To study the interaction between chloroquine/hydroxychloroquine and its monodiastereomer and angiotensin-converting enzyme 2

Chloroquine (CQ) and hydroxychloroquine (HCQ) are well-known antimalarials, and they were later found to also have immunomodulatory and anti-inflammatory effects and are widely used to treat rheumatic diseases such as rheumatoid arthritis and systemic lupus erythematosus
.
As early as 1983, CQ was reported to suppress influenza A and B viruses, and it was later found that CQ has an inhibitory effect
on the replication of human immunodeficiency virus (HIV) and SARS coronaviruses in vitro.
The use of CQ and HCQ as potential broad-spectrum antivirals has received widespread attention
in SARS-CoV-2 outbreaks.

Enantiomers were isolated from hydroxychloroquine (HCQ) and each enantiomer was retained on an angiotensin-converting enzyme 2 (ACE2)-HEK293T/cell membrane chromatography (CMC) column:

Chloroquine and hydroxychloroquine
have been studied since the early clinical treatment of the SARS-CoV-2 outbreak.
Considering that these two chiral drugs are currently used as racemates, a highly expressed angiotensin-converting enzyme 2 cell membrane chromatography method was established to study the difference
in the binding of the two paired enantiomers to the ACE 2 receptor.
Molecular docking assays and detection of SARS-CoV-2 spike pseudotype virus entering ACE 2-HEK293T cells were also performed for further investigation
.
In this study, a 2D LC system was established to isolate enantiomers of CQ/HCQ and to evaluate whether individual enantiomers interact
with ACE2 receptors.
ACE2-HEK293T CMC is used to assess the binding affinity of enantiomers acting on ACE2 receptors
.
The results showed that each enantiomer binds well to ACE 2, but in positive analysis, there is a difference
between the paired enantiomer and the corresponding racemate.
Compared with S-chloroquine/chloroquine (racemate), R-chloroquine exhibits better ACE 2 receptor binding capacity
.
S-hydroxychloroquine exhibits better ACE 2 receptor binding capacity
than R-hydroxychloroquine/hydroxychloroquine.
In addition, each enantiomer was shown to be effective compared to the control group; Compared to S-chloroquine or racemate, R-chloroquine exhibits better inhibition at the same concentration
.
As for hydroxychloroquine, R-hydroxychloroquine showed better inhibition than S-hydroxychloroquine, but slightly worse
than racemates.

Two molecular docking models of paired enantiomers to angiotensin-converting enzyme 2 (ACE2):

In summary, all four drugs interacted with the ACE2 receptor and showed potential activity to prevent the virus from binding to the ACE2 receptor, and the detection of SARS-CoV-2 spike pseudovirus entry validated its ability
.
ACE2-HEK293T/CMC is a reliable method
for screening potential active ingredients.
At the same time, R-CQ showed better inhibition than S-CQ and CQ racemates at the same concentration, and the enantiomers differed significantly
.
At the same concentration, the binding ability of S-HCQ to ACE2 receptors was better than R-HCQ/HCQ (racemate), while the effect of preventing SARS-CoV-2 pseudovirus from entering cells was weaker than that of R-HCQ/HCQ (racemate) at the same concentration
.

Parameters of the monoenantiomer of chloroquine (CQ)/hydroxychloroquine (HCQ) interaction with angiotensin-converting enzyme 2 (ACE2) receptors:

In this study, the researchers established a two-dimensional LC system, which would be a quick and convenient way to
isolate enantiomers and assess whether individual enantiomers interact with ACE2 receptors.
At the same time, the binding affinity of the enantiomer acting on the ACE2 receptor was evaluated by CMC
.
Each enantiomer of the dissociation equilibrium constant (_KD) was compared
for the first time with the ACE2 receptor.
In addition, the ability of two paired enantiomers to prevent SARS-CoV-2 spike pseudotype virus from entering cells was first studied and compared
.
This study provides new insights
into SARS-CoV-2 treatment using a single enantiomer of CQ/HCQ.

References: Jia Q, Fu J, Liang P, Wang S, Wang Y, Zhang X, Zhou H, Zhang L, Lv Y, Han S.
Investigating interactions between chloroquine/hydroxychloroquine and their single enantiomers and angiotensin-converting enzyme 2 by a cell membrane chromatography method.
J Sep Sci.
2022 Jan; 45(2):456-467.
doi: 10.
1002/jssc.
202100570.
Epub 2021 Nov 17.
PMID: 34729910; PMCID: PMC8661882.