Hydroxychloroquine and Azithromycin Molecular Action against SARS-CoV-2 Viral Protein: A Molecular Dynamic Study

Research Article

Austin J Nanomed Nanotechnol. 2021; 9(1): 1061.

Hydroxychloroquine and Azithromycin Molecular Action against SARS-CoV-2 Viral Protein: A Molecular Dynamic Study

Picaud F* and Herlem G

Nanomedicine Lab EA4662, University of Franche-Comté, France

*Corresponding author: Fabien Picaud, Nanomedicine Lab EA4662, University of Franche- Comté, UFR Sciences & Techniques, 16 Route de Gray, 25030 Besançon Cedex, France

Received: December 03, 2020; Accepted: December 28, 2020; Published: January 04, 2021

Abstract

For the past few months, the world has gone through hell with the emergence of the SARS-CoV-2 virus and the resulting pandemic. Faced with this disease, various therapeutic strategies have been developed to understand how to eradicate this virus. Here we present a molecular dynamics simulation study on the effect of a dual therapy (hydroxychloroquine and azithromycin) on the open and closed forms of a viral protein. We show in particular that hydroxychloroquine has no significant interaction with the viral receptor-binding domain RBD when it interacts with its host receptor. However, this molecule can, in the closed form of the virus, block the movement of these receptors and thus prevent the attachment of the virus to the host cell. The azithromycin molecule interacts very well with the open receptor but can also be inserted into the S2 domain of the protein. It therefore presents two potential mechanisms of action against the virus, mainly on the closed state of the viral protein.

Keywords: Hydroxychloroquine; Azythromicin; SARS-CoV-2 structure; MD-Simulation; Docking

Introduction

In 2002, the first emergence of a pathogenic coronavirus revealed to the world the possibility of a pandemic. The severe acute respiratory syndrome coronavirus, or SARS-CoV, was responsible for very important breathing syndromes [1-4] with, however, a small amount of death around the world (8000 persons) while the mortality rate reached 10%. More recently (2012), a severe pneumonia appeared in Saudi Arabia due to a novel coronavirus [5,6]. This one, called MERSCoV for Middle East Respiratory Syndrome Coronovirus, still exists but concerns only the Arabic peninsula. While very localized on a small area, this MERS-Cov is highly dangerous since its mortality rate is about 35% (1 over 3 patients). These two cases are not the only ones since periodically, other coronaviruses, while less virulent are appearing [7,8,9].

These first viral apparitions should have alerted us to the possibility, in the long term, of the emergence of a more virulent coronavirus clearly difficult to manage. At the end of 2019, a new coronavirus called severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), or COVID- 19 disease, developed as a human pathogen in a Chinese city (Wuhan). Despite strong resolutions in China and over several states (locking up millions of people), global economic and touristic development is leading to a general spread of the disease. Although SARS-CoV-2 has many points in common with SARS-CoV and MERS-CoV, it appears to be transmitted more easily and much faster than SARS-CoV [10,11] and MERS-CoV [12,13]. To date, more than 64 million cases of COVID-19 have been confirmed and at least 1.400,000 deaths have been recorded worldwide by the World Health Organization which declared the first real pandemic of March 21, 2020. Although all the cases have not been identified, these figures lead to a mortality rate close to 2.1%, i.e. 4 times the rate of the seasonal flu. While the peak of the pandemic seems to be behind us, the second wave forces us to find a therapeutic method to treat this virus.

Many drugs have been clinically tested in numerous clinical projects (“discovery” for France, “recovery” for England”) but no one has determined a truly effective treatment against the SARS-CoV-2 virus. In parallel, a protocol, confirmed by many other studies [14- 17], seems to be the most appropriate for combating the virus and reducing the degree of contagiousness. This protocol, that should be administered as soon as the first symptoms appear, combines both the antimalarial drug Hydroxychloroquine (HCQ) and the antibiotic Azithromycin (AZM). The results of the various studies lead to a sharp decrease of the mortality rate (under 0.5%). However, many other analyzes question the results of the therapies. In order to better understand the role of this double drug treatment against this new virus, an analysis of its action is necessary at the molecular level [18].

