# Fluorescent gammaPNA Miniprobes for Imaging Telomeric RNA

> **NIH NIH R21** · CARNEGIE-MELLON UNIVERSITY · 2022 · $226,898

## Abstract

Project Summary
Altered expression of non-coding RNAs are hallmarks of many cancers. One unique characteristic of telomerase-
free cancer cells that use an alternative lengthening of telomere (ALT) pathway for telomere maintenance is the
overexpression of telomere repeat-containing RNA (TERRA). TERRA is therefore an attractive target for ALT
cancer diagnosis and therapy. TERRA is transcribed from a subset of telomeres but forms foci on other telomeres
and non-telomere loci. Manipulation of TERRA binding proteins leads to mis-regulation in TERRA localization
and telomere dysfunction, suggesting TERRA function is highly linked to its localization. Visualizing TERRA in
fixed tissue and live cells would therefore provide insights needed for the development of TERRA as a biomarker
for ALT diagnosis or a target for ALT cancer therapy. Current methods for labeling TERRA suffer from either
high background, low efficiency (only label bright TERRA foci), or low selectivity (cannot differentiate TERRA
from telomere DNA). Our preliminary data suggest that fluorescent gPNA miniprobes can detect TERRA in fixed
cells with high efficiency. This is because the high affinity of gPNA allows us to use shorter probes, leading to
hybridization of more probes per TERRA and therefore improving the brightness of TERRA foci. This improved
brightness will also allow detection of shorter TERRA transcripts than is possible with conventional DNA TERRA
FISH probes. In this work, we aim to further improve this method for TERRA visualization in fixed cells and
extend the capacity into live cell imaging. In Aim 1, we will systematically vary gPNA probe length to achieve high
selectivity in labeling TERRA vs telomeric DNA, following up on promising preliminary data indicating that a 9mer
gPNA can accomplish this whereas a 12mer cannot. In Aim 2, we will achieve low background in TERRA labeling
by designing fluorescent gPNA pairs that will emit signal through Förster Resonance Energy Transfer (FRET)
only when they bind to TERRA close to each other. The unbound gPNA probes will not FRET and thus exhibit
low background. In Aim 3, we will use a synthetic cationic polymer vector to deliver gPNA to live cells to track
TERRA dynamics in real time. The optimized gPNA probes that detect TERRA with higher efficiency, high
selectivity and low background can be used to detect TERRA in fixed tissue to aid ALT cancer therapy. The
ability to monitor TERRA dynamics in live cells can be used to understand how TERRA contributes to telomere
elongation and other cellular functions in ALT cancer cells and inspire novel therapeutic strategies targeting
TERRA. Beyond TERRA, the design principles can be used to visualize other non-coding RNAs linked to various
cancers, particularly those implicated in dynamic processes such as chromatin organization that are challenging
to study with current methods. Simultaneously, by designing a library of probes for different non-coding RNAs,
our methods can be used t...

## Key facts

- **NIH application ID:** 10358270
- **Project number:** 1R21CA267260-01
- **Recipient organization:** CARNEGIE-MELLON UNIVERSITY
- **Principal Investigator:** Bruce A. ARMITAGE
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $226,898
- **Award type:** 1
- **Project period:** 2022-03-21 → 2025-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10358270

## Citation

> US National Institutes of Health, RePORTER application 10358270, Fluorescent gammaPNA Miniprobes for Imaging Telomeric RNA (1R21CA267260-01). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10358270. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*