Enhancement of 5-fluorouracil sensitivity by an rTS signaling mimic in H630 colon cancer cells

The rTS beta protein has been hypothesized to synthesize signaling molecules that can down-regulate thymidylate synthase. These molecules share biological and chemical properties with acyl-homoserine lactones (AHL), suggesting some AHLs might act as rTS signaling mimics and down-regulate thymidylate synthase. We have determined that the AHL, 3-oxododecanoyl homoserine lactone (3-oxo-C12-(L)HSL) can down-regulate thymidylate synthase protein at 10 mu mol/l, and reduce H630 (human colorectal cancer) growth by 50% at 23 mu mol/L (IC50) in cell culture. At its IC50 concentration, 3-oxo-CI2-(L)-HSL reduces the apparent IC50 of 5-fluorouracil (5-FU) from 1 mu mol/L to 80 nmol/L (12-fold) in a colony formation assay. 3-Oxo-C12-(L)-HSL enhances the activity of 5-fluorodeoxyuridine, tomudex, and taxol but not the activity of 5-fluorouridine, methotrexate or Adriamycin. The unexpected interaction with taxol probably results from effects of the AHL on tubulin expression. Differences in taxol sensitivity, tubulin, and cellular morphology between H630 and the thymidylate synthase and rTS beta-overproducing, 5-FU-resistant H630-1 cell line as determined by colony formation assays, Western analysis of one-dimensional and two-dimensional gels, and photomicroscopy confirm that cytoskeletal changes are induced by the AHL or by rTS signaling. lsozyme differences in thymidylate synthase and rTS also exist in the two cell lines. Phosphorylation of rTS amino acid S 121 is shown to occur and is decreased at least 10-fold in the drug-resistant cells. The data presented provide support for further investigations of rTS signaling mimics as enhancers to thymidylate synthase-directed chemotherapy, evidence that the phosphorylation state of rTS beta may be a marker for 5-FU resistance and a previously unrealized relationship between rTS signaling and the cytoskeleton