Serum IFN-γ and RNAemia temporal profiles as biomarkers of severe COVID-19 in solid organ transplant and immunocompetent patients

Abstract

•Highlights should be submitted in a separate editable file in the online submission system. Please use 'Highlights' in the file name and include 3 to 5 bullet points (maximum 85 characters, including spaces, per bullet point).•SOT recipients vs. no SOT patients have more undetectable IFN-γ and RNAemia rates.•Undetectable IFN-α rates are similar in SOT recipients and in no SOT patients.•In the no SOT patients, RNAemia and mortality decreased after 10 days of disease.•RNAemia is associated with unfavorable outcome in SOT recipients.•RNAemia and undetectable IFN-γ are associated with mortality in no SOT patients. Currently, the availability of SARS-CoV-2 vaccines, despite the great worldwide differences in vaccination rates,1WHO Coronavirus (COVID-19) dashboard. https://covid19.who.int (accessed 13 January 2023).Google Scholar have focused the impact of the pandemic in immunodepressed patients. Therefore, we read with interest the recent meta-analysis about COVID-19 vaccine in patients with solid malignancies2Chen D. Pan Y. Wang Y. Chen X. Zhang J. Zhang Y. et al.Serological response to COVID-19 vaccine and its predictors in patients with solid malignancies: a systematic review and meta-analysis of 16 cohort studies.J Infect. 2022; S0163-4453 (Epub ahead of print): 00673-00679https://doi.org/10.1016/j.jinf.2022.11.012Abstract Full Text Full Text PDF Google Scholar which found that 42% and 86% of patients achieved serological response after one and two doses, respectively. Other authors have found that the pooled odds ratio for developing anti-SARS-CoV-2 spike protein IgG was higher in the control group than in solid organ transplant (SOT) recipients.3Marra A.R. Kobayashi T. Suzuki H. Alsuhaibani M. Tofaneto B.M. Bariani L.M. et al.Short-term effectiveness of COVID-19 vaccines in immunocompromised patients: A systematic literature review and meta-analysis.J Infect. 2022; 84: 297-310https://doi.org/10.1016/j.jinf.2021.12.035Abstract Full Text Full Text PDF PubMed Scopus (22) Google Scholar The infection by SARS-CoV-2 elicits an innate and specific cellular and humoral immune response. Interferons (IFN) are key in the innate immune response during the acute phase of the viral infection, as seen with plasmacytoid dendritic cells expressing high concentrations of types I and III IFNs in COVID-19 patients.4Onodi F. Bonnet-Madin L. Meertens L. Karpf L. Poirot J. Zhang S.Y. et al.SARS-CoV-2 induces human plasmacytoid predendritic cell diversification via UNC93B and IRAK4.J Exp Med. 2021; 218e20201387https://doi.org/10.1084/jem.20201387Crossref PubMed Google Scholar,5Severa M. Diotti R.A. Etna M.P. Rizzo F. Fiore S. Ricci D. et al.Differential plasmacytoid dendritic cell phenotype and type I interferon response in asymptomatic and severe COVID-19 infection.PLoS Pathog. 2021; 17e1009878https://doi.org/10.1371/journal.ppat.1009878Crossref Scopus (31) Google Scholar In acute COVID-19 and convalescent patients, intracellular cytokine staining after stimulation with SARS-CoV-2 peptide pools, has showed significant IFN-γ increases in CD8 + T-cells, which is associated with viral elimination, and without differences in both phases of the disease.6Rydyznski Moderbacher C. Ramirez S.I. Dan J.M. Dan J.M. Grifoni A. Hastie K.M. et al.Antigen-specific adaptive immunity to SARS-CoV-2 in acute COVID-19 and associations with age and disease severity.Cell. 2020; 183 (e19): 996-1012https://doi.org/10.1016/j.cell.2020.09.038Abstract Full Text Full Text PDF PubMed Scopus (889) Google Scholar,7Jasim S.A. Mahdi R.S. Bokov D.O. Najm M.A.A. Sobirova G.N. Bafoyeva Z.O. et al.The deciphering of the immune cells and marker signature in COVID-19 pathogenesis: an update.J Med Virol. 