Polyamide Backbone Modified Cell Targeting and Penetrating Peptides in Cancer Detection and Treatment.

Cell penetrating and targeting peptides (CPPs and CTPs) encompass an important class of biochemically active peptides owning the capabilities of targeting and translocating within selected cell types. As such, they have been widely used in the delivery of imaging and therapeutic agents for the diagnosis and treatment of various diseases, especially in cancer. Despite their potential utility, first generation CTPs and CPPs based on the native peptide sequences are limited by poor biological and pharmacological properties, thereby restricting their efficacy. Therefore, medicinal chemistry approaches have been designed and developed to construct related peptidomimetics. Of specific interest herein, are the design applications which modify the polyamide backbone of lead CTPs and CPPs. These modifications aim to improve the biochemical characteristics of the native peptide sequence in order to enhance its diagnostic and therapeutic capabilities. This review will focus on a selected set of cell penetrating and targeting peptides and their related peptidomimetics whose polyamide backbone has been modified in order to improve their applications in cancer detection and treatment.

Bifunctional Au-templated RNA nanoparticles enable direct cell uptake detection and GRP75 knockdown in prostate cancer.

Nucleic acids templated on gold (Au) surfaces have led to a wide range of functional materials ranging from microarrays, sensors and probes in addition to drug delivery and treatment. In this application, we describe a simple and novel method for templating amino-functionalized RNA onto Au surfaces and their self-assembly into small, discrete nanoparticles. In our method, sample hybridization with a complementary RNA strand with and without a fatty acid (palmitamide) appendage produced functionalized double-stranded RNA on the Au surface. The resulting Au-functionalized RNA particles were found to be stable under reducing conditions according to UV-Vis spectroscopy. Sample characterization by DLS and TEM confirmed self-assembly into primarily small (∼10-40 nm) spherical shaped nanoparticles expected to be amenable to cell biology. However, fluorescence emission (λexc: 350 nm, λem: 650 nm) revealed radiative properties which limited cell uptake detection. Introduction of FITC within the Au-functionalized RNA particles produced a bifunctional probe, in which FITC fluorescence emission (λexc: 494 nm, λem: 522 nm) facilitated cell uptake detection, in a time-dependent manner. The dual encapsulation-release profiles of the FITC-labeled Au-functionalized RNA particles were validated by time-dependent UV-Vis spectroscopy and spectrofluorimetry. These experiments respectively indicated an increase in FITC absorption (λabs: 494 nm) and fluorescence emission (λem: 522 nm) with increased sample incubation times, under physiological conditions. The release of Au-functionalized siRNA particles in prostate cancer (PC-3) cells resulted in concomitant knockdown of GRP75, which led to detectable levels of cell death in the absence of a transfection vector. Thus, the formulation of stable, small and discrete Au-functionalized RNA nanoparticles may prove to be valuable bifunctional probes in the theranostic study of cancer cells.

Direct Transfection of Fatty Acid Conjugated siRNAs and Knockdown of the Glucose-Regulated Chaperones in Prostate Cancer Cells.

The emerging field of RNAi nanotechnology has led to rapid advances in the applications of siRNAs in chemical biology, medicinal chemistry, and biotechnology. In our RNAi approach, bioconjugation of linear, V-, and Y-shaped RNA templates were designed using a series of saturated and unsaturated fatty acids to improve cell uptake and knockdown efficacy of the oncogenic glucose regulated proteins (GRPs) in prostate (PC-3) cancer cells. An optimized HCTU-coupling procedure was developed for tagging variable saturated and unsaturated fatty acids onto the 5'-ends of linear and V-shaped RNA templates that were constructed by semiautomated solid phase RNA synthesis. Hybridization and self-assembly of complementary strands yielded linear, V-, and Y-shaped fatty acid-conjugated siRNAs which were characterized by native PAGE. CD spectroscopy confirmed their A-type helix conformations. RP IP HPLC provided trends in amphiphilic properties, whereas DLS and TEM confirmed multicomponent self-assembled structures that were prone to aggregation. Subsequently, the fatty acid conjugated siRNA bioconjugates were tested for their RNAi activity by direct transfection within PC-3 cells known to overexpress oncogenic GRP activity. The siRNA bioconjugates with sense strand modifiers provided more potent GRP knockdown relative to the antisense modified siRNAs, but to a lesser extent when compared to the unconjugated siRNA controls that were transfected with the commercial Trans-IT X2 dynamic delivery system. Flow cytometry revealed that the latter may be at least in part attributed to limited cell uptake of the fatty acid conjugated siRNAs. Nonetheless, these new constructs represent an entry point in modifying higher-order siRNA constructs that may lead to the generation of more efficient siRNA bioconjugates for screening important oncogene targets and for cancer gene therapy applications.