| Cancer of the kidney accounts for approximately 3.5% of all human malignancies and it is the third most common cancer of the urinary tract. The incidence of renal cell carcinoma (RCC) has been steadily rising over the past 30 years. Furthermore, because metastatic RCC is notoriously resistant to most current therapies, such as chemotherapy and radiotherapy, the prognosis of patients with RCC is poor as one-third of patients already have metastatic disease at the initial diagnosis and 30-40% of them develop distant metastases after resection of the primary tumor.Evidence is accumulating that tumors contain a subpopulation of cancer stem cells (CSCs). Like normal stem cells, CSCs have the ability of self-renewing and are capable of initiating and maintaining the growth of the tumor. If CSCs were not completely eradicated by conventional antineoplastic treatments, they might be the origin of local recurrence and distant metastases. However, the isolation of CSCs from human RCC has not been well studied up till now.From the early 1990s, in the pre-tyrosine-kinase inhibitor (TKI) era, interferon-a and interleukin-2 immunotherapy has been the leading treatment for metastatic RCC. However, only 5% to 20% of cases of metastatic clear cell renal cell carcinoma (ccRCC) respond to interferon-a and interleukin-2 treatment, which is thought to result from the pervasive presence of tumor-driven immune suppression that allows tumor to escape and that has not been adequately targeted by current therapies. However, it is unknown whether CSCs contribute to the profound immune suppression in RCC patients.In this study, we sought to enrich CSCs from the RCC cell line SK-RC-42 using the sphere culture system, characterized their immunophenotype and explored immunosuppressive strategies that are mediated by RCC CSCs. We demonstrated that a subpopulation of SK-RC-42 cells were capable of growing as tumor spheres in serum-free medium supplemented with EGF and bFGF. The sphere-forming cells (SFCs) have many properties similar to CSCs:ability of self-renewing in vitro and in vivo, higher mRNA expression levels of several'sternness'genes, stronger tumorigenicity and resistance to chemotherapeutic agents and irradiation compared with the monolayer adherent cells (MACs).With regard to their immunophenotype, the SFCs express high levels of MHC classâ… but low levels of MHC classâ…¡, CD80 and CD86. In contrast with MACs, the SFCs have lower expression levels of FasL and Fas, Her2 and hTERT and activating natural killer receptors. Additionally, SK-RC-42 SFCs and MACs both express significant and comparable levels of the transcription factor forkhead box protein 3 (FoxP3) and membrane complement regulatory proteins (mCRPs). Furthermore, we find that the SFCs markedly trigger T-cell apoptosis, induce regulatory T cells and myeloid-derived suppressor cell (MDSCs).Taken together, these findings indicate that CSCs can be enriched from RCC by culturing the tumor cells as spheres. The immune phenotype of the SFCs demonstrated in this study suggests that CSCs might play an important role in the evasion of tumor growth from immune surveillance. Our findings of further functional experiment provide new insights into our understanding of immunosuppressive mechanisms in the tumor microenvironment and novel therapeutic approaches against CSCs in RCC.
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