Researchers have developed a novel CRISPR-based method to better understand genetic mutations in inherited platelet disorders (IPDs), a differential diagnosis for fetal and neonatal alloimmune thrombozytopenia (FNAIT), according to a recently published study in AJHG.
IDP disorders include Glanzmann thrombasthenia (GT), where mutations disrupt the function of a key receptor on platelets known as αIIb/β3, which is essential for platelet aggregation and stopping bleeding.
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A major challenge in diagnosing IPDs is the high number of genetic variants whose significance is unclear and are called “variants of uncertain significance” (VUSs). Nearly 98% of the variants found in IPD patients fall into this category, making it hard for doctors to confirm a diagnosis or provide personalized treatment.
To address this, researchers created a new experimental platform called CRIMSON HD. It uses the CRISPR/Cas9 gene-editing tool to introduce precise mutations directly into immature human platelets (megakaryocytes), derived from CD34⁺ blood stem cells. This approach allows to test how each mutation affects the function of platelets in a setting that closely mimics the human body, avoiding the limitations of artificial lab systems that don’t reflect real cell behavior.
Using this technique, the team successfully recreated known GT disease patterns and reclassified several previously unclear variants. For example, they found that one common variant, αIIb Gly201Ala, caused nearly complete loss of the platelet receptor, confirming it is disease-causing. Another variant, β3 Arg119Gln, previously linked to FNAIT, was also shown to impair receptor function, offering new insights into its role in disease.
The experiments achieved high editing precision with over 90% efficiency, by delivering CRISPR machinery along with synthetic DNA templates into the cells using electroporation. Researchers then evaluated how much of the αIIb/β3 receptor was present on the surface and how well it responded to platelet-activating agents.
Ultimately, CRIMSON HD enabled researchers to distinguish between harmful and harmless variants with unprecedented accuracy. This method could pave the way for better diagnosis, risk assessment, and even personalized therapies in patients with rare platelet disorders.
“These findings demonstrate the importance of lineage-specific, physiologically relevant assays for the functional classification of platelet-related variants, providing mechanistic information and clinically meaningful insights for individuals with IPDs,” the authors concluded.
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