Bio-inspired trypsin-chitosan cross-linked enzyme aggregates: a versatile approach for stabilization through carrier-free immobilization

Mageed, Heidi; Ezz, Nermeen; Radwan, Rasha Ali

doi:10.5114/bta.2019.87589

Cited by 13 publications

(9 citation statements)

References 30 publications

(47 reference statements)

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“…19 The incorporation of nanotechnology introduces remarkable solutions for improvement of solubility, absorption, and stability of different agents. 31,39 Moreover, this process enhances the therapeutic activity of promising compounds through improving their solubility and adequate delivery. 22,40 Therefore, in this study, we formulated SIM in a SIM-CB formula.…”

Section: Discussionmentioning

confidence: 99%

Potentiating the Cytotoxic Activity of a Novel Simvastatin-Loaded Cubosome against Breast Cancer Cells: Insights on Dual Cell Death via Ferroptosis and Apoptosis

Elakkad¹,

Mohamed²,

Abuelezz³

2021

BCTT

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Purpose Female breast cancer is the most prevalent cancer worldwide. Emerging evidence shows that simvastatin (SIM) has promising anticancer activities. However, the underlying mechanisms are not fully elucidated. Increasing reports imply statins can modulate ferroptosis through disrupting reactive oxygen species (ROS) and glutathione peroxidase enzyme (GPX4) levels. However, whether ferroptosis contributes to SIM anticancer activity, especially regarding GPX4 is unclear. Moreover, poor aqueous SIM solubility hinders its delivery in adequate levels to tumor sites. Meanwhile, cubosomes are biocompatible nanocarriers that enhance lipophilic drug delivery. Therefore, in this study, we formulated a novel SIM-loaded cubosome (SIM-CB) and analyzed its cytotoxic activity on MCF-7 cancer cells in comparison with free SIM. Methods The present study tested the cytotoxic activity of SIM-CB on MCF-7 cells, in comparison with SIM using sulphorhodamine assay. We analyzed SIM-CB effect on apoptosis and cell cycle using flowcytometry, and investigated its effect on Bcl-2, caspase 3, ROS, reduced glutathione (GSH), lipid peroxides and GPX4 enzyme. Finally, we tested the persistence of SIM-CB apoptosis and ferroptosis activities on MCF-7 cells in presence of vitamin E, a potent antioxidant and ferroptosis inhibitor. Results SIM-CB was successfully formulated at the nano size. SIM-CB significantly increased simvastatin therapeutic activity, with IC 50 of SIM-CB 52% lower than SIM. 95% CI [1.8, 2.7], SD = 0.34 for SIM-CB, and [4.1, 5.2], SD = 0.45 for SIM. Compared with free SIM, SIM-CB doubled total deaths and increased apoptosis (p < 0.05). Moreover, SIM-CB remarkably increased caspase-3, ROS, and lipid peroxide levels but decreased antiapoptotic Bcl-2 protein, GSH, and GPX4 compared with free SIM. Notably, SIM-CB elicited a high distinguished resistance against the inhibitory effects of vitamin E. Conclusion To the best of our knowledge, this study is the first to present SIM-CB as a promising means to enhancing the therapeutic potential of simvastatin against breast cancer cells, through potentiating both apoptosis and ferroptosis.

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Section: Discussionmentioning

confidence: 99%

Potentiating the Cytotoxic Activity of a Novel Simvastatin-Loaded Cubosome against Breast Cancer Cells: Insights on Dual Cell Death via Ferroptosis and Apoptosis

Elakkad¹,

Mohamed²,

Abuelezz³

2021

BCTT

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“…Therefore, a co-aggregate compound rich in free amino groups, such as Chitosan, must be added before crosslinking. Chitosan as a co-aggregate can increase the effectiveness of crosslinking of CLEAs formation for various enzymes 14,15 . In this study, the Chitosan with high DD (>75%) was mixed with the enzyme to provide an adequate number of free amino groups for crosslinking.…”

Section: Discussionmentioning

confidence: 99%

“…The CLEAs formation method was adopted from Mageed et al 14 with slight modifications. A 0.5 mL chitosan solution was added to 1 mL of CCA aggregates suspension to obtain a final chitosan concentration in the range 1-5 mgmL -1 (x 1 ) according to the FFD factor level (Table 1).…”

