Bone marrow homing heptapeptide bears homology to CD84

G Nowakowski MD1, A Mihaliak BA2; H Valinski BA, MB2, PS Becker MD PhD2

1 Norwalk Hospital, Norwalk, CT; 2 University of Massachusetts Medical School, Worcester, MA

AIM: The goal of this study was to identify peptide sequences that selectively bound to primitive hematopoietic stem cells. METHODS: A phage display peptide library consisting of M13 bacteriophages bearing 7-residue sequences was used for in vivo and in vitro biopanning procedures to identify phages that homed to the bone marrow and bound to primitive murine hematopoietic lineage-depleted, Hoechst 33342dull/Rhodamine123dull stem cells. Immunofluorescent analysis was used to determine target organs. Affinity chromatography and Western blotting were performed to further characterize the protein receptor. Immunoprecipitation of bone marrow membrane proteins is currently in progress to further identify the peptide receptor. RESULTS: The majority of the heptapeptides, 50% of those that homed in vivo and 85% of those that bound to stem cells, consisted of lysine, proline, phenylalanine, two serines, and two threonines. Fluorescent analysis revealed intense staining within the bone marrow, minimal staining in the lungs, and no staining in other organs, indicating bone marrow specificity. Affinity chromatography using synthetic peptide and solubilized murine bone marrow membranes yielded an 82 kDa protein. Binding of specific phage to a Western blot of solubilized bone marrow membranes demonstrated a 37/33 kDa doublet that may represent a proteolytic cleavage product of the 82 kDa protein. The heptapeptide is homologous to N-terminal CD84, a cell surface glycoprotein expressed by lymphocytes, as determined by searching GenBank. CONCLUSION: The described heptapeptide specifically homes to bone marrow and binds to primitive stem cells, and potentially could be used in future clinical applications to target hematopoietic stem cells.

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Paper presented at the International Symposium on Predictive Oncology and Intervention Strategies; Paris, France; February 9 - 12, 2002; in the section on Stem Cell Biology.

This presentation received an honorable mention in our poster contest and was recognized with the Symposium Presidents' Award for Scientific Excellence.