The anticarcinogenic activity of t,t conjugated linoleic acid isomers mixture (designated t,t CLAs) was studied in mouse forestomach carcinogenesis model induced by benzo(a)pyrene (BP), referenced with c9,t11 CLA, t10,c12 CLA, and synthetic CLA. The t,t CLAs (99%), which was synthesized from synthetic CLA by methylation for 30 min with 14.0% BF3/methanol, consisted of 2.6% t12,t14; 10.7% t11,t13; 36.7% t10,t12; 38.2% t9,t11; 9.5% t8,t10; and 2.3% t7,t9 CLA. The purity of CLA samples was 95% synthetic CLA, 91% c9,t11 CLA, and 98% t10,c12 CLA. Female ICR mouse (6-7 weeks of age and 26±1 g) was given sample orally (0.1 mL + olive oil 0.1 mL) on Monday and Wednesday, and BP (2 mg/0.2 mL olive oil) on Friday. This cycle was repeated 4 times. Twenty-three weeks later, the experiment was terminated and analyzed with various parameters related to forestomach tumorigenesis. The t,t CLAs strongly inhibited the initiation of BP-induced female ICR mouse forestomach neoplasia relative to c9,t11 CLA, t10,c12 CLA, and synthetic CLA by the mechanistic action of apoptotic induction. The number of tumors per mouse by synthetic CLA, c9,t11 CLA, and t10,c12 CLA were 5.76±3.15, 5.96±3.47, and 5.36±2.99, respectively, but it was significantly reduced to 4.55±2.54 by t,t CLAs. Tumor incidence (84.6%) of mice treated with t,t CLAs was similar to 88.9% c9,t11 CLA and 89.3% t10,c12 CLA, but it was of significant difference, relative to 100% of control and linoleic acid-treated mice. The size of tumor was also reduced: 1.67±0.96 mm t,t CLAs; 1.85±0.69 mm synthetic CLA; 1.94±1.10 mm c9,t11 CLA; and 1.75±1.23 mm t10,c12 CLA. The t,t CLAs enhanced the fragmentation of DNA when analyzed by electrophoresis, and also enhanced the apoptotic index to 35.46±4.5%, measured by TUNEL method, as compared to 25.39±3.8 synthetic CLA, 21.56±5.2 c9,t11 CLA, 29.75±2.9 t10,c12 CLA, 4.50±2.3 BP, and 7.12±2.1 linoleic acid. The t,t CLAs reduced the expression of Bcl-2 gene analyzed by immunohistochemistry and western blotting, induced Bax gene expression and increased caspase-3 enzyme activity. cPLA2 activity was not affected by t,t CLAs. These results suggest that among CLA isomers, t,t CLAs showed the most potent anticarcinogenic activity for the BP-induced mouse forestomach carcinogenesis regimen, in part, due to induction of apoptosis. Key words: carcinogenesis, apotosis, bcl-2, forestomach

[Proc Amer Assoc Cancer Res, Volume 47, 2006]