# Development of an in vitro system to study heme-induced caspase activation

> **NIH NIH R03** · BAYLOR COLLEGE OF MEDICINE · 2024 · $80,250

## Abstract

PROJECT SUMMARY
Excessive intravascular hemolysis of red blood cells (RBCs) leads to numerous complications associated with
uncontrolled inflammation including pain, vaso-occlusion, and organ damage that underlie the pathophysiology
of several disease states including sepsis, intravascular hemorrhage, and sickle cell disease (SCD). The primary
mediator of this inflammation is heme released from damaged RBCs. The applicant’s goup recently reported
that heme activates the inflammatory caspases, caspase-1, caspase-4, and caspase-5 to coordinate the
cleavage and release of the apical pro-inflammatory cytokine IL-1β. Importantly, unlike caspase-1, caspase-4
and caspase-5 are activated independently of inflammasome assembly. Thus, heme is one of the first examples
of a DAMP (damage associated molecular pattern) that activates the non-canonical inflammasome pathway.
The applicant has shown that heme binds caspase-4 and caspase-5 and induces oligomerization of these
caspases. The central hypothesis is that heme directly binds to caspase-4 and caspase-5 to induce
oligomerization and activation. The hypothesis will be tested by the following Aims: 1) to determine the
biophysical composition of heme-induced inflammatory caspase complexes and 2) to identify the protein
components required for heme-induced inflammatory caspase activation. Under the first Aim full length
recombinant caspase proteins will be produced and the ability of heme to bind caspases and to induce caspase
oligomerization and cleavage will be assessed. Under the second aim the amino acid residues required for heme
binding, oligomerization and cleavage will be identified as well as the type of cytosolic factors, if any, contribute
heme-induced caspase activation. This work will provide an in vitro assay that will be the basis for future rational
drug design approaches and a source of purified caspases that can be used for future studies to determine the
structures of the heme-induced caspase complexes. Understanding the mechanisms of inflammatory caspase
activation with provide justification for targeting these proteins to improve clinical outcomes in hemolytic
conditions.

## Key facts

- **NIH application ID:** 10849821
- **Project number:** 5R03AI178641-02
- **Recipient organization:** BAYLOR COLLEGE OF MEDICINE
- **Principal Investigator:** Lisa Bouchier-Hayes
- **Activity code:** R03 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $80,250
- **Award type:** 5
- **Project period:** 2023-06-01 → 2025-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10849821

## Citation

> US National Institutes of Health, RePORTER application 10849821, Development of an in vitro system to study heme-induced caspase activation (5R03AI178641-02). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10849821. Licensed CC0.

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