Periodic Reporting for period 1 - PiloSyn (Elucidation of the biosynthesis pathway of the glaucoma drug pilocarpine in Pilocarpus plants)
Reporting period: 2023-05-01 to 2025-04-30
The medical drug pilocarpine is one of the treatments for glaucoma or dry mouth. Pilocarpine is an alkaloid made by Pilocarpus shrubs naturally grown in the Amazon rainforest. As it is not cost-effective to chemically synthesize pilocarpine, commercial demands are met by extracting it from its natural source, the leaves of Pilocarpus plants. Since cultivated Pilocarpus plants have lower pilocarpine content than wild plants, leaves of wild-grown Pilocarpus are preferred for production. Excessive leaf harvesting of wild populations has led to the official listing of several Pilocarpus species as threatened. Synthetic biology offers a sustainable approach for the production of pilocarpine through its synthesis in heterologous hosts, such as microbes. However, to realize this vision, a detailed knowledge about the biosynthetic pathway of pilocarpine in Pilocarpus plants is needed. The aim of PiloSyn is to understand how Pilocarpus plants make pilocarpine, thus laying the foundation for a more sustainable pilocarpine production.
PiloSyn had access to Pilocarpus microphyllus seeds harvested in the Amazon rainforest. Seed germination was challenging, but it was possible to grow a few dozen P. microphyllus plants under controlled conditions. PiloSyn analyzed the content of pilocarpine and related alkaloids in several P. microphyllus tissues of different ages. The analysis revealed that the plants only start to produce pilocarpine at a certain age. Furthermore, feeding studies with isotopically labeled compounds identified the biosynthetic tissue responsible for pilocarpine biosynthesis of related alkaloids. These results provide a roadmap for future RNAseq analysis aimed at identifying enzymes involved in pilocarpine biosynthesis.
PiloSyn provided valuable insights into how P. microphyllus plants make pilocarpine by identifying the relevant biosynthetic tissues and discovering that the plants only start to produce pilocarpine at a certain age. This lays the foundation for further studies on pilocarpine biosynthesis, especially via tissue- and age-specific RNASeq analysis. Beyond pilocarpine research, PiloSyn, obtained comprehensive metabolite datasets from different P. microphyllus tissues and ages. As the plant tissue is not widely accessible, the data sets are of scientific value. The scientific community can use the datasets to search for new molecules, which can be evaluated as e.g new active pharmaceutical ingredients (API).