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Authors: Poschet et al. Link to paper: https://doi.org/10.1101/2020.03.29.008631.

Journal/ Pre-Print: bioRxiv

Key Words: In-vitro, Cystic Fibrosis, Hyper-acidification, Secretory system

RESEARCH HIGHLIGHTS 

1. AZT and CPX can correct hyper-acidification of trans-Golgi network (TGN) and recycling endosomes (REs) in in vitro human CF cell models.

2. AZT enhances the binding of P. aeruginosa and cholera toxin to the CF epithelium through a concentration independent, non-antibiotic mechanism.

3. AZT reduces the pro-inflammatory (e.g. IL8) and pro-fibrotic (e.g. TGF-β) phenotype of CF epithelium through pH correction of hyper-acidified organelles of the secretory system.

SUMMARY 

The research examines the cellular and immune mechanisms of two FDA-approved antibiotics, azithromycin (AZT) and ciprofloxacin (CPX) in the treatment of cystic fibrosis (CF). The research confirmed the drugs’ effect in regulating abnormal organelle pH and enhanced innate immunity activity in different CF epithelium models, and draw similarities to the MOA of CQ. Both AZT & CPX are demonstrated to have concentration independent roles, which are explained by the accumulation of these drugs in acidified organelles following protonation. While authors suggest these two drugs might contribute to the COVID-19 treatment, there is no experimental evidence to support claims.

IMPACT FOR SARS-COV2/COVID19 RESEARCH EFFORTS

Understand the virology and/or cell biology of SARS-CoV2/COVID19

Paper emphasis on hyper-acidification of organelles. Link to COVID19 is suggestive only (no evidence!) – one extrapolation is that altered organelle pH by AZT/CPX & CQ may alter ability of PTM, such as glycosylation, which may impact upon ACE2 and spike protein glycosylation.

Treat of SARS-CoV2/COVID19 positive individuals

AZT, CPX & CQ highlighted as potential drugs that are able to reduce epithelial pro-inflammatory cytokine secretion and further fibrosis, meanwhile, inhibiting the infection. Model of CF. No model of COVID19 applied. Generalised role.

STUDY TYPE

· In vitro study

o Human CF cell line model

STRENGTHS AND LIMITATIONS OF THE PAPER

Novelty: > Though the article implied how AZT and CPX contribute to the treatment of CF, the underlying target of each phenotype is largely unknown.

> Many of the effects have been published before. Publication simply contributes different schemes of treatment or different models but not novel ideas.

>Application of drugs in treatment of SARS-CoV2 is only speculative, no experimental support.

>Largely based on role of CQ. Doesn’t provide any commentary on how AZT/CPX is advantageous/superior (or overcomes criticism) to these already established treatment routines with CQ/HCQ

Standing in the field: Some of the result is contradictory to the previously published ones. E.G. One of the major results is AZT can decrease the basal IL-8 level in CF cells (Figure 3), while in their cited article there should be no change even under several concentration (Azithromycin Treatment Alters Gene Expression in Inflammatory, Lipid Metabolism, and Cell Cycle Pathways in Well-Differentiated Human Airway Epithelia).

Appropriate statistics: Either ANOVA or not specified, N=6 for most replications.

Viral model used: Human Cystic Fibrosis Cell lines. (Including IB3-1 and pCEP-R cell lines) A normalised control CFTR-corrected IB3-1 cells referred to as the S9 cell line was used as a baseline control.

Translatability: No. Largely for reasons are above. Moreover, direct comparisons to HCQ and CQ are largely invalid due to both having a significantly higher pKa than either CPX or AZT (Table 1), therefore they are more potent to increase the pH of the TGN and RE in coronavirus-infected cells, thus decreasing the likely affectability of AZT/CPX in vivo.

Main limitations: >Poor model; CF based, not a SARS-CoV relevant model

>Research is based on CF study, whereby all CF models used display lower pH in TGN and RE than in normal cells, therefore cannot be certain of role in normal cells.

>Only confident result demonstrates that AZT and CPX can correct the pH in abnormal TGN and RE and the consequent immune phenotypes.

>We are not sure however if/how the drugs can further increase the organelle pH in coronavirus-infected cells. This undermines their further application.

>All links to SARS-CoV2 are speculative and not evidence driven & are very inflated. Lots of extrapolation.

>No MOA described, however bases a lot on CQ. It does not however address issues of CQ/HCQ SARS-CoV model, or suggest why this is a better model to use than CQ/HCQ regime.