We enzymatically synthesized c-myc mRNA fragment containing the translation initiation (TI) region (36mer) and examined the structure by CD, 1H-NMR and RNaseV1 digestion. These study demonstrated that the c-myc mRNA fragment forms a hairpin structure at 37 degrees C. Six 15-mer antisense oligodeoxynucleotides(ODNs), which hybridize to the c-myc mRNA fragment were designed, and synhesized. Thermostabilities of the antisense ODNs-mRNA fragment complexes were examined. Antisense A (complementary to the c-myc mRNA from -9 to +6)-c-myc mRNA fragment complex had the highest melting temperature(Tm), and Antisense D (complementary to the c-myc mRNA from +1 to +15)-c-myc mRNA fragment complex had the most different Tm from a predicted Tm. Furthermore, Antisense D exhibited high inhibitory effect on HL60 cell proliferation, whereas Antisense A did not. The results suggest that the difference in the antisense effect is not simply due to the complex thermostability.