# On-Demand Modulation of Extracellular Matrix Mechanics for Studying RhoA Activation in Primary and Metastatic Colorectal Cancer

> **NIH NIH R21** · UNIVERSITY OF WASHINGTON · 2024 · $428,361

## Abstract

PROJECT SUMMARY
Despite innovations in diagnosis and treatment, colorectal cancer (CRC) remains one the most common causes
of cancer-related deaths in the United States. During tumorigenesis, the extracellular matrix (ECM) drastically
stiffens, resulting in a self-amplifying feedback loop, furthering cancer progression; in CRC, increased matrix
elasticity is correlated with more advanced stages and worse treatment outcomes. Cells sense tissue stiffness
via integrins, which relay biochemical signals down many pathways, including the Rho/ROCK pathway. The
Rho/ROCK pathway has been shown to be highly dysregulated in many cancers, but its role in CRC is still highly
debated: some studies have shown elevated levels in metastatic lesions, whereas others have correlated
inactivation of this pathway with metastasis. Herein, we propose to study the mechanical contributions to RhoA
activation and how this may lead to increased proliferation and signaling, with a particular emphasis on
comparing the mechanical environments of the colon and the liver—the most common metastatic site, which has
been shown to be significantly stiffer than the primary tumor tissue. We will develop a novel hydrogel biomaterial
that can stiffen around cells from 1kPa (primary tumor) to 6kPa (liver metastatic tumor) upon exposure to 365nm
light over the course of the week, by employing photomediated oxime ligation, a chemistry we have pioneered
in our previous work for making photostiffening hydrogels as well as immobilizing full-length proteins in natural
and synthetic materials. To encode for subsequent cell release for downstream assays, we will develop a suite
of enzymatically sensitive peptide crosslinkers to enable bioorthogonal, user-defined hydrogel degradation. In
Aim 2, we will examine RhoA activation in primary patient-derived CRC tumor organoids as a function of
progressive stiffening of the matrix, and compare to levels of RhoA activation in metastatic cancer organoid lines
in stiff gels, via pull-down assays, Western blotting, and a lentivirally transduced biosensor construct. In Aim 3,
we will investigate the mechanical memory of RhoA activation in metastatic tumor organoid lines and whether is
reversable upon systematic softening of the matrix back to baseline, primary colon stiffnesses. We expect results
from these studies will result in novel insights into the role of matrix mechanics in CRC progression and
potentially open the door for biophysical routes of tumor resolution and treatment.

## Key facts

- **NIH application ID:** 10889787
- **Project number:** 1R21CA283686-01A1
- **Recipient organization:** UNIVERSITY OF WASHINGTON
- **Principal Investigator:** Cole A DeForest
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $428,361
- **Award type:** 1
- **Project period:** 2024-06-01 → 2026-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10889787

## Citation

> US National Institutes of Health, RePORTER application 10889787, On-Demand Modulation of Extracellular Matrix Mechanics for Studying RhoA Activation in Primary and Metastatic Colorectal Cancer (1R21CA283686-01A1). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10889787. Licensed CC0.

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