PDT applications.

Thiazolium-functionalized polystyrene monolithic microreactors for continuous-flow umpolung catalysis

Thiazolium salt pre-catalysts have been immobilized on silica and monolithic polystyrene and their activity was tested under batch conditions in three model umpolung reactions, namely the benzoin condensation of benzaldehyde, the acyloin-type condensation of biacetyl, and the Stetter reaction of biacetyl with trans-chalcone. A prerequisite of the study has been the utilization of environmentally benign water and ethanol solvents. After having established the higher performance of polystyrene monolithic thiazolium carbene catalysts, their effectiveness has been tested under the flow regime by fabricating the corresponding monolithic microreactors (pressure-resistant stainless-steel columns). Importantly, it has been demonstrated by a brief substrate scope study that the polymeric matrix and the continuous flow regime synergistically contribute to preserve the activity of the carbene catalysts over time, thus permitting the long-term operation (up to 7 days) of the prepared monolithic reactors for the production of valuable compounds via the umpolung strategy.

Polymer nanoparticles with electrostatically loaded multicargo for combined cancer phototherapy

We have developed a multifunctional polymer-based nanoplatform for bimodal cancer phototherapy. It was achieved by electrostatic entangling of two anionic photoactivable components, a commercial porphyrin and a tailored nitro-aniline derivative, within the cationic shell of polymeric nanoparticles (NPs) based on polymethyl methacrylate. The combination of steady-state and time-resolved spectroscopic and photochemical techniques shows that the two photoresponsive agents do not interfere with each other while being in close proximity in the same polymeric scaffold and can thus operate in parallel under the exclusive control of light stimuli. Specifically, visible light triggers satisfactory red fluorescence emission and generation of singlet oxygen (1O2) from one component and release of nitric oxide (NO) from the other. Fluorescence microscopy analysis provides unambiguous evidence for the internalization of the NPs within B78H1 melanoma cells, where they induce amplified mortality due to a combinatory effect of the two photogenerated cytotoxic species.

Sulfonates-PMMA nanoparticles conjugates: A versatile system for multimodal application

We report herein the viability of a novel nanoparticles (NPs) conjugated system, namely the attachment, based on ionic and hydrophobic interactions, of different sulfonated organic salts to positively charged poly(methylmethacrylate) (PMMA)-based core-shell nanoparticles (EA0) having an high density of ammonium groups on their shells. In this context three different applications of the sulfonates@EA0 systems have been described. In detail, their ability as cytotoxic drugs and pro-drugs carriers was evaluated in vitro on NCI-H460 cell line and in vivo against human ovarian carcinoma IGROV-1 cells. Besides, 8-hydroxypyrene-1,3,6-trisulfonic acid, trisodium salt (HPTS) was chosen for NPs loading, and its internalization as bioimaging probe was evaluated on Hep G2 cells. Overall, the available data support the interest for these PMMA NPs@sulfonates systems as a promising formulation for theranostic applications. In vivo biological data strongly support the potential value of these core-shell NPs as delivery system for negatively charged drugs or biologically active molecules. Additionally, we have demonstrated the ability of these PMMA core-shell nanoparticles to act as efficient carriers of fluorophores. In principle, thanks to the high PMMA NPs external charge density, sequential and very easy post-loading of different sulfonates is achievable, thus allowing the preparation of nanocarriers either with bi-modal drug delivery behaviour or as theranostic systems.

TPPS supported on core–shell PMMA nanoparticles: the development of continuous-flow membrane-mediated electrocoagulation as a photocatalyst processing method in aqueous media

Herein, the successful utilization of core shell poly-methyl methacrylate nanoparticles (CS-PMMA NPs) as a valuable support for organocatalyst immobilization in aqueous medium is presented along with an unprecedented direct current-based electrochemical method for processing a water colloid of the resulting nano-supported catalyst. The NP coagulation procedure relied on various electrocoagulation techniques and it was achieved through water electrolysis occurring in a dialysis membrane immersed in the colloid. Mass recoveries of up to 95% w/w were obtained for the NPs within 1 h of electrolysis. This was optimized at a constant current of 90 mA or a constant potential of 24 V. The disclosed NP separation procedure was effectively exploited in a continuous-flow reaction/separation/recycle sequence involving the tetraphenylporphyrin tetrasulfonate (TPPS) organo-photocatalyst immobilized on CS-PMMA NPs (NP2s). After an optimization study aimed at maximizing the chemical efficiency of the entire synthetic and purification process, the nano-supported catalyst NP2 (0.05 mol%) efficiently promoted the photoexcitation of triplet oxygen to singlet oxygen for the selective and environmentally benign sulfoxidation of a model sulfide in an aqueous medium.

