Time course of apoptotic tumor response after a single dose of chemotherapy: comparison with 99mTc-annexin V uptake and histologic findings in an experimental model.
In tumors the process of apoptosis occurs over an interval of time after chemotherapy. To determine the best timing for detecting apoptosis in vivo with (99m)Tc-annexin V after chemotherapy, we examined the changes in (99m)Tc-annexin V accumulation over time in comparison with those of caspase-3 and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) expression level after cyclophosphamide treatment in an experimental model.


METHODS
Hydrazinonicotinamide (HYNIC)-annexin V was labeled with (99m)Tc ((99m)Tc-annexin V). Rats were inoculated with allogenic hepatoma cells (KDH-8) into the left calf muscle. Eleven days after the inoculation, the rats were randomly divided into the group receiving a single dose of cyclophosphamide (150 mg/kg intraperitoneally) and the control group. (99m)Tc-Annexin V (18.5 MBq [0.5 mCi] per rat) was injected intravenously in the rats 4, 12, and 20 h after the treatment and also to the control rats (n = 5 in each group). Radioactivity in tissues was determined 6 h after (99m)Tc-annexin V injection. Immunostaining of caspase-3 and TUNEL were performed to detect apoptosis, and the rates of positively stained cells were calculated.


RESULTS
(99m)Tc-Annexin V accumulation in tumors significantly increased at 20 h (0.077 +/- 0.007 [%ID/g] x kg, where %ID/g = percentage injected dose per gram) but not at 4 or 12 h (0.048 +/- 0.008 and 0.052 +/- 0.014 [%ID/g] x kg, respectively) after cyclophosphamide treatment. (99m)Tc-Annexin V accumulation in tumors and the rate of apoptotic cells determined by caspase-3 immunostaining and TUNEL were significantly higher in treated rats 20 h after cyclophosphamide treatment as compared with control rats.


CONCLUSIONS
The effective detection of apoptotic tumor response with (99m)Tc-annexin V required 20 h after cyclophosphamide treatment in an experimental model. The present results provide an important basis for determining the best timing of annexin V imaging after the start of chemotherapy in a clinical setting.
Takei
Toshiki
T
Department of Nuclear Medicine, Graduate School of Medicine, Hokkaido University, Sapporo 060-8638, Japan.
Kuge
Yuji
Y
Zhao
Songji
S
Sato
Masayuki
M
Strauss
H William
HW
Blankenberg
Francis G
FG
Tait
Jonathan F
JF
Tamaki
Nagara
N
D000818:Animals; D017304:Annexin A5; D017209:Apoptosis / Q000187:drug effects*; D006528:Carcinoma, Hepatocellular / Q000000981:diagnostic imaging / Q000188:drug therapy*; D003520:Cyclophosphamide / Q000494:pharmacology*; D008297:Male; D011877:Radionuclide Imaging; D051381:Rats; D017556:Technetium Compounds; D014183:Transplantation, Heterologous; D014407:Tumor Cells, Cultured
D017304:Annexin A5; D017556:Technetium Compounds; D003520:Cyclophosphamide