Antitumor effects of in vivo caveolin gene delivery are associated with the inhibition of the proangiogenic and vasodilatory effects of nitric oxide

Abstract

In tumors, caveolin-1, the structural protein of caveolae, constitutes a key switch through its function as a tumor suppressor and a promoter of metastases. In endothelial cells (EC), caveolin is also known to directly interact with the endothelial nitric oxide synthase (eNOS) and thereby to modulate nitric oxide (NO)-mediated processes including vasodilation and angiogenesis. In this study, we examined whether the modulation of the stoichiometry of the caveolin/eNOS complex in EC lining tumor blood vessels could affect the tumor vasculature and consecutively tumor growth. For this purpose, we used cationic lipids, which are delivery systems effective at targeting tumor vs. normal vascular networks. We first documented that in vitro caveolin transfection led to the inhibition of both VEGF-induced EC migration and tube formation on Matrigel. The DNA-lipocomplex was then administered through the tail vein of tumor-bearing mice. The direct interaction between recombinant caveolin and native eNOS was validated in coimmunoprecipitation experiments from tumor extracts. A dramatic tumor growth delay was observed in mice transfected with caveolin- vs. sham-transfected animals. Using laser Doppler imaging and microprobes, we found that in the early time after lipofection (e.g., when macroscopic effects on the integrity of the tumor vasculature were not detectable), caveolin expression impaired NO-dependent tumor blood flow. At later stages post-transfection, a decrease in tumor microvessel density in the central core of caveolin-transfected tumors was also documented. In conclusion, our study reveals that by exploiting the exquisite regulatory interaction between eNOS and caveolin and the propensity of cationic lipids to target EC lining tumor blood vessels, caveolin plasmid delivery appears to be a safe and efficient way to block neoangiogenesis and vascular function in solid tumors, independently of any direct effects on tumor cells.

Pathogenic angiogenesis in IBD and experimental colitis: new ideas and therapeutic avenues

“…Additionally, Sonveaux et al (142) showed that Cav1 Ϫ/Ϫ mouse cells exhibit poor tube formation in cultured endothelium (32). Cav-1 expression has been shown to prevent NO production in response to VEGF-A activation of VEGFR2 by acting as a competitive inhibitor of eNOS (24,63). This is important for pathological angiogenesis, because NO has an inhibitory effect on leukocyte recruitment through downregulation of P-selectin, as a mechanism of decreasing inflammatory responses (2).…”

Altered Angiogenesis in Caveolin-1 Gene–Deficient Mice Is Restored by Ablation of Endothelial Nitric Oxide Synthase

“…Overexpressing caveolin-1 in cultured EC decreased their migration and in vitro capillary-like tube formation. 12 In vivo caveolin-1 gene delivery to angiogenic EC resulted in EC caveolin-1 overexpression and inhibition of angiogenesis in wild-type mice. 12 In mouse models where angiogenesis was stimulated by ischemia or an adenovirus encoding vascular endothelium growth factor (VEGF), endothelial-specific transgenesis of the caveolin-1 gene decreased neovascularization.…”

“…12 In vivo caveolin-1 gene delivery to angiogenic EC resulted in EC caveolin-1 overexpression and inhibition of angiogenesis in wild-type mice. 12 In mouse models where angiogenesis was stimulated by ischemia or an adenovirus encoding vascular endothelium growth factor (VEGF), endothelial-specific transgenesis of the caveolin-1 gene decreased neovascularization. 13 In agreement with these results, siRNA-mediated down-regulation of caveolin-1 increased EC transmigration in vitro subjected to angiogenic conditions.…”

“…30,31 Therefore, caveolin-1 is a tonic inhibitor of eNOS but is required for caveola formation, which in turn is necessary for signaling cascades leading to eNOS activation and NO production. The direct inhibitory effect of caveolin-1 on eNOS has been taken advantage of by two in vivo approaches, both successful, to inhibit tumor angiogenesis: overexpression of caveolin-1 in angiogenic EC, 12 and delivery of a peptide comprising the domain of caveolin-1 responsible for eNOS inhibition. 8 In agreement with caveolin-1 being a tonic inhibitor of eNOS, caveolin-1-null mice display excessive NO production as a result of persistent eNOS activation.…”

Caveolin-1 Is Critical for the Maturation of Tumor Blood Vessels through the Regulation of Both Endothelial Tube Formation and Mural Cell Recruitment

“…In the cardiovascular context, we and others have previously documented that the recombinant expression of caveolin 12 or the administration of caveolin-derived peptides 13 exert anti-angiogenic effects through the inhibition of the endothelial nitric-oxide synthase (eNOS). Caveolin has also been shown to co-purify with the VEGF-R2 receptor in caveolae and thereby to modulate VEGF signaling.…”