title: Novel tandem biotransformation process for the biosynthesis of a novel compound, 4-(2,3,5,6-tetramethylpyrazine-1)-4'-demethylepipodophyllotoxin.
abstract: According to the structure of podophyllotoxin and its structure-function relationship, a novel tandem biotransformation process was developed for the directional modification of the podophyllotoxin structure to directionally synthesize a novel compound, 4-(2,3,5,6-tetramethylpyrazine-1)-4'-demethylepipodophyllotoxin (4-TMP-DMEP). In this novel tandem biotransformation process, the starting substrate of podophyllotoxin was biotransformed into 4'-demethylepipodophyllotoxin (product 1) with the demethylation of the methoxyl group at the 4' position by Gibberella fujikuroi SH-f13, which was screened out from Shennongjia prime forest humus soil (Hubei, China). 4'-Demethylepipodophyllotoxin (product 1) was then biotransformed into 4'-demethylpodophyllotoxone (product 2) with the oxidation of the hydroxyl group at the 4 position by Alternaria alternata S-f6, which was screened out from the gathered Dysosma versipellis plants in the Wuhan Botanical Garden, Chinese Academy of Sciences. Finally, 4'-demethylpodophyllotoxone (product 2) and ligustrazine were linked with a transamination reaction to synthesize the target product 4-TMP-DMEP (product 3) by Alternaria alternata S-f6. Compared with podophyllotoxin (i.e., a 50% effective concentration [EC(50)] of 529 μM), the EC(50) of 4-TMP-DMEP against the tumor cell line BGC-823 (i.e., 0.11 μM) was significantly reduced by 5,199 times. Simultaneously, the EC(50) of 4-TMP-DMEP against the normal human proximal tubular epithelial cell line HK-2 (i.e., 0.40 μM) was 66 times higher than that of podophyllotoxin (i.e., 0.006 μM). Furthermore, compared with podophyllotoxin (i.e., log P = 0.34), the water solubility of 4-TMP-DMEP (i.e., log P = 0.66) was significantly enhanced by 94%. For the first time, the novel compound 4-TMP-DMEP with superior antitumor activity was directionally synthesized from podophyllotoxin by the novel tandem biotransformation process developed in this work.
authors.affiliation: Key Laboratory of Fermentation Engineering (Ministry of Education), Hubei Provincial Key Laboratory of Industrial Microbiology, College of Bioengineering, Hubei University of Technology, Wuhan 430068, China. yajietang@hotmail.com
authors.forename: Ya-Jie
authors.initials: YJ
authors.lastname: Tang
authors.forename: Wei
authors.initials: W
authors.lastname: Zhao
authors.forename: Hong-Mei
authors.initials: HM
authors.lastname: Li
mesh_terms: D000528:Alternaria / Q000302:isolation & purification / Q000378:metabolism*; D000970:Antineoplastic Agents / Q000378:metabolism* / Q000494:pharmacology; D028141:Berberidaceae / Q000382:microbiology; D001711:Biotransformation; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002681:China; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D004275:DNA, Ribosomal / Q000737:chemistry / Q000235:genetics; D020459:Genes, rRNA; D005874:Gibberella / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D020128:Inhibitory Concentration 50; D008969:Molecular Sequence Data; D011034:Podophyllotoxin / Q000031:analogs & derivatives* / Q000378:metabolism*; D011719:Pyrazines / Q000378:metabolism*; D012331:RNA, Fungal / Q000235:genetics; D012337:RNA, Ribosomal, 18S / Q000235:genetics; D017422:Sequence Analysis, DNA; D012988:Soil Microbiology
chemical_list: D000970:Antineoplastic Agents; D004271:DNA, Fungal; D004275:DNA, Ribosomal; D011719:Pyrazines; D012331:RNA, Fungal; D012337:RNA, Ribosomal, 18S; D011034:Podophyllotoxin