Abstract PD9-08: Breast cancer liver metastases vascularize by vessel co-option, not angiogenesis, and have a desert immune phenotype: A histopathological and gene expression study
Authors: Vermeulen PB , van Dam P-J , Daelemans S , Latacz E , Joye I , Kockx M , Dirix P , Verhoef K , Grunhagen D , Huget P , Van Laere S , Dirix LY 
Published: May 2019

Phase 3 trials of bevacizumab combined with chemotherapy in metastatic breast cancer have consistently failed to demonstrate a survival benefit for the addition of bevacizumab. When cancers metastasize to highly vascular organs (including the liver), they can utilize vessel co-option, instead of angiogenesis, as a mechanism to obtain a vascular supply (1). We have repeatedly shown by histopathological analyses that almost all (95%, 2 cohorts) breast cancer liver metastases utilize vessel co-option instead of angiogenesis to vascularize (2,3). The prevalence of vessel co-option in breast cancer could explain, at least in part, why anti-angiogenic therapy has been a disappointing therapeutic approach in metastatic breast cancer. Animal models of non-angiogenic liver and lung metastases also displayed resistance to anti-VEGF treatment (3,4).

We have now undertaken a gene expression study (mRNA sequencing) of targeted samples at the tumor-liver interface to discover gene expression patterns and signaling pathways that are associated with non-angiogenic growth of metastatic cancer in the liver (n = 70).

A network to detect biological themes of non-angiogenic growth was built by gene set enrichment analysis. Key components of this network are: cancer cell motility and invasion, epithelial-to-mesenchymal transition, stemness and proliferation. This contrasts with the network of angiogenic liver metastases of which the most important components are inflammation and ECM remodeling. Semi-automated image analyses of CD8-immunostained section of liver metastases confirms that non-angiogenic liver metastases have a significantly lower density of CD8-positive cytotoxic T-lymphocytes at the tumor-liver interface when compared with angiogenic liver metastases (300 cells/mm2 and 1000 cells/mm2, respectively (p&lt;0.0001)). In addition, a clear CXCL13-driven B-cell gene expression signature is associated with angiogenic growth of liver mets but is absent in non-angiogenic growth of breast cancer liver metastases. Gene expression patterns that may be play a role in vessel co-option are the up-regulation of LAMA3, LAMB3, LAMC2, coding for the 3 subunits of laminin-5, and of ITGA3, ITGB1, ITGA6 and ITGB4, coding for both integrin-receptors of laminin-5. This supports the concept of 'adhesive' vessel co-option during which cancer cells use the basement membrane of the co-opted blood vessels as a soil (5). In addition, the claudin-2 gene (CLDN2) is significantly overexpressed in non-angiogenic liver metastases which is consistent with earlier reports on the role of claudin-2 during breast cancer metastasis to the liver (6).

In conclusion, we provide evidence, based on morphology and gene expression, for the almost exclusive non-angiogenic growth of breast cancer liver metastases. In addition, non-angiogenic breast cancer liver metastases are characterized by a desert immune phenotype. Both observations can have an impact on the treatment strategy of patients with metastatic breast cancer.

References: 1. 10.1038/nrc.2018.14 – 2. 10.1038/sj.bjc.6601727 – 3. 10.1038/nm.4197 – 4. 10.1002/path.4845 – 5. 10.1097/NEN.0b013e318233afd7 – 6. 10.1038/onc.2010.518

Citation Format: Vermeulen PB, van Dam P-J, Daelemans S, Latacz E, Joye I, Kockx M, Dirix P, Verhoef K, Grunhagen D, Huget P, Van Laere S, Dirix LY. Breast cancer liver metastases vascularize by vessel co-option, not angiogenesis, and have a desert immune phenotype: A histopathological and gene expression study [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 PD9-08.