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Extracellular Nicotinamide Phosphoribosyltransferase Is a Therapeutic Target in Experimental Necrotizing
Melissa D Halpern1, Akash Gupta1, Nahla Zaghloul1
1Division of Neonatology, Department of Pediatrics, College of Medicine, University of Arizona, Tucson, AZ 85724, USA.
Abstract:
Necrotizing enterocolitis (NEC) is the most common gastrointestinal emergency of prematurity. Postulated mechanisms leading to inflammatory necrosis of the ileum and colon include activation of the pathogen recognition receptor Toll-like receptor 4 (TLR4) and decreased levels of transforming growth factor beta (TGFβ). Extracellular nicotinamide phosphoribosyltransferase (eNAMPT), a novel damage-associated molecular pattern (DAMP), is a TLR4 ligand and plays a role in a number of inflammatory disease processes. To test the hypothesis that eNAMPT is involved in NEC, an eNAMPT-neutralizing monoclonal antibody, ALT-100, was used in a well-established animal model of NEC. Preterm Sprague-Dawley pups delivered prematurely from timed-pregnant dams were exposed to hypoxia/hypothermia and randomized to control-foster mother dam-fed rats, injected IP with saline (vehicle) 48 h after delivery; control + mAB-foster dam-fed rats, injected IP with 10 µg of ALT-100 at 48 h post-delivery; NEC-orally gavaged, formula-fed rats injected with saline; and NEC + mAb-formula-fed rats, injected IP with 10 µg of ALT-100 at 48 h. The distal ileum was processed 96 h after C-section delivery for histological, biochemical, molecular, and RNA sequencing studies. Saline-treated NEC pups exhibited markedly increased fecal blood and histologic ileal damage compared to controls (q < 0.0001), and findings significantly reduced in ALT-100 mAb-treated NEC pups (q < 0.01). Real-time PCR in ileal tissues revealed increased NAMPT in NEC pups compared to pups that received the ALT-100 mAb (p < 0.01). Elevated serum levels of tumor necrosis factor alpha (TNFα), interleukin 6 (IL-6), interleukin-8 (IL-8), and NAMPT were observed in NEC pups compared to NEC + mAb pups (p < 0.01). Finally, RNA-Seq confirmed dysregulated TGFβ and TLR4 signaling pathways in NEC pups that were attenuated by ALT-100 mAb treatment. These data strongly support the involvement of eNAMPT in NEC pathobiology and eNAMPT neutralization as a strategy to address the unmet need for NEC therapeutics.
Insights
Necrotizing enterocolitis (NEC) involves inflammation and damage, potentially driven by extracellular nicotinamide phosphoribosyltransferase (eNAMPT). Neutralizing eNAMPT with ALT-100 significantly reduced NEC severity in a preclinical model.
Area of Science:
- Gastroenterology
- Immunology
- Neonatal Medicine
Background:
- Necrotizing enterocolitis (NEC) is a critical gastrointestinal condition in premature infants.
- Pathological mechanisms may involve Toll-like receptor 4 (TLR4) activation and reduced transforming growth factor beta (TGFβ).
- Extracellular nicotinamide phosphoribosyltransferase (eNAMPT), a damage-associated molecular pattern (DAMP) and TLR4 ligand, is implicated in inflammatory processes.
Purpose of the Study:
- To investigate the role of eNAMPT in NEC pathogenesis.
- To evaluate the therapeutic potential of an eNAMPT-neutralizing antibody (ALT-100) in a preclinical NEC model.
Main Methods:
- A validated animal model of NEC was employed, involving preterm pups exposed to hypoxia/hypothermia and formula feeding.
- Pups were randomized to receive either a saline vehicle or the ALT-100 monoclonal antibody (mAb).
- Ileal tissues were analyzed histologically, biochemically, and molecularly, including RNA sequencing.
Main Results:
- NEC induction led to significant ileal damage and increased fecal blood, which were markedly reduced by ALT-100 treatment.
- Ileal NAMPT levels and serum inflammatory markers (TNFα, IL-6, IL-8) were elevated in NEC pups but decreased with ALT-100 administration.
- RNA-Seq analysis revealed that ALT-100 attenuated dysregulated TGFβ and TLR4 signaling pathways in NEC.
Conclusions:
- These findings strongly support the involvement of eNAMPT in the pathophysiology of NEC.
- Neutralization of eNAMPT with ALT-100 presents a promising therapeutic strategy for NEC.
