Early Phase Preclinical Development of PACECOR, a Mutation-Independent Anti-SARS-Co2 Therapeutic Strategy

The proposed project is an integrated effort towards the prevention of infective spread of the SARS-CoV-2 virus and related viruses. To approach such challenge, the study will exploit the amalgamation of transversal competences, spanning from physical chemistry and bioinformatics to biomedicine, such as to warrant the operational multidisciplinary nature of the endeavours needed to take upon the various phases of the research programme. It is axiomatic that virus entrance into target cells is a key step to abrogate for halting infection, preventing the subsequent viremia in the host and viral dissemination. Current approaches in this direction have been biased towards targeting the virus, most conveniently through the generation of specific immunological tools. However, an alternative approach could be to devise tolerable and highly selective pharmacological probes aimed at obstructing ancillary components of the virus-host interaction, whille desensitising the impact of genotypic changes of the virus. Bearing this conceptual framework in mind, we propose to manufacture, and validate at an early preclinical level, an innovative bimodal anti-viral complex named PACECOR - "Pre-targeting ACE2-binding of CORona virus". The pharmacological probe is designed such as to exploit the targeting efficiency of a synthetic, next-generation antagonist of the angiotensin converting enzyme 2 (ACE2), which is functionalized with a biotin moiety in order to grant an irreversible engagement with an engineered avidin anchor of a second competitor of the Spike protein-ACE2 interaction. Bioinformatics approaches involving in silico structural modelling and computational molecular analyses will be employed, in conjunction with chemical derivation of small-molecule inhibitors of the catalytic ACE2 activity, for determination of structural-functional properties the ACE2 antagonists. Analogous in silico efforts will be conducted in parallel to support recombinant production of a set of chimeric polypeptides containing avidin isoforms linked to the reproduced receptor binding domain of corona viruses Spike proteins. Functional properties and lack of collateral effects of the PACECOR complex will be consolidated through biochemical/proteomic procedures and cell-based assays involving multimarker fluorescent platforms, flow cytometric technologies and high-resolution fluorescence microscopy. Anti-viral efficacy of the lead PACECOR complexes will be determined in infection assays in vitro using pseudotyped lentil viruses, SARS-CoV-2 and genetically modified cell lines. Effective PACECOR complexes will then be tested for their systemic anti-viral properties in the established SARS-CoV-2-infected and non-infected TG hACE2 transgenic “COVID-19 mouse model”. The outcome of the project will advance our understanding of the dynamics of SARS-CoV-2 infection and provide a prototype agent for a novel anti-viral therapeutic paradigm to be further developed preclinically.