Evaluation of the therapeutic efficacy of a VEGFR2-blocking antibody using sodium-iodide symporter molecular imaging in a tumor xenograft model

https://doi.org/10.1016/j.nucmedbio.2010.05.009Get rights and content

Abstract

Purpose

Vascular endothelial growth factor receptor 2-blocking antibody (DC101) has inhibitory effects on tumor growth and angiogenesis in vivo. The human sodium/iodide symporter (hNIS) gene has been shown to be a useful molecular imaging reporter gene. Here, we investigated the evaluation of therapeutic efficacy by molecular imaging in reporter gene transfected tumor xenografts using a gamma imaging system.

Methods

The hNIS gene was transfected into MDA-MB-231 cells using Lipofectamine. The correlation between the number of MDA-MB-231-hNIS cells and the uptake of 99mTc-pertechnetate or 125I was investigated in vitro by gamma imaging and counting. MDA-MB-231-hNIS cells were injected subcutaneously into mice. When the tumor volume reached 180-200 mm3, we randomly assigned five animals to each of three groups representing different tumor therapies; no DC101 (control), 100 μg, or 150 μg DC101/mouse. One week and 2 weeks after the first injection of DC101, gamma imaging was performed. Mice were sacrificed 2 weeks after the first injection of DC101. The tumor tissues were used for reverse transcriptase-polymerase chain reaction (RT-PCR) and CD31 staining.

Results

Uptake of 125I and 99mTc-pertechnetate into MDA-MB-231-hNIS cells in vitro showed correlation with the number of cells. In DC101 treatment groups, the mean tumor volume was smaller than that of the control mice. Furthermore, tumor uptake of 125I was lower than in the controls. The CD31 staining and RT-PCR assay results showed that vessel formation and expression of the hNIS gene were significantly reduced in the tumor tissues of treatment groups.

Conclusion

This study demonstrated the power of molecular imaging using a gamma imaging system for evaluating the therapeutic efficacy of an antitumor treatment. Molecular imaging systems may be useful in evaluation and development of effective diagnostic and/or therapeutic antibodies for specific target molecules.

Introduction

Tumor angiogenesis is essential for the growth and metastasis of solid tumors [1], [2]. A number of angiogenic inhibitors have been developed to treat tumors, and many of these inhibitors are directed against vascular endothelial growth factor (VEGF) or its receptors (VEGFRs), which are considered key regulators of tumor angiogenesis [3], [4], [5], [6]. DC101 is a monoclonal antibody that targets VEGFR2 and has previously been shown to have antitumor effects in vivo [7], [8], [9], [10].

Tumor imaging is important to monitor tumor progression and proliferative changes after therapy. Molecular imaging based on reporter gene expression is suitable for tumor imaging in living subjects. Molecular imaging enables the visualization, characterization, and quantification of biologic processes in living subjects [11], [12]. Reporter genes are of considerable interest to image molecular events in small animals. Noninvasive reporter gene expression imaging offers opportunities to understand tumor progression, metastasis, and therapy in whole animals [12], [13], [14].

The human sodium/iodide symporter (hNIS) gene is a promising reporter gene. Recently, the hNIS gene has been intensively investigated with regard to the diagnosis and treatment of thyroid diseases [15], [16], [17], [18], [19]. As compared with conventional imaging reporter genes, an NIS gene has the merit of wide availability of substrates, including radioiodine and 99mTc-pertechnetate, which have been used commonly in clinical fields. Moreover, this gene is unlikely to influence the underlying cell biochemistry, and iodide is not metabolized in most tissues [20]. The hNIS gene can be transfected into specific cancer cells, and tumor models of gene transfer can be monitored visually to assess tumor progression and therapeutic effects using radioiodine or 99mTc-pertechnetate after drug administration [12], [20]. The effectiveness of NIS as a noninvasive reporter gene has already been demonstrated by several studies of tumor models of gene transfer [21], [22], [23], [24], [25], [26].

In this study, we investigated the potential of molecular imaging to evaluate the therapeutic effect of a specific antibody, DC101, in vivo.

Section snippets

Materials

All laboratory chemicals, unless indicated otherwise, were of molecular biology grade and were purchased from Sigma-Aldrich (St. Louis, MO, USA) or Invitrogen (Carlsbad, CA, USA). Na125I was obtained from Perkin Elmer Life Sciences (Boston, MA, USA). 99mTc-pertechnetate was eluted from a technetium generator produced by Samyoung Unitech (Seoul, Korea).

Tumor cell culture and transfected cell line generation

MDA-MB-231 breast cancer cells were cultured in DMEM (Dulbecco's Modified Eagle Medium) medium containing 100 IU/ml penicillin, 100 μg/ml

Radioactivity increased according to cell number

We established an MDA-MB-231-hNIS cell line that stably expresses hNIS. Fig. 1 shows the results for 125I and 99mTc-pertechnetate uptake, both radiotracers transported by hNIS, in a dilution series of MDA-MB-231 and MDA-MB-231-hNIS cells. As the number of MDA-MB-231-hNIS cells increased, so did the uptake of 125I (Fig. 1A). Increased radioactivity was also observed in gamma camera images (Fig. 1B). Similarly, as the number of MDA-MB-231-hNIS cells increased, so did the accumulation of 99m

Discussion

Molecular imaging that enables the visualization, characterization and quantification of biologic processes in living subjects is highly valuable for preclinical tumor research and especially for the evaluation of new antitumor drugs.

Here, we describe a monitoring strategy using a reporter gene transfected into tumor cells to evaluate the therapeutic efficacy, including the appropriate dose and administration route, of an antibody. We demonstrated, through the use of tumor xenograft models

Conclusions

We determined the necessary therapeutic dose of an anti-angiogenesis antibody, DC101, for therapy by intravenous injection in an in vivo mouse model. We used molecular imaging of hNIS to evaluate the dose-dependent therapeutic efficacy of DC101. Molecular imaging evaluation systems are powerful tools that can be used to aid the development of effective therapeutic antibodies.

Acknowledgments

This study was supported by grants from Nuclear R&D program through the Korea Science and Engineering Foundation funded by the Ministry of Science & Technology (20090062447) and the National R&D Program for Cancer Control, Ministry of Health, Welfare and Family affairs, Republic of Korea (0620220 and 0720420). This article was also supported by research funds of Chonbuk National University in 2009.

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