Mast Cell-Derived Granzyme B Contributes To Resistance Against Anti-Angiogenic Therapy

Proceedings: AACR 107th Annual Meeting 2016; April 16-20, 2016; New Orleans, LASignificance: Targeted therapies have revolutionized the treatment of cancer. However, efficacy of anti-angiogenic therapies is limited due to significant resistance.Recent studies showed that the tumor microenvironment is involved in resistance towards targeted anti-angiogenic treatment. Based on the correlation of mast cell (MC) density with tumor growth and angiogenesis we put forward the hypothesis that MC might be implicated in anti-angiogenic therapy resistance.Methods: C57BL/6J, NSG or MC-deficient KitW-sh (Wsh) mice were subcutaneously injected with 5×105 (Panc02 and EL4) or 1×106 (TD2) cells +/- bone marrow derived MC. Tumors were treated with 20 mg/kg anti-VEGFR2 antibody (DC101) or 25 mg/kg cromoglicic acid (Cromo). BrdU was injected 12 h before sacrifice.Results: We show that MC alter the proliferative and organizational state of endothelial cells (EC). MC dose-dependently induced EC-proliferation (158 ± 12%; *pu003c0.05) and tube formation (290 ± 12%; *pu003c0.05). Furthermore, MC increased HUVEC migration by 1.4-fold (*pu003c0.05) and protected them from AAT in vitro. In MC-deficient mice, tumor growth was reduced by 36% (*pu003c0.05) and the efficacy of AAT was increased (WT + DC101: 1420 ± 134 mg; Wsh + DC101: 599 ± 107 mg; *pu003c0.05). Histomorphometric analyses unraveled that MC-deficiency decreased the numbers of mature pericyte-covered vessels by 80% (*pu003c0.05) rendering them more prone for therapy. Indeed, an additive anti-angiogenic effect of MC-deficiency and AAT was observed resulting in reduced microvessel density (MVD) and tumor cell proliferation. This “angiosensitizing” effect could be abrogated by adoptive transfer of bone marrow-derived MC into MC-deficient mice.In WT mice, AAT only initially reduced the proliferation of tumor vessels by 60% (*pu003c0.05), a process that got reverted after long-term treatment as a result of therapy resistance. Intriguingly, this pro-angiogenic rescue phenotype did not occur in MC-deficient mice. By blocking MC degranulation with Cromo we could increase the efficacy of AAT (DC101: 703 ± 48 mg; Cromo + DC101: 386 ± 92 mg; *pu003c0.05), leading to reduced vessel proliferation, MVD and tumor cell proliferation.Microarray analysis of tumor-resident MC unraveled increased expression levels of ECM-degrading granzyme b (gzmb) in response to therapy. MC-specific knock down of gzmb rendered tumors more susceptible for AAT and lowered the levels of alternative, pro-angiogenic mediators beside the VEGF-VEGFR2-axis in the tumor microenvironment.Conclusions: Our results indicate that tumor-resident MC interfere with AAT. We provide evidence that MC-derived gzmb liberates ECM-bound pro-angiogenic factors besides the targeted VEGF-VEGFR2 axis, thereby fine-tuning vessel maturation and proliferation, which ultimately decreases therapeutic efficacy. Importantly, knock down of gzmb and pharmacological inhibition of MC degranulation improved the therapeutic response towards AAT.Citation Format: Mark A. Wroblewski, Raimund Bauer, Miguel Cubas Cordova, Florian Udonta, Isabel Ben Batalla, Victoria Gensch, Stefanie Sawall, Jonas S. Waizenegger, Julian Pardo Jimeno, Klaus Pantel, Carsten Bokemeyer, Sonja Loges. Mast cell-derived granzyme b contributes to resistance against anti-angiogenic therapy. [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 3253.