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An LC-MS/MS-Based Quantification of Intracellular Anlotinib Reveals Exposure-Toxicity Relationships in Endothelial

Zhenhui Jiang1,2, Liya Ye1,2, Jianguo Sun3

  • 1College of Traditional Chinese Medicine, Yunnan University of Chinese Medicine, Kunming, Yunnan, China.

Biomedical Chromatography : BMC
|January 28, 2026
PubMed

Insights

This study developed a method to measure anlotinib inside cells. Intracellular anlotinib levels, not outside levels, correlated with cell toxicity, aiding research into drug side effects.

Area of Science:

  • Pharmacology
  • Analytical Chemistry
  • Cell Biology

Background:

  • Anlotinib treats non-small cell lung cancer but causes severe side effects like hypertension and diarrhea.
  • The link between intracellular anlotinib exposure and its toxicity is not well understood.
  • Understanding drug exposure within cells is crucial for managing therapeutic efficacy and adverse reactions.

Purpose of the Study:

  • To develop and validate a sensitive UHPLC-MS/MS method for quantifying anlotinib in human intestinal epithelial cells (HIECs) and human umbilical vein endothelial cells (HUVECs).
  • To investigate the relationship between intracellular and extracellular anlotinib concentrations and cytotoxicity in HIECs and HUVECs.
  • To establish a foundation for understanding anlotinib-induced adverse reactions by linking intracellular exposure to toxicity.

Main Methods:

  • Developed a rapid and sensitive UHPLC-MS/MS method using an Agilent 1290-6460A system.
  • Utilized pazopanib-d3 as an internal standard for accurate quantification of anlotinib.
  • Validated the method for specificity, linearity, accuracy, precision, stability, and dilution integrity in HIECs and HUVECs.

Main Results:

  • The UHPLC-MS/MS method accurately quantified anlotinib in the range of 10-4000 ng/mL.
  • The method was successfully applied to determine intracellular and extracellular anlotinib concentrations in HIECs and HUVECs.
  • Intracellular anlotinib concentrations, not extracellular levels, were significantly correlated with cytotoxicity in both cell types.

Conclusions:

  • A robust analytical method for intracellular anlotinib quantification was established.
  • Experimental evidence supports a positive correlation between intracellular anlotinib exposure and cytotoxicity.
  • This study provides a basis for further research into anlotinib's adverse reactions and personalized treatment strategies.
Keywords:
UHPLC–MS/MSadverse reactionanlotinibcell modelexposure concentration

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