IDENTIFICATION OF DIFFERENT PATHOGENICITY GENETIC PROFILES AND SCREENING OF POLYMORPHISMS F2 rs1799963 AND F5 rs6025 IN COVID-19 PATIENTS

Introduction and Aim. The clinical presentation of COVID-19, caused by the SARS-CoV-2 virus, widely varies, ranging from asymptomatic infections to severe respiratory distress, multi-organ failure, and death; in addition to significantly increased mortality rates and severe inflammatory response observed in COVID-19 patients, SARS-CoV-2 infections are also characterized by a high prevalence of thrombotic complications. While age, sex, and comorbidities such as hypertension and diabetes are well-established risk factors, host genetic variability has emerged as a critical determinant of individual susceptibility and disease progression. Additionally, hereditary thrombophilia, including Factor V Leiden and Prothrombin G20210A polymorphisms, has been linked to increased thrombotic complications in COVID-19 patients, particularly in patients with elevated D-dimer levels. The aim of the study was to identify genetic variants and profiles associated with severity of the disease and thrombotic events susceptibility. Methods. NGS analysis was conducted on a cohort of n=80 patients with COVID-19; genetic analysis included a sequencing panel of 11 genes (PROC, PROS1, FGA, FGB, FGG, SERPINC1, F2, F5, F10, PLAT, PLG) known to be involved in the coagulation processes. Moreover, a genotyping analysis of rs6025 (FV Leiden) and rs1799963 (Prothrombin G20210A) polymorphisms has been conducted through Real Time PCR on the whole cohort of n=994 patients hospitalized at Careggi Hospital with COVID-19. In these patients, coagulation parameters, which are routinely assessed upon patient admission to the emergency room or hospital ward, were also evaluated. Results. As regards NGS analysis, 69 rare variants (MAF≤1%) have been identified at the heterozygous state in 50 of the 80 COVID-19 patients studied. The majority (n=45/69, 65%) of rare variants were found in patients who experienced a more severe clinical course with respect to patients admitted to general wards (non-ICU, 24/69, 35%). 28/40 (70%) ICU/Death patients showed at least one rare variant with respect to Non-ICU patients (22/40, 55%), p<0.166. Concerning the distribution of the n=18 uncertain, potentially pathogenic or pathogenic variants (VUS/LP/P) out of 69 identified, 67% were in ICU/Death and 33% in Non-ICU patients, and 9/40 (23%) ICU/Death patients showed at least one rare variant with respect to non-ICU patients in which 6/40 (15%) showed at least one rare variant (p=0.390). In particular, patients with severe phenotypes presented LP/P mutations or burden of variants with a strong potential of pathogenicity. Concerning common genetic thrombophilia, in the whole n=994 COVID-19 patients cohort, n=45 were heterozygous for rs1799963 polymorphism and n=31 were heterozygous for rs6025 polymorphism. MAF for F2 rs1799963 and F5 rs6025 was comparable to that observed in the Tuscany population. We obtained verified clinical data on thrombotic complications for n=324 individuals; 19/324 patients (5.9%) experienced a thromboembolic event. No difference in MAF of F2 polymorphism was observed between COVID-19 patients with and without thrombotic events, whereas a trend towards a higher prevalence of FV Leiden was observed in patients with thrombotic events. The evaluation of coagulation parameters highlighted higher values of PT-INR and D-Dimer in patients with thromboembolic event, compared to patients in whom no complication was found. Conclusions. The presence of common genetic factors of hereditary thrombophilia does not seem to indicate a significant contribution in modulating the risk of developing thromboembolic complications in SARS-CoV-2 patients; on the other hand, elevated D-dimer and PT-INR levels emerged as a strong predictor of poor outcomes. As regard high-throughput sequencing, it revealed a higher prevalence of rare variants in genes involved in coagulation processes in patients with severe disease (ICU admission or death), suggesting a link between genetic predisposition and disease severity, in particular linked to thromboembolic complications. These findings support further research into genetic contributions to guide personalized risk assessment and patients’ management.