Abstract P1-01-01: The blood-Tumor barrier as a therapeutic target to improve therapy of brain metastases of breast cancer
Authors: Steeg PS , Lyle TR , Paranjapee A , Lockman PR , Duchnowska R , Brastianos PK , Peer C , Figg WD , Pauly GT , Schneider JP , Smith QR , Gril B 
Published: February 2018

Abstract
Objectives: Brain metastases of breast cancer demonstrate low and heterogeneous levels of permeability to drugs in mouse models and human craniotomies. The Blood-Brain Barrier (BBB), the protective lining of CNS blood vessels, impedes drug entry into the normal brain. When a metastasis forms, the BBB is locally altered to a poorly characterized Blood-Tumor Barrier (BTB). Quantitative experimental models indicate that most brain metastases have increased permeability over the normal BBB, but BTB permeability is both heterogeneous and ˜2 logs less than that of systemic metastases. We have interrogated three hematogenous models of brain metastasis of breast cancer to ask (1) whether the BTB is an ordered structure or a random breakdown of the BBB; (2) among brain metastases, whether consistent differences underlie the BTBs of lesions with low- and high permeabilities to fluorescent markers and drugs; (3) if alterations in BTB composition can functionally change its permeability. Our long term goal is to enhance uptake of drugs into brain metastases to effective levels.
Results: When uninvolved brain was compared with any brain metastasis, alterations in endothelial, pericytic, astrocytic, and microglial components of the BBB were observed. Both the pericyte and astrocyte components of the BTB were consistently altered with increased permeability: When metastases with relatively low and high permeability were compared, increased expression of a desmin+ subpopulation of pericytes was associated with higher permeability (231-BR6 P=0.0002; JIMT-1-BR3 P = 0.004; SUM190-BR3 P=0.008). A trend toward reduced CD13+ pericytes was observed in highly permeable metastases (231-BR6 P =0.014; JIMT-1-BR3 P =0.002, SUM190-BR3, NS). For GFAP+ astrocytes in the neuroinflammatory response surrounding metastases, no overall difference in cell number was observed between low and high permeability lesions. However, gene expression profiling of laser capture microdissected low and high permeabililty lesions demonstrated overexpression of the sphingosine-1 phosphate receptor 3 (S1P3) in the astrocytes of highly permeable lesions, which was confirmed at the protein expression level in all three models (231-BR6 P=0.034; JIMT-1-BR3 P = 0.01; SUM190-BR3 P=0.016). Inhibition of S1P3 via S1PR3 shRNA or a selective antagonist (TY-52156) functionally tightened the BTB in an in vitro model. Administration of TY-52156 to mice harboring 231-BR6 brain metastases had no effect on metastasis number, but decreased uptake of Texas Red Dextran dye into metastases (P=0.016). S1P3 mediated its effects on BTB permeability through astrocytic secretion of IL-6 and CCL2, which altered endothelial expression and localization of adhesive proteins, a potentially translatable pathway. Both desmin+ pericytes and S1P3+ astrocytes are present in human craniotomy specimens.
Conclusions: These experiments demonstrate that the BTB is a structure with consistent properties, and that further consistent changes underlie the transition from a low to high permeability BTB. While proof of principle, S1P3 inhibition studies indicate that the BTB permeability can be functionally modulated in vivo.
Citation Format: Steeg PS, Lyle TR, Paranjapee A, Lockman PR, Duchnowska R, Brastianos PK, Peer C, Figg WD, Pauly GT, Schneider JP, Smith QR, Gril B. The blood-Tumor barrier as a therapeutic target to improve therapy of brain metastases of breast cancer [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P1-01-01.