Spheroid-Derived Cells From Renal Adenocarcinoma Have Low Telomerase Activity and High Stem-Like and Invasive Characteristics.

Leili Saeednejad Zanjani,Raheleh Roudi,Øystein Fodstad,Yvonne Andersson,Arezoo Rasti,Gunhild Mari Mælandsmo,Mojgan Asgari,Maryam Abolhasani,Zahra Madjd,Kevin J Tam

doi:10.3389/fonc.2019.01302

Abstract

Cancer stem cells (CSCs) are a theorized small subpopulation of cells within tumors thought to be responsible for metastasis, tumor development, disease progression, treatment-resistance, and recurrence. The identification, isolation, and biological characterization of CSCs may therefore facilitate the development of efficient therapeutic strategies targeting CSCs. This study aims to compare the biology and telomerase activity of CSCs to parental cells (PCs) in renal cancer. Renal CSCs were enriched from the ACHN cell line using a sphere culture system. Spheroid-derived cells (SDCs) and their adherent counterparts were compared with respect to their colony and sphere formation, expression of putative CSC markers, tumorigenicity in non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice, and invasiveness. The expression of genes associated with CSCs, stemness, EMT, apoptosis, and ABC transporters was also compared between the two populations using quantitative real-time PCR (qRT-PCR). Finally, telomerase activity, hTERT expression, and sensitivity to MST-312, a telomerase inhibitor, was investigated between the two populations. We demonstrated that a subpopulation of ACHN cells was capable of growing as spheroids with many properties similar to CSCs, including higher clonogenicity, superior colony- and sphere-forming ability, and stronger tumorigenicity and invasiveness. In addition, SDCs demonstrated a higher expression of markers for CSCs, stemness, EMT, apoptosis, and ABC transporter genes compared to PCs. The expression of hTERT and telomerase activity in SDCs was significantly lower than PCs; however, the SDC population was more sensitive to MST-312 compared to PCs. These findings indicate that the SDC population exhibits stem-like potential and invasive characteristics. Moreover, the reduced expression of hTERT and telomerase activity in SDCs demonstrated that the expressions of hTERT and telomerase activity are not always higher in CSCs. Our results also showed that MST-312 treatment inhibited SDCs more strongly than PCs and may therefore be useful as a complementary targeted therapy against renal CSCs in the future.

Highlights

Renal cell carcinoma (RCC) is the most lethal neoplasm of all urologic cancers and is responsible for ∼90% of renal malignancies and 2–3% of total cancers in adults [1, 2]
The second generation of spheroid-derived cells (SDCs) from the ACHN cell line were used in the following experiment
The unique role that telomerase fills in conferring normal stem cells with immortality is well-established and brings about the possibility that it has a similar role in cancer stem cells (CSCs) [16]

Summary

Introduction

Renal cell carcinoma (RCC) is the most lethal neoplasm of all urologic cancers and is responsible for ∼90% of renal malignancies and 2–3% of total cancers in adults [1, 2]. The incidence of RCC has been increasing worldwide over the past 20 years [1]. It is estimated that there will be 65,340 new cases and 14,970 deaths from RCC in the United States in 2018. The 5 year relative survival rate of RCC in the United States was found to be 93% in localized disease, 67% in regional disease, and 12% in patients with metastases [3]. Despite improvements in diagnostic techniques, ∼25–30% of RCC patients present with metastases at the time of diagnosis.

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