# Wedelolactone, a Novel TLR2 Agonist, Promotes Neutrophil Differentiation and Ameliorates Neutropenia: A Multi‐Omics Approach to Unravel the Mechanism

## TL;DR

Wedelolactone, a natural compound, helps treat neutropenia by boosting neutrophil production through a key immune pathway.

## Contribution

Identifies TLR2 as a novel target for neutropenia treatment and reveals WED's mechanism via TLR2/MEK/ERK signaling.

## Key findings

- Wedelolactone promotes neutrophil differentiation and bactericidal activity in vitro and in animal models.
- TLR2 and the MEK/ERK pathway are essential for WED's effects, confirmed through DARTS and functional assays.
- Inhibition of TLR2 or MEK/ERK reduces WED-induced neutrophil development and transcription factor expression.

## Abstract

Neutropenia, a common complication in cancer patients undergoing radiotherapy, heightens the risk of infection and mortality, with limited treatment options. This study investigates wedelolactone (WED), a natural coumarin, as a potential therapeutic agent. WED is found to promote neutrophil differentiation and enhance bactericidal function in vitro. Its in vivo efficacy is validated in radiation‐induced neutropenic mouse and zebrafish models, where it facilitates rapid recovery of leukocyte and neutrophil levels. An integrated approach using the GEO database, RNA sequencing, molecular docking, and Drug Affinity Responsive Target Stability (DARTS) assays identifies TLR2 and its downstream MAPK signaling pathway as essential for WED's anti‐neutropenic effects. DARTS confirms significant binding of WED to TLR2. Knockdown of TLR2 with siRNA or inhibition of TLR2 with C29 reduces WED‐induced neutrophil differentiation, MEK1/2 and ERK1/2 phosphorylation, and expression of transcription factors (PU.1, CEBPβ). Similarly, ERK1/2 inhibition by SCH772984 impairs WED‐induced neutrophil differentiation and bactericidal activity, decreasing PU.1 and CEBPβ expression without affecting TLR2 levels. These findings position TLR2 as a key therapeutic target for neutropenia, with WED effectively promoting neutrophil differentiation via TLR2 and MEK/ERK pathway activation. This study highlights the therapeutic potential of targeting TLR2 to alleviate neutropenia and underscores the utility of a multi‐omics approach in uncovering drug mechanisms.

Wedelolactone (WED), a natural TLR2 agonist, promotes neutrophil differentiation and enhances bactericidal function, offering a potential therapeutic strategy for neutropenia. Using a multi‐omics approach, this study reveals that WED activates the TLR2/MEK/ERK pathway, upregulating key transcription factors (PU.1, CEBPβ) to drive neutrophil development. These findings highlight WED's potential as a novel treatment for neutropenia.