Application of synthetic biology in the construction of a genetic circuit to improve the anti-tumor effect of Salmonella spp

Author(s)

Kauan Ribeiro de Sena Gomes

• Document type: Master's Dissertation
• Press: Ribeirão Preto.
• Institution: Universidade de São Paulo (USP). Faculdade de Medicina de Ribeirão Preto (PCARP/BC)
• Defense date: 2020-03-26

Examining board members

Rafael Silva Rocha; Leticia Fröhlich Archangelo; Marco Antonio Stephano

• Advisor: Rafael Silva Rocha

Abstract

The engineering and construction of a biological machine, designed to develop a therapeutic activity, such as the fight against malignant cells, and designed to act within the complex space of a living individual, requires the concatenation of an intricate network of biological processes, internal and external . These processes have been described and annotated using a new scientific approach that employs the same logic as electronic computing systems to biological natural systems. This new biotechnological field, coined as Synthetic Biology, seeks to obtain genetic constructions capable of imbuing living organisms with new functionalities, obeying a defined, robust and precise blueprint. Bacteria are microorganisms capable of responding to environmental cues by changing the sets of genes required for proteins expression or triggering important metabolic pathways involved in the adaptative response to the new external conditions. Thus, through the methods offered by Synthetic Biology, our work sought to couple genetic elements capable of detecting extracellular oxygen levels to a genetic circuit containing a regulatory cascade that restricts the activation of our recombinant agent to hypoxic sites, such as those present in solid tumors, together with the presence of an external pharmacological agent appropriate for clinical application. Using a genetic regulator developed by our group, we were able to build a circuit capable of responding to the presence of aspirin, a clinically safe drug with an extensive history of use, and we selected promoters candidates to recognize low levels of oxygen as a pathophysiological signal to establish a logic AND gate that would allow greater safety and selectivity in the use of microbial agents as therapeutic tools to fight cancer. The in vitro validation of the response of our circuit to the condition of hypoxia has proved challenging, given the myriad of factors that influence the functioning of new genetic parts, such as inducible promoters, in synthetic constructs outside their native context, however the work offers an example of a simple and economical approach to investigate biological anaerobic sensors through the expression of reporter proteins. (AU)

• FAPESP's process: 18/03274-8 - Application of synthetic biology in the construction of genetic circuit to improve the antitumor effect of Salmonella spp.
• Grantee: Kauan Ribeiro de Sena Gomes
• Support Opportunities: Scholarships in Brazil - Master