Oligonucleotide

The non-nucleoside analog gemcitabine has been the standard of care for treating pancreatic cancer. The drug shows good potency in pancreatic cancer cells in vitro but, due to poor bioavailability, requires administration in large doses by infusion and this systemic exposure results in significant toxicity for the patient. Genes have been identified that, when silenced by siRNA, synergize with gemcitabine treatment and offer a means of reducing the gemcitabine dosage required for efficacy. However, benefiting from the synergism between the two agents requires that the gemcitabine and siRNA penetrate the same cells. To ensure co-delivery, we incorporated gemcitabine covalently within siRNAs against targets synergistic with gemcitabine (CHK1 or RAD17). We demonstrated that specific bases within an siRNA can be replaced with gemcitabine to increase efficacy. The result is a single drug molecule that simultaneously co-delivers gemcitabine and a synergistic siRNA. The siRNA–gemcitabine constructs demonstrate a 5–30-fold improvement in potency compared with gemcitabine alone. Co-delivering a CHK1 siRNA–gemcitabine construct together with a WEE1 siRNA resulted in a 10-fold improvement in IC50 compared with gemcitabine alone. These constructs demonstrate efficacy across a wide array of pancreatic tumor cells and may represent a novel therapeutic approach for treating pancreatic cancer.

Neurotoxic organophosphates (OP) have found widespread use in the environment for insect control. In addition, there is the increasing threat of use of OP based chemical warfare agents in both ground based warfare and terrorist attacks. Together, these trends necessitate the development of simple and specific methods for discriminative detection of ultra low quantities of OP neurotoxins. In our previous investigations a new biosensor for the direct detection of organophosphorus neurotoxins was pioneered. In this system, the enzymatic hydrolysis of OP neurotoxins by organophosphate hydrolase (OPH) generated two protons in each hydrolytic turnover through reactions in which P–X bonds are cleaved. The sensitivity of this biosensor was limited due to the potentiometric method of detection. Recently, it was reported that a change in fluorescence properties of a fluorophore in the vicinity of gold nanoparticles might be used for detection of nanomolar concentrations of DNA oligonucleotides. The detection strategy was based on the fact that an enhancement or quenching of fluorescence intensity is a function of the distances between the gold nanoparticle and fluorophore. While these reports have demonstrated the use of nanoparticle-based sensors for the detection of target DNA, we observed that the specificity of enzyme–substrate interactions could be exploited in similar systems. To test the feasibility of this approach, OPH-gold nanoparticle conjugates were prepared, then incubated with a fluorescent enzyme inhibitor or decoy. The fluorescence intensity of the decoy was sensitive to the proximity of the gold nanoparticle, and thus could be used to indicate that the decoy was bound to the OPH. Then different paraoxon concentrations were introduced to the OPH–nanoparticle–conjugate–decoy mixtures, and normalized ratio of fluorescence intensities were measured. The greatest sensitivity to paraoxon was obtained when decoys and OPH–gold nanoparticle conjugates were present at near equimolar levels. The change in fluorescence intensity was correlated with concentration of paraoxon presented in the solution.

Childhood Philadelphia chromosome positive (Ph(+)) acute lymphoblastic leukemia (ALL) has a poor prognosis. Because leukemia cell burden is reduced but not eradicated by polychemotherapy, improved treatment strategies should enhance those immune mechanisms responsible for the maintenance of complete remission. The aim of this study was to evaluate the protection of mice challenged with the syngeneic Ph(+) ALL cell line BM185 using genetically modified leukemia cell vaccines and immunomodulating oligonucleotides. Because retroviral vectors are ineffective at transducing nondividing primary cells from human hematopoietic malignancies, we first evaluated nonviral techniques (electroporation and ballistic transfer) using minimalistic immunogenically defined gene expression vectors to generate B7.1 or granulocyte macrophage colony-stimulating factor (GM-CSF)-expressing BM185 cells. Subsequently, protective vaccination experiments with these cells were performed in a leukemia challenge mo...

The bimetallic core–shell nanoparticles show unique plasmonic properties and their preparations and characterizations are currently under investigation. A new type of Au core–Ag shell (Au@Ag) nanoparticles is prepared by sandwiching the chemically attached Raman reporter molecules (RRMs) and a 12-base-long oligonucleotide between the 13 nm average size core-gold nanoparticles (AuNPs) and 9 nm and 21 nm average size of Ag shell. The synthesized Au@Ag nanoparticles are tested for their surface-enhanced Raman scattering (SERS) performance. It is found that the chemical attachment of the oligonucleotides along with the RRM improved the enhancement in Raman scattering more than one order of the magnitude with the Au@Ag nanoparticles with an average 9-nm shell thickness while the Au@Ag nanoparticles with 21 nm average shell thickness have poor SERS activity. A minimum enhancement factor of 1.0 × 107 is estimated for the SERS active oligonucleotide-mediated Au@Ag nanoparticles. The approach may provide new routes for preparation of highly sensitive new generation of bimetallic core–shell nanoparticles.