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Establishment and Characterization of Pemetrexed-resistant NCI-H460/PMT Cells

Yu-Lian Xu1, Xiao-Ming Jiang1, Le-Le Zhang1

  • 1State Key Laboratory of Quality Research in Chinese Medicine, Institute of Chinese Medical Sciences, University of Macau, Macau, China.

Anti-Cancer Agents in Medicinal Chemistry
|March 9, 2019
PubMed
Abstract

Insights

Overexpression of Thymidylate Synthase (TYMS) drives pemetrexed (PMT) resistance in Non-Small Cell Lung Cancer (NSCLC) cells. Targeting TYMS may overcome this resistance, while cross-resistance to other drugs necessitates careful consideration.

Area of Science:

  • Oncology
  • Molecular Biology
  • Cancer Research

Background:

  • Pemetrexed (PMT) is a crucial antifolate chemotherapy for Non-Small Cell Lung Cancer (NSCLC).
  • Acquired resistance to PMT poses a significant clinical challenge in NSCLC treatment.
  • Understanding the mechanisms of PMT resistance is vital for developing effective therapeutic strategies.

Purpose of the Study:

  • To investigate the molecular mechanisms underlying pemetrexed (PMT) resistance in Non-Small Cell Lung Cancer (NSCLC) cells.
  • To identify key molecular targets or pathways involved in conferring PMT resistance.
  • To evaluate the role of specific genes, such as ABCB1 and TYMS, in PMT resistance.

Main Methods:

  • Establishment of PMT-resistant NCI-H460/PMT cell lines through concentration escalation.
  • Assessment of cell proliferation, migration, and senescence using MTT assays, colony formation, immunofluorescence (Ki-67), and Transwell assays.
  • Quantitative PCR (qPCR) and Western blot analysis to determine mRNA and protein expression levels of target genes.
  • Gene knockdown experiments using small interfering RNA (siRNA) targeting ATP binding cassette subfamily B member 1 (ABCB1) and Thymidylate Synthase (TYMS).

Main Results:

  • PMT-resistant cells exhibited altered proliferation and enhanced migration, alongside cellular senescence.
  • Cross-resistance to multiple chemotherapeutics was observed, potentially linked to upregulated ABCB1 expression.
  • ABCB1 knockdown did not fully restore sensitivity to PMT.
  • Significant upregulation of Thymidylate Synthase (TYMS), a direct target of PMT, was identified in resistant cells.
  • Genetic silencing of TYMS partially reversed PMT resistance, indicating its critical role.

Conclusions:

  • Overexpression of Thymidylate Synthase (TYMS) is a key mechanism driving pemetrexed (PMT) resistance in KRAS-mutated NCI-H460 NSCLC cells.
  • The observed cross-resistance to other chemotherapeutics warrants consideration in clinical management of PMT-resistant NSCLC.
  • Targeting TYMS presents a potential therapeutic strategy to overcome PMT resistance in NSCLC.
Keywords:
ABCB1NSCLCTYMSchemotherapypemetrexedresistance.

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