Abstract 395: Host ephrinB2 regulates T-cells in tumor microenvironments

Abstract

Abstract Background: The tumor microenvironment consists of immune, stromal, and vascular cells. Proteins known to play roles in vascular development during embryogenesis also have been implicated in the control of leukocyte migration. The vascular patterning ligand ephrinB2, for example, influences T-cell migration in vitro. We therefore hypothesized that host ephrinB2 signaling regulates tumor immune microenvironments. Methods: EphrinB2+/lacZ mice were implanted with B16 melanoma, KR158 glioma, LLC lung, and EL4 T-lymphoma tumor cells. Macroscopic tumors were harvested and ephrinB2 expression assayed using x-gal staining. Mx1-cre/efnb2fl/fl mice were generated which allow for the post-natal deletion of ephrinB2 after pIpC treatment. Mx1-cre/efnb2+/+ mice served as controls. 1 million B16 or KR158 tumor cells were implanted in hindlimbs and tumor volume measured (V= 0.52*L*W2) at least twice weekly. Tumor tissues were stained with anti-CD3, F4/80, anti-CD34, and anti-NG2 antibodies. Experiments were repeated following T-cell depletion with i.p. injections of anti-CD4 and anti-CD8 antibodies 3x week. Finally, C57BL6 mice were transplanted with Mx1-cre/efnb2fl/fl or Mx1-cre/efnb2+/+ bone marrow and ephrinB2 was deleted after engraftment but prior to tumor growth experiments. Results: EphrinB2 was expressed in vascular structures all tumors. In addition we observed upregulation of ephrinB2 in microvascular structures in adjacent muscle during tumor invasion. Inducible deletion of host ephrinB2 significantly reduced B16 tumor growth (control tumor volume 2846 mm3 versus deleted tumor 869 mm3, p<0.01) and KR158 tumor growth (2028 mm3 versus 1403 mm3). EphrinB2 deletion did not affect CD34+ microvessel density, NG2+ pericyte or F4/80 macrophage infiltration. Rather, tumors grown in ephrinB2 deficient microenvironments demonstrated a significant increase in infiltrating CD3+ T-cells. In vivo depletion of T-cells abrogated the effect of ephrinB2 deletion on tumor growth delay demonstrating a role for immune cells in impairing tumor growth. EphrinB2 deletion in transplanted hematopoietic cells did not lead to tumor growth delay demonstrating a requirement for intact ephrinB2 signaling in vascular cells for T-cell regulation. Conclusion: EphrinB2 is upregulated in tumor vasculature and at sites of tumor invasion. Post-natal ephrinB2 deletion did not affect tumor angiogenesis but led to tumor growth delay via a T-cell mediated mechanism. These findings demonstrate a novel role for ephrinB2 specifically and vascular cells generally in regulating immune microenvironments. Therefore this study identifies a novel therapeutic strategy to increase the accumulation of T-cells in tumor tissues with relevance to combination immunotherapeutic strategies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 395. doi:1538-7445.AM2012-395