2022; 94: 5128-5148https://doi.org/10.1002/jmv.28000Crossref PubMed Scopus (0) Google Scholar In this context, to gain insight in the innate immune response in SOT recipients with COVID-19, compared with no SOT patients, we have assessed the IFN-α and IFN-ℽ serum levels and RNAemia at hospital admission and by days from the symptom's onset (DfSO; ≤1 to ≥15 days), as well as their association with unfavorable clinical outcomes (death and/or invasive mechanical ventilation [IMV]). With this aim, we conducted a multicentre prospective observational cohort study, including consecutive adult inpatients with confirmed COVID-19 (RT-PCR in nasopharyngeal swabs) and available samples for IFN-α/IFN-γ serum levels and RNAemia determinations (Methods, Supplementary material), from January 6th 2020 to August 13th 2021, followed until hospital discharge, death, or 30 days, whichever occurred first. The study was approved by The Ethics Committee of University Hospitals Virgen Macarena and Virgen del Rocío (C.I. 0771-N-20 and 0842-N-20). Data were separately analyzed for SOT recipients and no SOT patients. In addition, we performed a matched cohort analysis in which patients undergoing SOT were paired with those from the no SOT cohort (1:2) according to their propensity score (PS), using callipers of a 0.01 standard deviation, to control for residual confounders. Mortality in the matched pairs was compared using Cox regression. IFN-α and IFN-γ levels were analyzed as discrete (undetectable and detectable) and continuous (pg/mL) variables. Multivariate Cox regression and logistic regression analysis were performed to identify factors independently associated with 30-day all-cause mortality and unfavorable clinical outcomes (Methods, Supplementary material). Forty-seven (10.3%) SOT recipients and 408 (89.7%) no SOT patients (Supplementary Table 2) were recruited. The mean DfSO to hospital admission was 7.1 ± 4.3, without differences between groups. Undetectable IFN-α occurred in 8.5% and 13.5% (p = 0.36) of both groups, respectively (Fig. 1A), independently of the DfSO. In SOT recipients, IFN-α levels were higher with <7 DfSO than with ≥7 DfSO (p = 0.015), with a decrease from 19.5 pg/mL to 1.4 pg/mL. In no SOT, IFN-α levels were higher than in SOT recipients with ≥7 DfSO (Supplementary Table 3). Undetectable IFN-γ was more frequent in SOT recipients than in no SOT patients (42.6% and 19.4%, p < 0.001) and this difference was higher with ≥7 DfSO (Fig. 1A). IFN-γ levels were similar over the different time-periods, both in SOT and no SOT, and without differences between groups (Supplementary Table 3), which is consistent with other studies.6Rydyznski Moderbacher C. Ramirez S.I. Dan J.M. Dan J.M. Grifoni A. Hastie K.M. et al.Antigen-specific adaptive immunity to SARS-CoV-2 in acute COVID-19 and associations with age and disease severity.Cell. 2020; 183 (e19): 996-1012https://doi.org/10.1016/j.cell.2020.09.038Abstract Full Text Full Text PDF PubMed Scopus (889) Google Scholar,7Jasim S.A. Mahdi R.S. Bokov D.O. Najm M.A.A. Sobirova G.N. Bafoyeva Z.O. et al.The deciphering of the immune cells and marker signature in COVID-19 pathogenesis: an update.J Med Virol. 2022; 94: 5128-5148https://doi.org/10.1002/jmv.28000Crossref PubMed Scopus (0) Google Scholar RNAemia was more frequent in SOT recipients (57.4%) than in no SOT patients (18.9%, p < 0.001) (Fig. 1A, Supplementary Table 2). In SOT recipients, RNAemia detection was independent of the DfSO, and in no SOT patients decreased with ≥11 DfSO (p = 0.014) (Supplementary Table 3). Mortality was higher in SOT recipients than in no SOT patients with ≥4 DfSO (Fig. 1A). In SOT recipients, mortality was not associated to the DfSO at admission; however, in no SOT patients, mortality was much higher in patients with ≤3 DfSO, decreasing to 0% in patients with ≥11 DfSO (p < 0.001) (Fig. 1A).Fig. 1SOT recipients (n = 47) comparison with no SOT patients (n=408) regarding (A) undetectable IFN-α and IFN-γ serum levels, RNAemia, and mortality, by days from symptoms onset at hospital admission, and (B) Survival Kaplan Meier analysis of patients with and without undetectable IFN-γ serum levels and RNAemia detection.View Large Image Figure ViewerDownload Hi-res image Download (PPT) In the PS matched cohorts (Table 1), SOT recipients showed higher