The RGD domain of human osteopontin promotes tumor growth and metastasis through activation of survival pathways.
BACKGROUND
Human osteopontin (OPN), a known tumor associated protein, exists in different isoforms, whose function is unclear. It also possesses a RGD domain, which has been implicated in diverse function. Here, we use genetic approaches to systematically investigate the function of the RGD domain in different OPN isoforms on tumor progression and metastasis for 2 different solid tumor models.


RESULTS
Using isoform-specific qRT-PCR, we found that OPN-A and B were the main isoforms overexpressed in evaluated human tumors, which included 4 soft tissue sarcomas, 24 lung and 30 head and neck carcinomas. Overexpression of either OPN-A or B in two different cell types promoted local tumor growth and lung metastasis in SCID mouse xenografts. However, expression of either isoform with the RGD domain either mutated or deleted decreased tumor growth and metastasis, and resulted in increased apoptosis by TUNEL staining. In vitro, whereas mutation of the RGD domain did not affect cell-cell adhesion, soft agar growth or cell migration, it increased apoptosis under hypoxia and serum starvation. This effect could be mitigated when the RGD mutant cells were treated with condition media containing WT OPN. Mechanistically, the RGD region of OPN inhibited apoptosis by inducing NF-kappaB activation and FAK phosphorylation. Inhibition of NF-kappaB (by siRNA to the p65 subunit) or FAK activation (by a inhibitor) significantly increased apoptosis under hypoxia in WT OPN cells, but not in RGD mutant cells.


CONCLUSIONS
Unlike prior reports, our data suggest that the RGD domain of both OPN-A and B promote tumor growth and metastasis mainly by protecting cells against apoptosis under stressed conditions and not via migration or invasion. Future inhibitors directed against OPN should target multiple isoforms and should inhibit cell survival mechanisms that involve the RGD domain, FAK phosphorylation and NF-kappaB activation.
Department of Radiation Oncology, Stanford University, Stanford, California, United States of America.
Donald
D
Courter
Hongbin
H
Cao
Shirley
S
Kwok
Christina
C
Kong
Alice
A
Banh
Peiwen
P
Kuo
Donna M
DM
Bouley
Carmen
C
Vice
Odd Terje
OT
Brustugun
Nicholas C
NC
Denko
Albert C
AC
Koong
Amato
A
Giaccia
Quynh-Thu
QT
Le
D000818:Animals; D017209:Apoptosis; D045744:Cell Line, Tumor; D004195:Disease Models, Animal; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D051379:Mice; D009154:Mutation; D009362:Neoplasm Metastasis; D009368:Neoplasm Transplantation; D009842:Oligopeptides; D053495:Osteopontin / Q000737:chemistry* / Q000235:genetics*; D010766:Phosphorylation; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary
D009842:Oligopeptides; D020033:Protein Isoforms; D053495:Osteopontin; C047981:arginyl-glycyl-aspartic acid