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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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The majority of heterocycle compounds and typically common heterocycle fragments present in most pharmaceuticals currently marketed, alongside with their intrinsic versatility and unique physicochemical properties, have poised them as true cornerstones of medicinal chemistry. Apart from the already marketed drugs, there are many other being investigated for their promising activity against several malignancies. In particular, anticancer research has been capitalizing on the intrinsic versatility and dynamic core scaffold of these compounds. Nevertheless, as for any other promising anticancer drugs, heterocyclic compounds do not come without shortcomings. In this review, we provide for a concise overview of heterocyclic active compounds and families and their main applications in medicine. We shall focus on those suitable for cancer therapy while simultaneously addressing main biochemical modes of action, biological targets, structure-activity relationships as well as intrinsic limitation issues in the use of these compounds. Finally, considering the advent of nanotechnology for effective selective targeting of drugs, we shall discuss fundamental aspects and considerations on nanovectorization of such compounds that may improve pharmacokinetic/pharmacodynamic properties of heterocycles.

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Abstract A series of 4-ethynylaniline gold(I) complexes containing monophosphane (1,3,5-triaza-7-phosphaadamantane (pta; 2), 3,7-diacetyl-1,3,7-triaza-5-phosphabicyclo[3.3.1]nonane (3), and PR3, with R=naphthyl (4), phenyl (5), and ethyl (6)) and diphosphane (bis(diphenylphosphino)acetylene (dppa; 7), trans-1,2-bis(diphenylphosphino)ethene (dppet; 8), 1,2-bis(diphenylphosphino)ethane (dppe; 9), and 1,3-bis(diphenylphosphino)propane (dppp; 10)) ligands have been synthesized and their efficiency against tumor cells evaluated. The cytotoxicity of complexes 2–10 was evaluated in human colorectal (HCT116) and ovarian (A2780) carcinoma as well as in normal human fibroblasts. All the complexes showed a higher antiproliferative effect in A2780 cells, with the cytotoxicity decreasing in the following order 5>6=9=10>8>2>4>7>3. Complex 4 stands out for its very high selectivity towards ovarian carcinoma cells (IC50=2.3 μm) compared with colorectal carcinoma and normal human fibroblasts (IC50>100 μm), which makes this complex very attractive for ovarian cancer therapy. Its cytotoxicity in these cells correlates with the induction of the apoptotic process and an increase of intracellular reactive oxygen species (ROS). The effects of the nuclearity, rigidity, and solubility of these complexes on their biological activity were also analyzed. X-ray crystal structure determination allowed the identification of short N−H⋅⋅⋅π contacts as the main driving forces for the three-dimensional packing in these molecules.

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There is a strong interest in the use of biopolymers in the electronic and biomedical industries, mainly towards low-cost applications. The possibility of developing entirely new kinds of products based on cellulose is of current interest, in order to enhance and to add new functionalities to conventional paper-based products. We present our results towards the development of paper-based microfluidics for molecular diagnostic testing. Paper properties were evaluated and compared to nitrocellulose, the most commonly used material in lateral flow and other rapid tests. Focusing on the use of paper as a substrate for microfluidic applications, through an eco-friendly wax-printing technology, we present three main and distinct colorimetric approaches: (i) enzymatic reactions (glucose detection); (ii) immunoassays (antibodies anti-Leishmania detection); (iii) nucleic acid sequence identification (Mycobacterium tuberculosis complex detection). Colorimetric glucose quantification was achieved through enzymatic reactions performed within specific zones of the paper-based device. The colouration achieved increased with growing glucose concentration and was highly homogeneous, covering all the surface of the paper reaction zones in a 3D sensor format. These devices showed a major advantage when compared to the 2D lateral flow glucose sensors, where some carryover of the coloured products usually occurs. The detection of anti-Leishmania antibodies in canine sera was conceptually achieved using a paper-based 96-well enzyme-linked immunosorbent assay format. However, optimization is still needed for this test, regarding the efficiency of the immobilization of antigens on the cellulose fibres. The detection of Mycobacterium tuberculosis nucleic acids integrated with a non-cross-linking gold nanoprobe detection scheme was also achieved in a wax-printed 384-well paper-based microplate, by the hybridization with a species-specific probe. The obtained results with the above-mentioned proof-of-concept sensors are thus promising towards the future development of simple and cost-effective paper-based diagnostic devices.

