Global microRNA expression profile in laryngeal carcinoma unveils new prognostic biomarkers and novel insights into field cancerization

Abstract

Laryngeal carcinoma is still a worldwide burden that has shown no significant improvement during the last few decades regarding definitive treatment strategies. The lack of suitable biomarkers for personalized treatment protocols and delineating field cancerization prevents further progress in clinical outcomes. In the light of this perspective, MicroRNAs could be promising biomarkers both in terms of diagnostic and prognostic value. The aim of this prospective study is to find strong prognostic microRNA biomarkers for advanced laryngeal carcinoma and molecular signatures of field cancerization. Sixty patients were enrolled and four samples were collected from each patient: tumor surface and depth, peritumor normal mucosa, and control distant laryngeal mucosa. Initially, a global microRNA profile was conducted in twelve patients from the whole cohort and subsequently, we validated a selected group of 12 microRNAs with RT-qPCR. The follow-up period was 24 months (SD ± 13 months). Microarray expression profile revealed 59 dysregulated microRNAs. The validated expression levels of miR-93-5p (χ2(2) = 4.68, log-rank p = 0.03), miR-144-3p (χ2(2) = 4.53, log-rank p = 0.03) and miR-210-3p (χ2(2) = 4.53, log-rank p = 0.03) in tumor samples exhibited strong association with recurrence-free survival as higher expression levels of these genes predict worse outcome. Tumor suppressor genes miR-144-3p (mean rank 1.58 vs 2.14 vs 2.29, p = 0.000) and miR-145-5p (mean rank 1.57 vs 2.15 vs 2.28, p = 0.000) were significantly dysregulated in peritumor mucosa with a pattern of expression consistent with paired tumor samples thus revealing a signature of field cancerization in laryngeal carcinoma. Additionally, miR-1260b, miR-21-3p, miR-31-3p and miR-31-5p were strongly associated with tumor grade. Our study reports the first global microRNA profile specifically in advanced laryngeal carcinoma that includes survival analysis and investigates the molecular signature of field cancerization. We report two strong biomarkers of field cancerization and three predictors for recurrence in advance stage laryngeal cancer.

Peritumor mucosa in advanced laryngeal carcinoma exhibits an aberrant proangiogenic signature distinctive from the expression pattern in adjacent tumor tissue

“…According to it, on a molecular level, some processes of cell dysregulation characteristic of the malignancy are evident in clonal patches of the surrounding histologically normal mucosa around the tumor, and presumably, both have a common predecessor. Our team has already reported the existence of an aberrant microRNA signature in the peritumoral tissue surrounding the advanced laryngeal carcinoma that could be used as a biomarking tool for demarcating the eld cancerization [4]. In the current study, we report for the rst time in the literature the existence of an abnormal proangiogenic expression pro le present in the peritumoral mucosa in advanced laryngeal carcinoma when compared to paired distant laryngeal mucosa.…”

“…The study was approved by the Ethics Committee of Medical University -So a. The enrolled cohort was a single-surgeon consecutive series and the inclusion criteria were as described in a previous study [4]. All patients underwent postoperative radiotherapy or combined chemoradiotherapy according to the protocol.…”

“…The Qubit™ RNA HS Assay Kit, Qubit™ 2.0 Fluorometer (Life Technologies, Thermo Fisher Scienti c Inc.) was used for precise quanti cation of 50 ng/µl and 100 ng/µl RNA dilutions used further for miRNA array and RT-qPCR analysis, respectively. The RNA integrity numbers (RIN) were measured using the 2100 Bioanalyzer (Agilent, Santa Clara, CA, USA) as described in a previous study [4].…”

Diagnostic and Prognostic Value of microRNAs in Patients with Laryngeal Cancer: A Systematic Review

“…Of those, 166 were excluded for describing HNSCC cases without focus on the larynx ( n = 28), for being exclusively in vitro studies ( n = 39), for having a number of patients less than 10 or for using patient data from public databases ( n = 50), for not quantifying miRNAs in patient samples ( n = 43) and for other reasons, such as for not having statistically significant results or wrong miRNA sequence ( n = 6). Finally, the review was performed on a total of 177 studies [ 22 , 23 , 24 , 25 , 26 , 27 , 28 , 29 , 30 , 31 , 32 , 33 , 34 , 35 , 36 , 37 , 38 , 39 , 40 , 41 , 42 , 43 , 44 , 45 , 46 , 47 , 48 , 49 , 50 , 51 , 52 , 53 , 54 , 55 , 56 , 57 , 58 , 59 , 60 , 61 , 62 , 63 , 64 , 65 , 66 , 67 , 68 , 69 , 70 , 71 …”

“…Other tissue miRNAs were reported by five or more articles, including miR-145-5p [ 32 , 42 , 59 , 92 , 99 , 130 , 131 , 153 , 167 ], miR-375-3p [ 28 , 140 , 145 , 149 , 159 , 171 , 178 , 188 , 196 ], miR-155-5p [ 26 , 28 , 59 , 84 , 106 , 128 , 169 ], miR-195-5p [ 75 , 107 , 111 , 114 , 115 , 146 , 197 ], miR-106b-3p [ 26 , 127 , 132 , 145 , 146 , 196 ], miR-21-3p [ 32 , 42 , 59 , 97 , 146 , 168 ], miR-125b-5p [ 32 , 43 , 82 , 101 , 117 , 130 ], miR-204-5p [ 32 , 49 , 65 , 82 ,…”