p53-defective tumors with a functional apoptosome-mediated pathway: a new therapeutic target.
BACKGROUND
Although cancer cells appear to maintain the machinery for intrinsic apoptosis, defects in the pathway develop during malignant transformation, preventing apoptosis from occurring. How to specifically induce apoptosis in cancer cells remains unclear.


METHODS
We determined the apoptosome activity and p53 status of normal human cells and of lung, colon, stomach, brain, and breast cancer cells by measuring cytochrome c-dependent caspase activation and by DNA sequencing, respectively, and we used COMPARE analysis to identify apoptosome-specific agonists. We compared cell death, cytochrome c release, and caspase activation in NCI-H23 (lung cancer), HCT-15 (colon cancer), and SF268 (brain cancer) cells treated with Triacsin c, an inhibitor of acyl-CoA synthetase (ACS), or with vehicle. The cells were mock, transiently, or stably transfected with genes for Triacsin c-resistant ACSL5, dominant negative caspase-9, or apoptotic protease activating factor-1 knockdown. We measured ACS activity and levels of cardiolipin, a mitochondrial phospholipid, in mock and ACSL5-transduced SF268 cells. Nude mice carrying NCI-H23 xenograft tumors (n = 10) were treated with Triacsin c or vehicle, and xenograft tumor growth was assessed. Groups were compared using two-sided Student t tests.


RESULTS
Of 21 p53-defective tumor cell lines analyzed, 17 had higher apoptosome activity than did normal cells. Triacsin c selectively induced apoptosome-mediated death in tumor cells (caspase activity of Triacsin c-treated versus untreated SF268 cells; means = 1020% and 100%, respectively; difference = 920%, 95% CI = 900% to 940%; P<.001). Expression of ACSL5 suppressed Triacsin c-induced cytochrome c release and subsequent cell death (cell survival of Triacsin c-treated mock- versus ACSL5-transduced SF268 cells; means = 40% and 83%, respectively; difference = 43%, 95% CI = 39% to 47%; P<.001). ACS was also essential to the maintenance of cardiolipin levels. Finally, Triacsin c suppressed growth of xenograft tumors (relative tumor volume on day 21 of Triacsin c-treated versus untreated mice; means = 4.6 and 9.6, respectively; difference = 5.0, 95% CI = 2.1 to 7.9; P = .006).


CONCLUSIONS
Many p53-defective tumors retain activity of the apoptosome, which is therefore a potential target for cancer chemotherapy. Inhibition of ACS may be a novel strategy to induce the death of p53-defective tumor cells.
Mashima
Tetsuo
T
Cancer Chemotherapy Center, Japanese Foundation for Cancer Research, Tokyo, Japan.
Oh-hara
Tomoko
T
Sato
Shigeo
S
Mochizuki
Mikiko
M
Sugimoto
Yoshikazu
Y
Yamazaki
Kanami
K
Hamada
Jun-ichi
J
Tada
Mitsuhiro
M
Moriuchi
Tetsuya
T
Ishikawa
Yuichi
Y
Kato
Yo
Y
Tomoda
Hiroshi
H
Yamori
Takao
T
Tsuruo
Takashi
T
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis; D051033:Apoptosis Inducing Factor; D053477:Apoptotic Protease-Activating Factor 1; D015153:Blotting, Western; D001932:Brain Neoplasms / Q000378:metabolism; D001943:Breast Neoplasms / Q000378:metabolism; D002308:Cardiolipins / Q000097:blood; D020169:Caspases / Q000378:metabolism*; D003066:Coenzyme A Ligases / Q000037:antagonists & inhibitors*; D003110:Colonic Neoplasms / Q000378:metabolism; D045304:Cytochromes c / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D005420:Flavoproteins / Q000187:drug effects / Q000378:metabolism*; D018014:Gene Transfer Techniques; D006801:Humans; D008175:Lung Neoplasms / Q000378:metabolism; D008565:Membrane Proteins / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D008819:Mice, Nude; D008928:Mitochondria / Q000378:metabolism; D009374:Neoplasms, Experimental / Q000188:drug therapy / Q000201:enzymology / Q000378:metabolism*; D011506:Proteins / Q000037:antagonists & inhibitors; D034741:RNA, Small Interfering / Q000378:metabolism; D017422:Sequence Analysis, DNA; D013274:Stomach Neoplasms / Q000378:metabolism; D014162:Transfection; D014183:Transplantation, Heterologous; D014226:Triazenes / Q000494:pharmacology* / Q000627:therapeutic use; D016159:Tumor Suppressor Protein p53 / Q000187:drug effects / Q000378:metabolism*
C494829:AIFM1 protein, human; C506592:APAF1 protein, human; D000970:Antineoplastic Agents; C506593:Apaf1 protein, mouse; D051033:Apoptosis Inducing Factor; D053477:Apoptotic Protease-Activating Factor 1; D002308:Cardiolipins; D004791:Enzyme Inhibitors; D005420:Flavoproteins; D008565:Membrane Proteins; C494830:AIFM1 protein, mouse; D011506:Proteins; D034741:RNA, Small Interfering; D014226:Triazenes; D016159:Tumor Suppressor Protein p53; C034613:triacsin C; D045304:Cytochromes c; D020169:Caspases; D003066:Coenzyme A Ligases; C485465:ACSL5 protein, human