ABSTRACT
Introduction: Cell membrane-covered biomimetic nanosystems have allowed the development of homologous nanostructures to bestow nanoparticles with enhanced biointerfacing capabilities. The stability of these structures, however, still represents a challenge for the scientific community. This study is aimed at developing and optimizing cell derived membrane-coated nanostructures upon applying design of experiments (DoE) to improve the therapeutic index by homotypic targeting in cancer cells. Methods: Important physicochemical features of the extracted cell membrane from tumoral cells were assessed by mass spectrometry-based proteomics. PLGA-based nanoparticles encapsulating temozolomide (TMZ NPs) were successfully developed. The coating technology applying the isolated U251 cell membrane (MB) was optimized using a fractional two-level three-factor factorial design. All the formulation runs were systematically characterized regarding their diameter, polydispersity index (PDI), and zeta potential (ZP). Experimental conditions generated by DoE were also subjected to morphological studies using negative-staining transmission electron microscopy (TEM). Its short-time stability was also assessed. MicroRaman and Fourier-Transform Infrared (FTIR) spectroscopies and Confocal microscopy were used as characterization techniques for evaluating the NP-MB nanostructures. Internalization studies were carried out to evaluate the homotypic targeting ability. Results and Discussion: The results have shown that nearly 80% of plasma membrane proteins were retained in the cell membrane vesicles after the isolation process, including key proteins to the homotypic binding. DoE analysis considering acquired TEM images reveals that condition run five should be the best-optimized procedure to produce the biomimetic cell-derived membrane-coated nanostructure (NP-MB). Storage stability for at least two weeks of the biomimetic system is expected once the original characteristics of diameter, PDI, and ZP, were maintained. Raman, FTIR, and confocal characterization results have shown the successful encapsulation of TMZ drug and provided evidence of the effective coating applying the MB. Cell internalization studies corroborate the proteomic data indicating that the optimized NP-MB achieved specific targeting of homotypic tumor cells. The structure should retain the complex biological functions of U251 natural cell membranes while exhibiting physicochemical properties suitable for effective homotypic recognition. Conclusion: Together, these findings provide coverage and a deeper understanding regarding the dynamics around extracted cell membrane and polymeric nanostructures interactions and an in-depth insight into the cell membrane coating technology and the development of optimized biomimetic and bioinspired nanostructured systems.
ABSTRACT
COVID-19 has spread worldwide and early detection has been the key to controlling its propagation and preventing severe cases. However, diagnostic devices must be developed using different strategies to avoid a shortage of supplies needed for tests' fabrication caused by their large demand in pandemic situations. Furthermore, some tropical and subtropical countries are also facing epidemics of Dengue and Zika, viruses with similar symptoms in early stages and cross-reactivity in serological tests. Herein, we reported a qualitative immunosensor based on capacitive detection of spike proteins of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19. The sensor device exhibited a good signal-to-noise ratio (SNR) at 1 kHz frequency, with an absolute value of capacitance variation significantly smaller for Dengue and Zika NS1 proteins (|ΔC| = 1.5 ± 1.0 nF and 1.8 ± 1.0 nF, respectively) than for the spike protein (|ΔC| = 7.0 ± 1.8 nF). Under the optimized conditions, the established biosensor is able to indicate that the sample contains target proteins when |ΔC| > 3.8 nF, as determined by the cut-off value (CO). This immunosensor was developed using interdigitated electrodes which require a measurement system with a simple electrical circuit that can be miniaturized to enable point-of-care detection, offering an alternative for COVID-19 diagnosis, especially in areas where there is also a co-incidence of Zika and Dengue.
