Abstract 2651: Characterization of TIE2 function in cancer cell dormancy and bone metastases
Authors: Drescher Florian , Dueñas Salvador , Olvera-Rodriguez Felipe , Higuera Nicolás Serafín , Flores Samanta Jiménez , Juarez Patricia , Licea-Navarro Alexei , Fournier Pierrick G. 
Published: September 2020

Abstract
After arriving in bone, cancer cells can enter a dormancy state that can last for years, during which they are resistant to treatment. The bone microenvironment induces dormancy via mechanisms similar to the ones of hematopoietic stem cells (HSC). The angiopoietin receptor, TIE2, induces the dormancy of HSC and some cancer cells and is a target for anti-angiogenic treatments. However, its inhibition may restart the proliferation of cancer cells, posing the risk of bone metastasis growth, while increasing the sensitivity of cancer cells to chemotherapy. Thus, we aim to characterize the role of TIE2 in dormancy of bone metastases.
We found that higher levels of Tie2 in the primary tumor of breast cancer (BCa) patients are associated with a good prognosis in multiple cohorts. However, expression of Tie2 protein or mRNA was either very low or undetected in cultured BCa and prostate cancer (PCa) cell lines. BCa and PCa cells transduced to express TIE2 constitutively showed a gradual loss of TIE2+ cells, despite continuous antibiotic selection. Congruently, in 5 different patient datasets, Tie2 expression is significantly lower in invasive BCa compared to healthy breast tissue. These results suggest that there is no expression of TIE2 in growing cancer cells.
In MCF-7 TIE2tet BCa cells conditional TIE2 expression with a Tet-On system caused a significant decrease of cell proliferation and the percentage of cells in S and G2/M phase but not in MCF-7 GFPtet control cells. Expression of TIE2, but not GFP, significantly decreased mRNA levels of the proliferation marker Ki67 while increasing the dormancy markers p21 and p27. Additionally, TIE2 expression increased the resistance to the chemotherapeutic 5-Fluorouracil. 4T1 Tie2tet cells inoculated in the mammary fat pads of Balb/C mice receiving Dox showed significantly reduced tumor volume and weight of excised tumors compared to control mice, suggesting TIE2 caused a growth reduction. When intracardiacally inoculated, 4T1 Tie2tet cells caused bone metastases regardless of TIE2 expression. However, the osteolysis area was significantly decreased when TIE2 was expressed, indicating a delayed bone metastasis development.
To reverse dormancy and increase chemotherapy sensitivity, we selected a shark antibody (vNAR) that binds to TIE2 using a synthetic library and phage display. In silico modeling indicated this clone could bind to the FN2 domain and inhibit the multimerization of TIE2, which could prevent signaling.
Overall, we found that expression of TIE2 decreases BCa cell growth in vitro and in vivo, and their sensitivity to 5-Fluorouracil, while increasing the expression of dormancy marker in vitro, and the time to the development of metastasis and survival of BCa patients. Our results are then consistent with TIE2 inducing dormancy of cancer cells. Thus, neutralizing TIE2 with a vNAR in combination with cancer therapy could be used for the treatment of patients at risk of bone metastases.
Citation Format: Florian Drescher, Salvador Dueñas, Felipe Olvera-Rodriguez, Nicolás Serafín Higuera, Samanta Jiménez Flores, Patricia Juarez, Alexei Licea-Navarro, Pierrick G. Fournier. Characterization of TIE2 function in cancer cell dormancy and bone metastases [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2651.