Inherited Metabolic Disorders

Our research group studies rare genetic disorders which are the result of non-functional proteins that lead to the accumulation of substrates that are unable to be processed or degraded properly. We focus on two disorders, Niemann-Pick type C disease (NPC) and Glycogen Storage Disorder Type III (GSD III). To address the errors in substrate processing and degradation we are investigating protein homeostasis and mRNA replacement as two strategies to restore protein function in these monogenic disorders. Functional restoration of the aberrant proteins would allow for proper substrate processing and clearance, mitigating disease phenotypes.

Niemann-Pick type C1

Niemann-Pick type C1 is caused by variants in the NPC1 gene that mainly lead to the NPC1 protein misfolding. NPC1 is involved in exporting unesterified cholesterol from late endosomes and lysosomes. Lack of NPC1 protein function causes an accumulation of unesterified cholesterol mainly in the liver and brain. The most common disease-causing variant is an isoleucine-to-threonine substitution at position 1061 (I1061T). I1061T misfolds in the endoplasmic reticulum and is rapidly degraded by the proteosome. Importantly, I1061T is functional if it escapes proteasomal degradation and is trafficked to the lysosome, creating another avenue for therapeutic targeting being pursued by our lab.

Our group was the first to identify differences in protein trafficking between mouse and human NPC1. Interestingly, these differences were observed despite mouse and human proteins sharing 86% identity and 93% similarity. Importantly, we also demonstrated that these differences drive differential therapeutic response. We hypothesize that previous efforts to use proteostatic modulators as therapeutic agents for Niemann-Pick type C disease have been confounded by these species-specific differences in protein trafficking. We are actively investigating what are the molecular and regulatory pathways causing these differences.

An avenue of interest to begin parsing apart the species-specific differences are the glycosylation patterns of NPC1 between humans and mice. Glycans are topological regulators of protein folding, trafficking, and function. NPC1 is a heavily glycosylated protein however, little is known about how glycans impact NPC1 folding, trafficking, and degradation. As we have seen comparing mouse and human NPC1, differential glycosylation can drive differential protein trafficking and therapeutic response. 

As glycosylation can drive protein trafficking, an important part of this mechanism is the degradation pathway and how misfolded proteins are delt with. Our group was the first to report that I1061T-NPC1, the most common disease variant, is degraded by both MARCH6-dependent ERAD as well as FAM134B-dependent ER-phagy. Interestingly, FAM134B exists in multiple forms with different forms being predominant in hepatocytes compared to neurons. This observation provides further opportunities to explore tissue-specific degradation pathways. In order to address these questions, we have optimized protocols for differentiating iPSCs into both neurons and hepatic progenitor cells.

Glycogen Storage Disorder Type III

Glycogen Storage Disorder III is caused by a wide variety of loss of function variants in the gene AGL, which encodes the glycogen debranching enzyme (GDE). This enzyme is involved in the degradation of cytosolic glycogen in the cell. Without functional GDE, there is an accumulation of abnormally structured glycogen, mainly in the liver and skeletal muscle. Dietary intervention is the only strategy currently available to manage symptoms of GSD III but does not prevent long-term organ damage from occurring. 

The preliminary focus has been developing and characterizing relevant disease models for GSD III. We started by investigating the banked patient fibroblasts which were labeled as GSD III but lacked genomic sequencing. Our group was able to identify a misclassified GSD III patient fibroblast line highlighting the importance of characterizing disease models before using them in research. In addition to the patient fibroblast lines, we created an immortalized GSD III fibroblast line using CRISPR CAS9 to insert small deletions in AGL. This provides us with an isogenic cellular model to study GSD III. We recently published a paper on this work highlighting the importance of establishing well characterized cellular models and how patient fibroblast lines are a valuable tool to study diseases in a clinically relevant way, while immortalized fibroblasts provide cell lines to study the disease without confounding effects of genetic variation.

In addition to fibroblast models our lab is also using iPSCs to develop contracting myotubes to study GSD III in vitro. This model provides our lab with a rapid way to begin investigating the myopathy phenotype of GSD III in a more tissue specific manner.

Finally, the overall goal for investigating GSD III is to provide a viable therapeutic option for individuals with GSD III with an mRNA therapeutic. Work is going into investigating muscle-targeted delivery and the half-life of the protein to optimize mRNA delivery.