Evaluation of the apoptotic effects of the engineered nanoformulation in colon cancer cells
Restoration of tumor suppression and improved anticancer efficacy following nanoparticles-mediated combined anti-miRNA 21 and 5-fluorouracil therapy in colorectal cancer
Paper
2026
RSC Pharmaceutics |
DOI
Background
This study develops PLGA nanoparticles co-encapsulating 5-fluorouracil (5-FU) and a locked nucleic acid anti-miRNA 21 oligonucleotide to overcome chemoresistance in colorectal cancer. The optimized nanoplatform (~183 nm, low PDI, −25.7 mV zeta potential) achieved high entrapment efficiency for both agents and was efficiently internalized by HT-29 colon cancer cells. The dual therapy suppressed IL-8, restored PTEN tumor suppressor expression, reduced cell viability, and induced the strongest apoptotic response, outperforming 5-FU or anti-miRNA 21 monotherapies, supporting combinatorial nanoparticle-based RNAi/chemotherapy for colon cancer.
Key Findings
The optimized PLGA nanoparticles had a size of 183.3 ± 2.1 nm, PDI of 0.08, zeta potential of −25.7 mV, and encapsulation efficiencies of 90.4% (5-FU) and 83.2% (anti-miRNA 21); TEM and AFM confirmed uniform spherical morphology. Fluorescence imaging showed efficient cellular uptake in HT-29 cells with perinuclear cytoplasmic localization. MTT assays showed combinatorial nanoparticles produced the greatest reduction in cell viability at 24, 48, and 72 hours, while live/dead assays confirmed the highest dead-cell ratio (3.72 ± 0.21). The dual formulation suppressed IL-8 expression by 63.9 ± 3.8-fold and increased PTEN expression (2.12 ± 0.26) versus controls. Annexin V-FITC flow cytometry revealed apoptosis of 7.39 ± 0.46% with the combination, 1.33-fold higher than 5-FU nanoparticles (5.56%) and 2.06-fold higher than anti-miRNA 21 nanoparticles alone (3.59%), demonstrating a synergistic apoptotic effect.
Contribution
Creative Biogene supplied the customized FAM-tagged locked nucleic acid (LNA) anti-miRNA 21 oligonucleotide, the RNAi therapeutic component of the dual nanoformulation. This reagent enabled targeted silencing of overexpressed miRNA-21 in HT-29 colorectal cancer cells, underpinning the study's demonstration of enhanced apoptosis, IL-8 suppression, and restored PTEN tumor suppressor expression.
This study develops PLGA nanoparticles co-encapsulating 5-fluorouracil (5-FU) and a locked nucleic acid anti-miRNA 21 oligonucleotide to overcome chemoresistance in colorectal cancer. The optimized nanoplatform (~183 nm, low PDI, −25.7 mV zeta potential) achieved high entrapment efficiency for both agents and was efficiently internalized by HT-29 colon cancer cells. The dual therapy suppressed IL-8, restored PTEN tumor suppressor expression, reduced cell viability, and induced the strongest apoptotic response, outperforming 5-FU or anti-miRNA 21 monotherapies, supporting combinatorial nanoparticle-based RNAi/chemotherapy for colon cancer.
Key Findings
The optimized PLGA nanoparticles had a size of 183.3 ± 2.1 nm, PDI of 0.08, zeta potential of −25.7 mV, and encapsulation efficiencies of 90.4% (5-FU) and 83.2% (anti-miRNA 21); TEM and AFM confirmed uniform spherical morphology. Fluorescence imaging showed efficient cellular uptake in HT-29 cells with perinuclear cytoplasmic localization. MTT assays showed combinatorial nanoparticles produced the greatest reduction in cell viability at 24, 48, and 72 hours, while live/dead assays confirmed the highest dead-cell ratio (3.72 ± 0.21). The dual formulation suppressed IL-8 expression by 63.9 ± 3.8-fold and increased PTEN expression (2.12 ± 0.26) versus controls. Annexin V-FITC flow cytometry revealed apoptosis of 7.39 ± 0.46% with the combination, 1.33-fold higher than 5-FU nanoparticles (5.56%) and 2.06-fold higher than anti-miRNA 21 nanoparticles alone (3.59%), demonstrating a synergistic apoptotic effect.
Contribution
Creative Biogene supplied the customized FAM-tagged locked nucleic acid (LNA) anti-miRNA 21 oligonucleotide, the RNAi therapeutic component of the dual nanoformulation. This reagent enabled targeted silencing of overexpressed miRNA-21 in HT-29 colorectal cancer cells, underpinning the study's demonstration of enhanced apoptosis, IL-8 suppression, and restored PTEN tumor suppressor expression.
Figure 1. The figure illustrates CRISPR/Cas9 targeting of the CTLA4 locus, HDR donor design, and resulting editing efficiency and CTLA4 expression in edited versus unedited cells. (Fox TA, et al., 2022)