By Doug Macron
The National Institutes of Health this month earmarked more than $800,000 to fund three grants related to the delivery of siRNA-based therapeutics.
The first award went to Michigan State University's Stephen Walton to support a project focused on the general development of siRNA-design and -delivery approaches.
"Despite the intense study to date on the RNAi pathway and the use of siRNAs, the identification of the most active sequences and efficient delivery of those sequences to the cells of interest remain significant challenges," he wrote in his grant's abstract.
To address these issued, Walton aims to first characterize the binding interactions of siRNAs with key RNAi pathway proteins, the abstract states. He then plans to analyze "the interactions of siRNAs with delivery vehicles built from chemically diverse oligomeric and polymeric nanoparticles … to determine those structural features that encourage complex formation, protection of the siRNAs from degradation, and release of siRNAs upon entry into the cell.
"The polymeric nanoparticles to be studied are readily modified to provide a means of creating … a diverse array of vehicles that we will use to test variables such as amine density, polyethylene glycol modification, hydrophilicity, and binding cooperativity," it notes.
The grant, which is worth $235,202 in its first year, began on Sept. 1 and is set to run until the end of August 2014.
The second grant was awarded to Ernest Ramsay of the Medical University of South Carolina to fund his work on delivery vehicles for an siRNA-based pancreatic cancer treatment.
Camp's colleagues previously identified the cancer-associated Sm-like oncogene as a potential molecular target for pancreatic cancer, finding that its inhibition was able to slow tumor progression in mouse models of the disease.
"Recently, nanoparticle liposome-based complexes targeting the transferrin receptor single chain antibody fragment have been used to specifically target tumors in gene therapy," he wrote in his grant's abstract. "By combining these concepts, we hypothesize that CaSm functions as a master switch to destabilize multiple gene transcripts, contributing to the malignant phenotype observed in pancreatic cancer [that] will be effectively targeted by CaSm siRNA delivered by a novel transferrin-targeted nanovector system."
With the NIH funding, Camp aims to "define the critical molecular pathways involved in CaSm-mediated oncogenesis in pancreatic cancer … [and] establish the role of CaSm gene therapy in pancreatic cancer and serve as the foundation of future clinical trials in patients with pancreatic cancer."
His grant began on Sept. 1 and is slated to run through August 2015. It is worth $173,448 in its first year.
The final grant went to Peter Brink, a researcher at the State University of New York, Stony Brook, to support his research into the cellular delivery of siRNAs via gap junctions.
"Our previous studies have shown that gap junctions composed of connexin43 are permeable to siRNAs, and permeating siRNAs can subsequently reduce the mRNA levels of a specific gene," he wrote in the grant's abstract. However, gap junctions composed of Cx32 or Cx26 will not transfer siRNAs.
With the NIH award, Brink and colleagues will determine the permeability of gap junctions made of connexin40, connexin37 and connexin43, which are ubiquitously expressed in vivo in many organs, to siRNAs.
The team then aims to test the hypothesis that cellular delivery of siRNA via gap junction channels can silence the function of the pacemaker channel HCN2 in target cells by "characterizing the functional silencing of HCN2 via gap junction-mediated delivery of siRNA from [human mesenchymal stem cells] or other communication competent cells to a target cell expressing HCN2," according to the abstract.
"We follow HCN2 mRNA concentration using RT-PCR to allow an estimate of the relative content over time in the presence of siRNA," it adds. "We will also determine the concentrations of tagged … siRNAs to establish the effective concentration necessary to silence a gene, and also provide parameters for our 2D/3D model to determine penetration within a tissue."
Brink then plans to assess how far siRNAs penetrate multiple layers of a syncytium, and to derive a model for the transfer of siRNAs "along a simple linear chain of cells or geometries in two or three dimensions," which will help predict the number and position of siRNA-contain cells necessary to silence a target gene in a tissue or organ.
Lastly, the investigators will move in vivo, injecting GFP-containing cells intradermally or intramuscularly into nude mice, followed by human mesenchymal stem cells loaded with GFP-targeting siRNAs. "We will track the GFP fluorescence image over time using whole animal imaging," Brink notes in the abstract.
The grant began on Sept. 1 and will run until Aug. 31, 2014. It is worth $412,849 in its first year.