The design and optimization of polymeric gene and drug delivery systems require a delicate balance between transfection efficiency, cytotoxicity, and endocytic uptake. In this study, the influence of PPhe grafting length on the performance of PEI-co-PPhe copolymers was systematically investigated to identify the optimal composition for co-delivery applications. A series of copolymers with varying molar ratios of PEI-25k to Phe-NCA—1:26, 1:59, 1:85, 1:170, 1:230, and 1:267—were synthesized and characterized using 1H NMR and dynamic light scattering. The results indicated that increasing PPhe content led to a progressive reduction in zeta potential and particle size, suggesting improved charge shielding and enhanced complex stability. Notably, PEI-co-PPhe (1:170) exhibited the most favorable profile: moderate positive surface charge (~+15 mV), small particle size (~70 nm), and high colloidal stability, making it ideal for cellular internalization.

In vitro transfection assays using luciferase-expressing plasmid pGL3-DNA revealed that PEI-co-PPhe (1:170) outperformed both unmodified PEI-25k and other copolymer variants across a range of mass ratios. At a 1:5 DNA-to-copolymer ratio, the transfection efficiency reached 92% of that achieved by PEI-25k, while significantly reducing cytotoxicity. MTT assays confirmed this advantage: after 48 hours of incubation, cell viability remained above 85% at concentrations up to 500 µg/mL, compared to a sharp decline observed with PEI-25k. This improvement was attributed to the hydrophobic PPhe chains dispersing the dense positive charges of PEI, thereby minimizing membrane disruption and cellular damage.

Further evaluation focused on the co-delivery system comprising Bcl2-shRNA and CAD. When formulated at a 5:1:1 (w/w/w) mass ratio, the PEI-co-PPhe/Bcl2-shRNA/CAD complexes demonstrated a synergistic anti-tumor effect against B16F10 melanoma cells. Cytotoxicity testing showed that the combination therapy reduced cell viability to 38% at a CAD concentration of 1 µg/mL, significantly lower than either component alone.Crk-L Antibody Biological Activity Flow cytometry and confocal microscopy confirmed efficient co-uptake of both gene and drug into cells within 3 hours, with fluorescence signals intensifying over time and localizing near the nucleus.Cortactin Antibody In Vivo Additionally, pH-dependent drug release experiments verified that CAD facilitated rapid DOX release under acidic conditions (pH 5.PMID:34755650 0), with cumulative release reaching 55.2%, whereas only 27.3% was released at physiological pH (7.4). This targeted release mechanism enhances therapeutic specificity and minimizes off-target toxicity.

These findings underscore the importance of tailoring polymer architecture for biomedical applications. By optimizing the grafting density of PPhe on PEI-25k, this study successfully developed a low-toxicity, high-efficiency co-delivery platform capable of simultaneously delivering therapeutic genes and chemotherapeutics. The resulting PEI-co-PPhe (1:170)/Bcl2-shRNA/CAD system represents a promising strategy for overcoming resistance mechanisms in aggressive cancers such as melanoma, where dual inhibition of apoptosis and tumor proliferation is critical. Future work will explore in vivo efficacy and biodistribution to advance this platform toward clinical translation.MedChemExpress (MCE) offers a wide range of high-quality research chemicals and biochemicals (novel life-science reagents, reference compounds and natural compounds) for scientific use. We have professionally experienced and friendly staff to meet your needs. We are a competent and trustworthy partner for your research and scientific projects.Related websites: https://www.medchemexpress.com