*Review* **Variants of SARS-CoV-2: Influences on the Vaccines' Effectiveness and Possible Strategies to Overcome Their Consequences**

**Ali A. Rabaan 1,2,3,\*, Shamsah H. Al-Ahmed 4, Hawra Albayat 5, Sara Alwarthan 6, Mashael Alhajri 6, Mustafa A. Najim 7, Bashayer M. AlShehail 8, Wasl Al-Adsani 9,10, Ali Alghadeer 11, Wesam A. Abduljabbar 12, Nouf Alotaibi 13, Jameela Alsalman 14, Ali H. Gorab 15, Reem S. Almaghrabi 16, Ali A. Zaidan 17, Sahar Aldossary 18, Mohammed Alissa 19, Lamees M. Alburaiky 20, Fatimah Mustafa Alsalim 21, Nanamika Thakur 22,\*, Geetika Verma <sup>23</sup> and Manish Dhawan 24,25,\***

	- <sup>14</sup> Infection Disease Unit, Department of Internal Medicine, Salmaniya Medical Complex, Ministry of Health, Kingdom of Bahrain, Manama 435, Bahrain
	- <sup>15</sup> Al Kuzama Primary Health Care Center, Al Khobar Health Network, Eastern Health Cluster, Al Khobar 34446, Saudi Arabia
	- <sup>16</sup> Organ Transplant Center of Excellence, King Faisal Specialist Hospital and Research Center, Riyadh 11211, Saudi Arabia
	- <sup>17</sup> Gastroenterology Department, King Fahad Armed Forces Hospital, Jeddah 23831, Saudi Arabia
	- <sup>18</sup> Pediatric Infectious Diseases, Women and Children's Health Institute, Johns Hopkins Aramco Healthcare, Dhahran 31311, Saudi Arabia
	- <sup>19</sup> Department of Medical Laboratory Sciences, College of Applied Medical Sciences, Prince Sattam bin Abdulaziz University, Al-Kharj 11942, Saudi Arabia
	- <sup>20</sup> Pediatric Department, Safwa General Hospital, Eastern Health Cluster, Safwa 31921, Saudi Arabia
	- <sup>21</sup> Department of Family Medicine, Primary Health Care, Qatif Health Cluster, Qatif 32434, Saudi Arabia
	- <sup>22</sup> University Institute of Biotechnology, Department of Biotechnology, Chandigarh University, Mohali 140413, India
	- <sup>23</sup> Department of Experimental Medicine and Biotechnology, Post Graduate Institute of Medical Education and Research (PGIMER), Chandigarh 160012, India
	- <sup>24</sup> Department of Microbiology, Punjab Agricultural University, Ludhiana 141004, India
	- <sup>25</sup> Trafford College, Altrincham, Manchester WA14 5PQ, UK
	- **\*** Correspondence: arabaan@gmail.com (A.A.R.); nanamika.e13132@cumail.in (N.T.); dhawanmanish501@gmail.com (M.D.)

**Abstract:** The immune response elicited by the current COVID-19 vaccinations declines with time, especially among the immunocompromised population. Furthermore, the emergence of novel SARS-CoV-2 variants, particularly the Omicron variant, has raised serious concerns about the efficacy of currently available vaccines in protecting the most vulnerable people. Several studies have reported that vaccinated people get breakthrough infections amid COVID-19 cases. So far, five variants of concern (VOCs) have been reported, resulting in successive waves of infection. These variants have

**Citation:** Rabaan, A.A.; Al-Ahmed, S.H.; Albayat, H.; Alwarthan, S.; Alhajri, M.; Najim, M.A.; AlShehail, B.M.; Al-Adsani, W.; Alghadeer, A.; Abduljabbar, W.A.; et al. Variants of SARS-CoV-2: Influences on the Vaccines' Effectiveness and Possible Strategies to Overcome Their Consequences. *Medicina* **2023**, *59*, 507. https://doi.org/10.3390/ medicina59030507

