Sousa Ângela, Almeida Ana M, Faria Rúben, Konate Karidia, Boisguerin Prisca, Queiroz João A, Costa Diana
CICS-UBI - Centro de Investigação em Ciências da Saúde, Universidade da Beira Interior, Av. Infante D. Henrique, 6200-506 Covilhã, Portugal.
Centre de Recherche en Biologie cellulaire de Montpellier, CNRS UMR 5237, Université de Montpellier, 1919 Route de Mende, 34293 Montpellier Cedex 5, France.
Colloids Surf B Biointerfaces. 2019 Nov 1;183:110417. doi: 10.1016/j.colsurfb.2019.110417. Epub 2019 Aug 6.
The field of gene therapy still attracts great interest due to its potential therapeutic effect towards the most deadly diseases, such as cancer. For cancer gene therapy to be feasible and viable in a clinical setting, the design and development of a suitable gene delivery system is imperative. Peptide based vectors, in particular, reveal to be promising for therapeutic gene release. Following this, two different peptides, RALA and WRAP5, have been investigated mainly regarding their ability to form complexes with a p53 encoding plasmid (pDNA) with suitable properties for gene delivery. To address this issue, and after an initial screening study focused on the dependence of pDNA complexation capacity with the nitrogen to phosphate groups (N/P) ratio, a design of experiments (DoE) tool has been employed. For each peptide/pDNA system, parameters such as, the buffer pH and the N/P ratio were considered the DoE inputs and the vector size, zeta potential and pDNA complexation capacity (CC) were monitored as DoE outputs. The main goal was to find the optimal experimental conditions to minimize particle sizes, as well as, to maximize the positive surface charges of the formulated nanosystems and maximize the pDNA CC. Through the DoE method applied, the optimal RALA/pDNA and WRAP5/pDNA formulations were revealed and show interesting features related to peptide structure and pDNA complexation ability. This work illustrates the great utility of experimental design tools in optimizing the formulation of peptide/pDNA vectors in a minimum number of experiments providing relevant knowledge for the development of more suitable and efficient gene delivery systems. The new insights achieved on these carriers clearly instigate deeper research on gene therapy.
基因治疗领域因其对诸如癌症等最致命疾病的潜在治疗效果，仍然吸引着极大的关注。为了使癌症基因治疗在临床环境中可行且切实可行，设计和开发合适的基因递送系统至关重要。特别是基于肽的载体，显示出在治疗性基因释放方面很有前景。在此之后，主要研究了两种不同的肽，RALA和WRAP5，关于它们与编码p53的质粒（pDNA）形成具有适合基因递送特性的复合物的能力。为了解决这个问题，在最初一项侧重于pDNA络合能力与氮磷基团（N/P）比率相关性的筛选研究之后，采用了实验设计（DoE）工具。对于每个肽/pDNA系统，诸如缓冲液pH值和N/P比率等参数被视为DoE输入，而载体大小、zeta电位和pDNA络合能力（CC）则作为DoE输出进行监测。主要目标是找到最佳实验条件，以最小化颗粒大小，同时最大化所制备纳米系统的正表面电荷并最大化pDNA CC。通过应用DoE方法，揭示了最佳的RALA/pDNA和WRAP5/pDNA配方，并显示出与肽结构和pDNA络合能力相关的有趣特征。这项工作说明了实验设计工具在以最少的实验优化肽/pDNA载体配方方面的巨大效用，为开发更合适、更有效的基因递送系统提供了相关知识。在这些载体上获得的新见解显然促使对基因治疗进行更深入的研究。
