Gene therapy targeting survivin selectively induces pulmonary vascular apoptosis and reverses pulmonary arterial hypertension
M. Sean McMurtry, … , Lakshmi Puttagunta, Evangelos D. Michelakis
M. Sean McMurtry, … , Lakshmi Puttagunta, Evangelos D. Michelakis
Published June 1, 2005
Citation Information: J Clin Invest. 2005;115(6):1479-1491. https://doi.org/10.1172/JCI23203.
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Gene therapy targeting survivin selectively induces pulmonary vascular apoptosis and reverses pulmonary arterial hypertension

Abstract

Pulmonary arterial hypertension (PAH) is characterized by genetic and acquired abnormalities that suppress apoptosis and enhance cell proliferation in the vascular wall, including downregulation of the bone morphogenetic protein axis and voltage-gated K+ (Kv) channels. Survivin is an “inhibitor of apoptosis” protein, previously thought to be expressed primarily in cancer cells. We found that survivin was expressed in the pulmonary arteries (PAs) of 6 patients with PAH and rats with monocrotaline-induced PAH, but not in the PAs of 3 patients and rats without PAH. Gene therapy with inhalation of an adenovirus carrying a phosphorylation-deficient survivin mutant with dominant-negative properties reversed established monocrotaline-induced PAH and prolonged survival by 25%. The survivin mutant lowered pulmonary vascular resistance, RV hypertrophy, and PA medial hypertrophy. Both in vitro and in vivo, inhibition of survivin induced PA smooth muscle cell apoptosis, decreased proliferation, depolarized mitochondria, caused efflux of cytochrome c in the cytoplasm and translocation of apoptosis-inducing factor into the nucleus, and increased Kv channel current; the opposite effects were observed with gene transfer of WT survivin, both in vivo and in vitro. Inhibition of the inappropriate expression of survivin that accompanies human and experimental PAH is a novel therapeutic strategy that acts by inducing vascular mitochondria-dependent apoptosis.

Authors

M. Sean McMurtry, Stephen L. Archer, Dario C. Altieri, Sebastien Bonnet, Alois Haromy, Gwyneth Harry, Sandra Bonnet, Lakshmi Puttagunta, Evangelos D. Michelakis

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Figure 5

Selective expression of survivin mutant in resistance PAs causes an increase in PASMC outward K+ current.

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Selective expression of survivin mutant in resistance PAs causes an incr...
(A) In FBS-rich conditions (10% FBS in the medium, a condition known to increase endogenous survivin), infection with Ad-GFP-S-M causes augmentation of K+ currents and decreased capacitance (Cm, a measure of cell size), consistent with apoptosis; the opposite is seen with Ad-GFP-S infection. In contrast, in serum-deprived conditions (0.1% FBS), infection with Ad-GFP-S causes a decrease in K+ currents, consistent with apoptosis resistance. Since in these conditions endogenous survivin is absent, infection with Ad-GFP-S-M has no effect on K+ current. Cells carrying the transgenes were selected by the green fluorescence. Mean data for current density over voltage are shown on the right (n = 6 cells per group; *P < 0.01 vs. control). (B) Both GFP immunofluorescence microscopy and quantitative RT-PCR of laser-capture-microdissected resistance PAs demonstrate efficient delivery of the transgenes, particularly to the very small (less than 50 μm) resistance PAs (arrows). (C) In our inhaled gene therapy approach, the expression of the transgenes is restricted to the lungs, as shown by the expression of GFP, measured by quantitative RT-PCR. The expression of endogenous survivin in nontreated rats is minimal in all organs studied, except the spleen. Our WT-survivin primer also detects the survivin mutant, as shown by the increased lung signal in the treated rats. Data from 5 rats per group are shown.