Current status and future prospects in cannabinoid production through in vitro culture and synthetic biology

For centuries, cannabis has been a rich source of fibrous, pharmaceutical, and recreational ingredients. Phytocannabinoids are the most important and well-known class of cannabis-derived secondary metabolites and display a broad range of health-promoting and psychoactive effects. The unique characteristics of phytocannabinoids (e.g., metabolite likeness, multi-target spectrum, and safety profile) have resulted in the development and approval of several cannabis-derived drugs. While most work has focused on the two main cannabinoids produced in the plant, over 150 unique cannabinoids have been identified. To meet the rapidly growing phytocannabinoid demand, particularly many of the minor cannabinoids found in low amounts in planta, biotechnology offers promising alternatives for biosynthesis through in vitro culture and heterologous systems. In recent years, the engineered production of phytocannabinoids has been obtained through synthetic biology both in vitro (cell suspension culture and hairy root culture) and heterologous systems. However, there are still several bottlenecks (e.g., the complexity of the cannabinoid biosynthetic pathway and optimizing the bioprocess), hampering biosynthesis and scaling up the biotechnological process. The current study reviews recent advances related to in vitro culture-mediated cannabinoid production. Additionally, an integrated overview of promising conventional approaches to cannabinoid production is presented. Progress toward cannabinoid production in heterologous systems and possible avenues for avoiding autotoxicity are also reviewed and highlighted. Machine learning is then introduced as a powerful tool to model, and optimize bioprocesses related to cannabinoid production. Finally, regulation and manipulation of the cannabinoid biosynthetic pathway using CRISPR- mediated metabolic engineering is discussed.