In partial fulfillment of the requirements for the degree of
Doctor of Philosophy in Bioinformatics
in the Department of Biomedical Engineering
Anastasia Zhurikhina
Defends her thesis:
Integrative computational-experimental study of the biomechanics of Cerebral Cavernous Malformation and its pharmacological rescue
Thursday, August 15, 2024
2:00pm Eastern Time
In-person location = EBB Krone 3029
Virtual = https://gatech.zoom.us/j/93279427638?pwd=zej5sA6jkOVQEjUDyuDvSRF1ZVAA2s.1
Meeting ID: 932 7942 7638
Passcode: 361141
Thesis Advisor:
Dr. Denis Tsygankov, Department of Biomedical Engineering
Committee Members:
Dr. Mark Borodovsky, School of Computational Sciences & Engineering, Department of Biomedical Engineering
Dr. Cheng Zhu, Department of Biomedical Engineering
Dr. Eberhard Voit, Department of Biomedical Engineering
Dr. Melissa Kemp, Department of Biomedical Engineering
Abstract:
Cerebral Cavernous Malformations (CCMs) are vascular abnormalities in the brain and spinal cord characterized by clusters of dilated capillaries with thin, fragile walls prone to leaking. These vascular lesions can lead to critical neurological issues such as seizures, focal neurological deficits, and potentially fatal brain hemorrhages. Current treatment options are predominantly surgical, involving the removal of the accessible regions of the affected tissue. However, surgery carries significant risks, especially for deep-seated lesions in critical brain areas. Moreover, not all patients are suitable candidates for surgery, and those who undergo surgery can develop new lesions over time. Despite advances in medical research, there remains a pressing need for non-surgical treatment options that can effectively manage or cure CCM disease.
Identifying pharmaceutical strategies is crucial for providing safer, more accessible treatments that can address the underlying molecular and cellular causes of CCMs, prevent lesion progression, and improve the quality of life for affected individuals. This thesis aims to unveil the dysregulations of molecular pathways and biomechanical deficiencies in cell function, resulting from CCM mutations and suggests two small-molecular inhibitors as novel pharmacological agents for the disease intervention.
To this end, we developed a multi-scale computational model to reproduce healthy, diseased, pharmacologically rescued endothelial cell behaviors during vascular network formation, accounting for the viscoelastic properties of cell bodies, mechanosensing, dynamics of cellular protrusions, and response of cell contacts to mechanical loading. The model replicates distinct multicellular patterns observed in wild-type and CCM knockdown cultures, revealing the critical role of the balance between cell-cell and cell-extracellular matrix (ECM) interactions in maintaining vascular integrity. Experimental validation using the tube formation assay confirmed the model's predictions, showing that CCM1 knockdown disturbs the balance by affecting mainly cell-ECM adhesion, while CCM3 knockdown disrupts the balance by significantly weakening cell-cell contacts.
Our RNA-seq analysis of CCM phenotypes identified significant changes in gene expression profiles, particularly in genes related to cell adhesion, ECM organization, and cytoskeletal regulation. The key targets identified by the analysis include genes associated with the WNT signaling pathway, such as AKT2 and GNAI2. To validate these targets, we cross-referenced our findings with publicly available datasets from CCM patient samples and CCM3-deficient mouse models. This comparative analysis confirmed the relevance of the WNT pathway and the involvement of AKT2 and GNAI2 in CCM pathology.
Finally, we employed pharmacological inhibitors to rescue the severely disrupted patterns in CCM1 and CCM3 knockdown cultures. LY20909314, a GSK3B inhibitor similar in its action to AKT2, and XAV939, a Tankyrase inhibitor that stabilizes Axin similar to GNAI2, were used to rescue both CCM phenotypes. These inhibitors demonstrated a high efficiency in restoring normal (wild-type) collective behavior of the CCM1 and CCM3 knockdown cells.
In conclusion, this thesis provides a comprehensive mechanobiological framework for understanding the molecular and cellular mechanisms driving CCM pathology. By integrating computational modeling, RNA-seq analysis, and experimental validation, we identified novel therapeutic candidates and demonstrated their efficiency in cell cultures. Our findings suggest that targeting CCM rescue through WNT signaling modulation may offer effective strategies for treating CCM, ultimately improving patient outcomes and reducing reliance on surgical interventions.
