title: Synergistic cytotoxicity of artemisinin and sodium butyrate on human cancer cells.
abstract: BACKGROUND
  Butyric acid is a short chain fatty acid produced by large bowel bacterial flora. It serves as an antiinflammatory agent and nutrient for normal colon cells. Butyric acid has also been shown to induce apoptosis in colon and many other cancer cells. Artemisinin is a compound extracted from the wormwood Artemisia annua L. It has been shown to selectively kill cancer cells in vitro and to be effective in treating animal and human cancer. We and others have found that the artemisinin analog, dihydroartemisinin (DHA), kills cancer cells by apoptosis. In the present study, the efficacy of a combined treatment of DHA and butyric acid at low doses in killing cancer cells was investigated.
  
  
  METHODS
  Molt-4 cells (a human lymphoblastoid leukemia cell line) and freshly isolated human lymphocytes, cultured in complete RPMI-1640 medium, were first incubated with 12 microM of human holotransferrin at 37 degrees C in a humid atmosphere of 5% CO2 for one hour to enhance the iron concentration in the cells. Cells from each cell type were then divided into 20 flasks. These flasks were grouped into four sets of five cultures each. Zero, 5, 10 or 20 microM of DHA was added, respectively, to these sets and the cells were incubated at 37 degrees C for one hour. Zero, 1, 5, 10, or 20 mM of sodium butyrate was then added to the five cultures of each set, respectively. Thus, the treatments involved a combination of 4 doses of DHA and 5 doses of sodium butyrate. The cells were counted immediately before the addition of DHA, and at 24 and 48 hours after the addition of sodium butyrate.
  
  
  RESULTS
  DHA alone at the 24-hour time-point and 20 microM concentration significantly reduced the number of Molt-4 cells in the culture by approximately 40% (p < 0.001, compared to non-treated control), whereas it did not significantly affect the number of normal human lymphocytes. Similarly, 1 mM sodium butyrate alone at 24 hours reduced the number of Molt-4 cells by approximately 32% (p < 0.001, compared to non-treated control), without significantly affecting normal human lymphocytes. The combination of 20 microM DHA and 1 mM sodium butyrate killed all Molt-4 cells at the 24-hour time-point and did not significantly affect lymphocytes.
  
  
  CONCLUSIONS
  DHA in combination with butyric acid acts synergistically at low doses. The combination may provide a less toxic, inexpensive and effective cancer chemotherapy.
authors.affiliation: Department of Bioengineering, Box 357962, University of Washington, Seattle, WA 98195-7962, USA. Narendra@u.washington.edu
authors.forename: Narendra P
authors.initials: NP
authors.lastname: Singh
authors.forename: Henry C
authors.initials: HC
authors.lastname: Lai
mesh_terms: D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology*; D037621:Artemisinins / Q000008:administration & dosage / Q000494:pharmacology*; D002087:Butyrates / Q000008:administration & dosage / Q000494:pharmacology*; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D006801:Humans; D008214:Lymphocytes / Q000187:drug effects; D012717:Sesquiterpenes / Q000008:administration & dosage / Q000494:pharmacology*
chemical_list: D037621:Artemisinins; D002087:Butyrates; D012717:Sesquiterpenes; C039060:artenimol