Preclinical strategic advances in the therapeutic landscape of intracerebral hemorrhagic stroke

Abstract

Intracerebral hemorrhage (ICH) is a form of stroke caused by bleeding into the brain parenchyma following blood vessel rupture. It is associated with high mortality and morbidity and, unlike ischemic stroke (IS), still lacks an approved, curative treatment despite several promising preclinical candidates. While IS therapies aim to restore blood flow by resolving vascular blockages, ICH management is focused on blood pressure control and preventing hematoma expansion. Critically, no approved therapy addresses the secondary injury caused by clot breakdown products, despite several candidates have been identified in preclinical studies._x000D_ _x000D_ One factor contributing to poor translation may be the use of preclinical models with limited predictive value. Rodent models, although useful in early-stage research, have lissencephalic brains and different gray-white matter architecture compared to humans. In contrast, large animals (e.g. pigs) provide a more suitable model that would help overcome critical shortcomings in ICH preclinical research._x000D_ A core focus of this thesis work is the evaluation of human apotransferrin (hATf) as a therapeutic for ICH. hATf significantly improved sensorimotor performance at early time points in mice without affecting hematoma size, edema, or coagulation. hATf rapidly reduced plasma transferrin saturation (TSAT) within 24 hours, accumulated in the injured brain, and reduced oxidative stress (4-HNE). These effects were independent of canonical ferroptosis pathways (GPX4/xCT or FSP1). Instead, hATf preserved the iron chaperone PCBP2, decreased expression of TfR1 (a hallmark ferroptosis marker), and blocked caspase-2 activation, indicating combined antiferroptotic and anti-apoptotic action. _x000D_ _x000D_ Notably, hATf reduced TSAT and improved neurological outcomes more rapidly than previously described for deferoxamine, which has shown a delayed effect on TSAT and only modest benefits in the iDEF and TANDEM trials. hATf’s demonstrated safety and efficacy in both ICH and ischemic rodent models support its potential as a broadly applicable stroke therapy._x000D_ This study also highlights that in ICH, functional recovery rather than hematoma volume is the most relevant and reliable preclinical endpoint. To improve its evaluation, an automated DeepLabCut-based motion tracking tool was developed to analyze the neurobehavioral performance of mice in the adhesive tape removal test. This system provided a sensitive, objective assessment of neurobehavioral deficits and accurately classified stroke severity in both ischemic and hemorrhagic models. The tool is currently being adapted for pig studies to enhance translational value._x000D_ _x000D_ To further bridge the gap between preclinical and clinical research, a novel, minimally invasive swine ICH model was developed using autologous injection of blood, avoiding surgical trauma. Although the study pig population should be expanded, early MRI imaging and biomarker shifts validated the model’s relevance._x000D_ _x000D_ Together with prior work, this thesis builds a compelling case for hATf as a rapid-acting, multi-target stroke therapy. Its efficacy in both stroke types supports its potential as a pre-hospital intervention.

Date of Award	15 Dec 2025
Original language	English
Awarding Institution	Universitat Autònoma de Barcelona (UAB)
Supervisor	Octavi Marti Sistac (Director) & Teresa Gasull Dalmau (Director)