Abstract P1-05-01: Breast cancer cell-derived IL-1B drives metastasis and colonisation of the bone microenvironment
Authors: Ottewell PD , Lefley DV , Freeman K , Gregory WM , Hanby AM , Spicer-Hadlington AR , Liu X , Bradbury SM , Hambley L , Allocca G , Cookson V , Kruithof de-Julio M , Coleman RE , Brown JE , Holen I , Tulotta C 
Published: May 2019

Abstract
Background: Breast cancer bone metastases are incurable and new therapeutic targets needed. After homing and colonising bone, cancer cells remain dormant, until signals from the microenvironment stimulate their proliferation to form overt metastases. We have recently identified interleukin-1B (IL-1B) as a potential marker for predicting breast cancer patients at increased risk of skeletal relapse and established a role for IL-1 signalling in tumour dormancy in bone. Here we present novel data to support that tumour cell-derived IL-1B plays major roles in breast cancer metastasis and growth in bone.
Methods: Tumour/stromal IL-B and IL-1R1 expression was assessed in samples from the AZURE study following immunohistochemical staining. A humanised mouse model of MDA-MB-231 bone metastasis was used to assess effects of the IL-1R antagonist, Anakinra, on tumour growth and spontaneous metastasis. Effects of tumour cell-derived IL-1B on parameters associated with epithelial to mesenchymal transition and metastasis were measured by ELISA, QPCR, Western blot, transwell and scratch assays in MDA-MB-231, MCF7 and T47D cells transfected with IL-1B/control. Homing of breast cancer cells to bone was monitored in BALB/c nude mice intra venously injected with IL-1B overexpressing/control cells. To assess anti-tumour effects of IL-1 inhibition combined with standard of care, BALB/c or C57BL/6 mice were injected with 4T1 or E0771 cells (intra cardiac or intra ductal) 7-days prior to administration of Anakinra, doxorubicin, zoledronic acid (ZA) or placebo, alone or in combination for 14 days.
Results: In tissue samples from &gt;1300 patients with stage II/III breast cancer, active IL-1B in tumour cells correlated with relapse in bone (hazard ratio 1.85; 95% CI 1.05-3.26; P=0.02) and other sites (hazard ratio 2.09; 95% CI 1.26-3.48; P=0.0016). In a model of spontaneous human breast cancer metastasis to human bone, Anakinra significantly reduced metastasis to bone (from 80% in control animals to, 20% in Anakinra treated mice) and reduced the number of tumour cells shed into the circulation. Genetic manipulation of breast cancer cells to overexpress IL-1B demonstrated that exogenous production of IL-1B promoted EMT (decreased E-Cadherin, N-Cadherin and G-Catenin), invasion, migration and organ-specific homing in ER-ve (MDA-MB-231) and ER+ve (T47D and MCF7) cells in vitro and in vivo.
Contact between tumour cells and osteoblasts or bone marrow cells increased IL-1B secretion from all three cell types. Exposure of tumour cells to IL-1B in the absence of bone cells did not stimulate tumour cell proliferation. Instead, elevated concentrations of IL-1B caused expansion of the bone metastatic niche (increased osteoblasts and blood vessels) that in turn stimulated tumour proliferation.
Adding Anakinra to chemotherapy and ZA to mice injected with E0771 cells completely prevented bone metastasis formation and reduced extra-skeletal metastasis by 33%. In mice injected with 4T1 cells, the triple combination therapy reduced bone metastasis by 50% and extra-skeletal metastases by 65%.
Conclusion: Our novel data demonstrate that IL-1B/IL-1R1 signalling plays an important role in breast cancer metastasis to bone. Pharmacological inhibition of IL-1B has potential as a novel treatment.
Citation Format: Ottewell PD, Lefley DV, Freeman K, Gregory WM, Hanby AM, Spicer-Hadlington AR, Liu X, Bradbury SM, Hambley L, Allocca G, Cookson V, Kruithof de-Julio M, Coleman RE, Brown JE, Holen I, Tulotta C. Breast cancer cell-derived IL-1B drives metastasis and colonisation of the bone microenvironment [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P1-05-01.