ENAMPT neutralization reduces preclinical ARDS severity via rectified NFkB and Akt/mTORC2 signaling

Tadeo Bermudez,Matthew Hufford,Alexander N Garcia,Yansong Li,Nancy G Casanova,Jessica Burt,Jin H Song,Vivian Reyes Hernon,Viswanathan Natarajan,Ankit A Desai,Ruslan Rafikov,Christian Bime,Saad Sammani,Donna D Zhang,Jeffrey R Jacobson,Carrie L Kempf,Leopoldo C Cancio,Sara M Camp,Julie Álvarez ,Anne E Cress,Steven M Dudek,Joe G.n Garcia

doi:10.1038/s41598-021-04444-9

Abstract

Despite encouraging preclinical data, therapies to reduce ARDS mortality remains a globally unmet need, including during the COVID-19 pandemic. We previously identified extracellular nicotinamide phosphoribosyltransferase (eNAMPT) as a novel damage-associated molecular pattern protein (DAMP) via TLR4 ligation which regulates inflammatory cascade activation. eNAMPT is tightly linked to human ARDS by biomarker and genotyping studies in ARDS subjects. We now hypothesize that an eNAMPT-neutralizing mAb will significantly reduce the severity of ARDS lung inflammatory lung injury in diverse preclinical rat and porcine models. Sprague Dawley rats received eNAMPT mAb intravenously following exposure to intratracheal lipopolysaccharide (LPS) or to a traumatic blast (125 kPa) but prior to initiation of ventilator-induced lung injury (VILI) (4 h). Yucatan minipigs received intravenous eNAMPT mAb 2 h after initiation of septic shock and VILI (12 h). Each rat/porcine ARDS/VILI model was strongly associated with evidence of severe inflammatory lung injury with NFkB pathway activation and marked dysregulation of the Akt/mTORC2 signaling pathway. eNAMPT neutralization dramatically reduced inflammatory indices and the severity of lung injury in each rat/porcine ARDS/VILI model (~ 50% reduction) including reduction in serum lactate, and plasma levels of eNAMPT, IL-6, TNFα and Ang-2. The eNAMPT mAb further rectified NFkB pathway activation and preserved the Akt/mTORC2 signaling pathway. These results strongly support targeting the eNAMPT/TLR4 inflammatory pathway as a potential ARDS strategy to reduce inflammatory lung injury and ARDS mortality.

Highlights

Despite encouraging preclinical data, therapies to reduce ARDS mortality remains a globally unmet need, including during the COVID-19 pandemic
Intravenous administration of either the extracellular nicotinamide phosphoribosyltransferase (eNAMPT)-neutralizing pAb (4 mg/kg) or humanized mAb (0.4 mg/kg), given concurrently with LPS, significantly reduced histologic (Fig. 1C,D) and BAL evidence of inflammatory injury reflected by BAL protein levels and protein measurements and BAL cell counts (PMNs) counts (Fig. 1E,F) with the eNAMPT mAb providing superior protection compared to the eNAMPT pAb
Our studies are highly consistent with hypothesis that ARDS-related multi-organ failure and mortality are directly influenced by evolutionarily-conserved inflammatory cascades triggered by bacterial/viral pathogens, trauma and ventilator-mediated mechanical stress involving the pathogen-receptor recognition Toll–like receptor 4 (TLR4) pathway[3,5,28]

Summary

Introduction

Therapies to reduce ARDS mortality remains a globally unmet need, including during the COVID-19 pandemic. COVID-19- and non-COVID-ARDS mortality is linked to multi-organ failure and mechanistically to unremitting lung and systemic inflammation[3] generated by pathogen-activated innate immunity pathways (designed to contain the infection) and by damage-associated molecular pattern proteins DAMPs)-mediated amplification of inflammatory cascades elicited by exposure to mechanical ventilation, i.e. ventilator-induced lung injury (VILI)[1,3]. The current study is designed to assess the research hypothesis that targeting the eNAMPT/TLR4 inflammatory cascade via an eNAMPT-neutralizing mAb will significantly reduce the severity of preclinical acute lung injury in rat and porcine ARDS/VILI models These studies which have not been previously reported in either ARDS/VILI-challenged rats or pigs, attempt to maximize the translation of preclinical studies to successful human ARDS trials by extending prior murine results to a large animal model of ARDS/VILI. The combination of the eNAMPT-neutralizing mAb with predictive eNAMPT plasma biomarker and NAMPT genotyping assays provides an eNAMPT-focused personalized medicine platform to strategically identify high-risk subjects for ARDS clinical trials

Methods

Results

Conclusion