Comprehensive screening of genes resistant to an anticancer drug in esophageal squamous cell carcinoma

Mai Tsutsui, Hirofumi Kawakubo, Tetsu Hayashida, Kazumasa Fukuda, Rieko Nakamura, Tsunehiro Takahashi, Norihito Wada, Yoshiro Saikawa, Tai Omori, Hiroya Takeuchi, Yuukou Kitagawa

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Drug resistance to chemotherapy is a major issue in esophageal cancer management. Drug resistance may be mediated by genetic changes in the tumor; therefore, the identification of gene mutations may lead to better therapeutic outcomes. We used a novel method involving transposons to screen and identify drug-resistant genes. Transposons are DNA sequences that move from one location on the gene to another. A modified piggyBac transposon was designed as an insertion mutagen, and a cytomegalovirus (CMV) promoter sequence was added to induce strong transcription. When the transposon is inserted to the upstream of a certain gene, the gene will be overexpressed while when intserted down or intragenically, it will be downregulated. After establishing a transposon-tagged cell library, we treated cell lines derived from esophageal squamous cell carcinomas (ESCC) [Tohoku esophagus (TE)] with cisplatin (CDDP). We performed splinkerette PCR and TOPO cloning on the resistant colonies. Bacterial colonies were sequenced, and next-generation sequencing was used to identify the overexpressed/downregulated sequences as candidate genes for CDDP resistance. We established 4 cell lines of transposon-tagged cells, TE4, 5, 9 and 15. We treated the two relatively viable cell lines, TE4 and TE15, with CDDP. We identified 37 candidate genes from 8 resistant colonies. Eight genes were overexpressed whilst 29 were downregulated. Among these genes was Janus kinase 2 (JAK2) that is implicated in the progression of myeloproliferative neoplasms. We identified 37 candidate genes responsible for CDDP resistance in the two cell lines derived from ESCC cells. The method is inexpensive, relatively simple, and capable of introducing activating and de-activating mutations in the genome, allowing for drug-resistant genes to be identified.

Original languageEnglish
Pages (from-to)867-874
Number of pages8
JournalInternational Journal of Oncology
Volume47
Issue number3
DOIs
Publication statusPublished - 2015 Sep 1

Fingerprint

Pharmaceutical Preparations
Genes
Cell Line
Down-Regulation
Drug Resistance
Esophageal Squamous Cell Carcinoma
Janus Kinase 2
Mutation
Mutagens
Esophageal Neoplasms
Cytomegalovirus
Cisplatin
Esophagus
Organism Cloning
Neoplasms
Genome
Drug Therapy
Polymerase Chain Reaction
Therapeutics

Keywords

  • Cisplatin
  • Drug resistance
  • Esophageal cancer
  • Gene
  • Transposon

ASJC Scopus subject areas

  • Cancer Research
  • Oncology

Cite this

Comprehensive screening of genes resistant to an anticancer drug in esophageal squamous cell carcinoma. / Tsutsui, Mai; Kawakubo, Hirofumi; Hayashida, Tetsu; Fukuda, Kazumasa; Nakamura, Rieko; Takahashi, Tsunehiro; Wada, Norihito; Saikawa, Yoshiro; Omori, Tai; Takeuchi, Hiroya; Kitagawa, Yuukou.

In: International Journal of Oncology, Vol. 47, No. 3, 01.09.2015, p. 867-874.

Research output: Contribution to journalArticle

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