Benzethonium chloride: a novel anticancer agent identified by using a cell-based small-molecule screen

Clin Cancer Res. 2006 Sep 15;12(18):5557-69. doi: 10.1158/1078-0432.CCR-06-0536.


Purpose: This study aims to identify a novel therapeutic agent for head and neck cancer and to evaluate its antitumor efficacy.

Experimental design: A cell-based and phenotype-driven high-throughput screening of approximately 2,400 biologically active or clinically used compounds was done using a tetrazolium-based assay on FaDu (hypopharyngeal squamous cancer) and NIH 3T3 (untransformed mouse embryonic fibroblast) cells, with secondary screening done on C666-1 (nasopharyngeal cancer) and GM05757 (primary normal human fibroblast) lines. The "hit" compound was assayed for efficacy in combination with standard therapeutics on a panel of human cancer cell lines. Furthermore, its mode of action (using transmission electron microscopy and flow cytometry) and its in vivo efficacy (using xenograft models) were evaluated.

Results: Benzethonium chloride was identified as a novel cancer-specific compound. For benzethonium (48-hour incubation), the dose required to reduce cell viability by 50% was 3.8 micromol/L in FaDu, 42.2 micromol/L in NIH 3T3, 5.3 micromol/L in C666-1, and 17.0 micromol/L in GM05757. In vitro, this compound did not interfere with the effects of cisplatin, 5-fluorouracil, or gamma-irradiation. Benzethonium chloride induced apoptosis and activated caspases after 12 hours. Loss of mitochondrial membrane potential (DeltaPsiM) preceded cytosolic Ca2+ increase and cell death. In vivo, benzethonium chloride ablated the tumor-forming ability of FaDu cells, delayed the growth of xenograft tumors, and combined additively with local tumor radiation therapy. Evaluation of benzethonium chloride on the National Cancer Institute/NIH Developmental Therapeutics Program 60 human cancer cell lines revealed broad-range antitumor activity.

Conclusions: This high-throughput screening identified a novel antimicrobial compound with significant broad-spectrum anticancer activity.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antineoplastic Agents / isolation & purification*
  • Antineoplastic Agents / pharmacology
  • Antineoplastic Agents / therapeutic use
  • Apoptosis / drug effects
  • Benzethonium / isolation & purification*
  • Benzethonium / pharmacology
  • Benzethonium / therapeutic use
  • Calcium / metabolism
  • Caspases / metabolism
  • Cell Proliferation / drug effects
  • Cell Proliferation / radiation effects
  • Cell Survival / drug effects
  • Drug Screening Assays, Antitumor / methods*
  • Drug Therapy, Combination
  • Female
  • Head and Neck Neoplasms / drug therapy
  • Head and Neck Neoplasms / radiotherapy
  • Humans
  • Membrane Potentials / drug effects
  • Mice
  • Mice, Inbred BALB C
  • Mice, SCID
  • Mitochondrial Membranes / drug effects
  • Models, Biological
  • NIH 3T3 Cells
  • Neoplasms / drug therapy
  • Neoplasms / radiotherapy
  • Radiation-Sensitizing Agents / pharmacology
  • Radiation-Sensitizing Agents / therapeutic use
  • Tissue Array Analysis / methods*
  • Tumor Cells, Cultured
  • Xenograft Model Antitumor Assays / methods


  • Antineoplastic Agents
  • Radiation-Sensitizing Agents
  • Benzethonium
  • Caspases
  • Calcium