Neutralization of adrenomedullin inhibits the growth of human glioblastoma cell lines in vitro and suppresses tumor xenograft growth in vivo.
Presently, there is no effective treatment for glioblastoma, the most malignant and common brain tumor. Growth factors are potential targets for therapeutic strategies because they are essential for tumor growth and progression. Peptidylglycine alpha-amidating monooxygenase is the enzyme producing alpha-amidated bioactive peptides from their inactive glycine-extended precursors. The high expression of peptidylglycine alpha-amidating monooxygenase mRNA in glioblastoma and glioma cell lines points to the involvement of alpha-amidated peptides in tumorigenic growth processes in the brain. After screening of amidated peptides, it was found that human glioblastoma cell lines express high levels of adrenomedullin (AM) mRNA, and that immunoreactive AM is released into the culture medium. AM is a multifunctional regulatory peptide with mitogenic and angiogenic capabilities among others. Real-time quantitative reverse transcriptase-polymerase chain reaction analysis showed that AM mRNA was correlated to the tumor type and grade, with high expression in all glioblastomas analyzed, whereas a low expression was found in anaplastic astrocytomas and barely detectable levels in low-grade astrocytomas and oligodendrogliomas. In the present study we also demonstrate the presence of mRNA encoding the putative AM receptors, calcitonin receptor-like receptor/receptor activity-modifying protein-2 and -3 (CRLR/RAMP2; CRLR/RAMP3) in both glioma tissues and glioblastoma cell lines and further show that exogenously added AM can stimulate the growth of these glioblastoma cells in vitro. These findings suggest that AM may function as an autocrine growth factor for glioblastoma cells. One way to test the autocrine hypothesis is to interrupt the function of the endogenously produced AM. Herein, we demonstrate that a polyclonal antibody specific to AM, blocks the binding of the hormone to its cellular receptors and decreases by 33% (P < 0.001) the growth of U87 glioblastoma cells in vitro. Intratumoral administration of the anti-AM antibody resulted in a 70% (P < 0.001) reduction in subcutaneous U87 xenograft weight 21 days after treatment. Furthermore, the density of vessels was decreased in the antibody-treated tumors. These findings support that AM may function as a potent autocrine/paracrine growth factor for human glioblastomas and demonstrate that inhibition of the action of AM (produced by tumor cells) may suppress tumor growth in vivo.
Ouafik
L'Houcine
L
Laboratoires de CancérologieExpérimentale and Transfert d'OncologieBiologique, Faculté de MédecineSecteur Nord, IFR Jean Roche, Marseille, France. ouafik.h@jean-roche.univ-mrs.fr
Sauze
Samantha
S
Boudouresque
Françoise
F
Chinot
Olivier
O
Delfino
Christine
C
Fina
Frédéric
F
Vuaroqueaux
Vincent
V
Dussert
Christophe
C
Palmari
Jacqueline
J
Dufour
Henri
H
Grisoli
François
F
Casellas
Pierre
P
Brünner
Nils
N
Martin
Pierre-Marie
PM
D053607:Adrenomedullin; D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D005909:Glioblastoma / Q000378:metabolism / Q000473:pathology*; D005910:Glioma / Q000378:metabolism / Q000473:pathology; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D006899:Mixed Function Oxygenases / Q000235:genetics; D009097:Multienzyme Complexes / Q000235:genetics; D009368:Neoplasm Transplantation; D010446:Peptide Fragments / Q000494:pharmacology; D010455:Peptides / Q000235:genetics / Q000276:immunology / Q000378:metabolism / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism; D014183:Transplantation, Heterologous; D014407:Tumor Cells, Cultured
D000906:Antibodies; D009097:Multienzyme Complexes; D010446:Peptide Fragments; D010455:Peptides; D012333:RNA, Messenger; C104782:adrenomedullin (22-52); D053607:Adrenomedullin; D006899:Mixed Function Oxygenases; C035189:peptidylglycine monooxygenase