TY - JOUR
T1 - An Improved, Chemically Modified RNA Encoding BMP-2 Enhances Osteogenesis in Vitro and in Vivo
AU - Zhang, Wen
AU - De La Vega, Rodolfo E.
AU - Coenen, Michael J.
AU - Müller, Sebastian A.
AU - Peniche Silva, Carlos J.
AU - Aneja, Manish K.
AU - Plank, Christian
AU - Van Griensven, Martijn
AU - Evans, Christopher H.
AU - Balmayor, Elizabeth R.
N1 - Funding Information:
The support received from the Deutsche For-schungsgemeinschaft (DFG) via the Excellence Cluster ‘‘Nanosystems Initiative Munich (NIM)’’ (grant No. EXC 4/ 3) is greatly appreciated. W.Z. acknowledges the support received by the PhD program ‘‘Medical Life Science and Technology’’ at the Technical University of Munich. The authors would like to thank Zeljka Trepotec for the valuable assistance rendered while drafting the mRNA sequences, and Johannes Geiger during the cmRNAs production.
Publisher Copyright:
© Copyright 2019, Mary Ann Liebert, Inc., publishers 2019.
PY - 2019/1
Y1 - 2019/1
N2 - The first therapeutic application of messenger RNA (mRNA) was suggested more than two decades ago. However, its application was constrained by the ability of mRNA to activate the innate immune response, cytotoxicity, and poor potency. We and others recently demonstrated that these undesirable properties of mRNA may be overcome by alterating its structure. In this study, we developed a new chemically modified mRNA coding for BMP-2 with improved osteogenic features. To develop this new construct, we removed from the mRNA sequence the following undesirable elements: an upstream open reading frame in the 5′-untranslated region (UTR) and a polyadenylation element together with an AU-rich tract in the 3′UTR. In addition, a translation initiator of short UTRs (TISU) was introduced together with 5-iodo modified pyrimidine nucleotides. The new TISU BMP-2 chemically modified RNA (cmRNA) showed robust BMP-2 production in vitro in cell lines (HEK293 and MC3T3) and primary cells (muscle-derived mesenchymal stem cells). Stem cells additionally showed upregulation of osteogenic and angiogenic genes as a result of the TISU BMP-2 cmRNA transfection. The in vivo osteogenic properties of TISU BMP-2 cmRNA were explored in a critical-sized femoral defect in the rat. For this, the TISU BMP-2 cmRNA was loaded into collagen sponges to form transcript-activated matrices. Animals treated with TISU BMP-2 cmRNA showed superior bone formation that seemed to recapitulate endochondral ossification. The higher of the two doses examined in this model showed more robust new tissue formation. Finally, improved vascularization was detected in the healing area for animals treated with TISU BMP-2 cmRNA. The use of chemically modified RNA (cmRNA) with increased stability using translation initiator of short untranslated regions (TISU) offers the prospect of finally allowing us to unlock the potent osteogenic properties of BMP-2 in a clinically expedient manner. As noted, delivery of recombinant BMP-2 protein has had modest clinical efficacy, whereas gene delivery is effective but very difficult to translate into human clinical use. This study shows the great potential of cmRNA encoding BMP-2 with TISU in a long-bone critical-sized rat model.
AB - The first therapeutic application of messenger RNA (mRNA) was suggested more than two decades ago. However, its application was constrained by the ability of mRNA to activate the innate immune response, cytotoxicity, and poor potency. We and others recently demonstrated that these undesirable properties of mRNA may be overcome by alterating its structure. In this study, we developed a new chemically modified mRNA coding for BMP-2 with improved osteogenic features. To develop this new construct, we removed from the mRNA sequence the following undesirable elements: an upstream open reading frame in the 5′-untranslated region (UTR) and a polyadenylation element together with an AU-rich tract in the 3′UTR. In addition, a translation initiator of short UTRs (TISU) was introduced together with 5-iodo modified pyrimidine nucleotides. The new TISU BMP-2 chemically modified RNA (cmRNA) showed robust BMP-2 production in vitro in cell lines (HEK293 and MC3T3) and primary cells (muscle-derived mesenchymal stem cells). Stem cells additionally showed upregulation of osteogenic and angiogenic genes as a result of the TISU BMP-2 cmRNA transfection. The in vivo osteogenic properties of TISU BMP-2 cmRNA were explored in a critical-sized femoral defect in the rat. For this, the TISU BMP-2 cmRNA was loaded into collagen sponges to form transcript-activated matrices. Animals treated with TISU BMP-2 cmRNA showed superior bone formation that seemed to recapitulate endochondral ossification. The higher of the two doses examined in this model showed more robust new tissue formation. Finally, improved vascularization was detected in the healing area for animals treated with TISU BMP-2 cmRNA. The use of chemically modified RNA (cmRNA) with increased stability using translation initiator of short untranslated regions (TISU) offers the prospect of finally allowing us to unlock the potent osteogenic properties of BMP-2 in a clinically expedient manner. As noted, delivery of recombinant BMP-2 protein has had modest clinical efficacy, whereas gene delivery is effective but very difficult to translate into human clinical use. This study shows the great potential of cmRNA encoding BMP-2 with TISU in a long-bone critical-sized rat model.
KW - BMP-2
KW - bone healing
KW - mRNA
KW - modified mRNA
KW - transcript therapy
KW - vascularization
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U2 - 10.1089/ten.tea.2018.0112
DO - 10.1089/ten.tea.2018.0112
M3 - Article
C2 - 30009674
AN - SCOPUS:85060210835
SN - 1937-3341
VL - 25
SP - 131
EP - 144
JO - Tissue Engineering - Part A
JF - Tissue Engineering - Part A
IS - 1-2
ER -