Academic Editors: Yusra Habib Khan, Tauqeer Hussain Mallhi, Tahir Mehmood Khan and Muhammad Salman

Received: 6 February 2023 Revised: 27 February 2023 Accepted: 3 March 2023 Published: 5 March 2023

**Copyright:** © 2023 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

shown a variable amount of resistance towards the neutralising antibodies (nAbs) elicited either through natural infection or the vaccination. The spike (S) protein, membrane (M) protein, and envelope (E) protein on the viral surface envelope and the N-nucleocapsid protein in the core of the ribonucleoprotein are the major structural vaccine target proteins against COVID-19. Among these targets, S Protein has been extensively exploited to generate effective vaccines against COVID-19. Hence, amid the emergence of novel variants of SARS-CoV-2, we have discussed their impact on currently available vaccines. We have also discussed the potential roles of S Protein in the development of novel vaccination approaches to contain the negative consequences of the variants' emergence and acquisition of mutations in the S Protein of SARS-CoV-2. Moreover, the implications of SARS-CoV-2's structural proteins were also discussed in terms of their variable potential to elicit an effective amount of immune response.

**Keywords:** SARS-CoV-2; COVID-19; breakthrough infections; neutralizing antibodies (NAbs); Omicron; vaccines; variants

### **1. Introduction**

Safe and effective vaccination has been critical in the ongoing battle against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). The development of precise vaccine platforms in such a short period is a testament to global scientific prowess, and, as of 9 June, 2021, more than 2,156,550,767 doses of the COVID-19 vaccine have been given across the five continents [1]. Unfortunately, reports on variants of SARS-CoV-2 brought about by mutations with enhanced virulence, pathogenicity, and the ability to detrimentally affect host immune systems, especially the antibodies produced after COVID-19 vaccination, is a matter of concern and scientific deliberation.

However, the available published data divulge that the current vaccines could still be effective in preventing severe infection and death in people infected with the recent variants of SARS-CoV-2, such as Omicron and Delta [2–6]. Multiple studies have shown several advantages of the numerous mutations of the Omicron variant of the SARS-CoV-2 virus [7–9]. The Omicron variant and its subvariants evolved by evolutionary processes that may lead to a number of significant modifications in the virus's characteristics, such as immunological escape from the nAbs produced by the administration of the vaccines [10–12]. The high frequency of mutations has also been linked to improved proteolytic priming with transmembrane serine protease 2 (TMPRSS2) and the increased binding capacities of S Protein to the angiotensin converting enzyme 2 (ACE2) receptor [9,10,13–15]. The higher number of mutations in the Omicron variant have also been associated with improved resistance to endosomal restriction factors, specifically IFITM proteins, which enables the variant's more effective cellular invasion via the endocytic route [16]. Additionally, the modifications may make it more likely for spike protomers to adopt an up configuration to interact with ACE2, and may increase the stability of a down configuration to prevent contact with nAbs [12,17,18].

A variant can be defined as an isolate whose genome sequence differs from that of the reference virus. Thus, the variants share an identical inherited set of distinct mutations and are classified based on a lineage, i.e., the type of mutations that resulted in the origination of a new lineage of SARS-CoV-2. From this perspective, it is crucial to understand the mutational dynamics of SARS-CoV-2 and its effects on the vaccines that are currently available [19]. Studies have deciphered that a typical SARS-CoV-2 virus accrues, on average, one or two single-nucleotide genomic mutations in a period of 30 days [3,4]. This is just 50% of the rate of the mutational dynamics of influenza and 25% of the AIDS human immunodeficiency virus (HIV). The retarded mutational dynamics of SARS-CoV-2 could perhaps be credited to the specific exoribonuclease (ExoN) present in the genome of coronaviruses (CoVs), since inactivation of this ExoN has demonstrated a twenty-fold increase in the mutation rates [4,5].

