Oral and Inhaled Fosamprenavir Reverses Pepsin‐Induced Damage in a Laryngopharyngeal Reflux Mouse Model

Johnston, Nikki; Samuels, Tina L.; Goetz, Christopher J.; Arnold, Leggy A.; Smith, Brian C.; Seabloom, Donna E.; Wuertz, Beverly R.; Ondrey, Frank G.; Wiedmann, Timothy S.; Vuksanovic, Nemanja; Silvaggi, N.R.; Mackinnon, Alexander C.; Miller, James A.; Bock, Jonathan M.; Blumin, Joel H.

doi:10.1002/lary.30242

Cited by 7 publications

(29 citation statements)

References 138 publications

(330 reference statements)

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“…We recently identified an HIV protease inhibitor (amprenavir, commonly administered as fosamprenavir) which binds and inhibits pepsin in the nanomolar range and rescued histologic indications of peptic injury in a mouse LPR model. 54 Many LPR episodes are weakly acidic and weakly acidic pepsin elicits microscopically evident cell dissociation in vitro 43 akin to dilated intercellular spaces which are a hallmark of LPR and GERD.…”

Section: Discussionmentioning

confidence: 99%

Amprenavir inhibits pepsin‐mediated laryngeal epithelial disruption and E‐cadherin cleavage in vitro

Samuels

Blaine-Sauer

Yan

et al. 2023

Laryngoscope Investig Oto

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BackgroundLaryngopharyngeal reflux (LPR) causes chronic cough, throat clearing, hoarseness, and dysphagia and can promote laryngeal carcinogenesis. More than 20% of the US population suffers from LPR and there is no effective medical therapy. Pepsin is a predominant source of damage during LPR which disrupts laryngeal barrier function potentially via E‐cadherin cleavage proteolysis and downstream matrix metalloproteinase (MMP) dysregulation. Fosamprenavir (FDA‐approved HIV therapeutic and prodrug of amprenavir) is a pepsin‐inhibiting LPR therapeutic candidate shown to rescue damage in an LPR mouse model. This study aimed to examine amprenavir protection against laryngeal monolayer disruption and related E‐cadherin proteolysis and MMP dysregulation in vitro.MethodsLaryngeal (TVC HPV) cells were exposed to buffered saline, pH 7.4 or pH 4 ± 1 mg/mL pepsin ± amprenavir (10–60 min). Analysis was performed by microscopy, Western blot, and real time polymerase chain reaction (qPCR).ResultsAmprenavir (1 μM) rescued pepsin acid‐mediated cell dissociation (p < .05). Pepsin acid caused E‐cadherin cleavage indicative of regulated intramembrane proteolysis (RIP) and increased MMP‐1,3,7,9,14 24‐h postexposure (p < .05). Acid alone did not cause cell dissociation or E‐cadherin cleavage. Amprenavir (10 μM) protected against E‐cadherin cleavage and MMP‐1,9,14 induction (p < .05).ConclusionsAmprenavir, at serum concentrations achievable provided the manufacturer's recommended dose of fosamprenavir for HIV, protects against pepsin‐mediated cell dissociation, E‐cadherin cleavage, and MMP dysregulation thought to contribute to barrier dysfunction and related symptoms during LPR. Fosamprenavir to amprenavir conversion by laryngeal epithelia, serum and saliva, and relative drug efficacies in an LPR mouse model are under investigation to inform development of inhaled formulations for LPR.

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Section: Discussionmentioning

confidence: 99%

Amprenavir inhibits pepsin‐mediated laryngeal epithelial disruption and E‐cadherin cleavage in vitro

Samuels

Blaine-Sauer

Yan

et al. 2023

Laryngoscope Investig Oto

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“…However, the significant correlation of decreased distal esophageal MNBI with an increased number of proximal acidic impedance events, as seen in Table II, would favor the reflux theory. The lack of differences in pharyngeal MNBI be explained by the majority of pharyngeal reflux events being weakly acidic or non‐acidic as they are brief and quickly buffered by salivary bicarbonate and compensatory swallowing 28 . Sakin et al 29 found that patients with LPR symptoms had a significantly decreased pharyngeal MNBI compared to patients with GERD without LPR symptoms, despite a similar pharyngeal reflux rate on impedance monitoring.…”

Section: Discussionmentioning

confidence: 99%

“…The lack of differences in pharyngeal MNBI be explained by the majority of pharyngeal reflux events being weakly acidic or non-acidic as they are brief and quickly buffered by salivary bicarbonate and compensatory swallowing. 28 Sakin et al 29 found that patients with LPR symptoms had a significantly decreased pharyngeal MNBI compared to patients with GERD without LPR symptoms, despite a similar pharyngeal reflux rate on impedance monitoring. Although there were no control subjects included in the study, the decreased pharyngeal MNBI in LPR versus GERD patients would demonstrate chronic reflux into the hypopharynx, as described in the reflux theory.…”

