No abstract

Background: Among head and neck squamous cell carcinomas (HNSCCs), hypopharyngeal squamous cell carcinoma (HPSCC) has the worst prognosis. Iron metabolism, which plays a crucial role in tumor progression, is mainly regulated by alterations to genes and post-transcriptional processes. The recent discovery of the N6-methyladenosine (m 6 A) modification has expanded the realm of previously undiscovered post-transcriptional gene regulation mechanisms in eukaryotes. Many studies have demonstrated that m 6 A methylation represents a distinct layer of epigenetic deregulation in carcinogenesis and tumor proliferation. However, the status of m 6 A modification and iron metabolism in HPSCC remains unknown. Methods: Bioinformatics analysis, sample analysis, and transcriptome sequencing were performed to evaluate the correlation between m 6 A modification and iron metabolism. Iron metabolic and cell biological analyses were conducted to evaluate the effect of the m 6 A reader YTHDF1 on HPSCC proliferation and iron metabolism. Transcriptome-wide m 6 A-seq and RIP-seq data were mapped to explore the molecular mechanism of YTHDF1 function in HPSCC. Results: YTHDF1 was found to be closely associated with ferritin levels and intratumoral iron concentrations in HPSCC patients at Sir Run Run Shaw Hospital. YTHDF1 induced-HPSCC tumorigenesis depends on iron metabolism in vivo in vitro . Mechanistically, YTHDF1 methyltransferase domain interacts with the 3'UTR and 5'UTR of TRFC mRNA, then further positively regulates translation of m 6 A-modified TFRC mRNA. Gain-of-function and loss-of-function analyses validated the finding showing that TFRC is a crucial target gene for YTHDF1-mediated increases in iron metabolism. Conclusion: YTHDF1 enhanced TFRC expression in HPSCC through an m 6 A-dependent mechanism. From a therapeutic perspective, targeting YTHDF1 and TFRC-mediated iron metabolism may be a promising strategy for HPSCC.

Background: Peripheral blood inflammation factor neutrophil-lymphocyte ratio (NLR), platelet count (PLT) and nutritional factor serum albumin (ALB) have been proposed as prognostic markers of head and neck squamous carcinoma cancer (HNSCC) in recent years. In the current study, nomogram predict models based on pre-treatment hematological parameters and a modified risk-stratified score system have been built. Methods: A total of 197 patients with oropharyngeal, hypopharyngeal and laryngeal cancers receiving multimodality treatment between 2012 and 2014 were included. The pre-treatment ALB, neutrophil, lymphocyte and platelet count (PLT) were detected. Cancer-specific survival and locoregional recurrence (LRC) by 5 years' follow-up in the cases were obtained. To integrate clinical characteristics, we propose a modified risk-stratified score system. Kaplan-Meier method, proportional hazards COX model, logistic models were used to establish nomograms within external validation. Results: Five-year LRC was decreased (p=0.004) for 140 patients with pre-treatment NLR <2.77. Five-year LRC and 5-year cancer-specific survival were decreased (p=0.031, p=0.021) with pre-treatment PLT ≥248×10 9 /L. Comparison of univariate parametric models demonstrated that pre-treatment NLR evaluation and PLT>248×10 9 /L were better among tested models. On Bayesian information criteria (BIC) analysis, the optimal prognostic model was then used to develop nomograms predicting 3-and 5-year LRC. The external validation of this predictive model was confirmed in 57 patients from another hospital. Conclusion: Pre-treatment NLR elevation and PLT>248×10 9 /L are promising predictors of prognosis in patients with operable HNSCC. Nomograms based on the pre-treatment hematological markers and modified risk-stratified score system provide distinct risk stratifications. There results provided the feasibility of anti-inflammatory and antiplatelet treatments for HNSCC patients.

Background: Having emerged as the most abundant posttranscriptional internal mRNA modification in eukaryotes, N6-methyladenosine (m 6 A) has attracted tremendous scientific interest in recent years. However, the functional importance of the m 6 A methylation machinery in ferroptosis regulation in hypopharyngeal squamous cell carcinoma (HPSCC) remains unclear. Methods:We herein performed bioinformatic analysis, cell biological analyses, transcriptome-wide m 6 A sequencing (m 6 A-seq, MeRIP-seq), RNA sequencing (RNAseq), and RNA immunoprecipitation sequencing (RIP-seq), followed by m 6 A dot blot, MeRIP-qPCR, RIP-qPCR, and dual-luciferase reporter assays. Results:The results revealed that ALKBH5-mediated m 6 A demethylation led to the posttranscriptional inhibition of NFE2L2/NRF2, which is crucial for the regulation of antioxidant molecules in cells, at two m 6 A residues in the 3′-UTR. Knocking down ALKBH5 subsequently increased the expression of NFE2L2/NRF2 and increased the resistance of HPSCC cells to ferroptosis. In addition, m 6 A-mediated NFE2L2/NRF2 stabilization was dependent on the m 6 A reader IGF2BP2. We suggest that ALKBH5 dysregulates NFE2L2/NRF2 expression in HPSCC through an m 6 A-IGF2BP2dependent mechanism. Conclusion:Together, these results have revealed an association between the ALKBH5-NFE2L2/NRF2 axis and ferroptosis, providing insight into the functional importance of reversible mRNA m 6 A methylation and its modulators in HPSCC.