Liquid lncRNA Biopsy for the Evaluation of Locally Advanced and Metastatic Squamous Cell Carcinomas of the Head and Neck

Abstract

Background: Long non-coding RNA (lncRNA) are RNA molecules that are more than 200 nucleotides long and have the ability to modify the activity of genes. They can be found in both healthy and cancer tissues, as well as in plasma, saliva and other bodily fluids. They can also be used as biomarkers of early detection, prognosis and chemotherapy resistance in several cancer types. Treatment of head and neck squamous cell carcinoma (HNSCC) patients with locally advanced disease is still difficult, and choice of treatment should be based on more precise and available biomarkers, such as those obtained from a liquid biopsy. For improvement of treatment efficacy, identification and clinical implementation of new biomarkers are of the utmost importance. Methods: Plasma samples drawn before (p1) and three cycles post (p2) (TPF: docetaxel, cisplatin, 5-fluorouracil/PF: cisplatin, 5-fluorouracil) chemotherapy from 53 HNSCC patients (17 with locally advanced and 36 with metastatic disease) and 14 healthy volunteers were studied. Expression levels of 90 lncRNA expression were analyzed using the qRT-PCR method, and the obtained results were compared between proper groups. Statistical analyses were carried out using Jupyter Notebooks (5.7.2), Python (ver. 3.6) and GraphPad Prism 8. Results: The study demonstrated the differences between the expressions of several lncRNA in cancer patients’ and healthy volunteers’ plasma, as well as between locally advanced and metastatic patients’ groups. A correlation between the response to systemic therapy and lncRNA expression levels was observed. Patients with a (high/low) expression of Alpha 250 and Emx2os showed statistically significant differences in progression free survival (PFS), as well as for overall survival (OS) depending on the level of Alpha 250, snaR, SNHG1. The univariate and multivariate Cox regression model showed Alpha 250 as the best prognostic factor for HNSCC patients. Conclusions: Liquid biopsies based on lncRNAs are promising diagnostic tools that can be used to differentiate between those with cancer and healthy individuals. Additionally, they can also serve as biomarkers for chemotherapy resistance. An identified, circulating lncRNA Alpha 250 seems to prove the best prognostic biomarker, associated with extended PFS and OS, and should be validated in a larger cohort in the future.

Long Intergenic Non-Coding RNAs in HNSCC: From “Junk DNA” to Important Prognostic Factor

“…The lincRNA regulator of reprogramming (lincRNA-RoR) is an oncogene involved in EMT along with drug resistance in different malignancies, e.g., NPC, BC, and HNSCC [142][143][144]. Interestingly, we have observed that its expression level increases after exposure to cisplatin [52]. One of the most poorly studied lincRNA is maternally expressed 9 (MEG9), speculated to be induced by hypoxia in a mouse model by Voellenkle et al [145].…”

“…Its up-regulation is associated with intensified tumor proliferation and migration along with a poor prognosis for ovarian cancer patients [127]. Moreover, we have also proved that high Emx2os expression levels can negatively affect progression-free survival (PFS) of individuals with HNSCC [52]. Growth arrest-specific 5 (GAS5) lincRNA was identified as a poorly conserved tumor suppressor that also acts as a decoy for the glucocorticoid receptor (GR) [47,128,129].…”

“…Liquid biopsy may be a good response to the aforementioned need for a non-invasive, accurate method of detecting the disease, assessing its stage, and monitoring the course of its treatment. For this type of analysis, non-solid biological materials, such as blood and its fractions, are obtained [49][50][51][52]. They contain many biomarkers that allow the assessment of changes in the cancer phenotype (modifications of the genome, epigenome, and transcriptome), immunophenotype, response to therapy, and the incidence of infection [49][50][51][52][53][54][55][56].…”

“…For this type of analysis, non-solid biological materials, such as blood and its fractions, are obtained [49][50][51][52]. They contain many biomarkers that allow the assessment of changes in the cancer phenotype (modifications of the genome, epigenome, and transcriptome), immunophenotype, response to therapy, and the incidence of infection [49][50][51][52][53][54][55][56]. Such characterization cannot always be performed by traditional biopsy, because it often does not fully reflect tumor heterogeneity [57,58] or cannot be obtained at all, as in the case when neoadjuvant therapy reduces the volume of the neoplastic lesion to an undetectable size [50].…”

“…Previously, our team performed a liquid biopsy on HNSCC patients and healthy individuals to analyze lncRNA plasma expression differences between these two groups and to determine their diagnostic potential. In agroup of 90 lncRNA transcripts, 20 lincR-NAs, including ANRIL, Dios3os, Emx2os, GAS5, H19, HAR1B, HULC, Jpx, lincRNA-RoR, MALAT1, MEG9, ncR-uPAR, NEAT1, NRON, RNCR3, SNHG1, SNHG6, Tsix, UCA1, and Zfas1, had significantly higher expression levels in metastatic and/or recurrent patients' samples in comparison to healthy individuals [52]. We checked whether lncRNA is lincRNA in the LNCipedia database (version 5.2) [119].…”