Concise Review: Application of In Vitro Transcribed Messenger RNA for Cellular Engineering and Reprogramming: Progress and Challenges

Abstract

Several diseases are caused by missing or defective synthesis of proteins due to genetic or acquired disorders. In recent years, in vitro transcribed (IVT) messenger RNA (mRNA)-based therapy for de novo protein expression in cells has increased in importance. Thereby, desired proteins can be produced in cells by exogenous delivery of IVT mRNA, which does not integrate into the host genome and results in transient production of target proteins. Due to the lack of genomic integration, the risk of mutation and tumor development is minimized. Different approaches using IVT mRNA have been applied to alter the expression profiles of cells by the production of proteins. IVT mRNAs encoding transcription factors have led to the highly efficient induction of pluripotency in somatic cells and generated induced pluripotent stem cells that are free of viral vector components. Furthermore, specific IVT mRNA cocktails containing more than one specific IVT mRNA can be used to directly induce the differentiation into a desired cell type. In theory, every desired mRNA can be produced in vitro and used to enable extrinsic biosynthesis of target proteins in each cell type. Cells can be engineered by IVT mRNA to express antigens on dendritic cells for vaccination and tumor treatment, surface receptors on stem cells for increased homing to distinct areas, and to produce industrial grade human growth factors. In this review, we focus on the progress and challenges in mRNA-based cell engineering approaches. STEM CELLS 2017;35:68-79 SIGNIFICANCE STATEMENTThe use of synthetic messenger RNA (mRNA) to produce desired proteins in cells and thereby reprogramming and transdifferentiation of cells is a very promising technology to enable the application of these engineered cells in clinic. Synthetic mRNA does not integrate into the genome and it is not necessary to enter the nucleus. The protein is directly produced in the cytosol. In this review, we summarize the progress of the synthetic mRNA-based cell engineering strategies and highlight the challenges, which have to be overcome to improve the application.

Exploitation of Synthetic mRNA To Drive Immune Effector Cell Recruitment and Functional Reprogramming In Vivo

“…In the current study, although we did observe transient recruitment of neutrophils, the monocytes that were recruited appeared to be predominantly in a "resting" or nonactivated state. Synthetic mRNAs have the potential to activate mammalian cells through pattern recognition receptors such as TLR 7/8, RIG-1, and NOD2, discussed by Steinle and colleagues (56). However, the modifications introduced into the synthetic mRNA used in the current study appear to minimize cellular activation.…”

Silicon‐Nanotube‐Mediated Intracellular Delivery Enables Ex Vivo Gene Editing

“…These findings confirmed that SiNTs can mediate efficient cargo delivery while maintaining their biological functionality. Delivery of mRNA by SiNTs may open new opportunities for transient/nonintegrating transfection methods, [ 25 ] reducing the risk of mutation and tumor development associated with viral vectors, which is extremely important in clinical studies.…”

Improving mRNA-Based Therapeutic Gene Delivery by Expression-Augmenting 3′ UTRs Identified by Cellular Library Screening

“…The potency of the antigen-specific immune response correlates positively with the persistence of the synthetic mRNA in the hDCs and the amount of translated protein. 7,8 Regenerative medicine is another field in which mRNA is used for the conversion of cell fate, including the generation of induced pluripotent stem cells (iPSCs) from easily accessible adult somatic cells (reviewed in Steinle et al 9 ). iPSCs resemble human embryonic stem cells (hESCs) but are not associated with the respective ethical concerns.…”