MicroRNAs (miRNA) are single-stranded RNA molecules of 21-23 nucleotides length which down-regulate gene expression by annealing with the 3'UTR of target mRNAs, repressing translation and inducing mRNA degradation and de-adenylation. The imprecise miRNA recognition of their mRNA targets allows post-transcriptional regulation over hundreds of target mRNA during homeostasis or in response to stimuli. The microRNA miR-126 represents the most abundant endothelial miRNA upon expression profiling, and is expressed in a pan-endothelial fashion during embryogenesis. We have created knockout mice lacking miR-126 which exhibit 50% embryonic lethality associated with edema, hemorrhage, and angiogenic delay. In surviving miR-126 ko mice, adult angiogenesis is delayed for instance in corneal micropocket assays. Interestingly, miR-126 is present in intron 7 of a host gene, Egfl7. The miR-126 ko phenotype actually recapitulates previously described Egfl7 ko phenotypes, and previously described Egfl7 ko mice are now understood to have inadvertently disrupted miR-126 expression. An essential role for a miRNA during tumor angiogenesis has not been previously demonstrated. The current application explores the role of miR-126 in tumor angiogenesis based upon strong miR-126/Egfl7 expression in tumor endothelium and preliminary data in the MMTV- PyMT transgenic model of breast cancer.
Aim 1 investigates whether constitutive genetic deletion of miR-126 inhibits tumor progression and angiogenesis and extends survival in the MMTV-PyMT model.
Aim 2 evaluates the therapeutic potential of miR-126 inhibition through temporally conditional miR-126 ko with our floxed mouse allele in pre-established MMTV-PyMT tumors. Conditional miR-126 ko will serve as a reference standard for pharmacologic miR-126 inhibition in the MMTV-PyMT model via both antagomirs, as well as a novel 2'-F/methoxyester anti-miR chemistry. Further, miR-126 inhibition by either genetic deletion or antagomir/anti-mir treatment will be compared and combined with VEGF inhibition. Finally, Aim 3 investigates mechanisms of miR-126 action during breast tumorigenesis through compartment-specific deletion in endothelium, in vitro characterization of miR-126 ko endothelium, and endothelial tip- and stalk cell phenotypes. Overall, these investigations utilize complementary approaches of rigorously characterized miR-126 genetic knockout mice and novel antagomir and 2'-F/methoxyester anti-miR therapeutic strategies to explore the first functional linkages between an endothelial microRNA and tumor angiogenesis. The demonstration of a functional requirement for miR-126 during tumor angiogenesis and progression would have significant implications for design of future anti-angiogenic therapies.

Public Health Relevance

The inhibition of tumor blood vessels, termed anti-angiogenic therapy, is a promising new modality for cancer therapy. The majority of current anti-angiogenic therapies target a blood vessel growth factor termed Vascular Endothelial Growth Factor. Here, we explore the relevance of a RNA molecule, miR-126, as a novel factor regulating tumor blood vessels, with implications for development of new anti-angiogenic therapies.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA158528-04
Application #
8620617
Study Section
Tumor Progression and Metastasis Study Section (TPM)
Program Officer
Mohla, Suresh
Project Start
2011-03-04
Project End
2016-02-28
Budget Start
2014-03-01
Budget End
2015-02-28
Support Year
4
Fiscal Year
2014
Total Cost
$351,301
Indirect Cost
$136,606
Name
Stanford University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
009214214
City
Stanford
State
CA
Country
United States
Zip Code
94305
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