Chemical Dimerization of Fibroblast Growth Factor Receptor-1 Induces Myoblast Proliferation, Increases Intracardiac Graft Size, and Reduces Ventricular Dilation in Infarcted Hearts

Abstract

The ability to control proliferation of grafted cells in the heart and consequent graft size could dramatically improve the efficacy of cell therapies for cardiac repair. To achieve targeted graft cell proliferation, we created a chimeric receptor (F36Vfgfr-1) composed of a modified FK506-binding protein (F36V) fused with the cytoplasmic domain of the fibroblast growth factor receptor-1 (FGFR-1). We retrovirally transduced mouse C2C12 and MM14 skeletal myoblasts with this construct and treated them with AP20187, a dimeric F36V ligand ("dimerizer"), in vitro and in vivo to induce receptor dimerization. Dimerizer treatment in vitro activated the mitogen-activated protein kinase pathway and induced proliferation in myoblasts expressing F36Vfgfr-1 comparable with the effects of basic FGF. Wild-type myoblasts did not respond to dimerizer. Subcutaneous grafts composed of myoblasts expressing F36Vfgfr-1 showed a dose-dependent increase in DNA synthesis with dimerizer treatment. When myoblasts expressing F36Vfgfr-1 were injected into infarcted hearts of nude mice, dimerizer treatment resulted in a dose-dependent increase in graft size, from 20 +/- 3 to 42.9 +/- 4.3% of the left ventricle. Blinded echocardiographic analysis demonstrated that larger graft size was associated with a dose-dependent reduction in ventricular dilation after myocardial infarction, although animals with the largest grafts showed an increased incidence of ventricular tachycardia. Thus, selective proliferation of genetically modified graft cells can be induced with a systemically administered synthetic molecule in vitro or in vivo. Control of intramyocardial graft size by this approach may allow optimization of cell-based therapy to obtain desired cardiac function postinfarction.

Systems approaches to preventing transplanted cell death in cardiac repair

“…Several conditions could explain this. First, C2C12 cells are particularly ischemia-resistant; large grafts have been produced in mouse hearts infarcted by permanent coronary artery occlusion [106] without any pro-survival interventions. Additionally, hESC-derived cardiomyocyte viability by Trypan blue exclusion after long periods on ice is lower than C2C12s (60% after 5 hours versus 90% for C2C12s), and plating efficiency of the human cardiomyocytes at the end of a grafting experiment is correspondingly lower.…”

Selective control of endothelial cell proliferation with a synthetic dimerizer of FGF receptor-1

“…Studies of myoblasts transduced with our F36Vfgfr-1 construct grafted into infarcted hearts recently showed larger grafts and improvement in cardiac function following dimerizer administration, demonstrating that this system can be used to regulate therapeutic cell proliferation in vivo. 19 Future in vivo studies with F36Vfgfr-1 endothelial cells will test whether angiogenesis in ischemic tissues can be controlled with the dimerizer system.…”

“…24 Dimerization of FGF receptor-1 (FGFR1) in skeletal myoblasts induced their proliferation and prevented differentiation similar to bFGF. 16,19 Cell proliferation via chemically induced FGF receptor dimerization has also been shown in mammary ductal epithelial cells 25 and prostate cancer cells. 26 In the present study, we describe a method to selectively induce FGFR1 signaling in human endothelial cells.…”

“…In a system that has been described for a number of different cell/receptor combinations, the cytoplasmic signaling domain of the growth factor receptor of interest is fused to a modified drugbinding domain. [14][15][16][17][18][19][20][21] Cells expressing this chimeric receptor are then exposed to a small synthetic bifunctional ligand (dimerizer), which binds to the drug-binding domains of two receptor molecules. The growth factor signaling domains are thus brought into close proximity, mimicking the effect of endogenous receptor activation.…”

FGFR1 (Fibroblast Growth Factor Receptor 1)

“…Myoblasts expressing a chimeric Fgfr1 were injected into infarcted hearts of mice. The treatment induced an increase in graft size, showing that selective proliferation can be induced (Stevens et al, 2007). Oncogenesis FGFR1 over-expression was detected in 2 out of 2 embryonal rhabdomyosarcomas, 6 out of 6 alveolar rhabdomyosarcomas, and in the one pleomorphic adult variant tested, as well as in the RMS cell lines tested.…”