A preliminary study of imaging paclitaxel-induced tumor apoptosis with (99)Tc(m)-His10-Annexin V.
BACKGROUND
In tumors the process of apoptosis occurs over an interval of time after chemotherapy. It is important to determine the best time for detecting apoptosis by in vivo imaging. In this study, we evaluated the dynamics and feasibility of imaging non-small cell lung cancer (NSCLC) apoptosis induced by paclitaxel treatment using a (99)Tc(m)-labeled Annexin V recombinant with ten consecutive histidines (His10-Annexin V) in a mouse model.


METHODS
(99)Tc(m)-His10-Annexin V was prepared by one step direct labeling; radio-chemical purity (RCP) and radio-stability was tested. The binding of (99)Tc(m)-His10-Annexin V to apoptotic cells was validated in vitro using camptothecin-induced Jurkat cells. In vivo bio-distribution was determined in mice by dissection. The human H460 NSCLC tumor cell line (H460) tumor-bearing mice were treated with intravenous paclitaxel 24, 48 and 72 hours later. (99)Tc(m)-His10-Annexin V was injected intravenously, and planar images were acquired at 2, 4 and 6 hours post-injection on a dual-head gamma camera fitted with a pinhole collimator. Tumor-to-normal tissue ratios (T/NT) were calculated by ROI analysis and they reflected specific binding of (99)Tc(m)-His10-Annexin V. Mice were sacrificed after imaging. Caspase-3, as the apoptosis detector, was determined by flow cytometry, and DNA fragmentation was analyzed by the terminal deoxynucleotidytransferase mediated dUTP nick-end labeling (TUNEL) assay. Nonspecific accumulation of protein was estimated using bovine serum albumin (BSA). The imaging data were correlated with TUNEL-positive nuclei and caspase-3 activity.


RESULTS
(99)Tc(m)-His10-Annexin V had a RCP > 98% and high stability 2 hours after radio-labeling, and it could bind to apoptotic cells with high affinity. Bio-distribution of (99)Tc(m)-His10-Annexin V showed predominant uptake in kidney, relatively low uptake in myocardium, liver and gastrointestinal tract, and rapid clearance from blood and kidney was observed. The T/NT was significantly increased after paclitaxel treatment, whereas it was low in untreated tumors (T/NT = 1.43 ± 0.18). The %ID/g activity in Group 2 (24 hours), Group 3 (48 hours) and Group 4 (72 hours) after treatment was 2.55 ± 0.73, 3.35 ± 1.10, and 3.4 ± 0.96, respectively. Whereas in the non-treated group, Group 1, %ID/g was 1.10 ± 0.18. The radiotracer uptake was positively correlated to the apoptotic index (r = 0.852, P < 0.01), as well as caspase-3 activity (r = 0.816, P < 0.01).


CONCLUSIONS
This study addresses the dynamics and feasibility of imaging non-small cell lung tumor apoptosis using (99)Tc(m)- His10-Annexin V.
Department of Nuclear Medicine, China-Japan Friendship Hospital, Beijing 100029, China.
Yu-min
YM
Zheng
Feng
F
Wang
Wei
W
Fang
Zi-chun
ZC
Hua
Zi-zheng
ZZ
Wang
Qing-le
QL
Meng
Jue
J
Yan
D000818:Animals; D017304:Annexin A5; D000972:Antineoplastic Agents, Phytogenic / Q000627:therapeutic use*; D017209:Apoptosis; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy / Q000473:pathology*; D045744:Cell Line, Tumor; D004195:Disease Models, Animal; D006639:Histidine; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy / Q000473:pathology*; D051379:Mice; D015609:Organotechnetium Compounds; D017239:Paclitaxel / Q000627:therapeutic use*; D019275:Radiopharmaceuticals
D017304:Annexin A5; D000972:Antineoplastic Agents, Phytogenic; D015609:Organotechnetium Compounds; D019275:Radiopharmaceuticals; C417036:technetium Tc 99m annexin V; D006639:Histidine; D017239:Paclitaxel