Seite - 13 - in Cancer Nanotheranostics - What Have We Learnd So Far?

Condeetal. Biofunctionalizationandsurfacechemistryof inorganicnanoparticles FIGURE3 |Fluorescent-AuNPs. (A) Chemical structure of a FAM–DNA–AuNP and schematic illustration of its FRET-based operating principles. (B–E) Confocal fluorescence and phase-contrast images of living cells. (B) Fluorescence image of macrophages incubated with the probe for 30min at 37◦C. (C) Fluorescence image of probe-stained macrophages stimulated with PMA for 1h at 37◦C. (D) Bright field image of live macrophages shown in (C), confirming their viability. (E) AO staining of probe-loaded macrophages, confirming their viability (Tang et al., 2008). Reproduced with permission from Tang et al. (2008), Copyright 2013. biologicalmatrices,suchasserum(Deetal.,2009).Bycorrelating the variation in fluorescence intensity with specific proteins of interest, they were able to identify proteins such as fibrinogen, human serum albumin and immunoglobin G, among others withover97%accuracy. In essence, fluorescent-nanoparticle systems can be used for sensing by exploring a typical FRET in order to provide effi- cient in vivo detection and tumor targeting. These nanocarriers symbolize an important class ofmaterials with unique features suitable for biomedical imaging applications such as increased sensitivity indetection,highquantumyields forfluorescenceand a bounty of novel applications in optics and nanophotonics for moleculardiagnostics (Condeetal., 2012d). QDsareoftenusedasfluorescentmoleculesper se, since they are semiconductornanoparticleswithnarrow, tunable, symmet- rical emission spectra and high quantum yields (Weller, 1993; Bruchezetal., 1998).ThesecharacteristicswereevidencedbyWu etal.usingQDsmodifiedwithdifferentcellularantigensenabling the simultaneous detection of two different targets in the same cell (Wu et al., 2003). It was also shown their higher brightness and photobleaching resistance when compared to organic dyes. These properties make QDs exceptional substitutes as fluores- cence labels (Xinget al., 2006; Smith et al., 2010; SmithandNie, 2012). The inclusion of dyes onto MNPs allows the creation of multifunctional NPs, which might be used for MRI and opti- cal imaging. These dual MNPs allow for multimodal imaging, which implies that the limitationsofone imagingmodalitycould be compensated by the other, creating a complementary effect (Louie, 2010). For instance,Medarova and co-workers reported the synthesis of a multifunctional MNP that included near- infrared optical imaging dye, peptides formembrane transloca- tion and synthetic siRNA targeting a specific gene (Medarova et al., 2007). In vivo accumulation of theMNPswas assessed by MRI and optical imaging and the silencing efficiency was also probedby invivooptical imaging. Nucleicacids WatsonandCrickfirstdescribedDNAas twohelical chains each coiled around the same axis, consisting of simple and repeating unitscallednucleotideswithbackbonesmadeofsugarsandphos- phategroupsjoinedbyesterbondsthatruninoppositedirections toeachother.The importanceof thismoleculewithin livingcells isundisputable(WatsonandCrick,1953).Besidestheirbiological function, nucleic acids canbe employed as polymericmolecules which will bind specifically to targets thanks toWatson–Crick basepairing(FichouandFerec,2006). Mirkin et al. (1996) described the use of a cross-linking method that relies on the detection of single-stranded oligonu- cleotidetargetsusingtwodifferentgoldnanoprobes,eachofthem functionalized with a DNA-oligonucleotide complementary to onehalf of the given target. This functionalizationwas achieved using the strong affinity of thiol or disulfide groups to the gold surface of the NPs, forming quasi-covalent bonds. By modify- ing a nucleic acidmolecule with a thiol group in either the 5′ or the 3′ end it is possible to fine-tune theDNA assembly into the gold surface (Hurst et al., 2006), controlling variables such as salt concentration, oligo/NP ratio or nanoparticle size. This phenomenon indicates the potential of AuNPs modified with DNAs tobe applied inbiosensingor asDNAprobes fordiagno- sis (Cao et al., 2005). These assays became an importantmark in detection once they have PCR-like sensitivity, selectivity for target sequences, capacity for massive multiplexing, and most importantly,have theability tobeperformedat thepointofcare. Usingafluorescence-basedmethod,Demersetal.,havedeter- mined the number of thiol-derivatized single-stranded oligonu- cleotides bound to AuNPs and their extent of hybridization withcomplementaryoligonucleotides in solution (Demers et al., 2000). Also, using a fluorescencemethod,Conde et al. reported thepotential of a singlemolecularnanoconjugate to intersect all RNA pathways: from gene specific downregulation to silencing the silencers, i.e., siRNA and miRNA pathways, by using gold nanobeacons (Figure4). These nanoconjugates functionalized Frontiers inChemistry | ChemicalEngineering July2014 |Volume2 |Article48 | 13