Abstract 2845: Dominant-negative TGFβRII (DNIIR) in c-neu-induced mammary carcinoma cells increases metastasis by disturbing vasculogenesis and pericyte balance
Authors: Novitskiy Sergey , Gorska Agnieszka , Pickup Michael , Chytil Anna , Aakre Mary , Moses Harold L. 
Published: June 2012

Abstract
In a mouse model with knockout of the type II TGF-β receptor (Tgfbr2) in mammary carcinoma cells (MMTV-PyMT), we have shown that TGF-β suppresses the expression of chemokines from tumor cells that are responsible for the expansion and recruitment of MDSCs in tumor bearing mice and these cells play a key role in tumor growth and metastasis. With the combination of the MMTV-c-neu oncogene and MMTV-DNIIR, tumors developed with a median latency of 41 weeks, 7 weeks later than mice expressing the MMTV-c-neu transgene alone. This indicates that c-neu signaling may cooperate to some degree with TGF-β signaling to induce tumor formation. This is different from the results with Tgfbr2 KO in combination with the PyMT transgene where the median latency was much shorter than with the MMTV-PyMT transgene alone. Significantly, expression of the DNIIR along with c-neu increases pulmonary metastases four fold similar to the effect of knockout of Tgfbr2 in the PyMT model. This model is important, because expression of the DNIIR transgene increases latency of mammary tumor formation in parallel with increases pulmonary metastases.
Because of the long latency in the c-neu model, we used of established c-neu and c-neu/DNIIR (cDNIIR) cell lines in collagen plugs orthotopically implanted in syngeneic mice. We found that cDNIIR implants have a longer latency than the c-neu cells, similar to that observed with the spontaneous tumors. However, by 11 weeks, the cDNIIR tumors are the same size as the c-neu tumors. Importantly, the cDNIIR tumors metastasize more readily to the lung than the c-neu tumors similar to the spontaneous model. We found that cDNIIR carcinoma cells, unlike the PyMT-Tgfbr2 KO cells, actually express less CXCL1/5 in parallel with less abundance of myeloid cells in tumor tissue. Protein analysis showed that cDNIIR cells express significantly more MCP-1, IL-6 and VEGF than c-neu cells. Also, we found that TGF-β has a very strong effect for up-regulation of VEGF secretion from cDNIIR cells compared with c-neu, which we expect is the primary mechanism for increased metastasis. It has been shown by other investigators that TGF-β plays an important role in pericytes and activation of this signaling pathway in these cells is necessary to support angio-and vasculogenesis. We hypothesize that increased binding of TGF-β to cDNIIR will lead to diminished TGF-β in tumor microenvironment and result in a decreased number of pericytes. By immunofluorescent staining of tumor tissue for blood vessels (CD31) and pericytes (desmin, NG2) we found fewer vessels wrapped by pericytes in cDNIIR mice.
We conclude that the tumor microenvironment of mammary carcinomas expressing DNIIR have increased vasculogenesis by up-regulation of VEGF in tumor cells and reduced pericytes. As a result these mice have more metastasis by increased permeability and leakage of tumor vessels.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2845. doi:10.1158/1538-7445.AM2011-2845