Microalgae as an Efficient Vehicle for the Production and Targeted Delivery of Therapeutic Glycoproteins against SARS-CoV-2 Variants

Abstract

Severe acute respiratory syndrome-Coronavirus 2 (SARS-CoV-2) can infect various human organs, including the respiratory, circulatory, nervous, and gastrointestinal ones. The virus is internalized into human cells by binding to the human angiotensin-converting enzyme 2 (ACE2) receptor through its spike protein (S-glycoprotein). As S-glycoprotein is required for the attachment and entry into the human target cells, it is the primary mediator of SARS-CoV-2 infectivity. Currently, this glycoprotein has received considerable attention as a key component for the development of antiviral vaccines or biologics against SARS-CoV-2. Moreover, since the ACE2 receptor constitutes the main entry route for the SARS-CoV-2 virus, its soluble form could be considered as a promising approach for the treatment of coronavirus disease 2019 infection (COVID-19). Both S-glycoprotein and ACE2 are highly glycosylated molecules containing 22 and 7 consensus N-glycosylation sites, respectively. The N-glycan structures attached to these specific sites are required for the folding, conformation, recycling, and biological activity of both glycoproteins. Thus far, recombinant S-glycoprotein and ACE2 have been produced primarily in mammalian cells, which is an expensive process. Therefore, benefiting from a cheaper cell-based biofactory would be a good value added to the development of cost-effective recombinant vaccines and biopharmaceuticals directed against COVID-19. To this end, efficient protein synthesis machinery and the ability to properly impose post-translational modifications make microalgae an eco-friendly platform for the production of pharmaceutical glycoproteins. Notably, several microalgae (e.g., Chlamydomonas reinhardtii, Dunaliella bardawil, and Chlorella species) are already approved by the U.S. Food and Drug Administration (FDA) as safe human food. Because microalgal cells contain a rigid cell wall that could act as a natural encapsulation to protect the recombinant proteins from the aggressive environment of the stomach, this feature could be used for the rapid production and edible targeted delivery of S-glycoprotein and soluble ACE2 for the treatment/inhibition of SARS-CoV-2. Herein, we have reviewed the pathogenesis mechanism of SARS-CoV-2 and then highlighted the potential of microalgae for the treatment/inhibition of COVID-19 infection.

Keywords: ACE2; COVID-19; S-glycoprotein; SARS-CoV-2; biopharmaceuticals; edible vaccine; microalgae; synthetic biology; targeted delivery.

Figure 1

Schematic representation of the SARS-CoV-2 virus and its mechanism of entry into the human cells. The spike glycoproteins (S-glycoprotein) that cover all surfaces of SARS-CoV-2 bind to ACE2 receptors that are primarily expressed in alveolar epithelial cells in the lung and enterocytes of the small intestine. The S-glycoprotein is composed of the S1 and S2 subunits, the latter consisting of the FP, HR1, HR2, and TD domains. Upon interaction of S-glycoprotein with ACE2, TMPRSS2, and in some cases cathepsin L or B activates S-glycoprotein by proteolytic cleavage and facilitates SARS-CoV-2 virus entry by endocytosis. FP: fusion peptide domain, HR1: heptad-repeat domain I, HR2: heptad-repeat domain II, and TD: transmembrane domain.

Figure 2

Differences between ACE2 and ACE functions. ACE is the major component of the renin-angiotensin-aldosterone system (RAAS) and is involved in the inflammation and fibrosis pathways. ACE2 is a homolog of ACE that counteracts the physiological function of ACE in order that the protein has anti-inflammation and anti-fibrosis properties. In addition, ACE2 has anti-proliferation properties that can significantly inhibit SARS-CoV-2 infection. Moreover, vitamin D is involved in the suppression of ACE and also in the increase of ACE2 expression. Therefore, the administration of this molecule could be used to prevent SARS-CoV-2 infection. Ang; angiotensin, AT1R; angiotensin II type I receptor and MasR; Mas receptor.

Figure 3

The mechanism of action of an edible microalgae vaccine against SARS-CoV-2 is composed of the S1 subunit. The S1 subunit is expressed in a soluble form by the microalgae nuclear genome (I and II). Then, the transformants are lyophilized (III) and administrated orally to patients with COVID-19 infection (IV). The transgenic microalgae can successfully pass through the gastric system (V and VI) due to their thick and rigid cell wall. Upon reaching the small intestine (one of the routes of entry for SARS-CoV-2), the cell wall of microalgae is hydrolyzed by bacterial enzymes, resulting in the release of produced recombinant antigens. The released antigens (i.e., S1 subunit) are taken up by M cells and subsequently captured by antigen-presenting cells such as DC, eliciting humoral immune responses against SARS-CoV-2 by triggering various cytokines, including Th0, Th1, Th2, Th17, IL-4, IL-6, IL-23, TFG-β, and IFN-γ. Both humoral immune responses could produce secretory IgA and IgG against the SARS-CoV-2 virus. DC: dendritic cells, Th: T helper type, IL: interleukin, TFG-β: transforming growth factor beta, IFN-γ: interferon-gamma, FoxP3: Forkhead box protein P3, Tregs: regulatory T cells.

Figure 4

Targeted delivery of recombinant human ACE2 (rhACE2) produced by microalgae into the small intestine. Given the high expression of ACE2 in small intestine enterocytes and the presence of SARS-CoV-2 in this region, rhACE2 released from microalgal cells can competitively bind to SARS-CoV-2 and prevents virus entry into target cells. Viruses inactivated by rhACE2 can then be excreted from the human body via feces.