# Suppressing oncogenic RNA regulons using engineered zinc finger ribonucleases

> **NIH NIH R21** · UNIVERSITY OF MARYLAND BALTIMORE · 2021 · $216,686

## Abstract

Gene expression is extensively reprogrammed in cancer. These dysregulated genes include
subpopulations called RNA regulons that are coordinately regulated by posttranscriptional mechanisms at the
RNA level and control key features of tumor aggressiveness. To understand the molecular consequences of
dysregulated RNA regulons in cancer, the goal of this exploratory, high-risk/high-reward R21 proposal is to
develop a zinc finger-directed RNA-cleaving agent to suppress RNA regulons that are upregulated in many
tumors. Our prototype links the tandem zinc finger (TZF) domain from tristetraprolin (TTP) to the
endoribonuclease RNase4 (R4). In cells, the chimeric TZF-R4 protein is expected to bind and rapidly degrade
mRNA substrates of TTP, but our design will allow substrate specificity to be systematically modified. The TTP
TZF domain was chosen for our prototype because it has been evolutionarily selected to target an RNA
regulon containing AU-rich elements (AREs), which includes many mRNAs that encode regulators of the cell
cycle, angiogenesis, and metastasis. Furthermore, TTP expression or activity is frequently suppressed in
human cancers; in particular, low TTP levels in breast tumors are associated with poor patient prognosis.
 This proposal is aimed at providing the “proof of concept” that a TZF-R4 chimera can function as a guided
RNA degradation system in cancer cells to suppress a pro-tumorigenic RNA regulon and attenuate associated
tumor cell phenotypes. Purified TZF-R4 shows selective RNA recognition and cleavage in vitro, and
suppressed two known TTP substrate mRNAs when transiently transfected into cells. Furthermore, TZF-R4
dramatically slows cell proliferation when expressed in a clonal, stably-transfected breast cancer cell line.
Building upon these key preliminary results, two specific aims will be pursued. First, we will use transcriptome-
wide approaches to define the RNA regulon that is targeted and destabilized by TZF-R4 when stably
expressed in aggressive breast cancer cell models and demonstrate that TZF-R4 suppresses multiple mRNA
targets more efficiently than current technologies. Second, we will assess the impact of TZF-R4 on breast
cancer cell proliferation, stemness, invasion, migration, and in vivo tumor development.
 Several future applications of this technology are envisioned, including: (i) discovery tools for
characterizing RNA-mediated biological pathways, (ii) developing methods for promoting uptake of purified
TZF-R4 into cells, bypassing transfection and opening possibilities for direct in vivo administration of this
reagent, (iii) restoration or augmentation of TTP function to suppress tumor aggressiveness or inflammatory
signaling, and (iv) expanding the specificity of the TZF-R4 platform by altering its RNA-targeting specificity.
Strategies to broaden the scope include the iterative or combinatorial modification of the TZF moiety and
substitution of other RNA-binding domains to `guide' the chimeric protein, creat...

## Key facts

- **NIH application ID:** 10107292
- **Project number:** 1R21CA247647-01A1
- **Recipient organization:** UNIVERSITY OF MARYLAND BALTIMORE
- **Principal Investigator:** Gerald M. Wilson
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $216,686
- **Award type:** 1
- **Project period:** 2021-03-09 → 2024-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10107292

## Citation

> US National Institutes of Health, RePORTER application 10107292, Suppressing oncogenic RNA regulons using engineered zinc finger ribonucleases (1R21CA247647-01A1). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10107292. Licensed CC0.

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