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Tuberculosis, still one of the leading human infectious diseases, reported 8.7 million new cases in 2011 alone. Also, the increasing rate of multidrug-resistant tuberculosis (MDRTB) and its treatment difficulties pose a serious public health threat especially in developing countries. Resistance to isoniazid and rifampicin, first line antibiotics, is commonly associated with point mutations in katG, inhA and rpoB genes of Mycobacterium tuberculosis complex (MTBC). Therefore, the development of cheap, fast and simple molecular methods to assess susceptibility profiles would have a huge impact in the capacity of early diagnosis and treatment of MDRTB.

Gold nanoparticles functionalized with thiol-modified oligonucleotides (Au-nanoprobes) have shown the potential to provide a rapid and sensitive detection method for MTBC and single base mutations associated with antibiotic resistance, namely the characterization of the three most relevant codons in rpoB gene associated to rifampicin resistance. Here we extend the Au-nanoprobe approach towards discriminating specific mutations within inhA and rpoB genes in PCR amplified DNA from isolates. Using a multiplex PCR reaction for these two genes, it is possible to assess both loci in parallel, and extend the potential of the Au-nanoprobe method to MDRTB molecular characterization with special application in the most frequent Portuguese genotypes.

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Identification of specific nucleic acid sequences mediated by gold nanoparticles derivatized thiol-modified oligonucleotides (Au-nanoprobes) has been proven to be a useful tool in molecular diagnostics. Here, we demonstrate that, on optimization, detection may be simplified via the use of a single Au-nanoprobe to detect a single nucleotide polymorphism (SNP) in homo- or heterozygote condition. We validated this non-cross-linking approach through the analysis of 20 clinical samples using a single specific Au-nanoprobe for an SNP in the FTO (fat mass and obesity-associated) gene against direct DNA sequencing. Sensitivity, specificity, and limit of detection (LOD) were determined, and statistical differences were calculated by one-way analysis of variance (ANOVA) and a post hoc Tukey's test to ascertain whether there were any differences between Au-nanoprobe genotyped groups. For the first time, we show that the use of a single Au-nanoprobe can detect SNP for each genetic status (wild type, heterozygous, or mutant) with high degrees of sensitivity (87.50%) and specificity (91.67%).

The design of small interfering RNA (siRNA) delivery materials showing efficacy in vivo is at the forefront of nanotherapeutics research. Polyurea (PURE-type) dendrimers are ‘smart’ biocompatible 3D polymers that unveil a dynamic and elegant back-folding mechanism involving hydrogen bonding between primary amines at the surface and tertiary amines and ureas at the core. Similarly, to a biological proton pump, they are able to automatically and reversibly transform their conformation in response to pH stimulus. Here, we show that PURE-G4 is a useful gene silencing platform showing no cellular toxicity. As a proof of concept we investigated the PURE-G4-siRNA dendriplex, which was shown to be an attractive platform with high transfection efficacy. The simplicity associated with the complexation of siRNA with polyurea dendrimers makes them a powerful tool for efficient cytosolic siRNA delivery.

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Traditional methods of drug delivery present several drawbacks, mainly due to off-target effects that may originate severe side and toxic effect to healthy tissues. Parallel to the development of novel more effective drugs, particular effort has been dedicated to develop and optimize drug delivery vehicles capable of specifically targeting the required tissue/organ and to deliver the cargo only where and when it is needed. New drug delivery systems based on nanoscale devices showing new and improved pharmacokinetic and pharmacodynamics properties like enhanced bioavailability, high drug loading or systemic stability have surged in the past decade as promising solutions to the required therapeutic efficacy. Amongst these nanoscale vectors, nanoparticles for drug delivery, such as polymeric, lipidbased, ceramic or metallic nanoparticles, have been at the forefront of pharmaceutical development. The interest in nanomedicine for treatment and diagnosis is clearly reflected on the increasing number of publications and issued patents every year. Here, we provide a broad overview of novel nanoparticle based drug delivery systems, ranging from polymeric systems to metal nanoparticles, while simultaneously listing the most relevant related patents.