GammaPNA Miniprobes for Telomere Analysis and RNA FISH

DESCRIPTIONprovided by applicantThe proposed Phase II project will continue development of gammaPNA miniprobe technology originally developed at Carnegie Mellon University and the University of Pittsburgh and subsequently transferred to PNA InnovationsInca small business spun out of Carnegie Mellon UniversityThe basis of gammaPNA miniprobes is the high affinity with which gammaPNA hybridizes to complementary DNAThe specific applications addressed in this proposal are telomere analysiswhich is currently done using fluorescent PNA probesbases in lengthwhich hybridize toconsecutive repeats of the human telomere sequenceAATGGGand mRNA labeling by complementary fluorescent probesIn Phase Iwe demonstrated that the higher affinity of gammaPNA allows shorterbase telomere probes to be usedresulting in more fluorescent dyes being delivered to a telomere of a given lengthThis allow more reliable analysis of the shortesti ecritically shorttelomereswhich are implicated in a variety of conditions including aging related diseases and cancerWe published a paper describing our results in Organic and Biomolecular Chemistry and we launched a marketing campaign around our Telo MiniprobesTMleading to our first sales within this product lineThe proposed research will have four Specific AimsThe first aim extends our Phase I work in three waysFirst we will study several additional cell lines This will help us toadetermine the range of variability in miniprobe performance andbpotentially identify other versions of the miniprobe that work in varied cell linesSecondwe wil develop miniprobes that target the C rich telomere strandThirdwe will build on promising preliminary results for synthesizing internally labeled miniprobes that will double or triple the brightness of our current best probeThe second and third aims are directed toward new applicationsspecifically in development of a high throughput telomere assay based on our miniprobes and testing of fresh and archived tissue sampleswhich are currently difficult to study by FISH due to autofluorescenceThe fourth aim will significantly extend gammaPNA FISH probes into RNA labelingWe will synthesize sets of gammaPNAs targeted to different sites on a single mRNAbut rather than covalently label the probeswhich is costlywe will use an innovative and economical co hybridization approach to label our probesThe synthesis of the gammaPNA monomers and oligomers will be done at PNA InnovationsBiophysical characterization and telomere staining will be done at academic laboratories at Carnegie Mellon and the University of Pittsburgh where the gammaPNA miniprobe technology was inventedOptimized miniprobes will then be sent to independent beta testing laboratories that currently use conventional PNA probes for telomere analysis or DNA based molecular beacons for mRNA labeling