SARS-CoV-2 project 2(AbSano)

Thymidine phosphorylase (TYMP), which facilitates platelet activation and thrombosis, is significantly increased in COVID-19 patients. We hypothesize that TYMP mediates SARS-CoV-2 spike protein (SP)-induced thrombosis.

Plasmids encoding wildtype SP or empty vector (p3.1) were transfected into COS-7 cells, and cell lysates were prepared as a reservoir for SP or p3.1 (control), respectively. K18-hACE2^TG and K18-hACE2^TG/Tymp^-/- mice were treated with a single dose of SP or p3.1 by intraperitoneal injection and then subjected to thrombosis studies three days later. The role of SP on inflammatory signaling activation was assessed in BEAS-2B cells.

SARS-CoV-2 SP increased the expression of TYMP, resulting in the activation of STAT3 and NF-κB in BEAS-2B cells. A siRNA-mediated knockdown of TYMP attenuated SP-enhanced activation of STAT3. SP significantly promoted arterial thrombosis in K18-hACE2^TG mice. SP-accelerated thrombosis was attenuated by inhibition or genetic ablation of TYMP. SP treatment did not influence ADP- or collagen-induced platelet aggregation but significantly increased platelet adhesion to fibrinogen. SP treatment also significantly shortened activated partial thromboplastin time, which was reversed and even prolonged by TYMP deficiency. Additionally, SP binds to platelet factor 4 (PF4) and TYMP. TYMP does not bind PF4 but enhances the formation of the SP/PF4 complex, which may augment the procoagulant and prothrombotic effect of PF4.

We conclude that SP is prothrombotic and upregulates TYMP expression, and TYMP inhibition or knockout mitigates SP-enhanced thrombosis. These findings suggest that inhibition of TYMP may be a novel therapeutic strategy for COVID-19-associated thrombosis.

The coronavirus that causes serious acute respiratory syndrome. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is still a major problem in public health and biomedicine. Even if there is no cure for it, the infection is still progressing naturally, and the only time that optimal treatment choices, such as doxycycline, work is at the beginning of the infection. Our project is structured into two critical parts: the first focuses on the identification of potential drug targets, and the second on vaccine design, both aimed at exploring new ways to treat the disease. Initially, cytoplasmic proteins identified through subtractive analysis underwent comprehensive evaluation for potential drug targeting, focusing on metabolic pathways, homology prediction, drugability assessment, essentiality, and protein-protein interactions. Subsequently, surface proteins underwent rigorous assessment for allergenicity, antigenicity, physiochemical attributes, conserved regions, protein interactions, and identification of B and T cell epitopes. Molecular docking and immunological simulation analyses were then employed to develop and characterize a multi-epitope vaccine, integrating findings from the aforementioned evaluations. Findings from the study point to six proteins as potential critical therapeutic targets for SARS-CoV-2, each of which is involved in a distinct metabolic process. The reverse vaccinology analysis suggested that the following proteins could be used as vaccine candidates: sp|P05106, sp|O00187, sp|Q9NYK1, sp|P05556, sp|P09958, and sp|Q9HC29. Four multi-epitope vaccine named as SARS-COV-2-, C1, C2, C3, and C4 was designed by utilizing different adjuvants and eighteen B cell overlapped epitopes which were predicted from top ranked protiens. Based on immune simulation study, the vaccine exhibited adequate immune-reactivity and favorable encounters with toll-type receptors (TLR4, TLR8, HLA, etc ACE), Among them the SARS-COV-2-C2 showed best binding affinity of which all receptors. Findings from this study could be a game-changer in the quest to develop a vaccine and medication that effectively combat SARS-CoV-2. It is necessary to do additional experimental analyses, nevertheless.