SMAD4 Expression in Renal Cell Carcinomas Correlates With a Stem-Cell Phenotype and Poor Clinical Outcomes.

Arezoo Rasti,Sadegh Babashah,Maryam Abolhasani,Zahra Madjd,Mitra Mehrazma,Leili Saeednejad Zanjani,Mojgan Asgari

doi:10.3389/fonc.2021.581172

Abstract

Renal cell carcinoma (RCC) is the most lethal neoplasm of common urologic cancers with poor prognoses. SMAD4 has a principal role in TGF-β (Transformis growth factorβ)-induced epithelial to mesenchymal transition (EMT) as a key factor in gaining cancer stem cell (CSC) features and tumor aggressiveness. This study aimed to evaluate the expression patterns and clinical significance of SMAD4 in RCC and the impact of its targeting on stem cell/mesenchymal cells and EMT characteristics in renal spheroid derived cells (SDCs) compared to parental cells (PCs) in RCC. The expression pattern and clinical significance of SMAD4 was evaluated in RCC. SDCs were enriched using a sphere culture system. Then SDCs and their PCs were compared with respect to their sphere and colony formation, expression of putative CSC markers, invasiveness as well as expression of genes, including stemness/mesenchymal, SMAD4 and TGFβ1genes. Finally, the effect of SMAD4 knockdown on SDCs was analyzed. We demonstrated that SMAD4 is positively correlated with decreased disease specific survival (DSS) in RCC patients and clear cell RCC (ccRCC) subtype and associates with poor DSS in patients with RCC, especially in ccRCC as the most metastatic RCC subtype. SDCs exhibited higher stem cell/mesenchymal properties. Inhibition of SMAD4 in PCs accelerated the dissociation of SDCs and decreased their clonogenicity, invasiveness, expression of mesenchymal markers and expression of SMAD4 and TGFβ1 genes compared to SDCs before transfection. We suggest that targeting SMAD4 may be useful against renal CSCs and may improve RCC prognosis.

Highlights

Renal cell carcinoma (RCC) is the most popular epithelial malignancy of human adult kidneys and has the highest mortality among all urologic cancers (1)
Despite the advances in traditional therapies that aim to eliminate the bulk of the tumor, poor cure rates may result from the ability of cancer to repopulate and spread after initial therapies due to the presence of cancer stem cell (CSC) (26)
The resistance of renal CSCs to chemotherapy and radiotherapy prepared the rationale for novel therapeutic strategies targeting this invasive cell population, inducting cell differentiation, and blocking CSCs maintenance pathways (27)

Summary

Introduction

Renal cell carcinoma (RCC) is the most popular epithelial malignancy of human adult kidneys and has the highest mortality among all urologic cancers (1). Based on the cancer stem cell (CSC) model, a small subset of cancer cells have stem cell properties, such as expression of stem cells markers, clonogenic ability, lack of differentiative epithelial markers and sphere formation in the absence of adhesive cell culture medium (4). They are responsible for disease progression, tumor development, metastasis, recurrence and resistance to anticancer therapy (5). It has been proved that epithelial to mesenchymal transition (EMT) has a key role in getting CSC features in self-renewing SDCs from RCC cell lines (7). Tumor cells often miss the ability to respond to TGF-b–mediated growth inhibition and instead apply TGF-b signaling to raise EMT invasion and metastasis (9)

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