Abstract
Abstract
The interactions between neoplastic cells and their tumor microenvironment (TME) have become increasingly evident. Cancer-associated fibroblasts (CAFs) and M2 macrophages (M2 Mθ) are important components of the TME that have been recognized to orchestrate tumor-promoting inflammation and thereby tumor growth and progression. Here, we aim to determine the presence and role of CAFs and M2 Mθ in high grade cervical intraepithelial neoplasia (CIN 2/3) and invasive cervical carcinomas (SCCx).
We optimized immunohistochemistry (IHC) markers for M2 Mθ (CD163, ARG1), CAFs (αSMA) and Th1 T cells (TBX21) in paraffin-embedded tissue sections from CIN2/3 (n=6) and SCCx cases (n=4). We found an increased infiltration of TBX21+ cells in the epithelium and stroma of CIN2/3 and SCCx cases compared to their normal adjacent stroma. This suggests that despite increased Th1 T cells, the progression from normal to CIN2/3 to SCCx might be facilitated by M2 Mθ and CAFs. Moreover, by performing double staining Immunofluorescence (IF) we found that M2 Mθ (CD163+) were the cells responsible for ARG1 production, an immunesuppressive mechanism utilized by these cells. We recently stained by IHC a cervical tissue microarray (TMA) of normal, CIN2/3 and SCCx cases (n=63) with CD163, αSMA and TBX21, and we are currently analyzing the correlation of these markers with clinicopathological data available for these cases (HPV status, tumor stage, therapy response, overall survival).
With the hypothesis in mind that CAFs induce the polarization of macrophages into a M2 Mθ phenotype, we isolated fibroblasts from normal (n=2), CIN2/3 (n=1) and SCCx (n=2) fresh tissue explants obtained from surgery. We performed Immunocytochemistry (ICC) with αSMA, and observed a progressive increase in αSMA+ fibroblasts from normal to CIN2/3 and from CIN2/3 to SCCx fibroblasts, suggesting that CAFs might play an important role in the transformation of normal cervix into CIN2/3 and SCCx. Furthermore, we are currently performing co-culture experiments with normal, CIN2/3 and SCCx isolated fibroblasts and peripheral blood monocytes obtained from healthy donors. We will perform triple staining IF with vimentin, αSMA and CD163 in the slides generated from the co-culture, to evaluate if monocytes acquired a M2 Mθ phenotype with the presence of any of these 3 subsets of fibroblasts. Moreover, we will perform a cytokine analysis in the supernatants of these co-cultures to evaluate for potential cytokines produced by CAFs, implicated in M2 Mθ polarization.
In this study we provide evidence regarding the role of CAFs in SCCx initiation and progression. Although the mechanisms that regulate fibroblast activation have not been elucidated, it is possible that CAFs might serve as novel therapeutic targets in cancer, either alone or in combination with existing anti-cancer therapies.
Citation Format: Leonel Maldonado, Teresa Diaz-Montes, Abdulrahman Sinno, Edward J. Tanner, Christopher VandenBussche, Richard Roden, Iveta Yotova, Benjamin Tycko, Cornelia L. Trimble. Role and presence of cancer associated fibroblasts and M2 macrophages in high grade cervical intraepithelial neoplasia and invasive cervical carcinomas. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3672. doi:10.1158/1538-7445.AM2014-3672