Project 3 - Nanomaterials for gene imaging and regulation in lung injury
Project Leader - Steven L. Brody, M.D.
The goal of this Project is to develop multifunctional NPs optimized for intracellular delivery of payloads that target genetic elements for therapeutic modulation and diagnostic imaging of gene expression in lung injury.
Acute lung injury is a common clinical problem with high mortality. Lung injury is caused by trauma, infection, allergic and other stimuli that result in a complex inflammatory environment, marked by increased inflammatory cells, cytokines, and other mediators, including NO. In lung injury, NO is produced primarily by iNOS. With marked injury, iNOS can drive an over-exuberant response. Importantly, genetic and pharmacologic models of iNOS inhibition alter lung injury and inflammation. One source of high levels of iNOS is alveolar macrophages, where downregulation ameliorates lung injury. Thus, we have investigated the use of NPs loaded with siRNA or PNA to bind iNOS mRNA to modulate and image acute lung injury. Development of NPs for targeting this molecule will open the door for the use of NPs to carry payloads for monitoring and regulating the expression of other genes identified as biomarkers or therapeutic targets in lung injury.
We hypothesize that functionalized NPs provide:
To accomplish our goal, we have demonstrated: Efficient intracellular delivery in lung epithelial cells and macrophages after a single bolus intratracheal injection of SCKs labeled with the protein transduction domain, TAT (SCK-TAT) or PEGylated cationic SCKs (cSCK-5kDaPEG) (Figure); enhanced cell entry via PEGylation of cSCKs; validation of PNA and siRNA sequences that bind and suppress iNOS; functional payload release of PNA bound to SCKs and cSCKs; low immune responses in mice (for SCKs) and methods for tracking particle fate in vitro and in vivo.
- A therapeutic strategy for decreasing inflammation and injury in lung disease models by carrying RNA inhibitors (siRNA and PNA);
- A diagnostic probe when integrated with peptide nucleic acids (PNA) to bind specific mRNA to image mRNA in vivo as a “molecular signature”.
We, therefore, propose the following specific aims:
- Characterize and optimize the features of NPs required for efficient, non-toxic, intracellular delivery of PNA and siRNA;
- Use NPs to deliver PNAs and siRNAs in vivo to image and modulate transcription of iNOS in models of lung injury.