ProLung ™- budesonide Inhibits SARS-CoV-2 Replication and Reduces Lung Inflammation

Abstract

Background

Inhaled budesonide benefits patients with COVID-19. ProLung™-budesonide enables the sustained, low dose administration of budesonide within a delivery vehicle similar to lung surfactant. ProLung™-budesonide may offer anti-inflammatory and protective effects to the lung in COVID-19, yet it’s effect on SARS-CoV-2 replication is unknown.

Objective

To determine the efficacy of ProLung™-budesonide against SARS-CoV-2 infection in vitro, evaluate its ability to decrease inflammation, and airway hyperresponsiveness in an animal model of lung inflammation.

Methods

SARS-CoV-2-infected Vero 76 cells were treated with ProLung™-budesonide ([0.03– 100 μg/ml]) for 3 days, and virus yield in the supernatant was measured. Ovalbumin-sensitized C57BL/6 mice received aerosolized (a) ProLung™-budesonide weekly, (b) only budesonide, either daily or weekly, or (c) weekly empty ProLung™-carrier (without budesonide). All treatment groups were compared to sensitized untreated, or normal mice using histopathologic examination, electron microscopy (EM), airway hyperresponsiveness (AHR) to Methacholine (Mch) challenge, and eosinophil peroxidase activity (EPO) measurements in bronchioalveolar lavage (BAL).

Results

ProLung™-budesonide showed significant inhibition on viral replication of SARS-CoV-2-infected cells with the selectivity index (SI) value > 24. Weekly ProLung™-budesonide and daily budesonide therapy significantly decreased lung inflammation and EPO in BAL. ProLung™-budesonide localized in type II pneumocytes, and was the only group to significantly decrease AHR, and EPO in BAL with Mch challenge

Conclusions

ProLung™-budesonide significantly inhibited viral replication in SARS-CoV-2 infected cells. It localized into type II pneumocytes, decreased lung inflammation, AHR and EPO activity with Mch challenge. This novel drug formulation may offer a potential inhalational treatment for COVID-19.

SciScore for 10.1101/2021.05.05.442779: (What is this?)

Please note, not all rigor criteria are appropriate for all manuscripts.

Table 1: Rigor

Ethics	IACUC: All animal protocols were approved by the Animal Care Committee of the Medical College of Wisconsin and the Zablocki Veterans Administration Medical Center, in agreement with the National Institute of Health’s guidelines for the care and use of laboratory animals.
Sex as a biological variable	Animal Studies: Six-week-old male C57 black 6 mice (C57BL/6) were purchased from Charles River Laboratories, Inc., Wilmington, MA.
Randomization	not detected.
Blinding	Coded slides were examined by a veterinary pathologist in a blinded fashion for evidence of inflammatory changes.
Power Analysis	not detected.
Cell Line Authentication	not detected.

Table 2: Resources

Experimental Models: Cell Lines
Sentences	Res…

SciScore for 10.1101/2021.05.05.442779: (What is this?)

Please note, not all rigor criteria are appropriate for all manuscripts.

Table 1: Rigor

Ethics	IACUC: All animal protocols were approved by the Animal Care Committee of the Medical College of Wisconsin and the Zablocki Veterans Administration Medical Center, in agreement with the National Institute of Health’s guidelines for the care and use of laboratory animals.
Sex as a biological variable	Animal Studies: Six-week-old male C57 black 6 mice (C57BL/6) were purchased from Charles River Laboratories, Inc., Wilmington, MA.
Randomization	not detected.
Blinding	Coded slides were examined by a veterinary pathologist in a blinded fashion for evidence of inflammatory changes.
Power Analysis	not detected.
Cell Line Authentication	not detected.

Table 2: Resources

Experimental Models: Cell Lines
Sentences	Resources
Confluent or near-confluent cell culture monolayers of Vero 76 cells were prepared in 96-well microplates.	Vero 76 suggested: None
Experimental Models: Organisms/Strains
Sentences	Resources
Animal Studies: Six-week-old male C57 black 6 mice (C57BL/6) were purchased from Charles River Laboratories, Inc., Wilmington, MA.	C57 black 6 suggested: None
Sensitization: The C57BL/6 mice were sensitized with ovalbumin (OVA) as described in our previous studies (8).	C57BL/6 suggested: None
Software and Algorithms
Sentences	Resources
Animal Studies: Six-week-old male C57 black 6 mice (C57BL/6) were purchased from Charles River Laboratories, Inc., Wilmington, MA.	Charles River Laboratories suggested: (Charles River Laboratories, RRID:SCR_003792)
All treatment groups were compared to either Sensitized Untreated (SENS) or Untreated, Unsensitized (NORMAL)	NORMAL suggested: (NOrMAL, RRID:SCR_010889)
Data Analysis: One-way ANOVA with Tukey-Kramer multiple comparison data analysis was used for Mch responses using SigmaStat Statistical Software (Systat software Inc., San Jose, CA)	SigmaStat suggested: (SigmaStat, RRID:SCR_010285)

Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

Results from TrialIdentifier: No clinical trial numbers were referenced.

Results from Barzooka: We found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).

Results from JetFighter: We did not find any issues relating to colormaps.

Results from rtransparent:

Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
No protocol registration statement was detected.

Results from scite Reference Check: We found no unreliable references.

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