Overexpression of Murine Rnaset2 in a Colon Syngeneic Mouse Carcinoma Model Leads to Rebalance of Intra-Tumor M1/M2 Macrophage Ratio, Activation of T Cells, Delayed Tumor Growth, and Rejection

Abstract

Human RNASET2 acts as a powerful oncosuppressor protein in in vivo xenograft-based murine models of human cancer. Secretion of RNASET2 in the tumor microenvironment seems involved in tumor suppression, following recruitment of M1-polarized macrophages. Here, we report a murine Rnaset2-based syngeneic in vivo assay. BALB/c mice were injected with parental, empty vector-transfected or murine Rnaset2-overexpressing mouse C51 or TS/A syngeneic cells and tumor growth pattern and immune cells distribution in tumor mass were investigated. Compared to control cells, mouse Rnaset2-expressing C51 cells showed strong delayed tumor growth. CD86 + M1 macrophages were massively recruited in Rnaset2-expressing C51-derived tumors, with concomitant inhibition of MDSCs and CD206 + M2 macrophages recruitment. At later times, a relevant expansion of intra-tumor CD8 + T cells was also observed. After re-challenge with C51 parental cells, most mice previously injected with Rnaset2-expressing C51 cells still rejected C51 tumor cells, suggesting a Rnaset2-mediated T cell adaptive immune memory response. These results point at T2 RNases as evolutionary conserved oncosuppressors endowed with the ability to inhibit cancer growth in vivo through rebalance of intra-tumor M1/M2 macrophage ratio and concomitant recruitment of adaptive anti-tumor CD8 + T cells.Cancers 2020, 12, 717 2 of 18 in tumor suppression. Indeed, two independent in vivo xenograft-based murine models of human ovarian cancer have been previously used to demonstrate a marked RNASET2-mediated in vivo tumor suppression [1,2]. It has been previously shown that RNASET2 affects several important cancer-related parameters (such as modulation of cell proliferation, cytoskeletal re-organization, cell adhesion, motility, and angiogenesis) in a cell-autonomous manner, [3][4][5][6]. Moreover, further studies in both in vitro and in vivo experimental models have demonstrated a marked modulation of macrophage plasticity played by the RNASET2 protein, thus suggesting the occurrence of a RNASET2-mediated noncell-autonomous oncosuppressive role [7]. In fact, human RNASET2 overexpression in two independent human ovarian cancer cell lines, followed by challenging of immunodeficient mouse models with these cells, showed a marked tumor suppressive effect coupled with a RNASET2-mediated recruitment of M1-polarized host macrophages within the tumor mass [1,2]. These data, coupled to the well-established notion of T2 RNases as stress response genes [8,9], strongly suggest that human RNASET2 secretion by cancer cells might represent a "danger" signal for cells belonging to the monocyte/macrophage lineage, whose role would be to trigger an effective host immune response. Of note, the macrophage-activating role of T2 RNases has been recently confirmed in species far away from mammals in term of evolutionary distance, such as the invertebrate Hirudo verbana [10], thus pointing at T2 RNases as highly conserved immune system-related stress response proteins acting across many Phyla.The concept that the ...

M1 Macrophage and M1/M2 ratio defined by transcriptomic signatures resemble only part of their conventional clinical characteristics in breast cancer

“…Previously, polarized TAMs were recognized with oversimplified terms such as M1 macrophages and M2 macrophages based on the cell polarization concept of helper T cell type 1 and type 2 6,10,11 . However, with accumulating in vivo data from the studies of inflammatory diseases including cancer demonstrated that macrophage polarization is the representation of continuum of diverse functional state [6][7][8] . Therefore, currently the terms M1-like macrophages and M2-like macrophages are used to describe the phenotypes of polarized macrophages 9 .…”

Tumor-Associated Macrophages (TAMs) in Colorectal Cancer (CRC): From Mechanism to Therapy and Prognosis

“…As evolutionarily conserved tumor suppressors, T2 RNases can inhibit tumor growth in vivo by balancing the M1/M2 macrophage ratio in tumors and recruiting adaptive antitumor CD8 + T cells [143]. Furthermore, Halama, N. and his colleagues also confirmed that inhibiting CCR5 can repolarize the phenotype of TAMs from M2 to M1 by regulating the STAT3/SOCS3 signaling pathway in TAMs, thereby exerting antitumor effects in a phase I clinical trial of patients with CRC liver metastases [144].…”

Human RNASET2: A Highly Pleiotropic and Evolutionary Conserved Tumor Suppressor Gene Involved in the Control of Ovarian Cancer Pathogenesis

“…Noteworthy, a murine Rnaset2 syngeneic murine model was also recently reported by using mouse Rnaset2 -overexpressing C51 colon carcinoma and TS/A mammary adenocarcinoma syngeneic cells in comparison to control empty vector-transfected tumor cells [ 50 ]. Although the reported in vivo study was mainly focused on the C51 cancer cell model, a trend for a Rnaset2-mediated tumor suppressive effect was observed in TS/A cells as well [ 50 ].…”

“…Noteworthy, a murine Rnaset2 syngeneic murine model was also recently reported by using mouse Rnaset2 -overexpressing C51 colon carcinoma and TS/A mammary adenocarcinoma syngeneic cells in comparison to control empty vector-transfected tumor cells [ 50 ]. Although the reported in vivo study was mainly focused on the C51 cancer cell model, a trend for a Rnaset2-mediated tumor suppressive effect was observed in TS/A cells as well [ 50 ]. Of note, in the in vivo C51-based syngeneic model, a strong Rnaset2 -dependent retardation in tumor growth rate was observed, concomitant with early tumor infiltration by M1 macrophages, inhibition of M2 and myeloid-derived suppressor cells (MDSCs) cells and, very interestingly, a later expansion of CD8 + T lymphocytes with rejection capacity and antitumor recall immunity [ 50 ].…”