Mad2 checkpoint gene silencing using epidermal growth factor receptor-targeted chitosan nanoparticles in non-small
Ana Vanessa Nascimento1, Amit Singh, Hassan Bousbaa
1CESPU, Instituto de Investigação e Formação Avançada em Ciências e Tecnologias da Saúde, IINFACTS, Rua Central de Gandra 1317, 4585-116 Gandra PRD, Portugal.
Abstract:
RNA interference has emerged as a powerful strategy in cancer therapy because it allows silencing of specific genes associated with tumor progression and resistance. Mad2 is an essential mitotic checkpoint component required for accurate chromosome segregation during mitosis, and its complete abolition leads to cell death. We have developed an epidermal growth factor receptor (EGFR)-targeted chitosan system for silencing the Mad2 gene as a strategy to efficiently induce cell death in EGFR overexpressing human A549 non-small cell lung cancer cells. Control and EGFR-targeted chitosan nanoparticles loaded with small interfering RNAs (siRNAs) against Mad2 were formulated and characterized for size, charge, morphology, and encapsulation efficiency. Qualitative and quantitative intracellular uptake studies by confocal imaging and flow cytometry, respectively, showed time-dependent enhanced and selective intracellular internalization of EGFR-targeted nanoparticles compared to nontargeted system. Targeted nanoparticles showed nearly complete depletion of Mad2 expression in A549 cells contrasting with the partial depletion in the nontargeted system. Accordingly, Mad2-silencing-induced apoptotic cell death was confirmed by cytotoxicity assay and flow cytometry. Our results demonstrate that EGFR-targeted chitosan loaded with Mad2 siRNAs is a potent delivery system for selective killing of cancer cells.
Insights
EGFR-targeted chitosan nanoparticles effectively deliver Mad2-silencing RNA (siRNA) to lung cancer cells, inducing targeted cell death. This novel RNA interference strategy shows promise for treating EGFR-overexpressing non-small cell lung cancer.
Area of Science:
- Nanomedicine
- Cancer Therapy
- Molecular Biology
Background:
- RNA interference (RNAi) is a key strategy for cancer therapy by silencing genes driving tumor growth and resistance.
- Mad2 is crucial for accurate mitosis; its inhibition induces cancer cell death.
- EGFR overexpression is common in non-small cell lung cancer (NSCLC).
Purpose of the Study:
- To develop an EGFR-targeted chitosan nanoparticle system for Mad2 gene silencing.
- To induce selective cell death in EGFR-overexpressing A549 NSCLC cells.
Main Methods:
- Formulation and characterization of EGFR-targeted and control chitosan nanoparticles loaded with Mad2-specific siRNAs.
- Assessment of nanoparticle cellular uptake via confocal imaging and flow cytometry.
- Evaluation of Mad2 gene silencing, cell death, and apoptosis using cytotoxicity assays and flow cytometry.
Main Results:
- EGFR-targeted nanoparticles demonstrated enhanced and selective intracellular uptake in A549 cells.
- Targeted delivery led to near-complete depletion of Mad2 expression, unlike the nontargeted system.
- Mad2 silencing effectively induced apoptotic cell death in cancer cells.
Conclusions:
- EGFR-targeted chitosan nanoparticles loaded with Mad2 siRNAs represent a potent delivery system for selective cancer cell killing.
- This approach offers a promising strategy for treating EGFR-overexpressing NSCLC.
