# Mechanism of a novel approach for platelet cold storage

> **NIH NIH P01** · BOSTON CHILDREN'S HOSPITAL · 2022 · $655,826

## Abstract

ABSTRACT
Refrigerated storage reduces platelet life-span because it causes cytoskeletal rearrangements, de-
sialylated glycoprotein-Ib (GPIb) to cluster, form microdomains, shed and induces mitochondrial
dependent reactive oxygen species (ROS) and apoptosis, which may result in inflammatory response in
vulnerable patient populations. Recognition by host of clustered glycoproteins (GP) results in platelet
phagocytosis and clearance. As a consequence, cold stored platelets are only allowed for use in
trauma patient therapy and not for prophylaxis or treatment of stem cell transplant recipients and
hematology/oncology patients. The Rho family GTPases RHOA and RAC1 are central regulators of
cytoskeletal rearrangements, and have been shown to control lipid raft formation and composition;
changes in Rho GTPase activities may influence platelet membrane lipid raft assembly, post-
translational modifications of membrane glycoproteins, included GpIb and increased mitochondrial ROS
and apoptotic activity. Our preliminary, submitted and published data using genetic and
pharmacological means show that reversible RHOA GTPase inhibition results in an inhibition of myosin
activity and prevention of clathrin-independent formation and internalization of lipid rafts enriched in
active glycosyl-transferases (GT) and GPIb. RHOA GTPase inhibition prevents the metabolic
reprogramming effect and allows the maintenance of glycolytic flux and mitochondrial dependent
respiration and ROS production. Importantly, we further demonstrate that murine, human and Rhesus-
macaque platelets, when stored in refrigerated conditions for up to 14 days in the presence of a lead
RHOA inhibitor, G04, can retain survival function at a level similar to that of room-temperature stored
platelets and retain hemostatic activity in vivo, and an antioxidant phenocopies some of the effects of
G04. We hypothesize that RHOA controls the process of GP clustering during cold storage through the
regulation of actomyosin activity, vesicle trafficking and mitochondrial respiration. We will first identify
the mechanism by which RHOA regulates lipid raft formation, GP clustering and endocytosis in
platelets upon refrigeration. We will also determine the outcomes of pharmacologic inhibition of RHOA
in preventing the metabolic and mitochondrial damage of long-term cold stored platelets by analyzing
the effect of long-term storage on mitochondrial activity and the crosstalk between RHOA and the
master metabolic regulator AMPK in regulating platelet metabolism and mitophagy. Our studies will
provide the mechanism and a stringent proof-of-principle for the translational value of a novel approach
to refrigerated platelet storage.

## Key facts

- **NIH application ID:** 10494385
- **Project number:** 1P01HL158688-01A1
- **Recipient organization:** BOSTON CHILDREN'S HOSPITAL
- **Principal Investigator:** Jose A. Cancelas
- **Activity code:** P01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $655,826
- **Award type:** 1
- **Project period:** 2022-08-15 → 2027-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10494385

## Citation

> US National Institutes of Health, RePORTER application 10494385, Mechanism of a novel approach for platelet cold storage (1P01HL158688-01A1). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10494385. Licensed CC0.

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