Requisite endothelial reactivation and effective siRNA nanoparticle targeting of Etv2/Er71 in tumor angiogenesis

Abstract

Angiogenesis, new blood vessel formation from preexisting vessels, is critical for solid tumor growth. As such, there have been efforts to inhibit angiogenesis as a means to obstruct tumor growth. However, antiangiogenic therapy faces major challenges to the selective targeting of tumor-associated-vessels, as current antiangiogenic targets also disrupt steady-state vessels. Here, we demonstrate that the developmentally critical transcription factor Etv2 is selectively upregulated in both human and mouse tumor-associated endothelial cells (TAECs) and is required for tumor angiogenesis. Two-photon imaging revealed that Etv2-deficient tumor-associated vasculature remained similar to that of steady-state vessels. Etv2-deficient TAECs displayed decreased Flk1 (also known as Vegfr2) expression, FLK1 activation, and proliferation. Endothelial tube formation, proliferation, and sprouting response to VEGF, but not to FGF2, was reduced in Etv2-deficient ECs. ROS activated Etv2 expression in ECs, and ROS blockade inhibited Etv2 expression in TAECs in vivo. Systemic administration of Etv2 siRNA nanoparticles potently inhibited tumor growth and angiogenesis without cardiovascular side effects. These studies highlight a link among vascular oxidative stress, Etv2 expression, and VEGF response that is critical for tumor angiogenesis. Targeting the ETV2 pathway might offer a unique opportunity for more selective antiangiogenic therapies.

ETV2/ER71 Transcription Factor as a Therapeutic Vehicle for Cardiovascular Disease

“…Mechanistically, ETV2 was able to activate critical genes for vessel development and angiogenesis, such as Flk1, Cdh5 and Vegfa/b/c, as seen in developing embryos. Similarly, upregulated level of Etv2 in tumor associated endothelial cells (TAECs) has also been reported in a previous study 48. The authors demonstrated that the deletion of Etv2 in endothelial cells or systemic delivery of si Etv2 into tumor bearing mice impairs tumorigenesis and angiogenesis.…”

“…As discussed, ETV2 is specifically expressed in the vasculatures within a narrow developmental window between E8.5 and E10.5. In accordance with the transient expression pattern of ETV2, mice deficient in endothelial Etv2 (i.e., Tie2-Cre or Cdh5-Cre;Etv2 floxed/floxed mice ) are born alive and develop normal vascular structures 48, 49, suggesting that ETV2 is dispensable for steady-state vessel formation. There is a prevailing notion that embryonic events or signaling pathways become critical for the development of diseases or pathophysiological events in adults.…”

NF-κB Inhibition Suppresses Experimental Melanoma Lung Metastasis

“…To that end, we have designed a flexible siRNA nanoplex delivery system comprising an amphipathic peptide (“p5RHH”) capable of condensing siRNA into a stable 55 nm nanoparticle and preclinical validation studies have demonstrated that this peptide-based delivery platform is efficacious in atherosclerosis [ 18 ], necrotizing enterocolitis [ 19 ], pancreatic cancer [ 20 ], cancer angiogenesis [ 21 ], ovarian and uterine cancer [ 22 ], osteoarthritis [ 23 , 24 ], and rheumatoid arthritis [ 25 ]. Moreover, NF-κB suppression in inflamed targeted tissues remains localized and does not disrupt important signaling actions of NF-κB in off-target tissues/organs [ 25 ].…”

Precision delivery of RAS-inhibiting siRNA to KRAS driven cancer via peptide-based nanoparticles

“…In addition to the noted potential advantages in drug delivery, prior polymer and lipid based NP constructs are prone to cause generation of reactive oxygen species and calcium leakage, leading to off target effects [26–28, 52]. Our peptide based construct is predominantly cleared renally and has been found to have minimal off target effects both based on our data and our group’s prior work [26, 27, 29, 34]. One issue with prior peptide NPs had been endosomal uptake and sequestration, but our melittin derived p5RHH NP construct is by design endosomolytic and rapidly delivers siRNA to the cytoplasm of the target cell (Figure 1) [26].…”

“…Due to the use of CO2 euthanasia, serum potassium could not accurately be measured. The effect of our NP formulation on cardiac function was not assessed, as our p5RHH based NP has previously been shown to have no adverse effects on murine cardiac function in a model of atherosclerosis [34]. Finally, in studies looking at our p5RHH based NP in murine arthritis and atherosclerosis, after serial IV dosing, there was no activation of the innate or adaptive immune responses: namely, 1) no IgG or IgM to the intact peptide-siRNA NP or peptide itself; 2) no suppression of innate immune responsivity by splenocytes (anti-CD3–activated CD4 + T cells secreting normal levels of TNF-α, IFN-γ, IL-6, and IL-10); 3) no change in splenocyte subpopulations (CD4 + and CD8 + T cells, CD19 + B cells, NK1.1 + natural killer cells, and Foxp3 + T regulatory cells); and 4) no activation of complement (C3a, 5a) [27, 28, 35].…”