prevalence of undetectable IFN-γ than no SOT patients (39.4% vs. 10.6%, respectively; p = 0.001), lower plasma IFN-α and IFN-γ levels in those with RNAemia (p = 0.013 and p = 0.001, respectively; Supplementary Fig. 1B), higher RNAemia detection (57.6% vs. 13.6%; p < 0.001) and mortality (27.3% vs. 4.5%; p = 0.003).Table 1Comparison of solid organ transplant (SOT) recipients matched (1:2) with no SOT patients according to propensity score.aVariables included in the propensity score were sex, dyspnea, SpO2, neutrophil and lymphocyte counts, C-reactive protein, ferritin, D-dimer, and LDH.VariableSOT recipients (n = 33)No SOT patients (n = 66)P valueMale sex20 (60.6)37 (56.1)0.666Age >70 years4 (12.1)21 (31.8)0.033Dyspnoea15 (45.5)36 (54.5)0.394SpO2 <95%17 (51.5)26 (39.4)0.251Neutrophil count >7500/μL6 (18.2)6 (9.1)0.327Lymphocyte count <1000/µL18 (54.5)30 (45.5)0.394C-reactive protein >100 mg/L9 (27.3)20 (30.3)0.755Ferritin >1000 ng/mL6 (18.2)10 (15.2)0.699D-dimer >600 ng/mL29 (87.9)48 (72.7)0.087LDH >300 IU/L14 (42.4)33 (50.0)0.477IFN-α undetectable3 (9.1)8 (12.1)0.747IFN-α (pg/mL)bMedian (IQR). P values are calculated by the Mann-Whitney U test.1.43 (0.60–22.01)11.98 (3.24–23.11)0.163IFN-γ undetectable13 (39.4)7 (10.6)0.001IFN-γ (pg/mL)bMedian (IQR). P values are calculated by the Mann-Whitney U test.26.14 (0.00–240.96)145.35 (40.00–330.96)0.347RNAemia positive19 (57.6)9 (13.6)<0.001RNAemia (log10 copies/mL)bMedian (IQR). P values are calculated by the Mann-Whitney U test.2.38 (2.12–3.19)2.36 (1.92–2.98)0.921CCI ≥325 (75.8)37 (56.1)0.056CURB-65 ≥212 (36.4)12 (18.8)0.057WHO basal score 6–9cSeverity rating according to the WHO Clinical Progression Scale (doi: 10.1016/S1473-3099(20)30483-7), ranged from 0 (not infected) to 10 (dead), of which scores 6–9 represent severe disease.3 (9.1)4 (6.1)0.683IMV10 (30.3)5 (7.6)0.003Mortality at day 309 (27.3)3 (4.5)0.003WHO final score 7-10cSeverity rating according to the WHO Clinical Progression Scale (doi: 10.1016/S1473-3099(20)30483-7), ranged from 0 (not infected) to 10 (dead), of which scores 6–9 represent severe disease.13 (39.4)7 (10.6)0.001Data are presented as No. (%). P values are calculated by Cox regression.Abbreviations (in order of appearance): SpO2, peripheral capillary oxygen saturation; LDH, lactate dehydrogenase; IFN, interferon; CCI, Charlson Comorbidity Index (30); CURB-65 (31), Severity Score for Community-Acquired Pneumonia; IMV, invasive mechanical ventilation.a Variables included in the propensity score were sex, dyspnea, SpO2, neutrophil and lymphocyte counts, C-reactive protein, ferritin, D-dimer, and LDH.b Median (IQR). P values are calculated by the Mann-Whitney U test.c Severity rating according to the WHO Clinical Progression Scale (doi: 10.1016/S1473-3099(20)30483-7), ranged from 0 (not infected) to 10 (dead), of which scores 6–9 represent severe disease. Open table in a new tab Data are presented as No. (%). P values are calculated by Cox regression. Abbreviations (in order of appearance): SpO2, peripheral capillary oxygen saturation; LDH, lactate dehydrogenase; IFN, interferon; CCI, Charlson Comorbidity Index (30); CURB-65 (31), Severity Score for Community-Acquired Pneumonia; IMV, invasive mechanical ventilation. In SOT recipients, the multivariate logistic regression model selected RNAemia as predictor of unfavorable clinical outcome (Supplementary Table 6). Regarding no SOT patients, in the Cox regression multivariate analysis, 30-day all-cause mortality was associated with RNAemia and undetectable IFN-γ levels (Supplementary Table 7). In the Kaplan-Meier analysis, patients with RNAemia had lower survival, both in SOT (p < 0.0133) and no SOT (p = 0.001) groups (Fig. 1B). RNAemia has been associated with COVID-19 mortality.8Brunet-Ratnasingham E. Anand S.P. Gantner P. Dyachenko A. Moquin-Beaudry G. Brassard N. et al.Integrated immunovirological profiling validates plasma SARS-CoV-2 RNA as an early predictor of COVID-19 mortality.Sci Adv. 2021; 7: eabj5629https://doi.org/10.1126/sciadv.abj5629Crossref PubMed Scopus (14) Google Scholar The present data