Biology subjects

Sheehan, J.

Publications and source records attributed to Sheehan, J..

3 recordsLinked to original sources

bioRxiv2024

Focused Ultrasound Blood-Brain Barrier Opening Arrests the Growth and Formation of Cerebral Cavernous Malformations

BACKGROUNDCerebral cavernous malformations (CCM) are vascular lesions within the central nervous system, consisting of dilated and hemorrhage-prone capillaries. CCMs can cause debilitating neurological symptoms, and surgical excision or stereotactic radiosurgery are the only current treatment options. Meanwhile, transient blood-brain barrier opening (BBBO) with focused ultrasound (FUS) and microbubbles is now understood to exert potentially beneficial bioeffects, such as stimulation of neurogenesis and clearance of amyloid-{beta}. Here, we tested whether FUS BBBO could be deployed therapeutically to control CCM formation and progression in a clinically-representative murine model. METHODSCCMs were induced in mice by postnatal, endothelial-specific Krit1 ablation. FUS was applied for BBBO with fixed peak-negative pressures (PNPs; 0.2-0.6 MPa) or passive cavitation detection-modulated PNPs. Magnetic resonance imaging (MRI) was used to target FUS treatments, evaluate safety, and measure longitudinal changes in CCM growth after BBBO. RESULTSFUS BBBO elicited gadolinium accumulation primarily at the perilesional boundaries of CCMs, rather than lesion cores. Passive cavitation detection and gadolinium contrast enhancement were comparable in CCM and wild-type mice, indicating that Krit1 ablation does not confer differential sensitivity to FUS BBBO. Acutely, CCMs exposed to FUS BBBO remained structurally stable, with no signs of hemorrhage. Longitudinal MRI revealed that FUS BBBO halted the growth of 94% of CCMs treated in the study. At 1 month, FUS BBBO-treated lesions lost, on average, 9% of their pre-sonication volume. In contrast, non-sonicated control lesions grew to 670% of their initial volume. Lesion control with FUS BBBO was accompanied by a marked reduction in the area and mesenchymal appearance of Krit mutant endothelium. Strikingly, in mice receiving multiple BBBO treatments with fixed PNPs, de novo CCM formation was significantly reduced by 81%. Mock treatment plans on MRIs of patients with surgically inaccessible lesions revealed their lesions are amenable to FUS BBBO with current clinical technology. CONCLUSIONSOur results establish FUS BBBO as a novel, non-invasive modality that can safely arrest murine CCM growth and prevent their de novo formation. As an incisionless, MR image-guided therapy with the ability to target eloquent brain locations, FUS BBBO offers an unparalleled potential to revolutionize the therapeutic experience and enhance the accessibility of treatments for CCM patients.

bioengineering↗

bioRxiv2023

Molecular mechanisms of functional impairment for active site mutations in glucose-6-phosphatase catalytic subunit 1 linked to glycogen storage disease type 1a.

Mediating the terminal reaction of gluconeogenesis and glycogenolysis, the integral membrane protein G6PC1 regulates hepatic glucose production by catalyzing hydrolysis of glucose-6-phosphate (G6P) within the lumen of the endoplasmic reticulum. Consistent with its vital contribution to glucose homeostasis, inactivating mutations in G6PC1 cause glycogen storage disease (GSD) type 1a characterized by hepatomegaly and severe hypoglycemia. Despite its physiological importance, the structural basis of G6P binding to G6PC1 and the molecular disruptions induced by missense mutations within the active site that give rise to GSD type 1a are unknown. Exploiting a computational model of G6PC1 derived from the groundbreaking structure prediction algorithm AlphaFold2 (AF2), we combine molecular dynamics (MD) simulations and computational predictions of thermodynamic stability with a robust in vitro screening platform to define the atomic interactions governing G6P binding as well as explore the energetic perturbations imposed by disease-linked variants. We identify a collection of side chains, including conserved residues from the signature phosphatidic acid phosphatase motif, that contribute to a hydrogen bonding and van der Waals network stabilizing G6P in the active site. Introduction of GSD type 1a mutations into the G6PC1 sequence elicits changes in G6P binding energy, thermostability and structural properties, suggesting multiple pathways of catalytic impairment. Our results, which corroborate the high quality of the AF2 model as a guide for experimental design and to interpret outcomes, not only confirm active site structural organization but also suggest novel mechanistic contributions of catalytic and non-catalytic side chains.

biophysics↗

bioRxiv2017

Comprehensive Analysis of Constraint on the Spatial Distribution of Missense Variants in Human Protein Structures

The spatial distribution of genetic variation within proteins is shaped by evolutionary constraint and thus can provide insights into the functional importance of protein regions and the potential pathogenicity of protein alterations. Here, we comprehensively evaluate the 3D spatial patterns of constraint on human germline and somatic variation in 4,568 solved protein structures. Different classes of coding variants have significantly different spatial distributions. Neutral missense variants exhibit a range of 3D constraint patterns, with a general trend of spatial dispersion driven by constraint on core residues. In contrast, germline and somatic disease-causing variants are significantly more likely to be clustered in protein structure space. We demonstrate that this difference in the spatial distributions of disease-associated and benign germline variants provides a signature for accurately classifying variants of unknown significance (VUS) that is complementary to current approaches for VUS classification. We further illustrate the clinical utility of our approach by classifying new mutations identified from patients with familial idiopathic pneumonia (FIP) that segregate with disease.

genetics↗