OC 76.2 - Anti-proliferative, anti-angiogenic and anti-invasive effect of antithrombin in glioblastoma multiforme

Wednesday, June 26, 2024

10:30 – 10:45 ICT

Room: 210 A-D

Presenting Author(s)

Julia Peñas Martínez, PhD Student

PhD student
Department of Hematology and Medical Oncology, University General Hospital Morales Meseguer, Center of Hemodonation, University of Murcia, IMIB-Pascual Parrilla, 30003, Murcia, Spain
Murcia, Murcia, Spain

Co-Author(s)

David Zaragoza Huesca, Degree in Biology and Master

PhD student
Department of Hematology and Medical Oncology, University General Hospital Morales Meseguer, Center of Hemodonation, University of Murcia, IMIB-Pascual Parrilla, 30003, Murcia, Spain
Murcia, Murcia, Spain
SE

Salvador Espin

Technical suport
Department of Hematology and Medical Oncology, University General Hospital Morales Meseguer, Center of Hemodonation, University of Murcia, IMIB-Pascual Parrilla, 30003, Murcia, Spain
Murcia, Murcia, Spain
Constantino Martínez

Doctor
Department of Hematology and Medical Oncology, University General Hospital Morales Meseguer, Center of Hemodonation, University of Murcia, IMIB-Pascual Parrilla, 30003, Murcia, Spain
Murcia, Murcia, Spain
RG

Rocío González-Conejero

Doctor
Department of Hematology and Medical Oncology, University General Hospital Morales Meseguer, Center of Hemodonation, University of Murcia, IMIB-Pascual Parrilla, 30003, Murcia, Spain
Murcia, Murcia, Spain
ML

María Luisa Lozano

Hematologist
Servicio de Hematología, Hospital General Universitario Morales Messeguer, Centro Regional de Hemodonación, Universidad de Murcia, Universidad de Murcia, IMIB-Pascual Parrilla, CIBERER-ISCIII
Murcia, Murcia, Spain
VV

Vicente Vicente

Senior Researcher
Servicio de Hematología, Hospital General Universitario Morales Messeguer, Centro Regional de Hemodonación, Universidad de Murcia, Universidad de Murcia, IMIB-Pascual Parrilla, CIBERER-ISCIII
Murcia, Murcia, Spain
Irene Martínez Martínez

Researcher
Servicio de Hematología, Hospital General Universitario Morales Messeguer, Centro Regional de Hemodonación, Universidad de Murcia, Universidad de Murcia, IMIB-Pascual Parrilla, CIBERER-ISCIII
Murcia, Murcia, Spain

Background: Antithrombin (AT) has other functions beyond hemostasis. We have demonstrated that native and prelatent AT reduce migration and invasion as well as the expression and phosphorylation of VEGFA, STAT3, and ERK1/2 in U87 glioblastoma multiforme (GBM) cells. Regarding GBM, it is the most lethal primary malignant tumor of the central nervous system in adults, and the median survival is 12-15 months after diagnosis.

Aims: 1) investigate the role of AT on other pro-tumor pathways using in vitro and preclinical models of GBM; 2) identify the potential receptor of AT in GBM.

Methods: Native and prelatent AT were purified from healthy donor’s plasma. Prior to each experiment, U87 cells were treated with 2.16 µM of native, prelatent AT or PBS. A microarray analysis was performed on U87 (Human Clariom D). Differences were validated by western blot. Cell cycle and proliferation were assessed by flow cytometry. Three microRNAs (miR-A/B/C) with altered expression in GBM were transfected into U87 cells, and angiogenesis was evaluated by co-culture with endothelial cells. The receptor was determined by crosslinking and immunoprecipitation, followed by quantitative proteomics and confocal microscopy. Healthy human brain RFP-organoids were generated from iPSCs, and neurospheres from the biopsy of one GMB patient, at diagnosis (GFP-275) and at relapse (GFP-275-BIS). Anti-invasive AT effect was validated through organoids and neurospheres co-culture.

Results: Native AT reduced U87 S-phase and proliferation as well as E2F4 and pRb expression (Fig.1A). Moreover, AT increased miR-A expression (FC 1.61), which in turn reduced angiogenesis (Fig. 1B). We identified dystonin as the receptor of AT on U87 (Fig. 2A). Interestingly, AT partially reduced invasion of 275 neurospheres, and completely reduced 275-BIS neurospheres invasion of the brain-organoids (Fig. 2B).

Conclusion(s): AT has surprising and versatile anti-tumor properties on GBM. Our results support its potential therapeutic usefulness in GBM, a tumor in which new treatments are urgently needed.