Recent Developments in mRNA-Based Protein Supplementation Therapy to Target Lung Diseases

Abstract

Protein supplementation therapy using in vitro-transcribed (IVT) mRNA for genetic diseases contains huge potential as a new class of therapy. From the early ages of synthetic mRNA discovery, a great number of studies showed the versatile use of IVT mRNA as a novel approach to supplement faulty or absent protein and also as a vaccine. Many modifications have been made to produce high expressions of mRNA causing less immunogenicity and more stability. Recent advancements in the in vivo lung delivery of mRNA complexed with various carriers encouraged the whole mRNA community to tackle various genetic lung diseases. This review gives a comprehensive overview of cells associated with various lung diseases and recent advancements in mRNA-based protein replacement therapy. This review also covers a brief summary of developments in mRNA modifications and nanocarriers toward clinical translation.

Keywords: in vitro transcription; lung; mRNA; nanotransporter; protein supplement.

Figure 1

mRNA IVT, Modifications, and Function and Timeline Overview of milestones in protein supplementation therapy, in vitro transcription, and mRNA modification. White boxes, important milestones for the development of mRNA therapy; blue boxes, evolution of different cap structures;, , red, green, and gray boxes, 5′ UTR, 3′ UTR, and poly(A) tail, respectively, the addition of regulatory elements in the modification of mRNA;, , yellow boxes, nucleoside modifications and sequence optimizations in the development for mRNA therapy., , , , ,

Figure 2

Processing of IVT mRNA in a Cell (A) In vitro-transcribed (IVT) mRNA from linearized DNA or PCR-amplified fragment is used to transfect the cell of interest. Step 1: mRNA protection from RNase degradation and mRNA uptake are facilitated by various carriers. Step 2 of mRNA transport and release inside the cell is still unclear. Different capping modification can increase translation in step 3 and also protect from degradation. In steps 3 and 4, the translated protein from delivered mRNA gets transferred to various parts of the cell system based on post-translational modification. For an immunotherapeutic approach, the translated protein needs to get degraded by proteasome to antigen epitopes and delivered to MHC (major histocompatibility complex) class I located in the endoplasmic reticulum. MHC class I mediates surface presentation of the presented epitope to CD8+ cytotoxic T cells. The T cell further initiates the immune response by relocating the antigen and presenting in to MHC II. (B) IVT mRNA cause inflammatory responses and inhibition of mRNA replication as triphosphorylated mRNA or double-stranded RNA (dsRNA) can be recognized by Toll-like receptors 3, 7, and 8 (endosomal innate immune receptors), which can initiate inflammation associated with type 1 interferon (IFN), interleukin-6 and -12, and tumor necrosis factor (TNF). Cytoplasmic receptors, protein kinase R (PKR), retinoic acid-inducible gene I protein (RIG-I), melanoma differentiation-associated protein (MDA5), and 2′-5′-oligoadenylate synthase (OAS) can detect triphosphorylated mRNA or dsRNA and stalled translation through eIF2α, RNA degradation by ribonuclease L (RNase L), and inhibition of mRNA replication by IFN.,

Figure 3

Deposition of Nanoparticles for Delivery in the Lung after Intratracheal or Intravenous Administration Left: intratracheal instillation requires a particle size of 1–3 μm to reach the alveoli efficiently; particles from 4 to 7 μm are mainly distributed to the upper airways and main bronchioles, and particles smaller than 1 μm are exhaled again., Right: inhaled nanoparticles can enter bronchial as well as alveolar epithelium; nanoparticles can enter lymph and blood circulation to be delivered to secondary organs. Intravenous injection can systemically deliver nanoparticles to a limited part of the alveolar epithelium due to small and non-fenestrated endothelial cells in the capillaries in the lung.