Chemically modified RNA activated matrices enhance bone regeneration

Abstract

There exists a dire need for improved therapeutics to achieve predictable bone regeneration. Gene therapy using non-viral vectors that are safe and efficient at transfecting target cells is a promising approach to overcoming the drawbacks of protein delivery of growth factors. Here, we investigated the transfection efficiency, cytotoxicity, osteogenic potential and in vivo bone regenerative capacity of chemically modified ribonucleic acid (cmRNA) (encoding BMP-2) complexed with polyethylenimine (PEI) and made comparisons with PEI complexed with conventional plasmid DNA (encoding BMP-2). The polyplexes were fabricated at an amine (N) to phosphate (P) ratio of 10 and characterized for transfection efficiency using human bone marrow stromal cells (BMSCs). The osteogenic potential of BMSCs treated with these polyplexes was validated by determining the expression of bone-specific genes, osteocalcin and alkaline phosphatase as well as through the detection of bone matrix deposition. Using a calvarial bone defect model in rats it was shown that PEI-cmRNA (encoding BMP-2)-activated matrices promoted significantly enhanced bone regeneration compared to PEI-plasmid DNA (BMP-2)-activated matrices. Our proof of concept study suggests that scaffolds loaded with non-viral vectors harboring cmRNA encoding osteogenic proteins may be a powerful tool for stimulating bone regeneration with significant potential for clinical translation.

3D Bioprinting of Developmentally Inspired Templates for Whole Bone Organ Engineering

“…The approach could also be adapted to other biofabrication methods such as inkjet printing and electrospinning where cellular and extra cellular material can be arranged in complex patterns [31] . Future work will look at recapitulating biomolecule gradients that occur during skeletal developmental processes using bioprinted patterns of plasmid DNA encoding for vascular, chondrogenic and osteogenic factors such as VEGF, PDGF, TGF-β3 and BMP-2 [32][33][34][35] . We will also explore the spatial and temporal control of these and other factors [3,36] to help engineer the micro-environment of developing bones.…”

Human cellular CYBA UTR sequences increase mRNA translation without affecting the half-life of recombinant RNA transcripts

“…In another study by Kormann and colleagues, translation was maximized and innate immune response was reduced in a mouse lung disease model by introducing chemically-modified nucleotides during in vitro transcription of mRNA (IVT-mRNA), coding for surfactant protein B (SP-B) with a 25% replacement of cytidine and uridine by modified 5-methyl cytidine (5-methyl CTP) and 2-thio uridine (2-thio UTP), respectively19. Other studies have reported that modified nucleotide(s) within the mRNA structure have a strong impact on protein translation and enable reprogramming of human cells to pluripotency, vascular regeneration, as well as bone regeneration in vitro and in vivo 20212223. In the latter example, modified mRNA coding for human bone morphogenetic protein 2 (hBMP2) was used to induce bone regeneration2223.…”

Controlled and Sequential Delivery of Fluorophores from 3D Printed Alginate-PLGA Tubes

“…HEK293 cells were tested because they are routinely used in drug and gene delivery studies. Bone marrow stromal cells were tested due to the fact that they are widely used in tissue engineering, specifically for bone regeneration 4, 5 . Mechanical testing revealed that insertion of a PLGA core enhanced the mechanical strength of the alginate tubes by up to 4-fold.…”