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Silver Nanoparticles (AgNPs) Uptake by Caveolae-Dependent Endocytosis is Responsible for Their Selective Effect
Mariana Morais1,2, Francisca Dias1, Patrícia Figueiredo3
1Molecular Oncology and Viral Pathology Group, Research Center of IPO Porto (CI-IPOP) / RISE@CI-IPOP (Health Research Network), Portuguese Oncology Institute of Porto (IPO Porto) / Porto Comprehensive Cancer Center (Porto.ccc), Porto, Portugal.
Purpose:
Castration Resistant Prostate Cancer (CRPC) is characterized by poor prognosis and limited therapeutic options. AgNPs functionalized with glucose (G-AgNPs) were observed cytotoxic to CRPC cell lines (PC-3 and Du-145) and not LNCaP. This study aims to evaluate AgNPs and G-AgNPs' uptake mechanisms in these cells and understand their role in the selective effect against CRPC cells.
Methods:
Uptake of AgNPs and G-AgNPs was assessed through transmission electron microscopy (TEM). A microRNA (miRNAs) analysis approach was used to uncover the main molecular differences responsible for the endocytic mechanisms' regulation. Caveolin (Cav) 1 and 2 mRNA and protein levels were assessed in the three cell lines. Caveolae-dependent endocytosis was inhibited with genistein or siCav1- and siCav2- in PC-3 and Du-145 and resazurin assay was used to evaluate viability after AgNPs and G-AgNPs administration. Caveolae-dependent endocytosis was induced with Cav1+ and Cav2+ plasmids in LNCaP, resazurin assay was used to evaluate viability after AgNPs and G-AgNPs administration and TEM to assess their location.
Results:
AgNPs and G-AgNPs were not uptaked by LNCaP. miRNA analysis revealed 37 upregulated and 90 downregulated miRNAs. Functional enrichment analysis of miRNAs' targets resulted in enrichment of terms related to endocytosis and caveolae. We observed that Cav1 and Cav2 are not expressed in LNCaP. Inhibiting caveolae-dependent endocytosis in Du-145 and PC-3 led to a significative reduction of cytotoxic capacity of AgNPs and G-AgNPs and induction of caveolae-dependent endocytosis in LNCaP lead to a significative increase as well as their uptake by cells.
Conclusion:
This study shows the potential of these AgNPs as a new therapeutic approach directed to CRPC patients, uncovers caveolae-dependent endocytosis as the uptake mechanism of these AgNPs and highlights deregulation of Cav1 and Cav2 expression as a key difference in hormone sensitive and resistant PCa cells which may be responsible for drug resistance.
Insights
Silver nanoparticles (AgNPs) functionalized with glucose (G-AgNPs) show selective cytotoxicity against Castration Resistant Prostate Cancer (CRPC) cells. Uptake is mediated by caveolae-dependent endocytosis, linked to Cav1 and Cav2 expression, offering a potential targeted therapy for CRPC.
Area of Science:
- Nanomedicine
- Cancer Biology
- Molecular Biology
Background:
- Castration Resistant Prostate Cancer (CRPC) presents a significant clinical challenge with limited treatment options.
- Silver nanoparticles (AgNPs) functionalized with glucose (G-AgNPs) exhibit selective cytotoxicity towards specific CRPC cell lines (PC-3, Du-145) but not hormone-sensitive LNCaP cells.
Purpose of the Study:
- To investigate the cellular uptake mechanisms of AgNPs and G-AgNPs in CRPC and hormone-sensitive prostate cancer cell lines.
- To elucidate the molecular basis for the selective cytotoxic effect of AgNPs and G-AgNPs against CRPC cells.
Main Methods:
- Transmission electron microscopy (TEM) was employed to visualize nanoparticle uptake.
- MicroRNA (miRNA) analysis identified key molecular regulators of endocytosis.
- Caveolin (Cav) 1 and 2 expression levels were assessed via mRNA and protein analysis.
- Functional assays (resazurin) evaluated cell viability following inhibition or induction of caveolae-dependent endocytosis using chemical inhibitors (genistein) or genetic manipulation (siCav1/2, Cav1/2 plasmids).
Main Results:
- AgNPs and G-AgNPs were not internalized by LNCaP cells, which lack Cav1 and Cav2 expression.
- miRNA analysis revealed significant dysregulation of miRNAs involved in endocytosis and caveolae pathways.
- Inhibition of caveolae-dependent endocytosis in PC-3 and Du-145 cells markedly reduced AgNP and G-AgNP cytotoxicity.
- Induction of caveolae-dependent endocytosis in LNCaP cells enhanced nanoparticle uptake and cytotoxicity.
Conclusions:
- AgNPs and G-AgNPs demonstrate potential as targeted therapeutics for CRPC.
- Caveolae-dependent endocytosis is the primary mechanism for AgNP and G-AgNP cellular uptake.
- Differential expression of Cav1 and Cav2 is a key distinguishing factor between hormone-sensitive and resistant prostate cancer cells, potentially contributing to drug resistance.
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