Engineering biomolecular systems: Controlling the self-assembly of gelatin to form ultra-small bioactive nanomaterials

The size of nanocarriers determines the biological property of the materials, especially as it relates to intratumoral distribution. Previous research has shown that sizes of 10-50 nm penetrate deep inside the tumor, resulting in better efficacy. On the other hand, studies have shown that gelatin exhibits excellent biological properties, including compatibility, degradability, and toxicity. Therefore, FDA approved gelatin as a safe material to use as an excipient in injectables. The bottleneck is the nonexistence of smaller-sized gelatin nanoparticles (GNPs) to realize the full potential of these biomaterials. Yet, GNPs with sizes of less than 50 nm have not been reported; the synthetic strategy reported in the literature uses "uncontrolled crosslinking coupled with nanoprecipitation", resulting in larger particle size. We have developed a new method to self-assemble gelatin strands by using an anionic, phosphate-based crosslinker and controlled precipitation. The method we developed produced ultra-small gelatin nanoparticles (G(X)) of size 10 nm with a high degree of reproducibility, and it was characterized using dynamic light scattering (DLS), Energy-dispersive X-ray spectroscopy (EDS), High-resolution transmission, and scanning electron microscopy (HR-TEM/STEM). We also explored G(X) as a bioactive platform to encapsulate imaging and therapy agents within the cavity. Interestingly, we were able to encapsulate 2 nm size gold nanoparticles within the void of G(X). The versatile nature of the G(X) particles was further demonstrated by surface functionalizing with larger size gelatin nanoparticles to form core-satellite nanocomposites. Additionally, we studied the tumor penetrability of dye-tagged 10, 50, and 200 nm gelatin nanoparticles. The study showed that smaller size gelatin nanoparticles penetrate deeper tumor regions than larger particles. In general, G(X) was efficient in penetrating the inner region of the spheroids. The results demonstrate the potential capabilities of ultra-small G(X) nanoparticles for multi-staged payload delivery, diagnostics, and cancer therapy.