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L. A. Leach, K. Valyi-Nagy, V. Barak, J. Pe'er, I. Kalickman, N. Baumann, A. Y. Lin, M. Apushkin, A. J. Maniotis, R. Folberg; Screening for Biomarkers of Early Metastatic Uveal Melanoma in an Animal Model. Invest. Ophthalmol. Vis. Sci. 2007;48(13):4764.
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There is a clinical need to develop sensitive markers to detect early hepatic metastases in uveal melanoma. We have reported that osteopontin (OPN) is elevated in patients with metastatic uveal melanoma. We have also reported that highly invasive uveal melanomas synthesize keratin 18 (TPS). Using an animal model of metastatic uveal melanoma to the liver, we compared the sensitivity of OPN and TPS in detecting very early subclinical metastases.
Supernatant from cultured uveal melanoma cell lines was assayed for OPN and TPS by ELISA, as was pooled serum from 4 untreated SCID mice. Sera were then collected from 2 groups of 15 SCID mice each 2 weeks after injection of either tissue culture medium (control) or human metastatic uveal melanoma (MUM2B) into the mouse liver. For each group of mice, sera were pooled to measure OPN and TPS because the volume of blood in each mouse does not generate a sufficient volume of serum for the ELISA assays.
We detected 17 µg/L OPN and 333 µg/L TPS in the supernatants of MUM2B cells in 3D culture conditions. After 2 weeks, 11 of the 15 mice who were injected with MUM2B cells developed small hepatic nodules (0.1 - 2.8 mm; mean = 0.80 mm). Serum OPN in normal untreated mice, media-injected mice, and mice receiving MUM2B cells yielded levels <0.14 µg/L for each group. By contrast, serum TPS levels in normal untreated mice (50.6 µg/L) and media-injected mice (84.7 µg/L) were substantially lower than levels in MUM2B-injected mice (601 µg/L). The assay for OPN is human specific, and OPN was not detected in serum for untreated mice (<0.14 µg/L). The TPS levels in the control mice that did not receive MUM2B cells (50.6 µg/L and 84.7 µg/L) are most likely due to cross-reactivity of the assay with the mouse TPS protein (Endo B) which has 89.7% amino acid sequence identity to the human TPS protein.
Although both OPN and TPS discriminate clinically between patients with and without metastatic disease, the sharp elevation in TPS indicates that it may be a more sensitive marker than OPN for detecting low tumor burdens. Therefore, measurement of serial TPS levels may be helpful in detecting small, subclinical hepatic metastases. This study illustrates the use of the animal model of hepatic metastases in clinical screening for putative sensitive biomarkers to detect metastatic uveal melanoma in patients.
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