Binase Immobilized on Halloysite Nanotubes Exerts Enhanced Cytotoxicity toward Human Colon Adenocarcinoma Cells

Abstract

Many ribonucleases (RNases) are considered as promising tools for antitumor therapy because of their selective cytotoxicity toward cancer cells. Binase, the RNase from Bacillus pumilus, triggers apoptotic response in cancer cells expressing RAS oncogene which is mutated in a large percentage of prevalent and deadly malignancies including colorectal cancer. The specific antitumor effect of binase toward RAS-transformed cells is due to its direct binding of RAS protein and inhibition of downstream signaling. However, the delivery of proteins to the intestine is complicated by their degradation in the digestive tract and subsequent loss of therapeutic activity. Therefore, the search of new systems for effective delivery of therapeutic proteins is an actual task. This study is aimed to the investigation of antitumor effect of binase immobilized on natural halloysite nanotubes (HNTs). Here, we have developed the method of binase immobilization on HNTs and optimized the conditions for the enzyme loading and release (i); we have found the non-toxic concentration of pure HNTs which allows to distinguish HNTs- and binase-induced cytotoxic effects (ii); using dark-field and fluorescent microscopy we have proved the absorption of binase-loaded HNTs on the cell surface (iii) and demonstrated that binase-halloysite nanoformulations possessed twice enhanced cytotoxicity toward tumor colon cells as compared to the cytotoxicity of binase itself (iv). The enhanced antitumor activity of biocompatible binase-HNTs complex confirms the advisability of its future development for clinical practice.

Zeolites as Carriers of Antitumor Ribonuclease Binase

“…Zeolite-based nanoparticles used in generating time-controlled release of 5-fluorouracil from zeolite preparations showed anti-cancer effect toward Caco-2 monolayers (Spanakis et al, 2014). Earlier, we have demonstrated that binase-halloysite complex doubled anticancer efficiency of binase due to its perfect absorption by cells and longer release reducing the viability of human colon adenocarcinoma cells Colo320 by 60% (Khodzhaeva et al, 2017). The same level of toxicity toward human adenocarcinoma Caco2 cells was obtained for chabazite-binase complex.…”

Recent Strategies for the Immobilization of Therapeutic Enzymes

“…The size and shape of nanoparticles will affect the adsorption patterns, protein structure, and function. Many enzymes, such as lysozyme [ 70 ], lipase [ 71 , 72 ], nattokinase [ 73 ], asparaginase [ 74 ], SOD [ 75 , 76 ], β-galactosidase [ 77 ], glucocerebrosidase [ 78 ], binase [ 79 ], hyaluronidase (HAses) [ 80 ], GOX, and HRP [ 69 ], have been reported to be immobilized onto nanoparticles. Among these enzymes, the activity and stability of glucocerebrosidase are compromised in patients with Gaucher disease (metabolic storage disorder) because these patients do not properly degrade glucosylceramide (GlcCer), which accumulates and can lead to health problems, including bone lesions.…”

Application of Halloysite Nanotubes in Cancer Therapy—A Review

“…In addition to siRNA, ribonucleases (RNases) have also been proposed in anticancer therapy since they may destroy RNA in cancer cells and produce signals leading to cancer cells’ programmed death. Khodzhaeva et al [ 76 ] immobilized Bacillus pumilus RNase-binase-on HNTs. Cytotoxicity studies toward human colon adenocarcinoma Colo320 confirmed that while the cells’ viability after 24 h incubation with pristine HNTs was as high as 97%, the free binase and binase-loaded HNTs reduced the number of viable cells by 27% and 64%, respectively.…”