Monitoring therapeutic response of human ovarian cancer to trastuzumab by SPECT imaging with (99m)Tc-peptide-Z(HER2:342).

INTRODUCTION: Patients with human epidermal growth factor receptor 2 (HER2)-positive cancer are candidates for treatment with the anti-HER2 antibody trastuzumab. How to systemically assess tumor HER2 expression and identifying appropriate use of anti-HER2 therapies by noninvasive imaging in vivo is an urgent issue. The purpose of this study was to evaluate SPECT imaging of (99m)Tc-Gly-(D)Ala-Gly-Gly-Z(HER2:342) ((99m)Tc-peptide-Z(HER2:342)) for monitoring therapeutic response to trastuzumab in nude mice bearing HER2-positive SKOV-3 xenografts. METHODS: Nude mice bearing HER2-positive SKOV-3 xenografts were treated with trastuzumab (treatment group) or saline (control) with ten mice in each group. Mice in trastuzumab-treated group were given trastuzumab intraperiotoneally 4 mg/kg on day 1 and 2 mg/kg on day 8; Mice in control group were given physiological saline on day 1 and 8. Mice body weights and tumour volume were monitored every three days during treatment. In vivo SPECT imaging was performed in mice of the two groups using (99m)Tc-peptide-Z(HER2:342) before treatment, on day 8 and 15 after treatment. Radiolabeled probe uptake in tumours was measured as the ratio of radioactive counts in the tumour to that in the contralateral equivalent region (T/NT). After SPECT imaging on day 15, all the mice were euthanized, biodistribution studies of the SKOV-3 xenografts were carried out to validate the imaging results and HER2 expression of the transplanted tumours was analyzed by immunohistochemistry (IHC). Correlation analysis was performed between T/NT ratios acquired by in vivo SPECT imaging on day 15 and the HER2 level of tumours. In vitro cell binding capacity of (99m)Tc-Z(HER2:342) with SKOV-3 cells in the absence and presence of varying amount of trastuzumab were also conducted in the study. RESULTS: Twenty mice body weight in the two groups gradually increased during treatment, but there was no statistical difference (p > 0.05). Though volumes of SKOV-3 xenografts gradually increased in each group during the treatment, the transplanted tumours in trastuzumab-treated group had a slower growth than those in control group (p < 0.05). Compared with the baseline, the results of in vivo imaging showed that radionuclide accumulation in transplanted tumours reduced significantly in trastuzumab-treated group after treatment (p < 0.05), whereas the tumour accumulation in control group increased after treatment. Biodistribution studies demonstrated that the results corresponded well with in vivo imaging data. Immunohistochemical staining confirmed the significant reduction in tumor HER2 level upon trastuzumab treatment, and there was an obviously positive correlation between T/NT ratios and HER2 level of tumours with correlation coefficient rs = 0.919, p < 0.05. There was no significant significance in cell binding ratios between varying amount of trastuzumab and the absence of trastuzumab (p > 0.05). CONCLUSIONS: The early response to trastuzumab in mice bearing SKOV-3 xenografts was successfully monitored by SPECT imaging using (99m)Tc-peptide-Z(HER2:342). This approach may be valuable in monitoring the therapeutic response in HER 2-positive tumours under HER2-targeted therapy.

Preparation and Evaluation of (99m)Tc-Epidermal Growth Factor Receptor (EGFR)-Peptide Nucleic Acid for Visualization of EGFR Messenger RNA Expression in Malignant Tumors.

UNLABELLED: Epidermal growth factor receptor (EGFR) is overexpressed in many carcinomas and remains a prime target for diagnostic and therapeutic applications. There is a need to develop noninvasive methods to identify the subset of patients that is most likely to benefit from EGFR-targeted treatment. Noninvasive imaging of EGFR messenger RNA (mRNA) expression may be a useful approach. The aim of this study was to develop a method for preparation of single-photon-emitting probes, (99m)Tc-labeled EGFR mRNA antisense peptide nucleic acid (PNA) ((99m)Tc-EGFR-PNA), and nontargeting control ((99m)Tc-CTL-PNA) and to evaluate their feasibility for imaging EGFR mRNA overexpression in malignant tumors in vivo. METHODS: On the 5' terminus of synthesized single-stranded 17-mer antisense EGFR mRNA antisense PNA and mismatched PNA, a 4-amino-acid (Gly-(D)-Ala-Gly-Gly) linker forming an N4 structure was used for coupling (99m)Tc. Probes were labeled with (99m)Tc by ligand exchange. The radiochemical purity of these (99m)Tc-labeled probes was determined by reversed-phase high-performance liquid chromatography. Cellular uptake, retention, binding specificity, and stability of the probes were studied either in vitro or in vivo. Biodistribution and radionuclide imaging were performed in BALB/c nude mice bearing SKOV3 (EGFR-positive) or MDA-MB-435S (EGFR-negative) carcinoma xenografts, respectively. RESULTS: The average labeling efficiencies of (99m)Tc-EGFR-PNA and (99m)Tc-CTL-PNA were 98.80% ± 1.14% and 98.63% ± 1.36% (mean ± SD, n = 6), respectively, within 6 h at room temperature, and the radiochemical purity of the probes was higher than 95%. (99m)Tc-EGFR-PNA was highly stable in normal saline and fresh human serum at 37°C in vitro and in urine and plasma samples of nude mice after 2-3 h of injection. Cellular uptake and retention ratios of (99m)Tc-EGFR-PNA in SKOV3 cells were higher than those of (99m)Tc-CTL-PNA and the EGFR-negative control. Meanwhile, EGFR mRNA binding (99m)Tc-EGFR-PNA was blocked with an excess of unlabeled EGFR-PNA in SKOV3 cell lines. The biodistribution study demonstrated accumulation of (99m)Tc-EGFR-PNA primarily in the SKOV3 xenografts and in EGFR-expressing organs. Radionuclide imaging demonstrated clear localization of (99m)Tc-EGFR-PNA in the SKOV3 xenografts shortly after injection but not in (99m)Tc-CTL-PNA and the EGFR-negative control. CONCLUSION: (99m)Tc-EGFR-PNA has the potential for imaging EGFR mRNA overexpression in tumors.