Section: Cleas-cca Formation Methodsmentioning

confidence: 99%

“…The addition of co-aggregates, such as bovine serum albumin, poly-ethyleneimine, and poly-lysine, could form larger aggregates size and higher stability 9,12,13 . Chitosan is a relatively inexpensive biopolymer (polyaminosaccharides, copolymers of N-acetyl-D-glucosamine and N-amino-D-glucosamine) with high availability rich in reactive amino groups, potentials to mediate the formation crosslinks between the aggregates 14,15 .…”

Section: Introductionmentioning

confidence: 99%

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Optimization of cephalosporin C acylase immobilization using crosslinked enzyme aggregates technique

Julkipli

Syamsu

Wibisana

2022

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Cephalosporin C acylase (CCA) is an essential enzyme for the one-step conversion of cephalosporin C into 7-aminocephalosporanic acid (7-ACA), an intermediate compound used to synthesize various semi-synthetic cephalosporin antibiotics. The industrial process prefers to use enzymes in immobilized form rather than soluble. A crosslinked enzyme aggregate (CLEAs) is a potential matrix-less enzyme immobilization technique to produce stable immobilized enzymes with high activity and low production costs. This study aimed to optimize the CCA immobilization using the CLEAs technique with Chitosan as a co-aggregate. The CCA lysate was obtained from harvesting CCA fermentation broth using a mutant strain of Escherichia coli through cell separation and lysis steps. Partially purified CCA by ammonium sulfate addition was conducted to obtain an active fraction of 20-60% saturation, followed by co-aggregation with Chitosan to form physical CCA aggregates. The aggregates were then immobilized by a crosslinking technique using glutaraldehyde to form CLEAs-CCA. Optimization of the immobilization process was carried out by Response Surface Methodology in three steps, (i) screening of the influencing factors, (ii) determining the level of the significant factors, and (iii) optimizing the immobilization condition. The CLEAs-CCA activity was used as a response parameter. Under optimum conditions, CLEAs-CCA activity obtained was 85.91 Ug-1.

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“…A large number of free amino groups in chitosan mediated a substantial cross-linking, thus maintaining a high-enzymatic activity. 22 However, excessive chitosan competed with the targeted enzyme to cross-link with TPP, which caused a reduction in the lipase recovery activity.…”

Section: Cleas Preparationmentioning

confidence: 99%

Preparation and characterization of a nontoxic cross‐linked lipase aggregate by using sodium tripolyphosphate and chitosan

Yan

Shi

Ding

et al. 2021

J of Applied Polymer Sci

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Cross-linked enzyme aggregates (CLEAs) have the advantages of high-enzyme activity and stability, and have an application potential in food and feed processing. In order to address the food and feed safety of CLEAs and have ideal enzymatic properties, a new design of CLEAs of lipase from Aspergillus niger is developed by using sodium tripolyphosphate (TPP) as a cross-linking agent and chitosan as an auxiliary agent. The enzyme activity recovery of the developed CLEAs reaches 126%. The half-life of the CLEAs is 11.4-fold higher than that of free enzyme at the optimum reaction temperature of CLEAs. The surface structure of the CLEAs reveals cluster and porous structure. The variations of the secondary structures of the CLEAs indicate an increase of α-helix and decrease of β-sheet, β-turn, and random coil. The adding of chitosan is beneficial to enzyme activity recovery and thermal stability of the CLEAs. TPP and chitosan are safe and nontoxic, and the CLEAs prepared with TPP and chitosan is suitable for the food and feed industries.

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Bio-inspired trypsin-chitosan cross-linked enzyme aggregates: a versatile approach for stabilization through carrier-free immobilization

Cited by 13 publications

References 30 publications

Potentiating the Cytotoxic Activity of a Novel Simvastatin-Loaded Cubosome against Breast Cancer Cells: Insights on Dual Cell Death via Ferroptosis and Apoptosis

Potentiating the Cytotoxic Activity of a Novel Simvastatin-Loaded Cubosome against Breast Cancer Cells: Insights on Dual Cell Death via Ferroptosis and Apoptosis

Optimization of cephalosporin C acylase immobilization using crosslinked enzyme aggregates technique

Preparation and characterization of a nontoxic cross‐linked lipase aggregate by using sodium tripolyphosphate and chitosan

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