# Analysis of B-Raf and MPC1 mutations & their correlation with specific DNA methylation patterns using de-identified, FFPE clinically annotated colorectal-specific tumor samples.

> **NIH NIH R01** · IHC HEALTH SERVICES, INC. · 2020 · $290,000

## Abstract

Our long-term goal is to facilitate the development of new prevention and therapeutic measures for
colon cancer formation by understanding the earliest cellular perturbations that follow APC mutation. In
the previous funding periods, we revealed an innovative model wherein APC promotes enterocyte
differentiation by controlling retinoic acid biosynthesis. These findings now demonstrate that the
functions of APC are not limited to its well-established role in regulating canonical WNT signaling.
Mechanistically, we identified the transcriptional co-repressor, C-terminal binding protein (CtBP), as a
novel, APC-regulated protein that suppresses retinol dehydrogenases and intestinal cell differentiation
in concert with dysregulated LEF-1. Early adenomas taken from FAP patients showed elevated levels
of CtBP and LEF1 in comparison to adjacent, uninvolved tissues. Surprisingly, however, these same
sections showed little evidence of nuclear β-catenin indicating that loss of retinoic acid and cell
differentiation precedes nuclear accumulation of β-catenin. We further demonstrated that activation of
KRAS promoted the accumulation of nuclear β-catenin and subsequent intestinal cell proliferation. We
also described and uncovered an unexpected connection between APC and a novel DNA demethylase
system. This DNA demethylase is upregulated upon loss of APC and retinoic acid and maintains
intestinal cells in progenitor-like state. This failed differentiation precedes K-RAS induced proliferation
via β-catenin. It is currently unclear where signals for activating KRAS, and therefore, β-catenin
originate in the context of APC loss. Interestingly, adult zebrafish made deficient in retinoic acid
demonstrate both differentiation defects and a profound proliferation response that is coincident with
activation of inflammatory stromal cells. Our preliminary findings indicate that adult zebrafish
engineered to lack immune cells show intestinal differentiation defects, but no proliferative response
when made deficient in retinoic acid. We will now expand our studies to examine whether retinoic acid
plays roles in both epithelial cells and in intestinal stromal cells and whether interactions between these
two cell populations account for both initiation and progression of colon adenomas. We hypothesize
that APC controls the production of retinoic acid. Control of retinoic acid production in
intestinal epithelial cells regulates cell potential for differentiation by remodeling the epigenetic
landscape. In parallel, retinoic acid acts to suppress intestinal inflammatory responses that are
need to stimulate epithelial cell proliferation. This project will expand our understanding of how APC
and retinoic acid contribute to intestinal development and differentiation. This understanding could
support a testable clinical hypothesis aimed at pharmacological re-activation of retinoid signaling in
preventing and treating human colon cancers.

## Key facts

- **NIH application ID:** 10023165
- **Project number:** 5R01CA116468-16
- **Recipient organization:** IHC HEALTH SERVICES, INC.
- **Principal Investigator:** DAVID A JONES
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $290,000
- **Award type:** 5
- **Project period:** 2019-09-01 → 2021-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10023165

## Citation

> US National Institutes of Health, RePORTER application 10023165, Analysis of B-Raf and MPC1 mutations & their correlation with specific DNA methylation patterns using de-identified, FFPE clinically annotated colorectal-specific tumor samples. (5R01CA116468-16). Retrieved via AI Analytics 2026-05-28 from https://api.ai-analytics.org/grant/nih/10023165. Licensed CC0.

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