THUMP domain containing 2 protein possibly induces resistance to cisplatin and 5-fluorouracil in in vitro human esophageal squamous cell carcinoma cells as revealed by transposon activation mutagenesis

Background: Although chemotherapy is a core treatment for esophageal cancer, some patients develop drug resistance. Gene screening with transposons (i.e. mobile genetic elements) is a novel procedure for identifying chemotherapy-resistant genes. Transposon insertion can randomly affect nearby gene expression. By identifying the affected genes, candidate genes can be found. The present study aimed to identify cisplatin (CDDP)/5-fluorouracil (5-FU)-resistant genes in in vitro human esophageal squamous cell carcinoma with transposons. Methods: After establishing transposon-tagged cells, we obtained CDDP/5-FU-resistant colonies. A polymerase chain reaction and sequencing were used to identify the transposon inserted site and candidate CDDP/5-FU resistant genes. Focusing on one candidate gene, we confirmed CDDP/5-FU resistance by comparing the IC₅₀ between drug-resistant and wild-type cells. Furthermore, we investigated gene expression by a real-time polymerase chain reaction. Finally, we mediated the candidate gene level with small interfering RNA to confirm the resistance. Results: Thirty-nine candidate genes for CDDP/5-FU resistance were identified. Nineteen were for CDDP resistance and 27 were for 5-FU resistance. Seven genes, THUMP domain-containing protein 2 (THUMPD2), nuclear factor interleukin-3-regulated protein (NFIL3), tyrosine-protein kinase transmembrane receptor 2 (ROR2), C-X-C chemokine receptor type 4 (CXCR4), thrombospondin type-1 domain-containing protein 2 (THSD7B) alpha-parvin (PARVA) and TEA domain transcription factor 1 (TEAD1), were detected as candidate genes in both colonies. Regarding THUMPD2, its expression was downregulated and knocking down THUMPD2 suggested drug resistance in both drugs. Conclusions: Thirty-nine candidate genes were identified with transposons. The downregulation of THUMPD2 was suggested to play a role in multidrug resistance in in vitro esophageal squamous cell carcinoma.