# Phenotyping, Human Tissue and Biomarkers Core

> **NIH NIH P01** · DUKE UNIVERSITY · 2021 · $313,270

## Abstract

The cerebral cavernous malformation (CCM) is a common hemorrhagic vascular anomaly, caused by germ line
and/or endothelial somatic mutations in three known genes. It affects more than a million Americans,
predisposing them to a lifetime risk of stroke and epilepsy. A lesion is alarmingly more likely to rebleed after a
prior symptomatic hemorrhage, and there is currently no therapy to prevent the genesis or clinical progression
of lesions. In the past 5 years, our program project has clarified key mechanisms of CCM pathogenesis through
exceptional collaboration and synergy among four laboratories. These included the identification of
MEKK3/KLF2/4 signaling as a primary trigger of lesion development in the setting of Ccm loss, downstream
activation of RhoA kinase (ROCK), and roles of microbiome driven TLR4/CD14 signaling, intestinal mucin loss,
hypoxia, dietary supplements, thrombospondin-1, thrombomodulin and clonal expansion of mutated
endothelium. The Scientific Core at U Chicago optimized the high throughput assessment of lesion burden using
micro-CT, and the quantification of chronic hemorrhage using densitometry of non-heme iron Perls staining. We
developed familiarity with data structure for optimal sample sizes and statistics, and a discipline of prospective
articulation of outcomes and blinding in the various experiments. Protocols were harmonized, and models,
specimens and data were shared across projects and sites. The Core produced transcriptomic libraries of
mutated cells, C. elegans, and neurovascular units from human and murine lesions with various genotypes and
at different stages of development. We streamlined the collection and distribution of genotyped human CCM
lesions from excised surgical specimens to the project sites. And we assessed the peripheral blood of human
subjects for protein levels and microRNAs related to the signaling aberrations. Yet critical knowledge gaps
remain, as pilot data has motivated hypotheses about the roles of Ccm deficient endothelium in lesion genesis
versus maturation, PIK3 signaling, ADAMTS proteolysis of versican, and activated protein C anticoagulant and
cytoprotective pathways. Core services shall continue, further enhanced by new techniques aimed at facilitating
single-cell RNA and DNA analyses from lesions, morphometric assessment of acute hemorrhage (as a clinically
relevant phenotypic feature distinct from chronic bleeding), and micro-sampling and processing for plasma
biomarker discovery in mice. Each of the projects will be assisted by and take advantage of the proposed
Phenotyping, Human Tissue and Biomarkers Core, aimed at (1) phenotype assessment and human lesion
dissection, and (2) biomarker discovery and validation. The new hypotheses being probed will identify novel
therapies, and the resulting biomarkers will facilitate risk stratification and disease monitoring in patients. In a
strategic sense, the proposed Core will continue to enhance collaboration and synergy among the project...

## Key facts

- **NIH application ID:** 10220144
- **Project number:** 5P01NS092521-07
- **Recipient organization:** DUKE UNIVERSITY
- **Principal Investigator:** ISSAM A AWAD
- **Activity code:** P01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $313,270
- **Award type:** 5
- **Project period:** 2015-09-01 → 2025-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10220144

## Citation

> US National Institutes of Health, RePORTER application 10220144, Phenotyping, Human Tissue and Biomarkers Core (5P01NS092521-07). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10220144. Licensed CC0.

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