Expression of hypoxia-inducible carbonic anhydrase-9 relates to angiogenic pathways and independently to poor outcome in non-small cell lung cancer.
Carbonic anhydrase-9 (CA9), a transmembrane enzyme with an extracellular active site, is involved in the reversible metabolism of the carbon dioxide to carbonic acid. Up-regulation of CA by hypoxia and the hypoxia-inducible factor (HIF) pathway has been recently postulated (Wykoff et al. Cancer Res., 60: 7075-7083, 2000). In the present study we examined the expression of this enzyme in non-small cell lung cancer. Of 107 cases analyzed, 39 (36.4%) had strong membrane/cytoplasmic expression of CA9 and were grouped as positive. The staining was confined around areas of necrosis, and a significant association of CA9 expression with the extent of necrosis was noted (P = 0.004). Nevertheless, 38 of 74 cases with focal or extensive necrosis did not express CA9. CA9 expression was more frequent in the squamous cell histology (P = 0.001) and with advanced T stage (P = 0.009). A significant coexpression of CA9 with platelet-derived endothelial cell growth factor and basic fibroblast growth factor receptor expression was noted. Double staining of CA9 with anti-CD31 monoclonal antibody revealed an overall higher microvessel density in the areas expressing CA9 than in negative areas (P = 0.0005). Thirty-one of 38 CA9-positive cases were positive for HIF1a/HIF2a, but HIF positivity was a more common event (68 of 107) and their patterns of expression were diffuse (not confined in the necrotic areas). A direct association of CA9 expression with epidermal growth factor receptor, c-erbB-2, and MUC1 expression was also noted (P < 0.04). Survival analysis showed that CA9 expression is related to poor prognosis. CA9 expression in tumors with low vascularization defined a prognosis similar to the one of patients with highly angiogenic tumors. Multivariate analysis revealed that CA9 expression is a significant prognostic factor independent of angiogenesis. We conclude that CA9 is an important molecule in non-small cell lung cancer, the up-regulation of which occurs in highly hypoxic/necrotic regions of the tumors. The expression of CA9 is linked to the expression of a constellation of proteins involved in angiogenesis, apoptosis inhibition, and cell-cell adhesion disruption, which explains the strong association of CA9 with poor outcome.
Giatromanolaki
A
A
Tumour and Angiogenesis Research Group, Department of Radiotherapy/Oncology, Democritus University of Thrace, Alexandroupolis 68100, Greece. targ@her.forthnet.gr
Koukourakis
M I
MI
Sivridis
E
E
Pastorek
J
J
Wykoff
C C
CC
Gatter
K C
KC
Harris
A L
AL
D000230:Adenocarcinoma / Q000098:blood supply / Q000201:enzymology / Q000473:pathology; D000328:Adult; D000368:Aged; D000951:Antigens, Neoplasm; D051792:Basic Helix-Loop-Helix Proteins; D000071231:Carbonic Anhydrase IX; D002256:Carbonic Anhydrases; D002289:Carcinoma, Non-Small-Cell Lung / Q000098:blood supply* / Q000201:enzymology* / Q000473:pathology; D002294:Carcinoma, Squamous Cell / Q000098:blood supply / Q000201:enzymology / Q000473:pathology; D015687:Cell Hypoxia; D004790:Enzyme Induction; D066246:ErbB Receptors / Q000096:biosynthesis; D005260:Female; D016222:Fibroblast Growth Factor 2 / Q000096:biosynthesis / Q000502:physiology; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D008175:Lung Neoplasms / Q000098:blood supply* / Q000201:enzymology* / Q000473:pathology; D008297:Male; D008875:Middle Aged; D018396:Mucin-1 / Q000096:biosynthesis; D009336:Necrosis; D009363:Neoplasm Proteins / Q000096:biosynthesis*; D009389:Neovascularization, Pathologic / Q000201:enzymology*; D011379:Prognosis; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000096:biosynthesis; D015398:Signal Transduction / Q000502:physiology; D013939:Thymidine Phosphorylase / Q000096:biosynthesis / Q000502:physiology; D015534:Trans-Activators / Q000096:biosynthesis; D014157:Transcription Factors / Q000096:biosynthesis; D000099162:Endothelial PAS Domain-Containing Protein 1
D000951:Antigens, Neoplasm; D051792:Basic Helix-Loop-Helix Proteins; D000071231:Carbonic Anhydrase IX; D002256:Carbonic Anhydrases; D066246:ErbB Receptors; D016222:Fibroblast Growth Factor 2; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D018396:Mucin-1; D009363:Neoplasm Proteins; D018719:Erb-b2 Receptor Tyrosine Kinases; D013939:Thymidine Phosphorylase; D015534:Trans-Activators; D014157:Transcription Factors; D000099162:Endothelial PAS Domain-Containing Protein 1; C497442:HIF1A protein, human; C089226:CA9 protein, human