This project is aimed at developing plant tissue biotechnology for the production of taxol and taxol efflux blockers to improve the availability of taxol for further clinical evaluations and overcome taxol-resistance problems. This objective will be accomplished by a consortium which is multidisciplinary (analytical chemistry, biochemistry, natural product chemistry, molecular biology, and pharmacology) and multi-institutional (Purdue University and Upjohn Company). We propose to: (1) develop suspension tissue cultures and hairy root cultures of Taxus species; (2) analyze and select high yielding colonies by tubulin stabilization and tandem mass spectrometry/high performance liquid chromatography; (3) develop highly sensitive and compound-selective tandem mass spectrometric methods for taxol and other taxanes; (4) enhance production of taxol and desirable taxanes by induction with elicitors and transformation with various Agrobacterium strains; (5) prescreen potential taxol efflux blockers from tissue cultures; (6) perform biological and physical assay- directed isolation, and elucidate the structures of desirable constituents; (7) synthesize taxol from baccatin III or other related taxanes; and (8) evaluate antitumor activity of novel taxanes in vivo.
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