Purinergic mechanisms in breast cancer metastasis.

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229 Background: Breast cancers survive in patients in a dormant state only to grow as metastatic disease in the future. Surgery to remove the primary tumor often terminates dormancy resulting in accelerated relapses. Many breast cancer deaths might be averted if we understand the cellular mechanisms underlying escape from dormancy. We have proposed that extracellular nucleotide signaling constitutes a pathological axis from the transformed tumor cell to the endothelium in the service of intravasation, dissemination, extravasation and angiogenesis. We demonstrate a role for the dinucleotide kinase NM23/NDPK in the generation of signals that stimulate angiogenesis and escape from latency. Methods: We have employed protein purification and identification by Western blot, tubulogenesis, migration and proliferation assays in human endothelial cells, signaling assays of receptor activation, nucleotide and receptor transactivation/phosphorylation assays and tumor growth and metastasis imaging in immunocompromized mice (SCID) to establish a role of r nucleotides in breast cancer metastasis. Results: A panel of breast cancer cell lines, but not normal breast cells, shed/secrete large amounts of NM23 that is active as an ADP transphosphorylase. NM23 stimulates tubulogenesis, proliferation and migration in cultures of human endothelial cells in a manner blocked by purinergic receptor antagonist, inhibitors of the nucleoside diphosphate kinase activity of NM23, or by VEGFR2 antagonists. In SCID mice carrying MDA-MB-231 luciferase over-expressing breast cancers, NM23 appears in the blood early in tumorigenesis. Treatment with the P2Y1 receptor antagonist MRS2179, or the NM23 antagonist ellagic acid slows the growth of primary breast tumors and prevents metastatic tumor formation. Conclusions: NM23 secretion leading to nucleotide generation activates endothelial P2Y1 receptors resulting in transactivation of VEGFR2 and angiogenesis in support of metastasis. Identification of Nm23 as a marker of early of breast cancer development and blockade of the extracellular actions of NM23 offer a new approach to breast cancer detection and treatment.