Toehold switch-based bacterial sensor device for detection of cervical cancer microRNA biomarkers

• Applicant: Priyanka Dhakane
• Subject Area: Biomaterials ; General Genetics and Functional Genome Biology
• Term: since 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 520639326

The project aims to develop a bacterial biosensor device for multiplex detection of cervical cancer-specific micro-RNA biomarkers in a cost-effective and multiplexed manner. Recently, it was discovered that significant amounts of extracellular miRNAs circulate in bodily fluids with high stability. Notably, dysregulated miRNA profiles have been associated with many diseases including cervical cancer because of which miRNA profiling is being explored as a promising strategy for cancer diagnosis, prognosis and treatment. Cervical cancer (CC) is the leading cause of death by cancer in women from developing countries with over 600k new cases and 340k deaths in 2020 worldwide. Currently cytology (pap-smear) and histopathology are gold standards for detection of Human Papillomavirus (HPV) related dysplasia and CC. Although efficient, these methods have low sensitivity (50%) and heavily rely on interpretation and technician training, leading to limited accessibility for women worldwide. To catch the development of this disease at an early stage, a simple and sensitive self-test is greatly desirable. The cervical vaginal fluid (CVF) has been shown to contain stable (average 5 days) miRNA biomarkers in abundance, making it ideal for disease diagnosis. In this study, I will detect miR-126-3p, mi-20b-5p, mi-451a, and mi-144-3p, which are significantly upregulated in CC (2 - 3 orders of magnitude) and proven to have around 90% sensitivity in detection of cervical lesions and neoplasia. To achieve this, E.coli will be engineered with four toehold switches specific for target miRNA sequences. Once the target miRNA binds to the toehold switch, it will enable expression of the reporter protein. Efficacy and limit of detection of the bacterial biosensor will be tested using synthetic miRNAs. To adapt it for on-site detection of miRNAs from the CVF, engineered bacteria will be encapsulated in a tampon like device. Apart from cancer, miRNA profiles are dysregulated in wide variety of human diseases. miRNA detection has not been done in bacteria till date, therefore this novel sensing strategy can massively improve the applicability of bacterial biosensors for disease diagnosis.

• DFG Programme: Research Grants