Ex vivo and in vivo evaluation of overexpressed VLA-4 in multiple myeloma using LLP2A imaging agents

Deepti Soodgupta, Haiying Zhou, Wissam Beaino, Lan Lu, Michael Rettig, Mark Snee, James Skeath, John F. DiPersio, Walter J. Akers, Richard Laforest, Carolyn J. Anderson, Michael H. Tomasson, Monica Shokeen

Research output: Contribution to journalArticle

17 Scopus citations

Abstract

Very-late-antigen-4 (VLA-4, α4β1 integrin, CD49d/CD29) is a transmembrane adhesion receptor that plays an important role in cancer and immune responses. Enhanced VLA-4 expression has been observed in multiple myeloma (MM) cells and surrounding stroma. VLA-4 conformational activation has been associated with MM pathogenesis. VLA-4 is a promising MM imaging and therapeutic biomarker. Methods: Specificity of 64Cu-LLP2A (64Cu-CB-TE1A1PPEG4- LLP2A), a high-affinity VLA-4 peptidomimetic-based radiopharmaceutical, was evaluated in α4 knock-out mice and by competitive blocking in wild-type tumor-bearing mice. 64Cu-LLP2A PET/CT (static and dynamic) imaging was conducted in C57BL6/KaLwRij mice bearing murine 5TGM1-GFP syngeneic tumors generated after intravenous injection via the tail. Blood samples were collected for serum protein electrophoresis. Bone marrow and splenic cells extracted from tumor-bearing and control mice (n = 3/group) were coincubated with the optical analog LLP2A-Cy5 and mouse B220, CD4, Gr1, and Mac1 antibodies and analyzed by fluorescence-activated cell sorting. Human radiation dose estimates for 64Cu-LLP2A were extrapolated from mouse biodistribution data (6 time points, 0.78 MBq/animal, n = 4/group). Ten formalin-fixed paraffin-embedded bone marrow samples from deceased MM patients were stained with LLP2A-Cy5. Results: 64Cu-LLP2A and LLP2A-Cy5 demonstrated high specificity for VLA-4-positive mouse 5TGM1-GFP myeloma and nonmalignant inflammatory host cells such as T cells and myeloid/monocytic cells. Ex vivo flow cytometric analysis supported a direct effect of myeloma on increased VLA-4 expression in host hematopoietic microenvironmental elements. SUVs and the number of medullar lesions detected by 64Cu-LLP2A PET corresponded with increased monoclonal (M) protein (g/dL) in tumor-bearing mice over time (3.29 ± 0.58 at week 0 and 9.97 ± 1.52 at week 3). Dynamic PET with 64Cu-LLP2A and 18F-FDG demonstrated comparable SUV in the prominent lesions in the femur. Human radiation dose estimates indicated urinary bladder wall as the dose-limiting organ (0.200 mGy/MBq), whereas the dose to the red marrow was 0.006 mGy/MBq. The effective dose was estimated to be 0.017 mSv/MBq. Seven of the ten human samples displayed a high proportion of cells intensely labeled with LLP2A-Cy5 probe. Conclusion: 64Cu-LLP2A and LLP2A-Cy5 demonstrated binding specificity for VLA-4 in an immune-competent murine MM model. 64Cu-LLP2A displayed favorable dosimetry for human studies and is a potential imaging candidate for overexpressed VLA-4.

Original languageEnglish
Pages (from-to)640-645
Number of pages6
JournalJournal of Nuclear Medicine
Volume57
Issue number4
DOIs
StatePublished - Apr 1 2016

Keywords

  • Animal imaging
  • Dosimetry
  • LLP2A
  • Molecular imaging
  • Multiple myeloma
  • Radiobiology/dosimetry
  • VLA-4 specificity

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