Antibodies: A Double Leg Takedown Against COVID-19.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is responsible for a pandemic that has, thus far, caused over 400 million cases of coronavirus disease-19 (COVID-19) with a case/fatality rate of about 1.4% [1], depending on the viral variant involved [2]. The infection may be associated with symptoms ranging from low-grade upper respiratory symptoms and anosmia/dysgeusia to severe interstitial pneumonia, thromboembolism, myocarditis, acute kidney and liver injury, acute respiratory distress syndrome, multiorgan failure, and death [3]. Moreover, some survivors report manifestations that persist beyond the acute illness, the so-called long COVID that may often include a constellation of debilitating symptoms [4].

The pathogenesis of tissue damage in COVID-19 is complex and multifaceted. Akin to most coronaviruses, SARS-CoV-2 has developed evasion strategies to limit host control and enhance replication and transmission (reviewed in [5]). However, invasive SARS-CoV-2 infection is characterized by excessive immune activation leading to systemic inflammation and severe disease. Although innate immunity is thought to represent an essential first line of defense against SARS-CoV-2 infection, adaptive immunity is key to eliminate the virus. Of note, mildly symptomatic individuals develop highly functional virus-specific CD4+ and CD8+ T cells, suggesting that robust and coordinated T-cell responses provide protection against progression to severe disease 
[6, 7]. After natural infection some of these cells persist as memory T cells, which may also cross-react with those elicited by past coronavirus infections, such as those that cause the common cold [8], and with variants of concern [9]. Effector memory T cells travel between peripheral tissue and the lymph nodes, stimulating B cells in germinal centers [10]. B cells are the other pillar of adaptive immunity and differentiate into plasma cells secreting neutralizing antibodies, which may provide sterilizing immunity against certain viral infections [11]. Interestingly, nonneutralizing antibodies may also provide protection in different ways, that is by elimination of virus-infected cells via CD16 (FcγRIII)-mediated antibody-dependent cellular cytotoxicity (ADCC) that makes use of effector cells such as natural killer (NK) cells binding to the Fc portion of immunoglobulin G (IgG) which, in turn, recognize viral proteins expressed on target cells [12]. Beside NK cells, effectors may include other subsets of cells expressing FcγRIII such as monocyte/macrophages, NKT cells, or γδ T cells. Fc-mediated effector functions also include mechanisms other than ADCC, such as antibody-mediated complement activation, antibody-dependent cellular phagocytosis, antibody-dependent cell-
mediated virus inhibition, and antibody-mediated virus opsonization. All these functions could be beneficial in fighting viral infections. However, while ADCC has been clearly shown to contribute to killing of tumor targets, its role in viral infections has been only superficially investigated, with the notable exception of HIV [13].

In this issue of the Journal of Infectious Diseases, Rieke and colleagues address the potential role of circulating antibodies in mediating ADCC in patients with natural SARS-CoV-2 infection and SARS-CoV-2–vaccinated individuals [14]. Their data show that while serum concentrations of anti-spike antibodies tend to wane over time in convalescing and vaccinated individuals, ADCC persists more efficiently and for longer in the former compared with the latter. The data are intriguing because of possible implications on the hyperinflammatory state typical of severe COVID-19 or on protection from SARS-CoV-2 infection.

Emerging evidence indicates that both neutralizing and Fc-mediated effector functions of antibodies (including ADCC) contribute to protection against SARS-CoV-2. However, it is unclear whether Fc-effector functions alone can protect against SARS-CoV-2 or can do so in combination with neutralizing activity. Interestingly, some Fc mutations significantly diminish the affinity of antibodies for FcγRIIIa and also impact Fc-mediated effector functions. Of note, studies in animal models showed that selected mutations diminish the capacity of neutralizing antibodies to protect mice from a lethal SARS-CoV-2 challenge [15]. Similarly, 2 other studies examining humoral responses in acutely infected individuals found that Fc-mediated effector functions were associated with survival [16, 17]. Thus, efficient Fc-mediated effector functions may potently contribute to the in vivo efficacy of anti-SARS-CoV-2 antibodies.

In the setting of viral infections, ADCC assays are usually set up with virus-infected or viral protein-expressing cells as target. To this end, a cell line stably expressing a green fluorescent protein-tagged SARS-CoV-2 spike has recently been developed to measure ADCC in SARS-CoV-2 infection, either using spike-specific monoclonal antibodies or plasma from previously infected or vaccinated individuals [18]. For practical purposes, the ADCC assay employed in Rieke et al’s study was, instead, a surrogate assay that focused on the effector rather than on the target cells [14]. It followed that cytotoxicity was measured as effector cell degranulation, for example CD107a expression, and not as target cell killing. In essence, what the authors showed is antibody recognition of a cell-free soluble protein coated on plastic. This is because cell membrane expression of SARS-CoV-2 spike protein was reported to be inconsistent and variable; hence, experiments lacked reproducibility. Interestingly, the spike protein was shown to be expressed on infected human airway epithelial cells, and antibodies from infected individuals were shown to bind to them [19], suggesting that cell membrane-bound immunoglobulin may mediate ADCC or other Fc-mediated effector functions. In any case, there is currently no perfect assay that can recapitulate ADCC responses against all the different aspects of virus infections.

Despite the limitations of the assay used in this study, the authors were able to show persistence of immunoglobulin Fc-mediated effector function for longer in naturally infected individuals. Differences in fucosylation of vaccine-induced versus infection-induced antibodies have been proposed by the authors as being responsible for such a phenomenon but, unfortunately, no direct evidence in support of this interesting hypothesis was provided. In addition, all NK cells might not be equal in their potential to mediate ADCC, and how vaccines might engage NK cells with higher ADCC potential is of considerable interest to the field. Along these lines, a new subset of NK cells expressing low levels of intracellular γ-signaling chain of Fc receptor (FcεRIγ) has recently been identified 
[20, 21]. These cells, named adaptive/memory NK cells, have increased expression of NKG2C and killer-cell immunoglobulin-like receptors (KIRs) and reduced expression of sialic acid-binding Ig-like lectin 7 (Siglec-7), NKG2A, NKp30, T-cell immunoglobulin, and mucin domain-3, and show enhanced ADCC [22]. Adaptive NK cells represent a minor subset of bulk NK cells and can be expanded in cytomegalovirus-positive subjects and in other viral infections [22]. Reduced adaptive NK-cell function may be restored following viral cure, as in hepatitis C [23]. Adaptive NK cells also express CD57, which defines them as a highly differentiated NK population, and their frequency has been shown to be increased in patients with full-blown COVID-19, particularly in those with severe disease [24]. Selective expansion of adaptive NK cells may, therefore, provide at least a partial explanation for prolonged Fc-mediated antibody effector functions, namely ADCC, in the setting of SARS-CoV-2 infection. By pinpointing this important and often neglected function of antibodies, Rieke and colleagues have paved the way to further research in the field. Indeed, ADCC may be more efficient at virus elimination than neutralizing antibodies that can only operate in the extracellular environment. Combining neutralization and Fc-mediated effector function is like a double leg takedown to combat viral infections.