# Internal Toxin Neutralizer for Treating STEC-infection

> **NIH NIH R21** · TEXAS A&M UNIVERSITY HEALTH SCIENCE CTR · 2020 · $231,000

## Abstract

Abstract
The Shiga toxin-producing E. coli (STEC) is the most common cause of bloody diarrhea and afflicts an
estimated 73,000 people in the US annually, causing significant morbidity. The most recent and largest STEC
outbreak occurred in Germany in 2011, affecting >3,800 people, including 54 deaths. Currently there is no
effective treatment for STEC infection. The pathology of STEC infection derives from two exotoxins – Shiga
toxin 1 (Stx1) and Shiga toxin 2 (Stx2) – that are secreted by STEC in the gut. Although antibiotic treatment can
reduce the load of STEC, it also augments Shiga toxin release, leading to increased risk of developing the more
serious hemolytic uremic syndrome (HUS) and kidney failure (up to 25%). Consequently, the CDC recommends
that antibiotics not be used in STEC patients and that only supportive therapy (e.g. oral and i.v. fluid, pain
control) be used. Although anti-toxin antibodies have been identified, the inability of antibodies to cross the cell
membrane renders them powerless against toxins already absorbed by the host cells, limiting their clinical
application. We hypothesize that a cytosol-accessible anti-toxin should be able to neutralize both extracellular
and intracellular Shiga toxin, leading to a much-prolonged therapeutic window and better therapeutic efficacy.
The overall goal of this study is to engineer a panel of intracellular toxin neutralizers (ITNs) against Shiga toxin
2 (Stx2). As a scaffold for the proposed ITN, we will use a designed ankyrin repeat protein (DARPin). DARPins
represent a versatile class of binding proteins that have been engineered to bind diverse targets with up to
picomolar affinity and possess low immunogenicity. In this project, we will first isolate DARPins that bind and
neutralize Stx2 (Aim 1). Concurrently, we will screen a panel of cell-penetrating peptides (CPPs) for their ability
to transport ITNs into cells (Aim 2). In Aim 3, we will assemble anti-Stx2 ITNs using the best anti-Stx2 DARPin
and CPP and evaluate the therapeutic potential of these anti-Stx2 ITNs in vitro and in vivo. The approach of
using ITN to combat toxins in circulation offers a new paradigm for the treatment of both STEC and non-STEC
bacterial infections.

## Key facts

- **NIH application ID:** 9886184
- **Project number:** 5R21AI137803-02
- **Recipient organization:** TEXAS A&M UNIVERSITY HEALTH SCIENCE CTR
- **Principal Investigator:** Zhilei Chen
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $231,000
- **Award type:** 5
- **Project period:** 2019-03-05 → 2023-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9886184

## Citation

> US National Institutes of Health, RePORTER application 9886184, Internal Toxin Neutralizer for Treating STEC-infection (5R21AI137803-02). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/9886184. Licensed CC0.

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