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Profile breast tumor microenvironment heterogeneity using gene circuits-loaded nanoparticles
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O cancro da mama é a neoplasia maligna mais comum e a segunda principal causa de mortalidade feminina. A heterogeneidade genética e o microambiente tumoral complexo limitam a eficácia dos tratamentos convencionais, destacando a terapia génica como uma alternativa promissora. O objetivo deste trabalho foi o desenvolvimento e caracterização de nanopartículas (NPs) poliméricas e lipídicas para a entrega de microRNAs (miRNAs). Estas NPs foram complexadas com plasmídeos que contém uma sequência de miR-125b e promotores celulares como o ERBB2 (específico para células HER2+, pERBB2) ou CMV (pCMV). As NPs poliméricas que contém 100% de aminas à sua superfície apresentaram menor tamanho, carga positiva e melhor uniformidade, enquanto as de 25% originaram partículas maiores. No caso das NPs lipídicas, estas são mais compactas e monodispersas. Nos ensaios de viabilidade celular, as NPs lipídicas e as poliméricas nas concentrações mais baixas mostraram baixa toxicidade. As NPs lipídicas demonstraram maior eficiência na entrega do miR 125b, quer com pERBB2 quer com pCMV, embora com baixa internalização (7,5% em células SK-BR-3). Por último, na análise por PCR não se observaram alterações na expressão de miR-125b, indicando falhas na transcrição plasmídica. Assim, a otimização dos plasmídeos é essencial para assegurar expressão e eficácia terapêutica.
Breast cancer (BC) is the most common malignant neoplasm and the second leading cause of female mortality. Genetic heterogeneity and the complex tumor microenvironment limit the effectiveness of conventional treatments, highlighting gene therapy as a promising alternative. The aim of this work was the development and characterization of polymeric and lipid nanoparticles (NPs) for the delivery of microRNAs (miRNAs). These NPs were complexed with plasmids containing a miR-125b sequence and cellular promoters such as ERBB2 (specific for HER2+ cells, pERBB2) or CMV (pCMV). Polymeric NPs with 100% surface amines exhibited smaller size, positive charge, and better uniformity, whereas those with 25% amines generated larger particles. Lipid NPs, in contrast, were more compact and monodisperse. In cell viability assays, both lipid and poly meric NPs at lower concentrations showed low toxicity. Lipid NPs demonstrated higher effi ciency in miR-125b delivery with both pERBB2 and pCMV, although with low internaliza tion (7.5% in SK-BR-3 cells). Finally, PCR analysis revealed no changes in miR-125b expression, indicating failures in plasmid transcription. Thus, plasmid optimization is essential to ensure expression and therapeutic efficacy.
Breast cancer (BC) is the most common malignant neoplasm and the second leading cause of female mortality. Genetic heterogeneity and the complex tumor microenvironment limit the effectiveness of conventional treatments, highlighting gene therapy as a promising alternative. The aim of this work was the development and characterization of polymeric and lipid nanoparticles (NPs) for the delivery of microRNAs (miRNAs). These NPs were complexed with plasmids containing a miR-125b sequence and cellular promoters such as ERBB2 (specific for HER2+ cells, pERBB2) or CMV (pCMV). Polymeric NPs with 100% surface amines exhibited smaller size, positive charge, and better uniformity, whereas those with 25% amines generated larger particles. Lipid NPs, in contrast, were more compact and monodisperse. In cell viability assays, both lipid and poly meric NPs at lower concentrations showed low toxicity. Lipid NPs demonstrated higher effi ciency in miR-125b delivery with both pERBB2 and pCMV, although with low internaliza tion (7.5% in SK-BR-3 cells). Finally, PCR analysis revealed no changes in miR-125b expression, indicating failures in plasmid transcription. Thus, plasmid optimization is essential to ensure expression and therapeutic efficacy.
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Breast cancer gene therapy miR-125b PAMAM Lipofectamine plasmids