Early Prediction of Tumor Response to Treatment: Preclinical Validation of ^99mTc-Duramycin

Abstract

Noninvasive imaging of cell death can provide an early indication of the efficacy of tumor treatment, aiding clinicians in distinguishing responding patients from nonresponding patients early on. 99m Tcduramycin is a SPECT tracer for cell death imaging. In this study, our aim was to validate the use of 99m Tc-duramycin for imaging the early response of tumors to treatment. Methods: An in vitro binding assay was performed on COLO205 cells treated with 5-fluorouracil (3.1, 31, or 310 μM) and oxaliplatin (0.7 or 7 μM) or radiation (2 or 4.5 Gy). 99m Tc-duramycin cell binding and the levels of cell death were evaluated after treatment. In vivo imaging was performed on 4 groups of CD1-deficient mice bearing COLO205 human colorectal cancer tumors. Each group included 6 tumors. The first group was given irinotecan (100 mg/kg), the second oxaliplatin (5 mg/kg), the third irinotecan (80 mg/kg) plus oxaliplatin (5 mg/kg), and the fourth vehicle (0.9% NaCl and 5% glucose). For radiotherapy studies, COLO205 tumors received 4.5 Gy, 2 fractions of 4.5 Gy in a 24-h interval, pretreatment with an 80 mg/kg dose of irinotecan combined with 2 fractions of 4.5 Gy in a 24-h interval, or no treatment (n 5 5-6/group). Therapy response was evaluated by 99m Tc-duramycin SPECT 24 h after the last dose of therapy. Blocking was used to confirm tracer specificity. Radiotracer uptake in the tumors was validated ex vivo using γ-counting, cleaved caspase-3, and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) histology. Results: Chemotherapy and radiotherapy increased 99m Tc-duramycin binding to COLO205 cells in a concentration/ dose-and time-dependent manner, which correlated well with cell death levels (P , 0.05) as analyzed by annexin V and caspase 3/7 activity. In vivo, 99m Tc-duramycin uptake in COLO205 xenografts was increased 2.3-and 2.8-fold (P , 0.001) in mice treated with irinotecan and combination therapy, respectively. Blocking with unlabeled duramycin demonstrated specific binding of the radiotracer. After tumor irradiation with 4.5 Gy, 99m Tc-duramycin uptake in tumors increased significantly (1.24 ± 0.07 vs. 0.57 ± 0.08 percentage injected dose per gram in the unirradiated tumors; P , 0.001). γ-counting of radioactivity in the tumors positively correlated with cleaved caspase-3 (r 5 0.85, P , 0.001) and TUNEL (r 5 0.81, P , 0.001) staining. Conclusion: We demonstrated that 99m Tc-duramycin can be used to image induction of cell death early after chemotherapy and radiotherapy. It holds potential to be translated into clinical use for early assessment of treatment response.

99mTc-Duramycin SPECT Imaging of Early Tumor Response to Targeted Therapy: A Comparison with18F-FDG PET

“…Labeling with both SPECT ( 99m Tc) and PET ( 18 F, 68 Ga) radioisotopes and successful imaging of treatment-induced cell death in human lymphoma and lung and breast cancer (21) have been reported. Previously, our group has demonstrated the potential of 99m Tcduramycin to visualize radiotherapy- (15) and chemotherapyinduced (14,15) apoptosis. The sensitivity for apoptosis imaging was demonstrated by a 3-fold increase in 99m Tc-duramycin uptake (2 %ID/g) in colorectal cancer xenografts treated with irinotecan in combination with oxaliplatin versus vehicle-treated tumors, in good correlation with the tumor apoptotic response (4-to 7-fold increase).…”

“…After intravenous injection, 99m Tc-duramycin had favorable biodistribution, clearance, and pharmacokinetic profiles. Previously, we and others showed that 99m Tc-duramycin quickly cleared from circulation via the renal system, with low hepatic and gastrointestinal uptake (13)(14)(15). Radiotracer uptake in blood, muscle, liver, spleen, and kidneys was not significantly changed by conatumumab treatment (P .…”

“…In the current study, we used 99m Tc-duramycin, exploiting its high affinity and selectivity for phosphatidylethanolamine and optimal biodistribution profile (14,15), for in vivo SPECT imaging of cell death. Using small-animal SPECT and PET imaging of treatment-resistant and treatment-sensitive models of human colorectal cancer, we aimed at directly comparing the performance of 99m Tcduramycin with the current clinical gold standard 18 F-FDG for early therapy response evaluation using targeted therapy.…”

Noninvasive Whole-Body Imaging of Phosphatidylethanolamine as a Cell Death Marker Using 99mTc-Duramycin During TNF-Induced SIRS

“…Because of the known limitations associated with avidin-biotin systems (background) and green fluorescent protein (poor light penetration through tissues), 99m Tc is one of the most widely used radiolabels for duramycin. As such, 99m Tc-duramycin has been applied extensively to detect dead or dying cells in vivo, for example, in atherosclerotic plaques (23), in tumors challenged by different chemotherapy or radiotherapy schemes (24,25), or in different stages of lung injury (26), where it can be useful in the characterization of new cell death biomarkers.…”

Duramycin-porphyrin conjugates for targeting of tumour cells using photodynamic therapy

“…On the premise that PE and phosphatidylserine are regulated by the same mechanisms and that both are exposed during apoptosis [21] and after irradiation [1] a number of studies investigated whether PE exposure was upregulated after chemotherapy. It was demonstrated that externalisation of PE, detected by duramycin, could potentially be a marker of early apoptosis and could be utilised as a detection method for the response to chemotherapy treatment [22][23][24].…”