Functional heterogeneity of osteopontin isoforms in non-small cell lung cancer.
BACKGROUND
Osteopontin (OPN) is a multifunctional protein with an important but poorly understood role in non-small cell lung cancer (NSCLC) pathogenesis. Moreover, the role of the three known mRNA isoforms (OPNa, OPNb, and OPNc) has not been reported. We hypothesize that OPN isoforms play different roles in determining the metastatic potential of NSCLC.


METHODS
We amplified mRNA for each OPN isoform in NSCLC tumors and matched normal lung. The functional impact of each isoform was evaluated by transfecting cDNA plasmids specific to each isoform into NSCLC cell lines and comparing behavior to empty vector controls in scratch closure, cell proliferation, soft-agar colony formation, and Matrigel invasion assays. Gene array was used to evaluate differences in downstream targets and was compared with a panel of markers for epithelial-mesenchymal transition (EMT).


RESULTS
OPNa expression was increased in 91% of NSCLC tumors compared with matched lung. OPNa overexpression significantly increased activity in scratch closure, proliferation, soft-agar colony formation, and Matrigel invasion assays compared with controls in all cell lines. OPNb overexpression produced a less significant modulation of function. OPNc overexpression significantly decreased activity in proliferation, colony formation, and invasion assays compared with controls. Expression arrays revealed an increase in EMT with OPNa overexpression but not OPNc. Differences were validated by quantitative reverse transcriptase-polymerase chain reaction.


CONCLUSIONS
Overexpression of the individual OPN isoforms in NSCLC results in divergent functional phenotypes. OPNa produced an aggressive phenotype, whereas OPNc produced a more indolent phenotype. Exon 4, which is transcribed in OPNa but absent in OPNc, may be central to this phenomenon and could serve as a target for isoform-specific inhibition of OPN in NSCLC.
Goparaju
Chandra M V
CM
Department of Cardiothoracic Surgery, New York University School of Medicine, New York City, New York 10016, USA.
Pass
Harvey I
HI
Blasberg
Justin D
JD
Hirsch
Nathalie
N
Donington
Jessica S
JS
D017209:Apoptosis; D014408:Biomarkers, Tumor / Q000235:genetics* / Q000378:metabolism; D015153:Blotting, Western; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D002448:Cell Adhesion; D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D003114:Colony-Forming Units Assay; D058750:Epithelial-Mesenchymal Transition; D020869:Gene Expression Profiling; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D020411:Oligonucleotide Array Sequence Analysis; D053495:Osteopontin / Q000235:genetics* / Q000378:metabolism; D020033:Protein Isoforms; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014945:Wound Healing
D014408:Biomarkers, Tumor; D020033:Protein Isoforms; D012333:RNA, Messenger; D053495:Osteopontin