CXM: A New Tool for Mapping Breast Cancer Risk in the Tumor Microenvironment

Abstract

The majority of causative variants in familial breast cancer remain unknown. Of the known risk variants, most are tumor cell autonomous and little attention has been paid yet to germline variants that may affect the tumor microenvironment. In this study, we developed a system called the Consomic Xenograft Model (CXM) to map germline variants that impact only the tumor microenvironment. In CXM, human breast cancer cells are orthotopically implanted into immunodeficient consomic strains and tumor metrics are quantified (e.g., growth, vasculogenesis, and metastasis). Because the strain backgrounds vary, whereas the malignant tumor cells do not, any observed changes in tumor progression are due to genetic differences in the non-malignant microenvironment. Using CXM, we defined genetic variant(s) on rat chromosome 3 that reduced relative tumor growth and hematogenous metastasis in the SS.BN3IL2Rγ consomic model compared to the SSIL2Rγ parental strain. Paradoxically, these effects occurred despite an increase in the density of tumor-associated blood vessels. In contrast, lymphatic vasculature and lymphogenous metastasis were unaffected by the SS.BN3IL2Rγ background. Through comparative mapping and whole genome sequence analysis, we narrowed candidate variants on rat chromosome 3 to six genes with a priority for future analysis. Collectively, our results establish the utility of CXM to localize genetic variants affecting the tumor microenvironment which underlie differences in breast cancer risk.

Host genetic modifiers of nonproductive angiogenesis inhibit breast cancer

“…Likewise, HCC1806-RR showed significant growth inhibition in SS.BN3 IL2Rγ rats (5,904 ± 700 mm 3 , n = 6) compared with SS IL2Rγ rats (8,893 ± 984 mm 3 , n = 8) (Figure 1C). These data demonstrate that TME-specific modifier(s) residing on RNO3 inhibit breast cancer growth [5]. …”

“…All procedures were approved by the MCW IACUC committee. The generation of the SS IL2Rγ and SS.BN3 IL2Rγ rats has been described elsewhere [5]. Congenic strains were generated by crossing SS/Mcwi and the SS.BN3 consomic strain, followed by intercrossing the F1 progeny and F2 generation to capture different regions of RNO3 by marker-assisted selection, as described previously [13, 14].…”

“…The TNBC orthotopic models of firefly luciferase-tagged MDA-MB-231 (231 Luc+ ) cells and Renilla-tagged HCC-1806 (HCC1806-RR) were described previously [5, 15, 16]. Briefly, 231 Luc+ cells (6×10 6 ) or HCC1806-RR cells (4×10 6 ) were suspended in 50% Matrigel and implanted into the #3 mammary fat pad (MFP) of female SS IL2Rγ and SS.BN3 IL2Rγ rats (n = 5–8 per group).…”

“…Immunofluorescent staining was performed using antibodies against CD31 (R&D Systems; AF3628 or BD; 555025) and DLL4 (R&D Systems; AF1389), as described previously [5, 22]. Images were acquired at 200X and 400X magnification on a Nikon E400 microscope equipped with a Spot Insight camera (Nikon Instruments).…”

“…We developed CXM as the first strategy for mapping host TME modifiers [5]. In CXM, human breast cancer cells are orthotopically implanted into consomic and congenic xenograft host strains, which are derived from two parental strains with different susceptibilities to breast cancer (see Figure 1A for schematic).…”