Nanomaterials with easy surface functionalization and high biocompatibility have emerged as good candidates for theranostic applications. In particular the integration of multiple smart functions such as contrast agents (CA) and photosensitizers (PS) into a nanodevice platform can enhance diagnostic and therapeutic eﬃcacies and is nowadays a great scientific challenge. To obtain an efficient nanodevice for coupling optical imaging together with photodynamic therapy (PDT), it is necessary to control the location of the contrast agent and/or of the photosensitizer and to optimize their loadings and their dispersion in order to avoid molecular aggregation that is detrimental to both imaging and photodynamic therapy. Among a variety of nanoparticles that can be used as nanovectors, mesoporous silica nanoparticles (MSNs), prepared through well-established synthetic procedures, have attracted great scientific attention due to their high specific surface area, high pore volume and uniform pore size that allow hosting, through simple synthetic procedures, different guests. The unique topology of MSNs with distinct domains that can be independently functionalized, the hexagonal mesochannels and the nanoparticle’s outermost surface, allows controlling accurately the guest location. In addition, MSNs with facile surface functionalization, low cytotoxicity, enhanced therapeutic efficiency, good biocompatibility, and avid cell uptake were recently applied in in vivo assays in small-animal disease models.
Among the great variety of dyes that can be used as fluorescent probes, Rhodamine dyes, belonging to the family of xanthenes, are extensively employed. In particular, Rhodamine B (RhB) is one of the mostly used rhodamines due to its high absorption coeﬃcient in the visible region, moderate to high fluorescence quantum yield depending on the solvent (0.30–0.60) and high sensitivity to the solvent polarity. Herein, Rhodamine B was conjugated to amino-functionalized MSNs (NH2-MSNs) and used as a contrast agent for optical imaging. The precise localization of the contrast agent (RhB) and the accurate control of its dispersion are key factors to enhance the fluorescence quantum yield and to avoid the presence of aggregates in such a type of nanodevice; for these reasons samples with diﬀerent RhB loadings (from 10 to 100 mg g-1) in diﬀerent silica environments were synthesized. In particular, the eﬃcacy of the mesoporous silica support in the dispersion and protection of the fluorophores was verified by covalently binding the RhB molecules inside the mesochannels (RhB-in-MSNs) or on the outermost surface (RhB-out-MSNs) of MSNs: extracted (without the surfactant) and as-synthesized (with the surfactant) NH2-MSNs were used for these purposes (Scheme 1). When the surfactant is present, the channels of MSNs are non-accessible and the RhB molecules can in principle be attached only on the external surface of the MSNs. In Table 1, the acronyms used for all the samples, the RhB nominal loadings and the MSN supports used are listed.
A systematic and detailed spectroscopic characterization of these nanomaterials using a very sensitive technique like photo-luminescence, augmented by fluorescence lifetime measurements, has been used to select the nanomaterial with the best photophysical performances. This optimized nanodevice, due to the presence of residual amino groups, could be further functionalized with a photosensitizer (PS) to generate a multi-functional nanomedical platform to couple optical imaging with PDT. Light-triggered nanodevices are in fact attracting increasing scientific attention and, in particular, light has been used to release therapeutic agents from nanodevices and to activate agents that produce cytotoxic species. PDT is a promising therapeutic modality alternative to chemo- and radiotherapy due to its non-invasive and selective destruction of tumors: this is a light activated treatment modality based on the generation of reactive oxygen species (ROS) or singlet oxygen (1O2)from the PS by the light irradiation, leading to the selective and irreversible destruction of diseased cells. As a PS, verteporfin, a potent second-generation photosensitizing agent derived from benzoporphyrins, was used; it shows intense absorption bands in the red region of the visible spectrum (670–690 nm) and high quantum yield in the generation of singlet oxygen.
A lipid-based formulation of verteporfin (Visudyne) has been approved by Food and Drug Administration (FDA) in the US and in Europe in 2000 for treatment of age-related macular degeneration (AMD).Verteporfin is a hydrophobic sensitizer,whichinaqueousmedia easily forms aggregates with reduced efficiency in the generation of singlet oxygen; however, by covalently coupling verteporfin into mesoporous silica nanoparticles, it is possible to avoid aggregation. In particular, the aggregated form usually shows a lower singlet oxygen generation quantum yield and therefore a lower photodynamic efficacy: this occurs because dimmers are characterized by low triplet quantum yield. Verteporfin was covalently bound to the amino groups of the optimized
RhB-MSN sample to produce bifunctional nanodevices (Ver–RhB-MSNs).
Structural and morphological characterization methods of the monofunctional RhB-MSN and bifunctional Ver–RhB-MSN nanodevices were performed by X-ray powder diffraction (XRD), high-resolution transmission electron microscopy (HRTEM) and volumetric analysis combined with FTIR spectroscopy. To elucidate the RhB location and distribution inside or outside of the channels of MSNs with respect to the RhB loadings and the influence of encapsulation into the channels on their photo-emission properties, a detailed spectroscopic characterization was carried out by using Diffuse Reflectance (DR) UV-Vis and photoluminescence coupled with the fluorescence lifetime measurements. Due to the potential application of the bifunctional nanosystems containing RhB and verteporfin, the 1O2 generation efficiency was also evaluated by a chemical method using uric acid, which undergoes oxidation upon PS irradiation and 1O2 generation.
B. Martins Estevao, I. Miletto*, L. Marchese, E. Gianotti. Phys. Chem. Chem. Phys., 2016, 18, 9042—9052