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Many diseases have their treatment options narrowed and end up being fatal if detected during later stages. As a consequence, point-of-care devices have an increasing importance for routine screening applications in the health sector due to their portability, fast analyses and decreased cost. For that purpose, a multifunctional chip was developed and tested using gold nanoprobes to perform RNA optical detection inside a microfluidic chip without the need of molecular amplification steps. As a proof-of-concept, this device was used for the rapid detection of chronic myeloid leukemia, a hemato-oncological disease that would benefit from early stage diagnostics and screening tests. The chip passively mixed target RNA from samples, gold nanoprobes and saline solution to infer a result from their final colorimetric properties. An optical fiber network was used to evaluate its transmitted spectra inside the chip. Trials provided accurate output results within 3 min, yielding signal-to-noise ratios up to 9 dB. When compared to actual state-of-art screening techniques of chronic myeloid leukemia, these results were, at microscale, at least 10 times faster than the reported detection methods for chronic myeloid leukemia. Concerning point-of-care applications, this work paves the way for other new and more complex versions of optical based genosensors.

Infectious diseases remain one of the leading causes of morbidity and mortality worldwide. The WHO and CDC have expressed serious concern regarding the continued increase in the development of multidrug resistance among bacteria. Therefore, the antibiotic resistance crisis is one of the most pressing issues in global public health. Associated with the rise in antibiotic resistance is the lack of new antimicrobials. This has triggered initiatives worldwide to develop novel and more effective antimicrobial compounds as well as to develop novel delivery and targeting strategies. Bacteria have developed many ways by which they become resistant to antimicrobials. Among those are enzyme inactivation, decreased cell permeability, target protection, target overproduction, altered target site/enzyme, increased efflux due to over-expression of efflux pumps, among others. Other more complex phenotypes, such as biofilm formation and quorum sensing do not appear as a result of the exposure of bacteria to antibiotics although, it is known that biofilm formation can be induced by antibiotics. These phenotypes are related to tolerance to antibiotics in bacteria. Different strategies, such as the use of nanostructured materials, are being developed to overcome these and other types of resistance. Nanostructured materials can be used to convey antimicrobials, to assist in the delivery of novel drugs or ultimately, possess antimicrobial activity by themselves. Additionally, nanoparticles (e.g., metallic, organic, carbon nanotubes, etc.) may circumvent drug resistance mechanisms in bacteria and, associated with their antimicrobial potential, inhibit biofilm formation or other important processes. Other strategies, including the combined use of plant-based antimicrobials and nanoparticles to overcome toxicity issues, are also being investigated. Coupling nanoparticles and natural-based antimicrobials (or other repurposed compounds) to inhibit the activity of bacterial efflux pumps; formation of biofilms; interference of quorum sensing; and possibly plasmid curing, are just some of the strategies to combat multidrug resistant bacteria. However, the use of nanoparticles still presents a challenge to therapy and much more research is needed in order to overcome this. In this review, we will summarize the current research on nanoparticles and other nanomaterials and how these are or can be applied in the future to fight multidrug resistant bacteria.

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.

Mesenchymal stem cells are multipotent adult stem cells capable of generating bone, cartilage and fat, and are thus currently being exploited for regenerative medicine. When considering osteogenesis, developments have been made with regards to chemical induction (e.g. differentiation media) and physical induction (e.g. material stiffness, nanotopography), targeting established early transcription factors or regulators such as runx2 or bone morphogenic proteins and promoting increased numbers of cells committing to osteo-specific differentiation. Recent research highlighted the involvement of microRNAs in lineage commitment and terminal differentiation. Herein, gold nanoparticles that confer stability to short single stranded RNAs were used to deliver MiR-31 antagomiRs to both pre-osteoblastic cells and primary human MSCs in vitro. Results showed that blocking miR-31 led to an increase in osterix protein in both cell types at day 7, with an increase in osteocalcin at day 21, suggesting MSC osteogenesis. In addition, it was noted that antagomiR sequence direction was important, with the 5 prime reading direction proving more effective than the 3 prime. This study highlights the potential that miRNA antagomiR-tagged nanoparticles offer as novel therapeutics in regenerative medicine.

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Nanotechnology has become a powerful approach to improve the way we diagnose and treat cancer. In particular, nanoparticles possess unique features for enhanced sensitivity and selectivity for earlier detection of circulating cancer biomarkers. In vivo, nanoparticles enhance the therapeutic efficacy of anticancer agents when compared to conventional chemotherapy, improving vectorization and delivery, and helping to overcome drug resistance. Nanomedicine has been mostly focused on solid cancers due to take advantage from the enhanced permeability and retention (EPR) effect experienced by tissues in the close vicinity of tumors, which enhance nanomedicine’s accumulation and, consequently, improve efficacy. Nanomedicines for leukemia and lymphoma, where EPR effect is not a factor, are addressed differently from solid tumors. Nevertheless, nanoparticles have provided innovative approaches to simple and non-invasive methodologies for diagnosis and treatment in liquid tumors. In this review, we consider the state of the art on different types of nanoconstructs for the management of liquid tumors, from pre-clinical studies to clinical trials. We also discuss the advantages of nanoplatforms for theranostics and the central role played by nanoparticles in this combined strategy.