### *Effects of Mutations on Variants' Characteristics*

Some significant mutations documented in SARS-CoV-2 include the 'N501Y', in which the spike protein (S Protein) 501st amino acid is swapped from N (asparagine) to Y (tyrosine) and assists the virus to attach more rigidly to human cells. Substitution of histidine at 681 positions instead of proline results in a change in an amino acid on the stem region of the spike of SARS-CoV-2 that triggers infected host cells to give rise to new spike proteins. 'H69-V70' is yet another mutation caused by the deletion in the 69th and 70th position of the a/a in spike protein, and it changes the shape of the spike, facilitating the virus to escape from some antibodies [4,20,21].

The 'Y144/145' mutation caused by the elimination of the 144th or 145th amino acids (tyrosine, Y) in the S Protein area challenges the effectual attachment of antibodies with the SARS-CoV-2 virus. 'ORF8 Q27stop' is another important mutation that involves the ORF8, a 121 amino acid protein whose function is yet to be completely deciphered. The 'D614 G' mutation has a moderate documented impact on transmissibility brought about by the alteration in the spike protein, where G (glycine) is substituted by D (aspartic acid). 'E484K', often referred to as the "escape mutation," brought about by a swap wherein the glutamic acid (E) is substituted by lysine (K) at position 484, shields the virus from at least one type of monoclonal antibody. 'L452R', initially detected in Denmark, is yet another mutation that has been detected in various lineages [4,22,23]. Moreover, the presence of mutations such as H69/V70 deletions, the substitution of lysine instead of threonine (T478K), and the insertion of alanine at 484 positions instead of glutamic acid (E484A) in already reported variants of concern (VOC-) have been associated to the variants' enhanced capacity to evade the defense mechanism of the body [21]. The higher ACE2 receptor binding capacity of the S Protein has been linked to the N501Y mutation. Furthermore, the S Protein's capacity to attach to the ACE2 receptor was markedly enhanced by the Q498R mutation with N501Y. These modifications make it simple for the Omicron variant to penetrate the host cell [10].

Important mutations in the Omicron variant [7] include A76V, Y145del, G339D, N440K, G446S, E484A, Q493R, G496S, Q498R, Y505H, T547K, H655Y, N679K, N764K, D796Y, N856K, Q954H, N969K, and L981F [7]. It is interesting to note that similar changes have been observed in other types, albeit with varying effects. Moreover, the Omicron variant acquired new mutations that increased its ability to spread [24,25]. Key amino acid alterations in the RBD of the S Protein [7] include G339D, S371L, S373P, S375F, K417N, N440K, G446S, S477N, T478K, E484A, Q493R, G496S, Q498R, N501Y, and Y505H [7,24,25]. Such alterations may also be linked to elevated affinities of the S Protein for the ACE2 receptor [26]. The crucial step in obtaining access into the host cell is the binding of the ACE2 receptor to the S Protein of the SARS-CoV-2 [27]. Human transmembrane protease serine 2 (TMPRSS2) cleaves the S Protein after it interacts with ACE2 receptors on the cell membrane. The S Protein is split up into its S1 and S2 subunits by TMPRSS2, which in turn makes the RBD on the S1 subunit available for interactions [28–30]. Consecutively, the S2 domain undergoes structural modifications that assist in the union of viral and cellular membranes [31,32]. It is noteworthy that according to studies using electron microscopy, the SARS-CoV-2 S Protein has a binding affinity to ACE2 that is around 10–20 times larger than that of the S proteins from other SARS-CoVs [30,33].

