Page - 83 - in Cancer Nanotheranostics - What Have We Learnd So Far?

Conniotet al. Nanocarriers for immunecell targetingand tracking The use of targeted nanoplatforms for this purpose enables amore specific interactionwith the intended target, withmin- imal interference to the biological system (Ballou et al., 2004). Additionally, nanocarriersmay be functionalized with single or multiple ligands, which may be important for the design of complex experiments. The targeting of ligandsmay enhance the selective recognition of the nanodelivery systems by cells, facili- tatingtheirendocytosis.Thiswillallownanosystemstobeusedas non-invasive localization,monitoring andassessmentplatforms, for instance, for site-specific intracellular characterizations and real-timetracking(Ruanetal., 2007). Fluorescenceimagingtechniques Fluorescence imaging is an optical imaging method based on the excitation/emissionofmolecules (Cai andChen, 2007). The useoffluorescentmolecularprobes—asfluorescentdyesandflu- orescent proteins—has been widely applied in the labeling of biomolecules, cellsandtissues.Althoughtheseprobesarealready used in vivo, for instance in retinal angiography and visualiza- tionof arteries, they areunsuitable for real-time imaging assays, regarding their low photostability and sensitivity at the cellular andmolecular levels (Santra andMalhotra, 2011). The applica- tion of fluorophores in real-time in vivo imaging has also been limited by the high absorption of optical signal by tissues and body fluids in theUVand visiblewavelength. The light scatter- ing caused by tissues that attenuate the optical signal and the tissue auto-fluorescence that influences the background signal is alsoa limitation(SantraandMalhotra,2011).Additionally, some fluorescent probes may be toxic for cells and body (Li et al., 2013). Several NP-based strategies have been proposed to over- come the limitations of fluorescent dyes for real-time in vivo imaging (SupplementaryMaterial) (Santra andMalhotra, 2011; Wangetal., 2013b). Fluorescent-labeled NPs are more stable in the body and increase the detection sensitivity andphotostability. In the same platforms, a great number of probe molecules can be incor- porated, in opposite to a single conventional molecule. Also, in NPs, fluorescent dyes can be protected from quenching and degradation(SantraandMalhotra,2011;Wangetal., 2013b). The most extensively studied nanosystems for fluorescence imaging are quantum dots (QDs) (Cai and Chen, 2007), inor- ganicfluorescentNPs that canbebasedonmetallic or semicon- ductormaterials, such as CdSe and CdTe (Ballou et al., 2004). As reviewed by Cai and Chen, in ideal conditions, QDs can have better properties than organic fluorescence probes. These includehigh resistance todegradationandphotobleaching, high quantum yields, highmolar extinction coefficients, continuous absorption spectra covering fromUVtonear-infrared, longflu- orescence lifetimes (>10 ns), narrow emission spectra and very longeffectiveStokes shifts (Cai andChen,2007).QDshavebeen usedforinnumerousapplications,fromcelltracking(Vouraetal., 2004) tomapping of sentinel lymphnodes (Ballou et al., 2007). QDs can be used to identify several ligands in the same exper- iment, using multiple colors and intensities to detect different structures(Ballouetal.,2004).ThepotentialuseofDC-targeting QDs as both fluorescent NPs for in vivo and in vitro imaging, and antigen-delivery system has also been investigated. In this study, it was proved that QDs display promising properties for combinedprimingandimmunoimagingofDC(Senetal.,2008). Functionalization andmodifications of the surface ofQDswith PEGchainsand ligands foractive targeting, suchaspeptides and antibodies, havebeenunder research to improve the application of thesenanosystems in thebiomedical field (Ballou et al., 2007; Cai and Chen, 2007). QD conjugates are already commercially available for immunospecific labeling (Ballouet al., 2004).Thus, thedevelopmentofmultifunctionalnanoplatformsholds agreat promise for the futureofbiomedicine, since itwill bepossible to combine simultaneouslybothdiagnosis and therapy in the same nanostructure(Kimetal., 2008a). Several other groups have suggested the use of silica-based NPs (siNPs) as an interesting strategy toperform imaging assays using fluorescence (Santra et al., 2005; Kim et al., 2008a;Wang et al., 2013b). siNPs have been used for high sensitive and spe- cific in situ labeling and tracking of cell surface receptors (He et al., 2004, 2007).Relyingon theaffinityof antigen-antibodyor ligand-receptor interactions,NPswere functionalizedwith anti- bodies and ligands andappliedas an immunediagnosticmethod (He et al., 2002). siNPs have also been used as a non-invasive tool for intracellular labeling, trackingandsensing in livingcells, contributing with novel information about dynamic biological processes of subcellular structures, such as lysosomes and endo- somes (Shi et al., 2010). Finally, siNPs were applied to better understand the biodistribution and fate of NPs, in vivo (Wang etal., 2013b). Molecular imagingtechniques The key role of immune cells in the development of future immunotherapeutic approaches against chronic pathologies, mainly cancer diseases, has fostered the design and optimiza- tionofdifferentreal-timeimagingtechniques,avoidingtheclassic exvivohistologicanalysis (Kircheret al., 2011;AhrensandBulte, 2013;LiuandLi,2014). Infact,mostof the informationobtained for immune cell tracking has arisen from optical and confocal microscopyandflowcytometry.Two-photonmicroscopyallowed theobservationofdifferent immunecells in theirbiologicalenvi- ronment at real time (Progatzky et al., 2013). However, despite being a powerful tool to observe these highly motile cells and characterizetheir interactionwithnativeenvironment, this imag- ing technique is unsuitable fordetectionofdeeper events due to tissueopacity (Dzhagalovetal., 2012). Bioluminescence imaging techniques, on the other hand, enable deeper tissue penetrations while tracking immune cells in vivo. Even though, it is oneof themost commonlyused tech- niques for immune cell tracking in vivo, allowing whole-body non-invasive tomography. This technique is only useful for pre- clinical studies in small animals, due to the limits related to the attenuationof light in tissues (Kircheretal., 2011). Allnear-infrared(NIR)multiphotonmicroscopymethodsare potential techniques for deep tissue imaging but further studies are needed to better characterize the capabilities of these NIR- excitation techniques and background reduction (Joshi et al., 2013). Magnetic resonance imaging (MRI), ultrasound, positron emission tomography (PET) (Yaghoubi et al., 2009), single Frontiers inChemistry | ChemicalEngineering November2014 |Volume2 |Article105 | 83