Connective tissue growth factor and its role in lung adenocarcinoma invasion and metastasis.
BACKGROUND
Tumor invasion and metastasis cause most deaths in cancer patients. Connective tissue growth factor (CTGF), a secreted protein that binds to integrins, modulates the invasive behavior of certain human cancer cells, but few mechanistic details are known. We investigated the roles of CTGF and collapsin response mediator protein 1 (CRMP-1) in metastasis and invasion of human lung adenocarcinoma.


METHODS
We compared vector control-transfected cells with corresponding CTGF gene-transfected cells. Invasive activity was measured with a modified Boyden chamber assay, and metastatic activity was measured in an animal model. We used CTGF deletion mutants, CTGF and CRMP-1 antisense oligonucleotides, and anti-integrin and anti-CRMP-1 antibodies to investigate the functional relationship between CTGF and CRMP-1. Expression of CTGF protein in 78 lung adenocarcinoma specimens was investigated immunohistochemically. All statistical tests were two-sided.


RESULTS
Invasive (both P<.001) and metastatic (P<.001 and P=.003, respectively) activities were lower in cells that overexpress CTGF than in vector control cells. Expression of CRMP-1 was higher in CTGF-transfected clones than in vector control cells, and its level decreased after cells were treated with anti-integrin alpha(v)beta3 and alpha(v)beta5 antibodies. Reduced levels of CRMP-1 protein after the transfection of CRMP-1-specific antisense oligonucleotides, but not sense oligonucleotides, increased the invasiveness of CTGF-transfected cells (mean numbers of invasive CTGF-transfected cells treated with 20 microM CRMP-1-specific sense and antisense oligonucleotides were 327 and 516 cells, respectively [difference = 189 cells, 95% confidence interval [CI] = 156 to 221 cells; P<.001]). The CT module of CTGF was the region primarily responsible for the increased expression of CRMP-1 and the inhibition of invasion (mean numbers of invasive cells expressing full-length CTGF and CT module-deleted mutant were 148 and 385 cells, respectively [difference = 237 cells, 95% CI = 208 to 266 cells; P<.001]). Reduced expression of CTGF in lung cancer specimens was statistically significantly associated with the risk of more advanced-stage disease (stages III and IV versus stages I and II; P=.001), lymph node metastasis (P =.014), and shorter survival (median survival with high levels of CTGF = 66.7 months and median survival for low levels = 18.2 months; difference = 48.5 months, 95% CI = 33.5 to 63.5 months; P =.02).


CONCLUSIONS
CTGF inhibits metastasis and invasion of human lung adenocarcinoma by a CRMP-1-dependent mechanism.
Laboratory of Molecular and Cellular Toxicology, Institute of Toxicology, College of Medicine, National Taiwan University, Taipei, Taiwan.
Cheng-Chi
CC
Chang
Jin-Yuan
JY
Shih
Yung-Ming
YM
Jeng
Jen-Liang
JL
Su
Been-Zen
BZ
Lin
Szu-Ta
ST
Chen
Yat-Pang
YP
Chau
Pan-Chyr
PC
Yang
Min-Liang
ML
Kuo
D000230:Adenocarcinoma / Q000378:metabolism* / Q000517:prevention & control / Q000556:secondary*; D014408:Biomarkers, Tumor / Q000378:metabolism*; D045744:Cell Line, Tumor; D055513:Connective Tissue Growth Factor; D018450:Disease Progression; D017353:Gene Deletion; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics / Q000378:metabolism*; D007150:Immunohistochemistry; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D008175:Lung Neoplasms / Q000378:metabolism* / Q000473:pathology*; D008207:Lymphatic Metastasis; D009154:Mutation; D009361:Neoplasm Invasiveness; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D016376:Oligonucleotides, Antisense; D010750:Phosphoproteins / Q000378:metabolism*; D010957:Plasmids; D016016:Proportional Hazards Models; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016019:Survival Analysis; D014162:Transfection; D015854:Up-Regulation; D000098855:Collapsin Response Mediator Protein 1
D014408:Biomarkers, Tumor; C526317:CCN2 protein, human; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D009419:Nerve Tissue Proteins; D016376:Oligonucleotides, Antisense; D010750:Phosphoproteins; D000098855:Collapsin Response Mediator Protein 1; D055513:Connective Tissue Growth Factor