Differential microRNA expression profiles between malignant rhabdoid tumor and epithelioid sarcoma: miR193a-5p is suggested to downregulate SMARCB1 mRNA expression

Abstract

Malignant rhabdoid tumor and epithelioid sarcoma are classified as tumors of uncertain differentiation. However, it is controversial whether these tumors are distinct entities because they share similar histological and immunohistochemical features such as the existence of rhabdoid cells or complete loss of SMARCB1 protein expression. MicroRNAs are small non-coding RNAs, and it is suggested that knowledge of microRNA expression profiles in cancer may have substantial value for diagnostics. We first analyzed microRNA expression profiles in 13 frozen materials (five malignant rhabdoid tumors, two proximal type epithelioid sarcomas, and six conventional type epithelioid sarcomas) and subsequently examined the specific microRNA expressions in 29 paraffin-embedded materials (8 malignant rhabdoid tumors, 13 proximal type epithelioid sarcomas, and 8 conventional type epithelioid sarcomas) and 13 previously described frozen materials by quantitative RT-PCR. According to the unsupervised hierarchical clustering of microRNA, proximal type epithelioid sarcoma and conventional type epithelioid sarcoma were classified into the same category, whereas malignant rhabdoid tumor was a distinct category from both types of epithelioid sarcoma. In addition, when malignant rhabdoid tumor with SMARCB1 gene alterations and proximal type and conventional type epithelioid sarcoma with no SMARCB1 gene alterations were compared, 56 microRNAs were isolated as being significantly different (ANOVA, Po0.05). Among them, quantitative RT-PCR using frozen and paraffin-embedded materials demonstrated that expression levels of miR193a-5p (P ¼ 0.002), which has been suggested to downregulate SMARCB1 mRNA expression, showed statistically different expression levels between malignant rhabdoid tumor and epithelioid sarcoma with no SMARCB1 gene alterations. These results suggest that epithelioid sarcoma, especially proximal type epithelioid sarcoma, and malignant rhabdoid tumor are distinct tumors with respect to the microRNA expression profiles and that miR193a-5p may have an important role in the inhibition of SMARCB1 mRNA expression.

Oncogenic roles of SMARCB1/INI1 and its deficient tumors

“…The ratio of gene alteration at either the DNA or RNA level causing SMARCB1/INI1 protein inactivation varies widely between 0% and 58% in conventional‐type or between 19% and 100% in proximal‐type cases . In addition, it is suggested that microRNAs such as miR193a‐5p, miR‐206, miR‐381 and miR‐671‐5p may have the potential to inhibit SMARCB1 mRNA in epithelioid sarcoma …”

Reclassification of rhabdoid tumor and pediatric undifferentiated/unclassified sarcoma with complete loss of SMARCB1/INI1 protein expression: three subtypes of rhabdoid tumor according to their histological features

“…MicroRNAs exert multiple biological functions by negatively regulating the expression of their target genes involved in development, differentiation, apoptosis, and cell proliferation, and it is suggested that knowledge of microRNA expression profiles in cancer may have substantial value for diagnostic and prognostic determinations. 14,[24][25][26] In our current study, the microRNA profiles in the malignant Figure 4 The cytoplasms of tumor cells in the conventional-type category were positive for vimentin (a, 2-year-old male, kidney), CAM5.2 (c, 4-month-old male, liver) and CD34 (d, 5-month-old female, kidney). Focal vimentin immunoreactivity is evident in the nuclear periphery of small cell-type tumor cells (b, 4-year-old male, retroperitoneum).…”

“…13,14 Extracted total RNA was labeled with Hy5 using a miRCURY LNA Array miR Labeling Kit (Exiqon, Vedbaek, Denmark). Labeled RNAs were hybridized onto 3D-Gene Human miRNA Oligo chips containing 837 antisense probes printed in duplicate spots (Toray, Kamakura, Japan).…”

Epigenetic regulation of SMARCB1 By miR‐206, ‐381 and ‐671‐5p is evident in a variety of SMARCB1 immunonegative soft tissue sarcomas, while miR‐765 appears specific for epithelioid sarcoma. A miRNA study of 223 soft tissue sarcomas

“…Indeed, miRNAs have been shown to be relevant to the biology of many soft tissue sarcomas (Subramanian and Kartha ), including rhabdomyosarcoma (Armeanu‐Ebinger et al, ; Li et al, ; Missiaglia et al, ), liposarcoma (Borjigin et al, ), osteosarcoma (Chen et al, ), angiosarcoma (Italiano et al, ), Ewing sarcoma (Dylla et al, ), and SS (Hisaoka et al, ). However, only two studies (Kohashi et al, ; Papp et al, ) examined miRNAs in ES so far. In our previous work (Papp et al, ), we used in silico target prediction analysis (targeting SMARCB1 mRNA) which revealed eight candidate miRNAs, and quantitative PCR demonstrated significant ( P < 0.001) overexpression of four miRNAs in ESs: miR‐206, miR‐381, miR‐671‐5p, and miR‐765.…”

“…Using functional testing, we found that miR‐206, miR‐381, and miR‐671‐5p (functionally active miRNAs) could silence the SMARCB1 mRNA expression in cell cultures, but not miR‐765. Kohashi (Kohashi et al, ) using differential miRNA expression profiling, found that miR193a‐5p might downregulate SMARCB1 mRNA expression in ES, but they applied no functional test to prove it. In our present work, we examined whether overexpression of functionally active miRNAs in silencing SMARCB1 mRNA is characteristic or not for the other types of SMARCB1 immunonegative soft tissue sarcomas and whether the overexpression of miR‐765 is specific for ES or not.…”