Development of near-infrared photoactivable phthalocyanine-loaded nanoparticles to kill tumor cells: An improved tool for photodynamic therapy of solid cancers

Conventional photodynamic therapy has shown to be beneficial in the treatment of a variety of tumors. However, one of its major limitations is the inadequate penetration depth of visible light. In order to overcome this constraint, we developed 80nm poly-methylmethacrylate core-shell fluorescent nanoparticles (FNP) loaded with the photosensitizer tetrasulfonated aluminum phthalocyanine (Ptl). To demonstrate the efficacy of our Ptl@FNP we performed in vitro and in vivo studies using a human prostate tumor model. Our data reveal that Ptl@FNP are internalized by tumor cells, favour Ptl intracellular accumulation, and efficiently trigger cell death through the generation of ROS upon irradiation with 680nm light. When directly injected into tumors intramuscularly induced in SCID mice, Ptl@FNP upon irradiation significantly reduce tumor growth with higher efficiency than the bare Ptl. Collectively, these results demonstrate that the newly developed nanoparticles may be utilized as a delivery system for antitumor phototherapy in solid cancers.

Elucidating new structural features of the triazole scaffold for the development of mPGES-1 inhibitors

We report a new potent revisited version of a triazole-based inhibitor obtained by structure-based drug design on the human mPGES-1 crystal structure. Moreover, we disclosed the substitution with a halogen atom at position 5 as a new key factor influencing the biological activity on the mPGES-1 enzyme.

Core–shell poly-methylmethacrylate nanoparticles as effective carriers of electrostatically loaded anionic porphyrin

Among the medical applications of nanoparticles, their usage as photosensitizer (PS) carriers for photodynamic therapy (PDT) has attracted increasing attention. In the present study we explored the morphological and photophysical properties of core-shell PMMA nanoparticles (PMMA-NPs) electrostatically post-loaded with the synthetic, water soluble 5,10,15,20-tetrakis(4-sulphonatophenyl)-porphyrin (TPPS4). pH response and singlet oxygen analyses of differently loaded samples proved the high capability of the PMMA-NPs to shield the PS from the environment, while retaining the PS singlet oxygen production capability. Preliminary in vitro imaging and phototoxicity experiments on HepG2 cells demonstrated the efficacy of the system to trigger photoinduced cell death in the culture.

Thiophene-based compounds as fluorescent tags to study mesenchymal stem cell uptake and release of taxanes.

Human mesenchymal stem cells (hMSC) are multipotent cells that display the unique ability to home and engraft in tumor stroma. This remarkable tumor tropic property has generated a great deal of interest in many clinical settings. Recently, we showed that hMSC represent an excellent base for cell-mediated anticancer therapy since they are able to internalize paclitaxel (PTX) and to release it in an amount sufficient to inhibit tumor cell proliferation. In order to shed light on the dynamics of drug uptake and release, in the present paper we describe the synthesis of two novel thiophene-based fluorophore-paclitaxel conjugates, namely PTX-F32 and PTX-F35, as tools for in vitro drug tracking. We aimed to study the ability of these novel derivatives to be efficiently internalized by hMSC and, in a properly engineered coculture assay, to be released in the medium and taken up by tumor cells. In order to ensure better stability of the conjugates toward enzymatic hydrolysis, the selected oligothiophenes were connected to the taxol core at the C7 position through a carbamate linkage between PTX and the diamino linker. Antiproliferative experiments on both tumor cells and stromal cells clearly indicate that, in good correlation with the parent compound, cells are sensitive to nanomolar concentrations of the fluorescent conjugates. Moreover, in the coculture assay we were able to monitor, by fluorescence microscopy, PTX-F32 trafficking from hMSC toward glioblastoma U87 tumor cells. Our work paves the way for novel possibilities to perform extensive and high quality fluorescence-based analysis in order to better understand the cellular mechanisms involved in drug trafficking, such as microvescicle/exosome mediated release, in hMSC vehicle cells.