Parasporins of Bacillus thuringiensis Strain Exhibit Apoptosis-Mediated Selective Cytotoxicity to MDA-MB-231 Cells through Oxidative Stress

Abstract

Historically, the most important source of both antibiotics and anticancer medications has been microorganisms. Bacillus thuringiensis (Bt) is one of the most prominent bacterial species used as a therapeutic agent targeting cancerous cells in recent worldwide investigations. This study was designed to isolate, molecularly identify, and discover novel Saudi Arabian Bt strains that selectively exhibit cytotoxic properties against MDA-MB-231, a human triple-negative breast cancer (TNBC) cell model. The bacterial strain under investigation was biochemically typed using API 20E and API CH50 and molecularly typed using 16S rDNA sequencing. Flow cytometry and immunoblotting were performed to elucidate the mechanism-of-action (MOA). Molecular typing confirmed the identity of the isolated non-hemolytic strain to be Bt and was named Bt HAU-145. Microscopic examination showed that the strain possessed a parasporal (PS) crystal protein with a spherical morphology. Data of cytotoxicity assay based on MTT revealed that Bt HAU-145 strain exhibited selective and potent cytotoxicity against MDA-MB-231, with a 50 percent inhibition (IC50) of 28 µg/ml. FACS analysis revealed that PS proteins induced both late and early apoptosis in a ROS-dependent manner. Immunoblotting assays showed increased expression of caspase-3 in response to PS treatment, paralleled by a reduction in Bcl-2 expression. This is the first study to investigate the MOA of PS proteins from the Saudi Arabian Bt strain, showing an induction of apoptosis through a ROS-dependent mechanism in TNBC cells. It is hoped that PS-based therapeutic strategies will be investigated at the preclinical scale in non-human primates prior to the clinical scale in randomized clinical trials.

Keywords

• bacillus thuringiensis
• apoptosis
• anticancer
• non-insecticidal inclusions
• parasporins
• reactive oxygen species