Tumor growth inhibition in vivo and G(2)/M cell cycle arrest induced by antisense oligodeoxynucleotide targeting thymidylate synthase

Chemotherapeutic agents targeting thymidylate synthase (TS) are effective a gainst human tumors. Efficacy is limited by drug resistance, often mediated by TS overexpression. Treatment of HeLa cells in vitro with an antisense o ligodeoxynucleotide (ODN 83) targeting human TS mRNA reduces TS mRNA and pr otein levels, inhibits cell proliferation, and sensitizes cells to TS-targe ting drugs (Ferguson et al., 1999). The present study investigates the mech anism by which ODN 83 inhibits cell proliferation and examines its antitumo r efficacy in vivo. ODN 83 treatment did not induce apoptosis in HeLa cells in vitro but caused accumulation of cells at G(2)/M. In contrast, TS-targe ting chemotherapeutics arrest at G(1) or S. Antisense down-regulation reduc ed TS mRNA levels in human colon cancer (HT29) cells by 40% in vitro, resul ted in G(2)/M arrest, and reduced proliferation without enhanced cell death . Growth of HT29 tumors in immunocompromised mice was significantly inhibit ed when antisense ODN 83 treatment began promptly after tumor implantation and was accompanied by a 40% reduction in TS protein levels. Growth of tumo rs allowed to reach 400 mm(3) prior to ODN administration was unaffected by antisense ODN 83. Radiolabeled ODNs were localized to the tumor periphery but evenly distributed in normal tissue. Thus, down-regulation of TS mRNA a nd protein by antisense ODN treatment exerts a novel G(2)/M cell cycle bloc k without increasing cell death and inhibits HT29 tumor cell growth in vivo . Antisense ODN 83 may be an effective therapy for colon carcinoma, alone o r in combination with TS-targeting cytotoxic drugs.