By Doug Macron
The National Institutes of Health last month handed out nearly $1.3 million in grant funding to support four research projects focused on developing siRNA-delivery technologies.
The first grant went to Albert Einstein School of Medicine's Deborah Palliser to fund her efforts to develop an siRNA-based microbicide to combat herpes virus infection.
In 2006, Palliser and colleagues at Harvard Medical School reported that vaginally delivered siRNAs targeting herpes simplex virus 2 genes or host-encoded viral entry receptor genes could protect mice from the infection.
“These findings were encouraging, but we encountered issues that would need to be addressed for siRNAs to be developed for clinical use,” she wrote in her grant's abstract. “Problems included toxicity of a cationic lipid used to complex the siRNAs, transient protection conferred by virus-specific siRNAs, and siRNA instability in vaginal washes.”
The team was able to address these limitations in follow-up work, and with the NIH grant she aims to further enhance the siRNA microbicide's gene-silencing ability in order to optimize it for clinical applications.
“Using the murine HSV-2 infection model we will determine whether … immune responses, induced either by the siRNA or through viral challenge affect siRNA-mediated protection; [if] enhanced incorporation of siRNA into RISC can result in more potent or durable protection; and [if] efficient siRNA uptake and intracellular localization in the vaginal mucosa requires the expression of a specific receptor,” according to the abstract.
The grant runs from March 1 until Feb. 28, 2013, and is worth $416,667.
The second and third grants were awarded to Northwestern University's Amy Paller to support her work developing topical siRNA delivery vehicles for the treatment of epidermal hyperplasia and diabetic wound healing, respectively.
“Our laboratories have engineered a novel nanoparticle conjugate utilizing siRNA duplexes that are densely packed on the surface of gold nanoparticles, and which demonstrates a surprising ability to transit the mouse and human stratum corneum and suppress two tested targets, green fluorescent protein and the epidermal growth factor receptor,” she wrote in her first grant's abstract.
Notably, no toxicity has been observed when the nanoparticles were delivered either topically or intravenously at levels far greater than required for gene knockdown, she noted.
Through the grant project, she and her colleagues plan to identify the mechanism by which the nanoparticles penetrate into human skin in order to help optimize the delivery approach.
“Next, we will assess the ability of [the nanoparticles] to suppress Ras signaling and reverse epidermal hyperplasia in a mouse model of skin-specific, inducible over-expression of H-Ras,” Paller wrote. “Building on our in vitro and in vivo mouse studies … we will test if topical application of [the siRNA-loaded nanoparticles] to a human transplanted skin model of Ras over-expression will similarly suppress aberrant Ras signaling to cause clinically and histologically detectable epidermal normalization.”
The grant runs from March 1 until the end of February 2016. It is worth $342,174 in its first year.
Paller was also awarded an NIH grant to develop the siRNA/gold nanoparticles for the treatment of wound healing.
According to that grant's abstract, ganglioside GM3, a sialylated membrane glycosphingolipid, is a “critical mediator of insulin resistance,” and Paller's lab has shown that it accumulates in keratinocyte membranes in diabetic mice. Further, depletion of GM3 reverses a wound-healing defect in the animals.
“We will first evaluate the efficacy and safety of topically-applied GM3 synthase [siRNA nanoparticles], which deplete gangliosides, in accelerating healing in diabetic mouse models,” she wrote in the abstract.
Her team will also evaluate the impact of gangliosides on keratinocyte mortality, as well as on the activation of insulin receptor, insulin-like growth factor-1 receptor-integrin, and epidermal growth factor receptor, all of which impact wound healing.
“Finally, we will evaluate the impact of GM3 depletion on glucose-induced insulin resistance,” the abstract states. “Reversal of the wound healing defect in obese diabetic mice by topical administration of our nanoparticle-conjugated nucleic acid inhibitors of ganglioside synthesis will be an innovative means to promote wound healing in chronic wounds.”
Paller's second grant runs from March 1 until Feb. 28, 2014. It is worth $205,761 in its first year.
The final grant awarded by the NIH went to Colorado State University's Mark Zabel to help support his research into liposome/siRNA/peptide complexes for the treatment of transmissible spongiform encephalopathies such as Creutzfeldt-Jakob disease.
According to the grant's abstract, PrPRES, a proteinase K-resistant form of the normal prion protein, “appears central to the pathogenesis of these diseases,” and experimental studies by other groups have used lentivirus-expressed shRNAs to “suppress expression of a normal host protein, PrPC, in neurons and subsequent prion neuropathology, exploiting the phenomenon that disease severity and progression correlate with the amount of PrPC present in neurons.”
Still, these efforts have been hampered by modest efficacy and an invasive delivery method, as well as concerns over the irreversibility of PrPC suppression.
“We have recently developed a novel small interfering RNA delivery system using liposome/siRNA/peptide complexes to protect siRNAs from serum degradation, deliver siRNA specifically to neurons, and knockdown PrPC sufficiently to cure prion disease in two cell culture models,” Zabel wrote in the abstract.
With the NIH funding, he aims to extend these findings and cure two distinct prion diseases, scrapie and chronic wasting disease, in mice using the siRNA complexes delivered intravenously and into the neurons of the brain.
The grant runs from March 1 until Feb. 28, 2017, and is worth $317,719 in its first year.
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