SLC20A1 is a prospective prognostic and therapy response predictive biomarker in head and neck squamous cell carcinoma

Abstract

Background: SLC20A1, a prominent biomarker in several cancers, has been understudied in its predictive role in head and neck squamous cell carcinoma (HNSCC).

Methods: The Cancer Genome Atlas (TCGA) database was used to analyze HNSCC prognosis, SLC20A1 overexpression, and clinical characteristics. Quantitative real-time PCR and Western blot analysis confirmed SLC20A1 expression in HNSCC tissues. Cellular behaviors such as invasion, migration and proliferation were assessed using Transwell, wound healing and colony formation assays. Immune system data were obtained from the Tumor Immune Estimation Resource (TIMER) and CIBERSORT databases. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and Gene Set Enrichment Analysis (GSEA) were used to explore biological parameters and pathways associated with SLC20A1 overexpression in HNSCC.

Results: In 499 HNSCC samples, SLC20A1 mRNA and protein expression were significantly higher than in 44 normal counterparts, confirmed by 24 paired samples. Patients were categorized based on SLC20A1 levels, survival status and overall survival. High SLC20A1 expression correlated with advanced T stage, increased risk scores and decreased survival. Stage, age and SLC20A1 expression emerged as independent predictive factors for HNSCC in univariate and multivariate analyses. SLC20A1 overexpression, which is associated with poor prognosis, may influence cell proliferation, migration, invasion, chemotherapy response, and the immune milieu.

Conclusions: SLC20A1 overexpression in HNSCC, characterized by increased cellular invasion, migration and proliferation, is a potential prognostic biomarker and therapeutic response indicator.

Keywords: SLC20A1; TCGA; cancer; prognosis; squamous cell carcinoma.

Figure 1

Exaggerated SLC20A1 expression levels in HNSCC. (A, B) SLC20A1 mRNA expression between the HNSCC specimens and nearby sound tissues from TCGA database and qRT-PCR analysis. (C) SLC20A1 protein expression in the HNSCC specimens and nearby sound ones from Western blotting analysis. (D) DNA methylation modification associated with SLC20A1 expression in HNSCC. *, P<0.05.

Figure 2

HNSCC prognosis via SLC20A1. (A) Kaplan-Meier analysis of poor survival outcomes with SLC20A1 overexpression in HNSCC from TCGA database. (B) Risk score with survival time in TCGA database. (C, D) Forest plots representing the uni- and multivariate analysis of the significantly marked prognostics.

Figure 3

Silencing SLC20A1 inhibited cell proliferation in TU686 and CAL-27 cells lines. (A) CCK8 and (B) colony formation experiments showed that silencing SLC20A1 inhibited cell proliferation. *, P<0.05. **, P<0.01. ***, P<0.001.

Figure 4

Silencing SLC20A1 inhibited migration and invasion in HNSCC. (A) Wound-healing experiments showed that silencing SLC20A1 inhibited cell migration. (B) Silencing SLC20A1 inhibited the ability to invade CAL-27 and TU686 cell lines. *, P<0.05. **, P<0.01. ***, P<0.001.

Figure 5

A nomogram based on SLC20A1 expression and clinical features. (A) Nomogram to predict the overall survival (OS) of the 1^st, 3^rd, and 5^th years for HNSCC cases. (B) Calibration curve of the nomogram prognostic model. (C) ROC curve of the nomogram for predicting the overall survival (OS) of the 1^st, 3^rd, and 5^th years.

Figure 6

Chemotherapy and immunotherapy response were associated with SLC20A1 expression in HNSCC. (A) Chemotherapeutic responses with differential SLC20A1 expression in HNSCC. (B) Correlation between SLC20A1 expression and immunotherapy responses. *, P<0.05. **, P<0.01. ***, P<0.001. IC50, 50% inhibiting concentration.

Figure 7

Association between SLC20A1 expression and TIICs in HNSCC. (A) Abundance of TIICs infiltration between low and high SLC20A1 expression in HNSCC from CIBERSORT database. (B) Correlation between SLC20A1 expression and TIICs from TIMER database.