Abstract
BackgroundWe aimed to develop a mouse spontaneous liver metastasis model from an orthotopically implanted human colon cancer cell line stably expressing a human sodium/iodide symporter (NIS) reporter gene, which can be imaged with single-photon emission computed tomography (SPECT) using 99mTcO4−.MethodsA recombinant plasmid containing a constitutively driven NIS gene (pcDNA3-NIS) was transfected into the human colon cancer cell line HCT116, and stable cell lines were established. The stable cells were subcutaneously injected into the nude mice. When the diameter reached 10 mm, the xenografts were excised, cut into small fragments, and orthotopically implanted into the cecal walls of another nude mice. 99mTcO4− SPECT/CT imaging was initiated 8 weeks later and repeated every 1 to 2 weeks.ResultsThe production and function of NIS protein was confirmed in vitro by Western blotting and 99mTcO4− uptake assay. On SPECT/CT imaging, focal 99mTcO4− uptake was detected in the liver. Necropsy revealed local growth of the orthotopic colon xenografts with extensive invasion, microscopic serosal metastasis, and metastatic foci in the corresponding hepatic regions showing focal 99mTcO4− uptake. Immunohistochemistry revealed high levels of NIS expression in cells forming liver tumor, indicating that the liver tumor cells originated from the orthotopic colon xenografts.ConclusionsThe present proof-of-concept study provided a rationale for employing a radionuclide reporter gene for the specific visualization of spontaneous liver metastasis in living mice. This unique animal model of clinically relevant and externally detectable liver metastasis will be a powerful tool for investigating tumor biology and developing novel therapies for cancer metastasis.
Highlights
We aimed to develop a mouse spontaneous liver metastasis model from an orthotopically implanted human colon cancer cell line stably expressing a human sodium/iodide symporter (NIS) reporter gene, which can be imaged with single-photon emission computed tomography (SPECT) using 99mTcO4−
To reproduce clinically relevant spontaneous tumor metastasis, tumor cells should be implanted into the correct anatomical sites of immunodeficient mice. This animal model has a drawback - poor predictability of spontaneous metastasis formation from the orthotopic primary tumor. These tumor cells have sometimes been labeled with fluorescent [6,7] or bioluminescent [8,9] reporter genes to enable specific detection, but the poor penetration of light within the mammalian tissue limits the use of fluorescent and bioluminescent imaging to superficial lesions
Western blotting NIS protein was recognized at approximately 97 kDa on Western blots from the three selected stable cell lines A, B, and C, but not from the negative control cell line N (Figure 1A,B)
Summary
We aimed to develop a mouse spontaneous liver metastasis model from an orthotopically implanted human colon cancer cell line stably expressing a human sodium/iodide symporter (NIS) reporter gene, which can be imaged with single-photon emission computed tomography (SPECT) using 99mTcO4−. To reproduce clinically relevant spontaneous tumor metastasis, tumor cells should be implanted into the correct anatomical (orthotopic) sites of immunodeficient mice. This animal model has a drawback - poor predictability of spontaneous metastasis formation from the orthotopic primary tumor. These tumor cells have sometimes been labeled with fluorescent [6,7] or bioluminescent [8,9] reporter genes to enable specific detection, but the poor penetration of light within the mammalian tissue limits the use of fluorescent and bioluminescent imaging to superficial lesions. Cells expressing fluorescent proteins remain detectable even after the cells die, which may prevent from assessing therapeutic response of the metastatic tumor
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