N6-methyladenosine-associated prognostic pseudogenes contribute to predicting immunotherapy benefits and therapeutic agents in head and neck squamous cell carcinoma

Rationale: N6-methyladenosine (m(6)A) is involved in critical cancerous processes. Pseudogenes play various roles in carcinogenesis and progression. However, the functional roles of m(6)A-associated pseudogenes in head and neck squamous cell carcinoma (HNSCC) are largely unknown. Methods: We systematically analyzed the mRNA profile of 24 m(6)A regulators and 13931 pseudogenes from The Cancer Genome Atlas HNSCC dataset and ultimately identified 10 m(6)A-associated prognostic pseudogenes, which were validated in the Gene Expression Omnibus and our hospital datasets. Based on the risk score of m(6)A-associated pseudogenes, comprehensive analytical frameworks and experimental validation were implemented among pseudogene-defined low-/high-risk subtypes. Results: Here, we found expression pattern of m(6)A-associated pseudogenes was significantly associated with infiltrating immune cell compositions, and the expression of antitumor immune response markers, including T cell exhaustion, antigen presentation, interferon, and kinase genes. The m(6)A-associated pseudogenes, which had dramatic m(6)A peaks and higher m(6)A levels, could regulate the expression of targeted immune-involved genes through miRNAs. We experimentally validate the oncogene PDIA3P1, and tumor-suppressor RRN3P3, which promote the RNA and protein expression of their targeted immune-involved genes AKT1 and EZH2 via miR-34a-5p and miR-26b-5p, respectively. Moreover, HNSCC patients in the high-risk subtype could benefit more from immune checkpoint inhibitors therapy. Furthermore, doxorubicin and topotecan were considered to hold the most promising therapeutic potential robustly in silico evidence and in vitro experiments for HNSCC patients in the high-risk subtype. Conclusions: Our discovery revealed that the 10 m(6)A-associated prognostic pseudogenes significantly contribute to predicting immunotherapy benefits and therapeutic agents, which might bring some potential implications for both immunotherapy and chemotherapy in HNSCC.