Abstracts

American Thoracic Society 2025 International Conference; https://www.atsjournals.org/doi/abs/10.1164/ajrccm.2025.211.Abstracts.A3378

Abstract

Rationale: Airway neutrophilia is associated with several lung diseases, including acute lung injury, severe asthma, and chronic obstructive pulmonary disease (COPD). RLS-0071 (Pegtarazimod), a novel dual-targeting anti-inflammatory peptide, has been shown to reduce airway neutrophilia in a human endotoxin challenge model (Cunnion et al., ERJ Open 2024). ChipCytometry allows quantification of airway cells including intracellular staining of effector molecules. This study investigates the effect of RLS-0071 on airway neutrophils following inhaled endotoxin challenge by ChipCytometry.

Methods: RLS-0071 at two doses versus placebo was given in a randomized, double-blind, proof-of-mechanism study following inhaled endotoxin (LPS) challenge in 30 healthy participants (NCT05351671). Induced sputum (IS) was collected before, 6 and 24 hours (h) after LPS challenge. IS cells (250,000) were loaded on ChipCytometry-specific slides according to the manufacturer protocol. Cell surface markers (CD45, CD66b, CD14, CD3, CD11a, CD11b, CD89, CD15, CD62L, CD49d, CD35, CD16) and intracellular effector molecules (myeloperoxidase (MPO), neutrophil elastase (NE), eosinophil peroxidase (EPX)) were stained with corresponding antibodies and then analyzed by ChipCytometry.

Results: RLS-0071 treatment led to decreased neutrophil numbers in IS at 6h (placebo: 13.0±4.8, low-dose: 4.9±9.6, high-dose: 6.8±3.5 x106/g) and 24h (placebo: 7.9±5.8, low-dose: 3.8±8.9, high-dose: 6.2±9.1 x106/g) compared to placebo. Neutrophils from RLS-0071-treated participants showed a tendency towards reduced expression of neutrophil activation markers CD89, CD11a and CD11b at 6h compared to placebo. Intracellular levels of NE and MPO in neutrophils were decreased at 6h in both treatment groups compared to placebo (NE: p < 0.01 for both doses; MPO: p = 0.06 and 0.01 for low- and high-dose, respectively), and tended to be reduced at 24h.

Conclusion: ChipCytometry data confirm and extend previous proof-of-mechanism that RLS-0071 decreases neutrophil numbers in IS and acts on neutrophil activation and adhesion. In line to previous findings of reduced MPO and NE concentrations in sputum supernatant, we found decreased intracellular staining of MPO and NE in neutrophils. Overall, RLS-0071 demonstrates potential as a novel anti-inflammatory treatment for neutrophil-dominated lung diseases by reducing neutrophil activity and infiltration into the lungs.