Green fluorescent protein (GFP) is a widely used scaffold for
protein-based targeted nanomedicines because of its high biocompatibility, biological neutrality and outstanding structural stability. However, being immunogenicity a major concern in the development of
drug carriers, the use of exogenous
proteins such as GFP in clinics might be inadequate. Here we report a human
nidogen-derived
protein (HSNBT), rationally designed to mimic the structural and functional properties of GFP as a scaffold for nanomedicine. For that, a GFP-like β-barrel, containing the G2 domain of the human
nidogen, has been rationally engineered to obtain a biologically neutral
protein that self-assembles as 10nm-nanoparticles. This scaffold is the basis of a humanized
nanoconjugate, where GFP, from the well-characterized
protein T22-GFP-H6, has been substituted by the
nidogen-derived GFP-like HSNBT
protein. The resulting construct T22-HSNBT-H6, is a humanized CXCR4-targeted nanoparticle that selectively delivers conjugated genotoxic
Floxuridine into
cancer CXCR4+ cells. Indeed, the administration of T22-HSNBT-H6-FdU in a CXCR4-overexpressing
colorectal cancer mouse model results in an even more efficient selective antitumoral effect than that shown by its GFP-counterpart, in absence of systemic toxicity. Therefore, the newly developed GFP-like
protein scaffold appears as an ideal candidate for the development of humanized
protein nanomaterials and successfully supports the
tumor-targeted nanoscale drug T22-HSNBT-H6-FdU. STATEMENT OF SIGNIFICANCE: Targeted nanomedicine seeks for humanized and biologically neutral
protein carriers as alternative of widely used but immunogenic exogenous
protein scaffolds such as
green fluorescent protein (GFP). This work reports for the first time the rational engineering of a human homolog of the GFP based in the human
nidogen (named HSNBT) that shows full potential to be used in humanized
protein-based targeted nanomedicines. This has been demonstrated in T22-HSNBT-H6-FdU, a humanized CXCR4-targeted
protein nanoconjugate able to selectively deliver its genotoxic load into
cancer cells.