With regard to its genome sequence, SARS-CoV-2 belongs to the same beta-coronavirus family as SARS-CoV and MERS-CoV. These coronaviruses have a spherical envelope with a diameter close to 100 nm. The latter is composed of a N nucleoprotein surrounded by a lipid bilayer originating from the host cell. Three other proteins are then found on this surface, protein S (or spike protein), protein M (or membrane protein), and protein E (or envelope protein). Homotrimerization of S proteins [11] on the surface of the virion is the key step in viral infection. To decrease the viral progression, the drug must therefore target the S protein. However, the latter is separated into two domains which have a very specific role [19]. The S1 domain, mainly formed by a Receptor Binding Domain (RBD), binds to the host cell receptor [20] (called angiotensin converting enzyme 2 (ACE2) [21,22] while that the S2 domain is at the origin of the fusion of protein E with the host cell [19].

Therefore, the main objective of any viral treatment is to block the binding of the RBD domain to the ACE2 receptor in order to avoid the fusion of the virus with the host cell. Note that the drugs could prevent protein fusion with the host cell through structural changes.

Here we propose to determine what are the main sites of interaction of the HCQ and the AZM drug molecules on structural S protein. Our work, based on several molecular dynamics simulations will be separated into two parts. We will first determine the binding of each drug when RBD is associated with the ACE2 receptor. Then, in a second step, we will show that these molecules can also have another target to on the state close to the S protein.

Method

Hydroxychloroquine (HCQ) is (RS)-2-[[4-[(7-chloroquinoline- 4-yl)amino]pentyl](ethyl)amino] ethanol. Its 3D structure has been obtained through the Pub-Chem CID: 3652 file. Azithromycin (AZM) is (2R, 3S, 4R, 5 100 R, 8R, 10R, 11R, 12S, 13S, 14R)-11- [(2S, 3R, 4S, 6R)-4-(dimethylamino)-3-hydroxy-6-methyloxan-2-yl] oxy-2-ethyl-3, 4, 10-trihydroxy-13- [(2R, 4R, 5S, 6S)-5-hydroxyl- 4-methoxy-4,6-dimethyloxan-2-yl] oxy-3, 5, 6, 8, 10, 12, 14 - heptamethyl-1-oxa-6-azacyclopentadecan-15-one. Its 3D structure has been obtained through the Pub-Chem CID: 447043 file.

Our strategy was organized in two different steps. First of all, we focused on the simulation of the Receptor Binding Domain (RBD) bound to the ACE2 receptor of the host cell with the HCQ or the AZM molecules. The goal of these first calculations is to observe whether the molecules of the drug can interact directly with the viral protein when they are attached to its host cell. To simulate such arrangement of proteins, we use the 6M0J pdb structure. The relaxed crystal structures approaching the living organism as well as the effect of glycosylation on the stability of the structure were studied.

Then, in a second step, the full conformation of the spike glycoprotein trimer SARS-CoV-2 was simulated in presence of HCQ or AZM molecules. Its structure was obtained from pdb file #6VXX. It has a resolution of 2.80Å as determined from electron microscopy. It is composed of 3 chains intercalated with different respective domains such as the NTD (N-terminal domain) and the RBD (receptor binding domain) which belong to the S1 part of the protein.

Classical MD simulations were performed by constructing the full molecular force field for HCQ and AZM using the SwissParam Force Field Toolkit package [23,24].

For the protein, the molecular force field was constructed according to the CHARMM-GUI procedure in order to appropriately relax the different parts of the protein [25,26]. N-glycosylation of the proteins, when necessary, was achieved using the CHARMMGUI Glycolipid Modeler.[27] The structure of these proteins was first minimized and then progressively equilibrated (1ns) and run (40ns) using MD simulations (NAMD 2.12 package) for a total of 41 ns in saline solution media [28]. All the structures of proteins and molecules are shown in Figure 1a-d.