Regulación del microambiente tumoral por EGFR en glioblastoma: análisis del efecto terapéutico de dacomitinib.

López-Amo Calvo, Beatriz (2020). Regulación del microambiente tumoral por EGFR en glioblastoma: análisis del efecto terapéutico de dacomitinib.. Proyecto Fin de Carrera / Trabajo Fin de Grado, E.T.S. de Ingeniería Agronómica, Alimentaria y de Biosistemas (UPM), Madrid.

Description

Title: Regulación del microambiente tumoral por EGFR en glioblastoma: análisis del efecto terapéutico de dacomitinib.
Author/s:
  • López-Amo Calvo, Beatriz
Contributor/s:
  • Sánchez-Gómez, Pilar
  • Ramírez Castillejo, María del Carmen
Item Type: Final Project
Degree: Grado en Biotecnología
Date: October 2020
Subjects:
Faculty: E.T.S. de Ingeniería Agronómica, Alimentaria y de Biosistemas (UPM)
Department: Biotecnología - Biología Vegetal
Creative Commons Licenses: Recognition - No derivative works - Non commercial

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Abstract

Gliomas are primary brain tumors histologically similar to glial cells. The histological classification of gliomas from the World Health Organization (WHO) identifies 4 degrees of malignancy depending on the amount of atypia, mitotic cells, microvascular proliferation and necrosis. Grade IV gliomas, also known as glioblastomas (GBM) are infiltrating and diffuse tumors with nuclear atypia, large numbers of mitotic cells and broad areas of necrosis and vascular proliferation. GBMs are considered the most aggressive and common primary tumors in the Central Nervous System, with a bad prognosis and an average survival after diagnosis of 15 months. This short life expectancy is due to the lack of effective treatments and knowledge about this cancer at molecular and physiological levels. Recent genomic and transcriptomic analysis of gliomas have facilitated the study of this type of cancer. Common alterations in primary GBM include mutations and/or amplifications of Epidermal Growth Factor Receptor (EGFRmut or EGFRamp), a type 1 tyrosine kinase receptor that promotes cell migration, proliferation and survival. Moreover, it regulates the immune response when activated. Around 40 to 50% of GBMs present EGFRamp, inducing constitutive activation of EGFR signaling. In addittion, 70% of GBMs EGFRamp show EGFRmut, with disrupted activity that promotes tumor growth. The most common EGFR mutation is the variant III (EGFRvIII), detected in 50% of GBMs EGFRamp. EGFRvIII has a characteristic tertiary conformation that triggers its constitutive activation, resulting in a superior oncogenic potential of the tumor cells, which leads to a worse prognosis. Recent studies from our group and others show a critical role of EGFR in the modulation of the vascular tumor microenvironment. Therefore, therapies targeting EGFR may affect tumor vasculature. Moreover, neuronal degeneration seems to be linked to BBB permeability, determined by the vascular phenotype of the GBM. Additionally, it has been observed an immunosuppressed microenvironment inside GBMs, enriched in infiltrated macrophages, among other cell types. Preliminary studies have shown differences in the immune infiltrate of GBMs depending on the expression of EGFRmut. It could be hypothesized that EGFR alterations may control immune microenvironment of GBM though the modulation of the vascular network. Alternatively, EGFR may control the function of the hematopoietic cells that express the receptor such as macrophages and T lymphocytes. First line treatment of GBM includes surgical removal of the tumor followed by radiotherapy (RT) and Chemotherapy (CT). If the tumor recurs, a personalized treatment follows. Different drugs have been tested as second line treatment for recurrent GBM and more are in development. Tyrosine Kinase Inhibitors (TKIs) against EGFR are showing promising results in preclinical and clinical trials. TKIs are low molecular weight small molecules that have the capacity to go through the blood brain barrier (BBB) and block the ATP binding pocket of the tyrosine-kinase domain of EGFR. Dacomitinib is a second generation TKI that has shown great inhibition potential against EGFRvIII. In this project we have tried to verify the results of previous investigations regarding the role of EGFR in the control and development of the tumor microenvironment, focusing on the vascular and immune compartments, using murine models of GBM expressing either the wild-type form of EGFR or the variant EGFRvIII. We have also analyzed the possible relation between EGFR and disruption of the BBB and neurodegeneration. Moreover, we have deepened into the study of dacomitinib as a therapeutic drug for GBM. Neither protein expression analysis though Western Blotting nor qRT-PCR analysis of the expression of genes related to vasculature contributed with relevant data due to the model used to recreate the malignancy. However, the analysis of the expression of genes related to BBB breakage proved a leakier BBB in EGFRwt/amp tumors. This was corroborated with the immunofluorescent analysis of immune extravasation, which showed an increased IgG extravasation in tumors expressing EGFRwt/amp opposite to the little extravasation observed in EGFRvIII tumors. This data validate the results obtained in previous studies of the group. Although neuronal genes expression analysis showed too much heterogeneity to get any significant differences, histologically we could appreciate more neurodegeneration in EGFRwt/amp tumors, results that correlate with the increased breakage of BBB and extravasation of this type of GBM. In addition, the qRT-PCR analysis of genes related to microglia and tumor associated macrophages with anti-inflammatory phenotype proved an increased infiltration of monocytes and macrophages in EGFRwt/amp tumors, associated to an increased BBB permeability and hypoxic and necrotic areas. In relation to dacomitinib, the drug did not show significant effect regarding BBB breakage or neurodegeneration, which may suggest that this drug does not affect the action of EGFR in the regulation of vascular permeability and immune infiltration (EGFR action may depend on local levels of the protein). Alternatively, dacomitinib could have a transitory effect on the function of EGFR that we were not able to detect. Interestingly, an increase in the levels of T-cells was observed in the tumors treated with dacomitinib. This may suggest an anti-glioma response to dacomitinib mediated by the activation of the immune system. These data unleash the possibility of a new study to compare the intratumoral levels of lymphocytes in GBMs from patients with different genetic profile treated with dacomitinib in order to understand the role EGFR and TKIs may have over the immune environment of GBMs.

More information

Item ID: 66040
DC Identifier: http://oa.upm.es/66040/
OAI Identifier: oai:oa.upm.es:66040
Deposited by: Biblioteca ETSI Agrónomos
Deposited on: 03 Feb 2021 11:01
Last Modified: 03 Feb 2021 11:01
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