Microenvironment-Derived FGF-2 Stimulates Renal Cell Carcinoma Cell Proliferation through Modulation of p27^Kip1: Implications for Spatial Niche Formation and Functional Intratumoral Heterogeneity

Abstract

Background/Objective: Clear cell renal cell carcinoma (ccRCC) is characterized by a high degree of functional intratumoral heterogeneity (ITH). This is highlighted by the finding that tumor cell proliferation and intracellular signaling occur preferentially in the tumor periphery. The driving forces for such a spatial organization are largely unknown. Herein, we investigate the role of the tumor microenvironment in the control of tumor cell proliferation and functional ITH. Methods: Conditioned media (CM) derived from nonmalignant peritumoral kidney tissue were used to stimulate RCC cells in vitro. A neutralization assay was used to characterize the role of FGF-2 in the CM. The molecular mechanisms underlying the action of CM on RCC cells were investigated us-ing immunoblotting, flow cytometry and immunofluorescence microscopy. Lastly, a series of ccRCCs were stained for Ki-67 and p27 Kip1 , and expression was analyzed in both tumor periphery and center. Results: We show that CM derived from nonmalignant kidney cells adjacent to an RCC can downregulate the expression of the CDK inhibitor p27 Kip1 through enhanced protein degradation in an FGF-2-dependent fashion. FGF-2 functions mainly through the PI3K/AKT pathway downstream of its receptors, and RCC cells with constitutively high AKT activity show not only an enhanced degradation of p27 Kip1 through the Emi1-Skp2 axis, but also a subcellular mislocalization of p27 Kip1 to the cytoplasmic compartment. Such a mislocalization was also detected in the tumor periphery in vivo suggesting that p27 Kip1 plays an important role in shaping this spatial niche. Conclusions: Our results suggest that the tumor microenvironment is involved in shaping the tumor peripheral niche by stimulating the enhanced proliferation that is characteristic for this zone.

Identification of distinct tumor cell populations and key genetic mechanisms through single cell sequencing in hepatoblastoma

“…Clusters Tr0, 6, 7, 10, and 11 have high expression of BMP ligands with vast expression of BMP receptors in most clusters and support cells. FGF signaling has been reported in numerous other cancers to promote growth and tumor progression 35 , 36 . In line with these reports FGF2 is upregulated in Tr2, which has been reported to drive tumor and circulate in tumor microenvironment 36 .…”

“…FGF signaling has been reported in numerous other cancers to promote growth and tumor progression 35 , 36 . In line with these reports FGF2 is upregulated in Tr2, which has been reported to drive tumor and circulate in tumor microenvironment 36 . CXCL signaling is also of interest based on a reported role in metastasis and interaction with endothelial cells 37 .…”

Kidney Cancer Models for Pre-Clinical Drug Discovery: Challenges and Opportunities

“…One manifestation of functional ITH in ccRCC is the enhanced proliferation and intracellular signaling activity at the tumor invasion front ( 35 ). While no specific mutations could be pinpointed to explain this finding, some observations suggest that factors from the tumor microenvironment may be involved ( 36 ). The tumor periphery may hence represent a spatial niche that harbors tumor cells with an increased fitness or potentially higher aggressiveness.…”

“…While the tumor immune microenvironment conceivably plays an important role in shaping tumor progression, non-immune cells may likewise support clonal and subclonal expansion. Growth factors and cytokines secreted by non-immune cells of the tumor microenvironment with stimulatory effects on tumor growth include FGF-2, among others ( 36 , 49 , 50 ). Efforts to co-culture RCC cells with non-malignant cells of the microenvironment are therefore laudable, however, the functional state of such cells may not adequately reflect the situation in vivo , where cells of the tumor microenvironment may become re-programmed by numerous cellular interactions as well as extrinsic insults such as inflammation and necrosis ( 51 ).…”

Accumulation of CD45RO+CD8+ T cells is a diagnostic and prognostic biomarker for clear cell renal cell carcinoma

“…Immune-checkpoint blockades (ICBs) including the well-known PD-1/PD-L1 as well as CTLA-4 have produced durable response rates and survival improvement in RCC and monoclonal antibodies such as nivolumab, pembrolizumab, atezolizumab and avelumab are being tested in RCC [ 7 ]. Taken together, researchers tried to activate T cells and weaken tumor immune escape in immune environment [ 8 ]. Considering the significance of the anti-tumor response by CD8+ T lymphocytes, not merely the efficacy of T cells matters, the component especially the proportionality of mature T cells activated by MHC and co-stimulator should be focused on [ 9 ].…”