Genetic Engineering of Clitoria ternatea (Aparajita) for Enhanced Production of Anticancer Metabolites

Abstract

Clitoria ternatea, often referred to as Aparajita or Butterfly Pea, is a medicinal herb celebrated for its abundant array of bioactive constituents, such as flavonoids, anthocyanins, and cyclotides, which demonstrate notable anticancer effects. This research investigates the genetic modification of C. ternateaC. ternatea through CRISPR/Cas9 and RNA interference (RNAi) techniques to boost the production of secondary metabolites, consequently elevating the plant's medicinal worth. The main objective of this research is to enhance the synthesis of anticancer agents by implementing precise gene alterations that stimulate biosynthetic routes while inhibiting detrimental regulatory elements. The approach utilised Agrobacterium-mediated transformation of C. ternateaC. ternatea callus tissues employing CRISPR/Cas9 constructs aimed at flavonoid biosynthesis genes, alongside RNAi constructs designed to silence anthocyanin- degrading enzymes. Analyses conducted using HPLC and LC-MS revealed notable enhancements in the levels of flavonoids and anthocyanins within genetically altered plants. In vitro cytotoxicity evaluations utilising MCF-7, HCT-116, and A549 cancer cell lines revealed a heightened induction of apoptosis following treatment with plant extracts, corroborated by the elevated expression of pro-apoptotic indicators including Bax and cleaved caspase-3. The results demonstrate that CRISPR-driven activation and RNA interference-based inhibition significantly boost the anticancer capabilities of C. ternatea, positioning it as a highly promising option for plant-sourced chemotherapeutic agents. Nonetheless, obstacles persist in extensive manufacturing, obtaining regulatory clearances, and enhancing bioavailability. Subsequent investigations ought to concentrate on advancing metabolic engineering and conducting clinical assessments to utilise C. ternateaC. ternatea as a promising source of natural anticancer compounds. This research offers significant perspectives on genetic engineering approaches for enhancing the therapeutic capabilities of C. ternateaC. ternatea in cancer-related applications.