[Synthesis of chimeric anti-MDR1 ribozymes (tRNA-Rzs) and their biological activities in cell-free system].
OBJECTIVE
Development of multidrug resistance(MDR) is the major obstacle to successful cancer chemotherapy. One strategy to block the P-glycoprotein(P-gp)-mediated MDR is to use a ribozyme (Rz) target against MDR1 mRNA.


METHODS
Three kinds of anti-MDR1 chimeric hammerhead ribozymes, the first-one cleaving codon 196 of MDR1 mRNA (196MDR1-Rz), the second one, a stem-II base modified (U9-->G9, U13-->A13, G14-->A14, A18-->C18) Rz against codon 196 (196MDR1-sRz) and the third one, the stem-II base modified Rz directed against the -6(-)-4 GUC sequence of the translation initiation site of the MDR1 mRNA (iMDR1-sRz), were synthesized based on the cloned retroviral constructs: N2A + tRNAi(met)-196MDR1-Rz, N2A + tRNAi(met)-196MDR1-sRz, N2A + tRNAi(met)-iMDR1-sRz.


RESULTS
In a cell-free system, the chimeric tRNA-sRz molecules were more stable and had more efficient catalytic activities than the corresponding naked Rz molecules. The stem-II base modified Rz molecules were also more stable and efficient in catalytic activities than the unmodified ones. In control, the disabled tRNA-mut-iMDR1-sRz had no cleavage activity.


CONCLUSIONS
Base modification in the Rz's stem-II of ribozyme structure and the development of chimeric tRNA-ribozyme molecules are able to enhance the cleavage efficacy.
Wang
F
F
Beijing Institute of Infectious Diseases, Beijing 100039, China.
Lei
Z
Z
Jin
L
L
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics / Q000378:metabolism*; D002474:Cell-Free System; D018432:Drug Resistance, Multiple / Q000502:physiology; D020871:RNA Stability; D016337:RNA, Catalytic / Q000138:chemical synthesis* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D012343:RNA, Transfer / Q000737:chemistry*
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D016337:RNA, Catalytic; D012333:RNA, Messenger; D012343:RNA, Transfer