Mechanism of synergy of levamisole and fluorouracil: induction of human leukocyte antigen class I in a colorectal cancer cell line.
BACKGROUND
The use of the combination of fluorouracil (5-FU) and levamisole has been shown to improve the survival of patients with resected Dukes' stage C colon carcinoma. 5-FU is incorporated into RNA, which results in aberrant processing and turnover of RNA. Neither the mechanism of synergy between the two drugs nor the precise molecular mechanism of action of levamisole is known. Each drug has previously been shown to alter the expression of class I human leukocyte antigens (HLA class I) in colorectal cancer cell lines.


OBJECTIVE
The purpose of this study was to explore the mechanism of interaction between 5-FU and levamisole by investigating the effect of this combination on HLA class I gene expression in the colorectal cancer cell line WiDr.


METHODS
WiDr cells were treated either with 5-FU alone or with 5-FU and levamisole. Expression of HLA class I antigens was analyzed by flow cytometry using the monoclonal antibody W6/32. Specific DNA probes for HLA class I, beta 2-microglobulin, beta-actin, HLA class II, and p53 (also known as TP53) were used in Northern blot analysis of the steady-state level of messenger RNAs (mRNAs) and for "run-on" transcription analysis.


RESULTS
5-FU alone produced more than 50% increases in the expression of the HLA class I antigens, and levamisole caused a further 8%-18% increase. 5-FU caused the steady-state level of HLA class I mRNAs to increase by about 80%, and levamisole enhanced this effect of 5-FU by a further 70%. 5-FU did not increase the other mRNAs. In vitro run-on transcription revealed that 5-FU caused a 20%-57% reduction in RNA synthesis, while levamisole caused a 30%-190% increase in RNA synthesis. Levamisole therefore reversed the inhibition of RNA synthesis caused by 5-FU. Both drugs had a general effect on RNA synthesis that was not restricted to HLA class I transcription.


CONCLUSIONS
The apparent synergy between levamisole and 5-FU is a result of the incorporation of 5-FU, which may stabilize HLA class I mRNAs, leading to their accumulation, while levamisole augments the accumulation of these stable mRNAs by increasing the rate of transcription.


CONCLUSIONS
Levamisole reduces the toxicity of 5-FU caused by generalized inhibition of RNA synthesis, and at the same time augments the effects of 5-FU, which may be due to selective stabilization of certain mRNAs.
AbdAlla
E E
EE
Department of Biochemistry and Molecular Biology, University of Leeds, England.
Blair
G E
GE
Jones
R A
RA
Sue-Ling
H M
HM
Johnston
D
D
D015152:Blotting, Northern; D015179:Colorectal Neoplasms / Q000188:drug therapy* / Q000235:genetics* / Q000276:immunology; D015342:DNA Probes; D004357:Drug Synergism; D004359:Drug Therapy, Combination; D005434:Flow Cytometry; D005472:Fluorouracil / Q000494:pharmacology*; D005805:Genes, MHC Class I / Q000187:drug effects* / Q000235:genetics; D006801:Humans; D007978:Levamisole / Q000494:pharmacology*; D012333:RNA, Messenger / Q000187:drug effects; D012334:RNA, Neoplasm / Q000187:drug effects; D014407:Tumor Cells, Cultured; D001613:beta 2-Microglobulin / Q000187:drug effects
D015342:DNA Probes; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D001613:beta 2-Microglobulin; D007978:Levamisole; D005472:Fluorouracil