The S Protein of SARS-CoV-2 must be split at the S1-S2 and S2 locations in order to enter host cells [7]. Furin24, type II transmembrane serine protease (TMPRSS2), or cathepsin L are the enzymes responsible for this cleavage [34,35]. The TMPRSS2 and cathepsin L breakdown at the S2 site facilitate two distinct SARS-CoV-2 entry pathways [7]. However, mutations in certain variants are considered plausible reasons behind the changes in such entry pathways [7]. Due to it being expressed on the cell membrane, TMPRSS2 promotes the invasion via the plasma membrane as opposed to cathepsin L in the endosome, which favors the endosomal pathway [35,36]. Six different mutations in the subunit 2 (S2) of the S Protein of the Omicron variant, notably N764K, D796Y, N856K, Q954H, N969K, and L981F, were linked to variations in viral entrance into the cellular machinery and modes of transmission [37–39]. Recent investigations have shown that the Omicron variant favors

the endosomal entry pathway over the plasma membrane entrance route [40]. Infection by the Omicron spike pseudotyped virus was likewise shown to be restricted in cells that express the transcription factor TMPRSS2 but increased in cells that support an endosomal route for entry [7,40].

Recent findings suggest that genetic alterations to the Omicron S protein non-RBD may alter the mode of viral entry into host cells, which is associated with a shift in cellular tropism away from TMPRSS2-expressing cells. These findings also demonstrate why, in contrast to other VOCs, such as the Alpha, Beta, and Delta variants, Omicron replicates more rapidly in the upper respiratory system than in the lower respiratory tract [40–43]. It seems that the Omicron variety also has three significant mutations, including P681H, H655Y, and N679K, in the furin cleavage region. It is known that changes such as P681H at the polybasic cleavage site (PBCS), which are also present in other VOCs such as Alpha and Gamma, make it easier for the S protein to be digested by furin and may thus make the organism more pathogenic [44]. Hence, altogether, this information suggests that the mutations or alterations in the viral genome led to drastic changes in the characteristics of their nature to infect and disseminate among populations. As per the mutations and their impacts on the pathogenicity and transmission, many government bodies have classified the variants of SARS-CoV-2 into various categories. The following section will highlight the same.

### **2. Classification of Variants of SARS-CoV-2**

SARS-CoV-2 variants could be classified into four different groups, i.e., variants of interest (VOIs), variants of high consequence (VOHCs), variants under monitoring (VUMs), and variants of concern (VOCs) by the US Department of Health and Human Services [17–19], and all five VOCs have been further categorized as α, β, γ, δ, and Omicron variants by the World Health Organization (WHO). The Omicron variant has quickly competed with other VOCs and spread across the world [10] [Table 1].

### *2.1. Variants of Interest (VOIs)*

VOI is a variant that has genetic markers specifically linked with changes to host receptor binding, exhibiting reduced antibody neutralization production versus a previous infection by the reference virus or vaccination, and showing a reduced response to hitherto effective treatments, causing a potential diagnostic impediment, and carrying on it a label of predictive upsurge in infection. These include the B.1.525 lineage brought about by the spike protein (S protein) substitutions 69del, 144del, 70del, A67V, D614G, E484K, F888L, andQ677H, which was first detected in the United Kingdom and Nigeria in December 2020; the B.1.526 lineage brought about by spike protein substitutions A701V, D253G, D614G, E484K, L5F, T95I, and S477N, which was first detected in the United States in November 2020; the B.1.526.1 lineage brought about by spike protein (S protein) substitutions D80G, D614G, D950H, 144del, F157S, L452R, T791I, and T859N, which was first detected in the United States in October 2020; the B.1.617 lineage, brought about by spike protein (S protein) substitutions D614G, L452R, and E484Q, which was first noticed in India in February 2021; the B.1.617.1 lineage, brought about by spike protein (S protein) substitutions, i.e., D614G, E484Q, E154K, G142D, L452R, P681R, Q1071H, and T95I, which was first identified in India in December 2020; B.1.617.3 lineage, brought about by spike protein (S protein) substitutions D614G, D950N, E484Q, G142D, L452R, P681R, and T19R, which was first spotted in India in October 2020; and the P.2 lineage, brought about by spike protein (S protein) substitutions D614G, E484K, F565L, andV1176F, which was first identified in Brazil in April 2020 [45–51].

High Consequence (VOHCs)


**Table 1.** Showing the various categories of the variants of SARS-CoV-2 with their clades and origin information.
