Abstract 1460: Bone marrow-derived CD11b+/Podoplanin+ cells are lymphatic progenitors directly responsible for breast cancer lymphatic formation
Authors: Griggs Caitlin , Hall Kelly , Volk-Draper Lisa , Robinson Kathy , Ran Sophia 
Published: July 2016

Introduction: Lymphatic metastasis, a key factor for poor outcome of breast cancer (BC), strongly depends on lymphatic vessel (LV) formation. Recent studies suggest that lymphangiogenesis is strongly promoted by bone marrow (BM)-derived CD11b+ monocytes that differentiate into Lymphatic Endothelial Cell Progenitors dubbed here M-LECP. While BC are known to recruit BM monocytes, a specific BM subset responsible for tumor lymphatic formation has not been identified. We recently discovered that podoplanin (Pdpn) is the highest upregulated lymphatic marker that signifies transdifferentiation of either human or mouse immature myeloid cells into M-LECP. We hypothesized that BM-derived M-LECP are represented by a subset expressing podoplanin in CD11b+ myeloid cells that are capable of inducing tumor lymphatics. Methods: Tumor lymphatic vessels expressing myeloid markers were detected immunohistochemically by triple-staining in clinical specimens and experimental breast tumors. Expression of Vegfr-3, Lyve-1, Sca-1, and other markers in BM-derived CD11b+/Pdpn+ and Pdpn-negative subsets was determined by FACS. Expression of lymphatic-specific markers in CD11b+/Pdpn+ and Pdpn− myeloid cells isolated from MDA-MB-231 tumors was determined by RT-qPCR. CD11b+/Pdpn+ and Pdpn-negative subsets from BM of GFP-expressing mice were adoptively transferred to mice with orthotopic breast tumors followed by quantifying mobilized GFP+/CD11b+/Pdpn+ cells and the density of tumor LV. Results: M-LECP were absent in normal human breast tissues but highly present in tumors. Analysis of clinical BC specimens showed significant correlations between tumor-mobilized M-LECP, density of LV and metastasis to regional nodes. All examined BC models exhibited very high densities (60-90%) of double-positive macrophages expressing lymphatic markers, and lymphatic vessels expressing myeloid proteins. At least a half of CD11b+ cells isolated from tumors were positive for podoplanin and nearly 90% of CD11b+/Pdpn+ subset expressed markers of progenitor cells. Only CD11b+/Pdpn+ subset (but not other myeloid cells) expressed a broad panel of proteins specific to the lymphatic endothelial lineage. Adoptive transfer of BM-derived CD11b+/Pdpn+ or Pdpn-negative fractions into tumor-bearing mice showed that only Pdpn+ BM myeloid cells integrated into preexisting LV and caused a 10-fold increase in peritumoral and intratumoral LV density. Conclusion: We show for the first time in clinical BC samples that tumor-recruited M-LECP significantly correlate with LN metastasis. We also identified a phenotypically distinct BM fraction of CD11b+/Pdpn+ cells that directly promote generation of new tumor lymphatic vessels. Our data suggest that tumors induce myeloid BM cells to undergo differentiation into M-LECP that subsequently promote cancer lymphatics thereby promoting tumor spread.

Citation Format: Caitlin Griggs, Kelly Hall, Lisa Volk-Draper, Kathy Robinson, Sophia Ran. Bone marrow-derived CD11b+/Podoplanin+ cells are lymphatic progenitors directly responsible for breast cancer lymphatic formation. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1460.