Molecular imaging of very late antigen-4 (α4β1 integrin) in the premetastatic niche

Monica Shokeen, Alexander Zheleznyak, Jessica M. Wilson, Majiong Jiang, Ruiwu Liu, Riccardo Ferdani, Kit S. Lam, Julie K. Schwarz, Carolyn J. Anderson

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Despite advances in cancer treatment over the past few decades, metastatic disease remains the primary cause of morbidity and mortality. Recent reports suggest the formation of a "premetastatic niche" before the metastatic cascade, where niche is defined as the microenvironment for tumor cells to be able to engraft and proliferate at secondary sites. Bone marrow-derived (BMD) cells that express vascular endothelial growth factor receptor-1 and very late antigen-4 (VLA-4) have been shown to arrive at sites of metastasis to form a receptive environment for tumor cells. Here we describe experiments toward imaging of VLA-4-positive BMD cells using a high-affinity PET probe, 64Cu-labeled 11-bis(carboxymethyl)-1,4,8,11-tetraazabicyclo[ 6.6.2] hexadecane (CB-TE2A)-LLP2A. Methods: VLA-4-negative MDA-MB-231/firefly luciferase (fluc) human breast tumor cells were injected intraarterially in the left ventricle in nude mice. Tumor metastasis in mice was monitored for 30 d by bioluminescence imaging and small-animal PET/CT. Small-animal PET images were collected 2 h after mice were injected in the tail vein with 64Cu-CB-TE2A-LLP2A (5.6-11.1 MBq [150-300 μCi; specific activity, 400 μCi/mg]). Cellular uptake of 64Cu-CB-TE2A-LLP2A was determined in VLA-4- positive B16F10 mouse melanoma cells and VLA-4-negative MDA-MB-231/fluc human breast cancer tumor cells. Biodistribution experiments in nude mice bearing VLA-4-positive B16F10 subcutaneous tumors in the flank were conducted to validate targeting of VLA-4-positive cells in vivo. Results: Uptake of 64Cu-CB-TE2A-LLP2A was higher in VLA-4-positive human melanoma B16F10 cells than in VLA-4-negative MDAMB- 231 cells (P <0.05). In B16F10 tumor-bearing mice, 64Cu- CB-TE2A-LLP2A had high uptake in the VLA-4-rich organs marrow, spleen, and tumor (11.26% ± 2.59%, 8.36% ± 2.15%, and 3.09% ± 0.58% injected dose/g, respectively). Cumulative standardized uptake value data from 2 independent studies (n 5 7 and 8 mice) on nude mice implanted with VLA- 4-negative MDA-MB-231/fluc human breast tumor cells suggested an influx of VLA-4-positive BMD cells that corresponded to metastasis (P <0.05). Immunohistochemical analysis and flow cytometry also showed upregulation of VLA-4-positive cell clusters and BMD cells at the metastatic sites, providing evidence for noninvasive imaging of BMD cells in the premetastatic niche. Conclusion: The results of the study demonstrated the potential of PET with VLA-4-targeted 64Cu-CB-TE2A-LLP2A to visualize BMD cell reorganization and expansion noninvasively in vivo.

Original languageEnglish
Pages (from-to)779-786
Number of pages8
JournalJournal of Nuclear Medicine
Issue number5
StatePublished - May 1 2012


  • Cu
  • PET imaging
  • Premetastatic niche
  • Very late antigen-4

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    Shokeen, M., Zheleznyak, A., Wilson, J. M., Jiang, M., Liu, R., Ferdani, R., Lam, K. S., Schwarz, J. K., & Anderson, C. J. (2012). Molecular imaging of very late antigen-4 (α4β1 integrin) in the premetastatic niche. Journal of Nuclear Medicine, 53(5), 779-786.