Enhanced expression of immune checkpoint receptors during SARS-CoV-2 viral infection

Abstract

The immune system is tightly regulated by the activity of stimulatory and inhibitory immune receptors. This immune homeostasis is usually disturbed during chronic viral infection. Using publicly available transcriptomic datasets, we conducted in silico analyses to evaluate the expression pattern of 38 selected immune inhibitory receptors (IRs) associated with different myeloid and lymphoid immune cells during coronavirus disease 2019 (COVID-19) infection. Our analyses revealed a pattern of overall upregulation of IR mRNA during severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. A large number of IRs expressed on both lymphoid and myeloid cells were upregulated in nasopharyngeal swabs (NPSs), while lymphoid-associated IRs were specifically upregulated in autopsies, reflecting severe, terminal stage COVID-19 disease. Eight genes (BTLA, LAG3, FCGR2B, PDCD1, CEACAM1, CTLA4, CD72, and SIGLEC7), shared by NPSs and autopsies, were more expressed in autopsies and were directly correlated with viral levels. Single-cell data from blood and bronchoalveolar samples also reflected the observed association between IR upregulation and disease severity. Moreover, compared to SARS-CoV-1, influenza, and respiratory syncytial virus infections, the number and intensities of upregulated IRs were higher in SARS-CoV-2 infections. In conclusion, the immunopathology and severity of COVID-19 could be attributed to dysregulation of different immune inhibitors. Targeting one or more of these immune inhibitors could represent an effective therapeutic approach for the treatment of COVID-19 early and late immune dysregulations.

Keywords: CEACAM1; COVID-19; SARS-CoV-2; SIGLEC10; immune checkpoint inhibitors; immune inhibitory receptors; influenza A virus; lung autopsies; respiratory viral infection; sialic acid.

Graphical abstract

Figure 1

Gene expression of immune inhibitory receptors in nasopharyngeal swabs and lung autopsies of COVID-19 patients (A) Enhanced expression of 31 immune inhibitory receptors known to be associated with lymphoid and myeloid immune cells in nasopharyngeal swabs (n = 431 COVID-19 versus n = 54 controls, GEO: GSE152075). (B) Upregulation of nine immune inhibitors associated mostly with lymphoid immune cells in lung autopsies (n = 16 COVID-19 versus n = 5 control lung autopsies, GEO: GSE150316). (C) Genes that are shared between nasopharyngeal swabs and lung autopsies. Seven of the eight genes were more expressed in lung autopsies compared to swabs. Results are presented as fold change of gene expression between cases and controls.

Figure 2

Association between SARS-CoV-2 viral levels and gene expression of immune inhibitory receptors Gene expression of immune inhibitory receptors was positively correlated with viral load as shown in (A)–(H) (n = 431 COVID-19 versus n = 54 controls, GEO: GSE152075). An unpaired Student’s t test was used to compare between the independent groups (mRNA expression between different viral load groups). Results are presented as mean (±SEM) of mRNA expression. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001.

Figure 3

Single-cell expression of bronchoalveolar lavage and PBMC immune cells in patients with COVID-19 (A) and (B) present the bronchoalveolar lavage data (GEO: GSE145926), while (C) and (D) present the data from PBMCs (GEO: GSE149689). Single-cell RNA sequencing was performed on bronchoalveolar lavage fluid (BALF) from 6 severe and 3 moderate COVID-19 patients and 3 healthy controls, and on PBMCs from 5 flu patients, 11 COVID-19 patients, and 4 healthy controls. (A) Expression of immune inhibitors in M1- and M2-like macrophage groups, which are enriched in severe COVID-19 patients. Fold changes were generated for each group of macrophages relative to total macrophages. (B) Specific upregulation of KLRC1, CD244 (2B4), and PECAM in moderate CD8+ T cells, while CTLA4, HAVCR2 (TIM3), and TIGIT were enhanced in CD8⁺ T cells from severe COVID-19 patients. The fold change compares expression of IRs in CD8⁺ T cells between moderate and severe groups. (C) Differential expression of IRs on monocytes during severe and mild course of COVID-19 disease within the same patients (n = 1, GEO: GSE149689). (D) Upregulation of HAVCR2 (TIM3) and CTLA4 in COVID-19-specific CD8⁺ T cell cluster. None of the immune inhibitors appeared in IAV-specific clusters.

Figure 4

Expression of IRs during COVID-19 and other viral infections The number and intensity of IR upregulation is higher during COVID-19 compared to other respiratory viral infection. (A) Upregulation of immune inhibitory genes during different respiratory infections. The difference in gene expression of IRs of case and controls is provided as fold change. (B) Intersection of upregulated immune inhibitory signatures in four different respiratory viral infections (IAV, RSV, SARS-CoV-1, and SARS-CoV-2). (C) Expression of CEACAM1 gene shared between all viral groups. The following datasets were used; GEO: GSE17156 (n = 17 IAV versus n = 17 controls), GEO: GSE17156 (n = 20 RSV versus n = 20 controls), GEO: GSE1739 (n = 10 SARS-CoV-1 versus n = 4 controls), and EGAS00001004503 (n = 39 COVID-19 versus n = 10 controls). For all analyses, p < 0.05 was considered significant. IAV, influenza A virus; RSV, respiratory syncytial virus.