# Multiplexed, Continuous Reporting of Gene Expression via CRISPR-mediated Transcriptional Activation

> **NIH NIH R61** · YALE UNIVERSITY · 2024 · $234,919

## Abstract

PROJECT SUMMARY:
Although the detection of gene expression by quantitative PCR, bulk or single cell RNA sequencing represents
a cornerstone of biological inquiry, these approaches are limited in their ability to measure transcriptional
dynamics over time and throughout the spatial heterogeneity of tissues, and to detect low-abundance
transcripts including those from cytokines. Cytokine expression shapes the infiltration and activity of immune
cells in tumors and tissues and represents an important correlate of response to immunotherapy. However,
cytokine expression is context-dependent, pleiotropic, dynamic and asynchronous. As such, it is particularly
poorly suited to single ‘snapshot’ measurements in time, or the detection of a single or even a few
inflammatory mediators in isolation. To address these challenges, we will combine advances in CRISPR-
mediated transcriptional activation technology with genetically-encoded set reporting, to fluorescently report
the transcriptional activity of 9 or more cytokines simultaneously, dynamically and in situ in tissues. We have
previously validated the core capability of our system to report single gene expression in preliminary studies
and will now optimize its detection capabilities and implement a Cre-Lox-based randomized fluorescent
reporter set-encoding strategy to multiplex detection. If successful, our study will establish a novel approach to
multiplexed and high-sensitivity CRISPR-based detection of gene expression. It will also produce tools and
reagents that can be directly applied to interrogate the basis for tumor microenvironment inflammation and can
be adapted for functional genomic and in vivo studies.

## Key facts

- **NIH application ID:** 10918789
- **Project number:** 1R61CA291201-01
- **Recipient organization:** YALE UNIVERSITY
- **Principal Investigator:** Jeffrey J. Ishizuka
- **Activity code:** R61 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $234,919
- **Award type:** 1
- **Project period:** 2024-09-01 → 2027-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10918789

## Citation

> US National Institutes of Health, RePORTER application 10918789, Multiplexed, Continuous Reporting of Gene Expression via CRISPR-mediated Transcriptional Activation (1R61CA291201-01). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10918789. Licensed CC0.

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