A Multiplex Molecular Cell-Based Sensor to Detect Ligands of PPARs: An Optimized Tool for Drug Discovery in Cyanobacteria

Abstract

Cyanobacteria produce a wealth of secondary metabolites. Since these organisms attach fatty acids into molecules in unprecedented ways, cyanobacteria can serve as a novel source for bioactive compounds acting as ligands for Peroxisome Proliferator-Activated Receptors (PPAR). PPARs (PPARα, PPARβ/δ and PPARγ) are ligand-activated nuclear receptors, involved in the regulation of various metabolic and cellular processes, thus serving as potential drug targets for a variety of pathologies. Yet, given that PPARs’ agonists can have pan-, dual- or isoform-specific action, some controversy has been raised over currently approved drugs and their side effects, highlighting the need for novel molecules. Here, we expand and validate a cell-based PPAR transactivation activity biosensor, and test it in a screening campaign to guide drug discovery. Biosensor upgrades included the use of different reporter genes to increase signal intensity and stability, a different promoter to modulate reporter gene expression, and multiplexing to improve efficiency. Sensor’s limit of detection (LOD) ranged from 0.36–0.89 nM in uniplex and 0.89–1.35 nM in multiplex mode. In triplex mode, the sensor’s feature screening, a total of 848 fractions of 96 cyanobacteria extracts were screened. Hits were confirmed in multiplex mode and in uniplex mode, yielding one strain detected to have action on PPARα and three strains to have dual action on PPARα and -β.

Microfluidic Biosensor Based on Molybdenum Disulfide (MoS2) Modified Thin-Core Microfiber for Immune Detection of Toxoplasma gondii

“…According to the methods in Ref. [32] and Ref. [33], the limit of quantitation (LOQ) is calculated to be about 287 fg/mL.…”

“…According to the methods in Refs. [32,33], the limit of quantitation (LOQ) is calculated to be about 287 fg/mL. It is concluded that the biosensor has extremely low LOD and can be used to detect biomolecules with lower concentration.…”

Emerging Technologies for the Discovery of Novel Diversity in Cyanobacteria and Algae and the Elucidation of Their Valuable Metabolites

“…A biosensor is an analytical device that contains a biorecognition element, such as antibodies, enzymes, nucleic acids, and aptamers, as well as a transducer that converts the recognition element into a measurable signal using optical, electrochemical, thermometric, or piezoelectric technology [134,135]. Biosensors are being used in the discovery of drugs and bioactive compounds [135,136]. Páscoa and colleagues (2023) developed a novel wholecell biosensor that incorporated reporter genes for signal intensity and stability, a promoter to modulate Fluc expression, and assay multiplexing to improve efficiency and output.…”

“…The sensor's selectivity was analyzed using clotrimazole and validated by recognizing fragments of cyanobacterial extracts possessing peroxisome proliferator-activated receptors (PPARs) agonist properties. Multiplex and uniplex screening assays were conducted to determine which PPARs the bioactive extracts were targeting [136].…”