Silica nanoparticles and polyethyleneimine (PEI)-mediated functionalization: a new method of PEI covalent attachment for siRNA delivery applications.

Small-interfering RNA (siRNA) is a synthetic double-stranded RNA that consists of approximately 21 nucleotides (nts). It induces degradation of target mRNAs in a sequence-specific manner by the RNA interference (RNAi) mechanism. Thus, siRNAs offer a potential strategy for silencing mutated or defective genes that cause a variety of human diseases. The main obstacles of harnessing siRNAs as drugs are their inefficient delivery to cells and off-target effect making clinical applications very challenging. To address these issues, researchers have studied a variety of nanocarrier systems for siRNA delivery. This study presents the design, fabrication, and full characterization of innovative polyethyleneimine (PEI)-decorated polycationic 34.2 ± 4.2 nm silica (SiO2) NPs for siRNA-mediated gene silencing. More specifically, a new means of introduction (covalent mode of attachment) of the polycationic 25 kDa PEI polymer onto the SiO2 NP surface has been developed that makes use of an effective electrophilic double Michäel acceptor, divinyl sulfone (DVS). The resulting novel SiO2-PEI nanoparticles (SPEI NPs) have been fully characterized using a wide range of analytical, spectroscopic, and microscopic methods (TEM, DLS, ζ potential, elemental analysis (EA), XPS, TGA, and FTIR). Disclosing quite low cytotoxicity due to this unique mode of PEI covalent grafting, SPEI NPs/siRNA polyplexes have been successfully tested for the induction of gene silencing using dual-reporter luciferase transfected human osteosarcoma U2OS cells. The corresponding gene silencing data showed a clear correlation between PEI/siRNA ratios, siRNA concentration(s), and the level of gene silencing. Moreover, these SPEI NPs have been demonstrated to be thermodynamically stable with an ability to efficiently bind siRNAs and induce silencing for at least a one-year-long storage.

CD24 and APC Genetic Polymorphisms in Pancreatic Cancers as Potential Biomarkers for Clinical Outcome.

BACKGROUND: There are no validated biomarkers that correlate with the prognosis of pancreatic ductal adenocarcinoma (PDA). The CD24 and adenomatous polyposis coli (APC) genes are important in the malignant transformation of gastrointestinal cells. This study examined APC and CD24 genetic polymorphisms and their possible impact on survival of patients with PDA. METHODS: Clinical and pathological data as well as blood samples for extracting DNA were obtained for 73 patients with PDA. Real-time PCR assessed genetic variants of APC (I1307K and E1317Q), and four different single nucleotide polymorphisms (SNPs) in the CD24 gene: C170T (rs52812045), TG1527del (rs3838646), A1626G (rs1058881) and A1056G (rs1058818). RESULTS: The median age at diagnosis was 64 (41-90) years. Thirty-one patients (42.5%) were operable, 16 (22%) had locally advanced disease and 26 (35.5%) had disseminated metastatic cancer. The malignancy-related mortality rate was 84%. Median survival was 14 months (11.25-16.74). Survival was similar for wild-type (WT), heterozygous and homozygous variants of the APC or CD24 genes. The three most frequent CD24 SNP combinations were: heterozygote for A1626G and WT for the rest of the alleles (14% of patients), heterozygote for C170T, A1626G, A1056G and WT for the rest (14% of patients), and heterozygote for C170T, A1056G and WT for the rest (10% of patients). All patients were APC WT. The first two groups were significantly younger at diagnosis than the third group. CONCLUSIONS: Specific polymorphisms in the APC and CD24 genes may play a role in pancreatic cancer development. Correlation with survival requires a larger cohort.

Tetracycline nanoparticles as antibacterial and gene-silencing agents.

The spread of antibiotic-resistant bacteria and parasites calls for the development of new therapeutic strategies with could potentially reverse this trend. Here, a proposal is presented to exploit a sonochemical method to restore the antibiotic activity of tetracycline (TTCL) against resistant bacteria by converting the antibiotic into a nanoparticulate form. The demonstrated sonochemical method allows nanoscale TTCL assembly to be driven by supramolecular hydrogen bond formation, with no further modification to the antibiotic's chemical structure. It is shown that tetracycline nanoparticles (TTCL NPs) can act as antibacterial agents, both against TTCL sensitive and against resistant bacterial strains. Moreover, the synthesized antibiotic nanoparticles (NPs) can act as effective gene-silencing agents through the use of a TTCL repressor in Trypanosome brucei parasites. It is demonstrated that the NPs are nontoxic to human cells and T. brucei parasites and are able to release their monomer components in an active form in a manner that results in enhanced antimicrobial activity relative to a homogeneous solution of the precursor monomer. As the TTCL NPs are biocompatible and biodegradable, sonochemical formation of TTCL NPs represents a new promising approach for generation of pharmaceutically active nanomaterials.

Proteinaceous microspheres for targeted RNA delivery prepared by an ultrasonic emulsification method.

In the present work we used sonochemically prepared proteinaceous BSA spheres as a novel RNA-delivery system. The preparation of RNA-loaded BSA spheres was accomplished using an environmental friendly method termed the "ultrasonic emulsification method". It was demonstrated that ultrasonic waves do not cause the RNA chains to degrade and the RNA molecules remain untouched. The BSA-RNA complex was successfully introduced into mammalian (human) U2OS osteosarcoma cells and Trypanosoma brucei parasites. Using PVA coating of the RNA-BSA spheres we have achieved a significant increase in the number of microspheres penetrating mammalian cells. The mechanism of RNA encapsulation and the structure of the RNA-BSA complex are reported.