confirm it, with a higher sample size and including SOT recipients, in which the RNAemia impact had not yet been analyzed. The Kaplan-Meier analysis also showed an association of undetectable IFN-γ with lower survival in SOT recipients (p = 0.048) (Fig. 1B). Our results, showing an association of undetectable IFN-γ in serum with mortality, support the protective role of the specific T-cells response. The Kaplan-Meier analysis did not show association of undetectable IFN-α levels with the survival at 30 days, both in SOT and no SOT groups. It has been reported that inborn errors of type I IFN immunity accounts for life-threatening COVID-19 pneumonia9Bastard P. Rosen L.B. Zhang Q. Michailidis E. Hoffmann H.H. Zhang Y. et al.Autoantibodies against type I IFNs in patients with life-threatening COVID-19.Science. 2020; 370: eabd4585https://doi.org/10.1126/science.abd4585Crossref PubMed Scopus (1291) Google Scholar and that autoantibodies against type I IFNs increased the infection fatality rate.10Manry J. Bastard P. Gervais A. Le Voyer T. Rosain J. Philippot Q. et al.The risk of COVID-19 death is much greater and age dependent with type I IFN autoantibodies.Proc Natl Acad Sci U S A. 2022; 119e2200413119https://doi.org/10.1073/pnas.2200413119Crossref PubMed Scopus (25) Google Scholar However, we have not found association of serum undetectable IFN-α with unfavorable outcome, including immunosuppressed patients as the SOT recipients. A limitation is that we did not analyse IFN-stimulated genes to define a type I IFN signature nor interferon autoantibodies, because of our purpose was to identify easy-to-measure variables in the clinical setting. In summary, the present results support the RNAemia and IFN-γ serum levels determinations, at hospital admission, in all adult COVID-19 patients, to guide their management and to assess the antiviral therapy efficacy in the case of RNAemia. The authors have declared that no competing interests exist. JS-C, EC, JMC, and JP conceived and designed the study; obtained public funding from the Spanish Ministry of Economy, Industry, and Competitiveness; and took responsibility for the integrity of the data and the accuracy of its analysis. JP and SS-A did the scientific literature search. PC-M, JB-C, CI, RR-A, JA, PP-P, EG-D, CR, JPR, MJB-V, SS, RV-O, JNA, CG-G, MJB-V, DG-C, NM, GB, MAG-B, JMS, MA-G, RA-M, JG-A, JAO, ZRP-B, AP, JAL and JRB supported the inclusion of patients and the acquisition of data. JS-C, MC-L and JB-C processed the research clinical samples and obtained the data. SS-A, MC-L and JB-C processed the data, SS-A, MC-L and JP did the statistical analysis. SS-A, MC-L, JP, MA-G, JS-C and EC did the interpretation of data and wrote the draft of the manuscript. All authors critically revised the manuscript for important intellectual content and gave final approval for the version published. This study was supported by Plan Nacional de I+D+i 2013‐2016 and Instituto de Salud Carlos III, Subdirección General de Redes y Centros de Investigación Cooperativa, Ministerio de Ciencia, Innovación y Universidades, Spanish Network for Research in Infectious Diseases (REIPI RD16/0016/0001, RD16/0016/0009, RD16/0016/0013); co‐financed by European Development Regional Fund “A way to achieve Europe”, Operative Program Intelligence Growth 2014‐2020. SS-A was supported by a grant from the Instituto de Salud Carlos III, Ministerio de Ciencia e Innovación, Proyectos de Investigación sobre el SARS-CoV-2 y la enfermedad COVID-19 (COV20/00370, COV20/00580). JAL, JMC, MA-G, RA-M, JS-C, and EC (CB21/13/00006) and PP-P, ZRP-B, AP, and JRB (CB21/13/00012) also were supported by CIBERINFEC - Consorcio Centro de Investigación Biomédica en Red, Instituto de Salud Carlos III, Ministerio de Ciencia e Innovación and Unión Europea – NextGenerationEU. JS-C is a researcher belonging to the program “Nicolás Monardes” (C-0059-2018), Servicio Andaluz de Salud, Junta de Andalucía, Spain. The funders have not role nor any influence on the design, data analysis or interpretation. Download .docx (.56 MB) Help with docx files