Noninvasive imaging of apoptosis and its application in cancer therapeutics.
OBJECTIVE
Activation of the apoptotic cascade plays an important role in the response of tumors to therapy. Noninvasive imaging of apoptosis facilitates optimization of therapeutic protocols regarding dosing and schedule and enables identification of efficacious combination therapies.


METHODS
We describe a hybrid polypeptide that reports on caspase-3 activity in living cells and animals in a noninvasive manner. This reporter, ANLucBCLuc, constitutes a fusion of small interacting peptides, peptide A and peptide B, with the NLuc and CLuc fragments of luciferase with a caspase-3 cleavage site (DEVD) between pepANLuc (ANLuc) and pepBCLuc (BCLuc). During apoptosis, caspase-3 cleaves the reporter, enabling separation of ANLuc from BCLuc. A high-affinity interaction between peptide A and peptide B restores luciferase activity by NLuc and CLuc complementation. Using a D54 glioma model, we show the utility of the reporter in imaging of apoptosis in living subjects in response to various chemotherapy and radiotherapy regimens.


RESULTS
Treatment of live cells and mice carrying D54 tumor xenografts with chemotherapeutic agents such as temozolomide and perifosine resulted in induction of bioluminescence activity, which correlated with activation of caspase-3. Treatment of mice with combination therapy of temozolomide and radiation resulted in increased bioluminescence activity over individual treatments and increased therapeutic response due to enhanced apoptosis.


CONCLUSIONS
The data provided show the utility of the ANLucBCLuc reporter in dynamic, noninvasive imaging of apoptosis and provides a rationale for use of this technology to optimize dose and schedule of novel therapies or to develop novel combination therapies using existing drugs.
Coppola
Julia M
JM
Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
Ross
Brian D
BD
Rehemtulla
Alnawaz
A
D000818:Animals; D017209:Apoptosis; D019556:COS Cells; D053148:Caspase 3 / Q000378:metabolism*; D002522:Chlorocebus aethiops; D003131:Combined Modality Therapy; D003606:Dacarbazine / Q000031:analogs & derivatives / Q000627:therapeutic use; D005909:Glioblastoma / Q000201:enzymology / Q000473:pathology / Q000628:therapy*; D006801:Humans; D008163:Luminescent Measurements; D051379:Mice; D009368:Neoplasm Transplantation; D000077204:Temozolomide; D014183:Transplantation, Heterologous
D003606:Dacarbazine; D053148:Caspase 3; D000077204:Temozolomide