Determination of the toxicity of Bacillus thuringiensis δ-endotoxin against cancer cells
Bacillus thuringiensis (Bt) is a Gram-positive, endospore forming bacterium that produces crystal proteins containing various delta-endotoxins during sporulation. This organism is known for its insect pathogenicity and thus is of agricultural and public health interest, which has led to various studies to classify this organism. In the past decade, delta-endotoxins have been found to contain properties that are non-hemolytic but capable of preferentially killing cancer cells, thus named "parasporins." Previous findings show that six of these toxins have the ability to kill cancer cells: Parasporin-1 (HeLa), Parasporin-2 (HepG2 and MOLT-4), Parasporin-3 (HL60 and HepG2), Parasporin-4 (Caco-2, Sawano, and MOLT-4), Parasporin-5 (no specificity), and Parasporin-6 (HepG2, HeLa, and normal). Based on previous reports, the objective of this study was fueled by the possibility that there are strains of Bacillus thuringiensis carrying delta-endotoxins capable of causing death to HeLa cancer cells. The purpose of this study was to determine if the Bt isolates, 1-14, obtained from environmental samples collected in Middle Tennessee Counties carry delta-endotoxins that may be potentially classified in the parasporin-1 protein family. It was hypothesized delta-endotoxin would be highly toxic to HeLa (cervical cancer) cells compared to MCF-7 (breast cancer) cells and served as a good anti-cancer agent. Delta-endotoxin was extracted from the Bt strains and activated by trypsin. Based on protein profiles from SDS-Page analysis Bt isolates 4, 5, 6, 8 and 9 were chosen to test various delta-endotoxins on HeLa and MCF-7 cancer cells. Growth analysis was determined using a cell viability indicator, Alamar Blue, and a florescent plate reader. There was no major growth (> 40%) inhibition from any of the delta-endotoxins on HeLa cells. Thus, the toxins from these strains do not belong to the parasporins groups that inhibit HeLa cells. After 24h exposure to various delta-endotoxins MCF-7 cells' growth was found to be Bt 4 at 36%, Bt 5 at 4%, Bt 6 at 18%, Bt 8 at 25%, and Bt 9 at 81%. It was found that the delta-endotoxins in high concentrations 75µg/µl and 87.5µg/µl from Bt 5 and Bt 6 respectively, show 96% and 82% inhibition of growth against MCF-7 cells. It was concluded the delta-endotoxins from Bt 5 and 6 were not of the Parasporin 1 group and would serve as a novel anti-cancer agent on MCF-7 cells. However, further studies must investigate purified delta-endotoxins and their mechanism of action on various cell lines and determine the minimum concentration for growth inhibition.
Mailene Leachana King,
"Determination of the toxicity of Bacillus thuringiensis δ-endotoxin against cancer cells"
ETD Collection for Tennessee State University.