Mulibrey Nanism (MUL) Cell Models for Research

Mulibrey Nanism (MUL) is a rare autosomal recessive disorder with an estimated incidence of 1 in 50,000 to 1 in 100,000 worldwide, though higher prevalence is observed in Finland (1 in 10,000). The condition is caused by mutations in the TRIM37 gene, leading to growth retardation, distinctive facial features, pericardial constriction, and increased risk of tumors, particularly Wilms tumor and hepatocellular carcinoma. According to the National Cancer Institute (NCI), the overall survival of MUL patients is reduced, with a 5-year survival of approximately 80% in those without malignancies, but significantly lower in those with tumors. The disease burden is substantial due to chronic cardiac and metabolic complications, requiring lifelong multidisciplinary care.

MUL serves as an excellent model for studying the molecular mechanisms of growth regulation, tumorigenesis, and peroxisomal biology. The TRIM37 protein is a member of the TRIM family and functions as an E3 ubiquitin ligase, playing roles in peroxisomal protein import and DNA damage response. The availability of patient-derived cell lines and the relatively simple monogenic etiology make MUL ideal for CRISPR-based gene editing to create isogenic models. Open questions include the precise role of TRIM37 in tumor suppression and the potential for targeted therapies.

MUL is primarily driven by loss-of-function mutations in TRIM37, leading to dysregulation of several pathways:

1. Peroxisomal biogenesis: TRIM37 is involved in peroxisomal matrix protein import; its loss impairs peroxisomal function, leading to metabolic abnormalities.

2. DNA damage response: TRIM37 participates in homologous recombination repair; its deficiency results in genomic instability and increased susceptibility to cancer.

3. Ubiquitin-proteasome system: TRIM37 acts as an E3 ligase, targeting substrates for degradation; loss of this activity alters protein homeostasis.

4. Tumor suppression: TRIM37 has been implicated as a tumor suppressor, with loss of function promoting tumorigenesis in tissues like liver and kidney.

The loss of TRIM37 affects multiple signaling networks:

• • Wnt/β-catenin pathway: TRIM37 loss leads to stabilization of β-catenin, promoting cell proliferation.
• • PI3K/AKT/mTOR pathway: Altered peroxisomal metabolism may activate this pathway, supporting cell survival.
• • DNA repair pathways: Defective homologous recombination leads to reliance on error-prone repair, increasing mutation load.
• • Peroxisome proliferator-activated receptor (PPAR) signaling: Impaired peroxisomal function alters lipid metabolism and PPAR target gene expression.

Organoids derived from patient tissues (e.g., liver, kidney) provide a more physiologically relevant 3D model for studying tumor development and drug response.

• • Patient-derived xenografts (PDX): Implantation of MUL patient tumor cells into immunodeficient mice to study tumor growth and drug sensitivity.
• • Genetically engineered mouse models (GEMM): Knockout of Trim37 in mice recapitulates MUL phenotypes, including growth retardation and increased tumor incidence.
• • Induced models: CRISPR-mediated knockout of Trim37 in mouse embryonic stem cells to generate chimeric mice for studying tissue-specific effects.

CRISPR-based gene editing enables the creation of isogenic cell lines with precise TRIM37 mutations. For example, a TRIM37 knockout cell line can be generated in HEK293T or HepG2 cells to study loss-of-function effects. Alternatively, a knock-in of a specific pathogenic mutation (e.g., a nonsense mutation) can be introduced to model patient-specific genotypes. These sequence-verified models are commercially available and accelerate research by providing consistent, reproducible systems for mechanistic studies and drug screening.

Gene-edited cell lines are essential for functional genomics studies. For example, TRIM37 knockout lines can be used to identify downstream targets of TRIM37 via transcriptomic and proteomic profiling. Knock-in lines with specific mutations can be compared to wild-type to assess the impact of individual variants on protein function and cellular phenotype.

Isogenic pairs (wild-type vs. TRIM37 knockout) are powerful tools for drug screening. They allow identification of compounds that selectively kill TRIM37-deficient cells, exploiting synthetic lethality. Additionally, resistance mechanisms can be studied by exposing knockout cells to drugs and selecting for resistant clones, then analyzing genetic and epigenetic changes.

CRISPR-based synthetic lethality screens using TRIM37 knockout cells can identify genes whose loss is lethal only in the absence of TRIM37. These genes may serve as novel therapeutic targets or biomarkers for patient stratification.