Section: Discussionmentioning

confidence: 99%

Correlation of Esophageal Mean Nocturnal Baseline Impedance With Markers of Laryngopharyngeal Reflux

et al. 2022

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Objectives: Mean nocturnal baseline impedance (MNBI) is a measure of the esophageal epithelial barrier function calculated via high-resolution impedance manometry and can be used as a diagnostic tool and treatment response predictor for gastroesophageal reflux disease (GERD). However, its utility for laryngopharyngeal reflux (LPR) has been minimally studied. We aimed to investigate the relationship of MNBI between patients with suspected LPR, healthy controls, and their 24-h multichannel intraluminal impedance-pH (MII-pH) study results.Methods: Retrospective patient series analysis was performed of patients with suspected LPR and healthy controls who underwent 24-h MII-pH monitoring. MNBI values were calculated from impedance channels at the level of the hypopharynx, proximal esophagus, and distal esophagus. We compared these MNBI values between the subject groups with secondary analysis on MII-pH results, reflux symptom index, reflux findings score, DeMeester score, and salivary pepsin levels.Results: Twenty-three patients with suspected LPR and 14 healthy controls were enrolled. Decreased distal esophageal MNBI was found to be significantly decreased in patients with suspected LPR compared with healthy controls (p < 0.01) and in subjects with positive MII-pH studies compared to negative MII-pH studies (p < 0.01). There were no significant correlations of MNBI at the hypopharynx or proximal esophagus.Conclusion: Distal esophageal MNBI has significant correlations with many phenotypic and biological markers of LPR. These findings indicate that MNBI has the potential to be applied to LPR, similar to its emerging use as a diagnostic tool and treatment response predictor for GERD.

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“…The IC50 of amprenavir for porcine pepsin (3.56 uM) was determined in a prior study [ 37 ]. The dose for study herein was selected based upon the serum concentration observed in patients taking the manufacturer-recommended dose for the treatment of HIV [ 90 ].…”

Section: Methodsmentioning

confidence: 99%

“…Our group recently identified the HIV protease inhibitor, amprenavir, as a potential pepsin-targeting therapeutic for laryngopharyngeal reflux (LPR). Amprenavir inhibited pepsin at low micromolar concentrations and its prodrug fosamprenavir prevented histologic changes in an LPR mouse model [ 37 ]. In parallel work, we found that pepsin was required for esophageal and laryngeal epithelial barrier disruption at pH4 and that amprenavir preserved laryngeal epithelial integrity and prevented damage to a critical cell adhesion molecule elicited by pepsin at pH4 [ 38 , 39 ].…”

Section: Introductionmentioning

confidence: 99%

The Protease Inhibitor Amprenavir Protects against Pepsin-Induced Esophageal Epithelial Barrier Disruption and Cancer-Associated Changes

Blaine-Sauer

Samuels

Yan

et al. 2023

IJMS

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Gastroesophageal reflux disease (GERD) significantly impacts patient quality of life and is a major risk factor for the development of Barrett’s esophagus (BE) and esophageal adenocarcinoma (EAC). Proton pump inhibitors (PPIs) are the standard-of-care for GERD and are among the most prescribed drugs in the world, but do not protect against nonacid components of reflux such as pepsin, or prevent reflux-associated carcinogenesis. We recently identified an HIV protease inhibitor amprenavir that inhibits pepsin and demonstrated the antireflux therapeutic potential of its prodrug fosamprenavir in a mouse model of laryngopharyngeal reflux. In this study, we assessed the capacity of amprenavir to protect against esophageal epithelial barrier disruption in vitro and related molecular events, E-cadherin cleavage, and matrix metalloproteinase induction, which are associated with GERD severity and esophageal cancer. Herein, weakly acidified pepsin (though not acid alone) caused cell dissociation accompanied by regulated intramembrane proteolysis of E-cadherin. Soluble E-cadherin responsive matrix metalloproteinases (MMPs) were transcriptionally upregulated 24 h post-treatment. Amprenavir, at serum concentrations achievable given the manufacturer-recommended dose of fosamprenavir, protected against pepsin-induced cell dissociation, E-cadherin cleavage, and MMP induction. These results support a potential therapeutic role for amprenavir in GERD recalcitrant to PPI therapy and for preventing GERD-associated neoplastic changes.

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Oral and Inhaled Fosamprenavir Reverses Pepsin‐Induced Damage in a Laryngopharyngeal Reflux Mouse Model

Cited by 7 publications

References 138 publications

Amprenavir inhibits pepsin‐mediated laryngeal epithelial disruption and E‐cadherin cleavage in vitro

Amprenavir inhibits pepsin‐mediated laryngeal epithelial disruption and E‐cadherin cleavage in vitro

Correlation of Esophageal Mean Nocturnal Baseline Impedance With Markers of Laryngopharyngeal Reflux

The Protease Inhibitor Amprenavir Protects against Pepsin-Induced Esophageal Epithelial Barrier Disruption and Cancer-Associated Changes

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