Response to the Letter by Seropian et al
In response to the letter by Seropian et al, we wish to briefly emphasize some important points addressed in our original article.
Our study specifically focused on reparative angiogenesis in a mouse model of myocardial infarction (MI) by permanent coronary artery occlusion.1 We found that MI induces a marked upregulation of phosphoinositide-3 kinase (PI3K)γ in cardiomyocytes and endothelial cells of the periinfarct zone, associated with activation of Akt kinase activity and increased phosphorylation of glycogen synthase kinase-3β and endothelial nitric oxide synthase (eNOS). To ascertain the functional importance of PI3Kγ/Akt induction, we used AS605240, the most potent member of a new class of PI3Kγ-selective inhibitors recently introduced as powerful antiinflammatory agents. AS605240 was given before MI induction to simulate the condition of patients who experience an ischemic accident while under medication with such an agent. AS605240 strongly prevented the upregulation of PI3Kγ/Akt/eNOS signaling in infarcted hearts and also inhibited Pim-1, an essential downstream modulator of Akt-induced cardiomyocyte survival.2 These important results clearly show an adverse molecular action centered on PI3Kγ/Akt/Pim-1 and potentially deleterious for post-MI recovery.
Consistently, reparative neovascularization was drastically impaired in infarcted mice treated with AS605240, as well as in mice carrying a null or kinase-dead mutation of PI3Kγ, through a mechanism involving the Akt inhibition. Furthermore, both PI3Kγ pharmacological and genetic interference resulted in profound inhibition of monocytes/macrophages homing to the infarcted myocardium. After acute ischemia, the circulating monocyte fraction becomes enriched with proangiogenic progenitor cells. Previous studies showed that PI3Kγ-silenced human progenitor cells, as well as PI3Kγ-deficient murine bone marrow–derived proangiogenic cells, exhibit remarkably depressed migratory activity, reduced Akt and eNOS phosphorylation and decreased nitric oxide production, which jeopardize vasculogenesis in a model of peripheral ischemia.3,4 Altogether, these data confirm that interfering with spontaneous activation of PI3Kγ could have harmful consequences for postischemic tissue healing.
One puzzling feature of PI3Kγ kinase-dead mice is that the impairment of cardiac function inflicted by MI was less severe than that observed in PI3Kγ knockout or AS605240-treated mice.1 How can the different results in genetic and pharmacological models be reconciled? Different explanations are discussed in the article and can be reinforced here. First, it is possible that, in genetically modified animals, compensatory adaptation primed animals to overcome the loss of PI3Kγ function. Second, the inhibitor might have, in selected conditions, off-target effects toward p110α5 and/or distinct pharmacokinetic and pharmacodynamic profiles in different cell types. Third, the existence of kinase-dependent and -independent functions might complicate interpretation of knockout data. Nonetheless, it is important to note that, differently from what has been stated by Seropian et al, PI3Kγ kinase-dead mice did not show any statistically significant improvement versus wild-type controls and still exhibited the angiogenic defect.1
In contrast to a potential lack of selectivity of the AS605240, our study reports a series of additional controls where pharmacology and genetics provide converging results. For example, human endothelial cells transduced with an adenoviral construct expressing a small interfering (si)RNA against p110γ, the PI3Kγ catalytic subunit, showed markedly weakened angiogenic functions and increased apoptosis following hypoxia/starvation. AS605240 treatment of p110γ siRNA-transduced endothelial cells did not induce any further functional falloff, thus underscoring the compound specificity.1
With ischemic disease remaining a global burden, new potent and safe compounds targeting crucial mechanisms of repair are eagerly awaited. At present, PI3K inhibitors do not represent the best candidates for the treatment of MI or peripheral vascular disease, considering the proangiogenic and prosurvival action exerted by the PI3K/Akt signaling pathway in ischemia. The hypothesis that selective inhibitors might be endowed with a better therapeutic profile is attractive, yet remains to be validated. A recent study indicates that cardiomyocyte-specific knockout of PI3Kα is detrimental for cardiac function in a mouse MI model, while targeted PI3Kα overexpression is protective.6
While waiting for new chemical structures with improved selectivity profiles and devoid of the harmful effects shown by AS605240, caution is needed in the use of PI3K kinase “selective” or “global” inhibitors for the treatment of patients with ischemic disease.
Sources of Funding
Supported by the British Heart Foundation.
Original received June 1, 2010; revision received June 1, 2010; accepted June 4, 2010.
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