The combined therapeutic effects of bortezomib and fenretinide on neuroblastoma cells involve endoplasmic reticulum stress response.
OBJECTIVE
The proteasome inhibitor bortezomib inhibited cell growth and angiogenesis in neuroblastoma. Bortezomib has been shown to induce synergistic activity when combined with other antineoplastic agents. Here we have investigated the antitumor activity of bortezomib in combination with fenretinide, a synthetic retinoid, against neuroblastoma cells.


METHODS
Different neuroblastoma cell lines were tested for sensitivity to bortezomib and fenretinide, given alone or in different dose-dependent and time-dependent combination schedules. Cell proliferation, cell viability, and apoptosis were evaluated by measuring 3H-thymidine incorporation, trypan blue staining, DNA fragmentation, and western blot analysis. Angiogenesis was assessed by the chick embryo chorioallantoic membrane assay. An orthotopic neuroblastoma mouse model was used to examine in vivo sensitivity.


RESULTS
Each compound alone was able to induce a dose-dependent inhibition of cell proliferation, with a significant enhanced antiproliferative effect for the drugs used in combination. This inhibition was characterized by marked G2-M and G1 cell cycle arrest with nearly complete depletion of S phase. Bortezomib and fenretinide in association triggered an increased apoptosis through activation of specific genes of the endoplasmic reticulum stress compared with either drug tested alone. Tumor-bearing mice treated with bortezomib plus fenretinide lived statistically significantly longer than mice treated with each drug alone. Histologic evaluation and chorioallantoic membrane analysis of primary tumors showed that the combined therapeutic activity of bortezomib and fenretinide rested upon antitumor and antiangiogenic mechanisms.


CONCLUSIONS
These findings provide the rationale for the development of a new therapeutic strategy for neuroblastoma based on this pharmacologic combination.
Pagnan
Gabriella
G
Laboratory of Oncology, G. Gaslini Children's Hospital, Genoa, Italy. gabriellapagnan@ospedale-gaslini.ge
Di Paolo
Daniela
D
Carosio
Roberta
R
Pastorino
Fabio
F
Marimpietri
Danilo
D
Brignole
Chiara
C
Pezzolo
Annalisa
A
Loi
Monica
M
Galietta
Luis J V
LJ
Piccardi
Federica
F
Cilli
Michele
M
Nico
Beatrice
B
Ribatti
Domenico
D
Pistoia
Vito
V
Ponzoni
Mirco
M
D000818:Animals; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000627:therapeutic use*; D017209:Apoptosis / Q000187:drug effects; D001897:Boronic Acids / Q000008:administration & dosage*; D000069286:Bortezomib; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002642:Chick Embryo; D004721:Endoplasmic Reticulum / Q000187:drug effects* / Q000378:metabolism; D017313:Fenretinide / Q000008:administration & dosage*; D006801:Humans; D051379:Mice; D009447:Neuroblastoma / Q000188:drug therapy* / Q000401:mortality / Q000473:pathology; D011719:Pyrazines / Q000008:administration & dosage*; D023041:Xenograft Model Antitumor Assays
D001897:Boronic Acids; D011719:Pyrazines; D017313:Fenretinide; D000069286:Bortezomib