Identification of stably expressed reference small non‐coding RNA s for micro RNA quantification in high‐grade serous ovarian carcinoma tissues

Micro RNA s (mi RNA s) belong to a family of small non‐coding RNA s (snc RNA s) playing important roles in human carcinogenesis. Multiple investigations reported mi RNA s aberrantly expressed in several cancers, including high‐grade serous ovarian carcinoma ( HGS ‐OvCa). Quantitative PCR is widely used in studies investigating mi RNA expression and the identification of reliable endogenous controls is crucial for proper data normalization. In this study, we aimed to experimentally identify the most stable reference snc RNA s for normalization of mi RNA qPCR expression data in HGS ‐OvCa. Eleven putative reference snc RNA s for normalization (U6, SNORD 48, miR‐92a‐3p, let‐7a‐5p, SNORD 61, SNORD 72, SNORD 68, miR‐103a‐3p, miR‐423‐3p, miR‐191‐5p, miR‐16‐5p) were analysed on a total of 75 HGS ‐OvCa and 30 normal tissues, using a highly specific qPCR . Both the normal tissues considered to initiate HGS ‐OvCa malignant transformation, namely ovary and fallopian tube epithelia, were included in our study. Stability of candidate endogenous controls was evaluated using an equivalence test and validated by geNorm and NormFinder algorithms. Combining results from the three different statistical approaches, SNORD 48 emerged as stably and equivalently expressed between malignant and normal tissues. Among malignant samples, considering groups based on residual tumour, miR‐191‐5p was identified as the most equivalent snc RNA . On the basis of our results, we support the use of SNORD 48 as best reference snc RNA for relative quantification in mi RNA expression studies between HGS ‐OvCa and normal controls, including the first time both the normal tissues supposed to be HGS ‐OvCa progenitors. In addition, we recommend miR‐191‐5p as best reference snc RNA in mi RNA expression studies with prognostic intent on HGS ‐OvCa tissues.