title: Carbonic anhydrase IX promotes tumor growth and necrosis in vivo and inhibition enhances anti-VEGF therapy.
abstract: OBJECTIVE
  Bevacizumab, an anti-VEGFA antibody, inhibits the developing vasculature of tumors, but resistance is common. Antiangiogenic therapy induces hypoxia and we observed increased expression of hypoxia-regulated genes, including carbonic anhydrase IX (CAIX), in response to bevacizumab treatment in xenografts. CAIX expression correlates with poor prognosis in most tumor types and with worse outcome in bevacizumab-treated patients with metastatic colorectal cancer, malignant astrocytoma, and recurrent malignant glioma.
  
  
  METHODS
  We knocked down CAIX expression by short hairpin RNA in a colon cancer (HT29) and a glioblastoma (U87) cell line which have high hypoxic induction of CAIX and overexpressed CAIX in HCT116 cells which has low CAIX. We investigated the effect on growth rate in three-dimensional (3D) culture and in vivo, and examined the effect of CAIX knockdown in combination with bevacizumab.
  
  
  RESULTS
  CAIX expression was associated with increased growth rate in spheroids and in vivo. Surprisingly, CAIX expression was associated with increased necrosis and apoptosis in vivo and in vitro. We found that acidity inhibits CAIX activity over the pH range found in tumors (pK = 6.84), and this may be the mechanism whereby excess acid self-limits the build-up of extracellular acid. Expression of another hypoxia inducible CA isoform, CAXII, was upregulated in 3D but not two-dimensional culture in response to CAIX knockdown. CAIX knockdown enhanced the effect of bevacizumab treatment, reducing tumor growth rate in vivo.
  
  
  CONCLUSIONS
  This work provides evidence that inhibition of the hypoxic adaptation to antiangiogenic therapy enhances bevacizumab treatment and highlights the value of developing small molecules or antibodies which inhibit CAIX for combination therapy.
authors.affiliation: Molecular Oncology Laboratories, Department of Medical Oncology, Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, United Kingdom.
authors.forename: Alan
authors.initials: A
authors.lastname: McIntyre
authors.forename: Shalini
authors.initials: S
authors.lastname: Patiar
authors.forename: Simon
authors.initials: S
authors.lastname: Wigfield
authors.forename: Ji-Liang
authors.initials: JL
authors.lastname: Li
authors.forename: Ioanna
authors.initials: I
authors.lastname: Ledaki
authors.forename: Helen
authors.initials: H
authors.lastname: Turley
authors.forename: Russell
authors.initials: R
authors.lastname: Leek
authors.forename: Cameron
authors.initials: C
authors.lastname: Snell
authors.forename: Kevin
authors.initials: K
authors.lastname: Gatter
authors.forename: William S
authors.initials: WS
authors.lastname: Sly
authors.forename: Richard D
authors.initials: RD
authors.lastname: Vaughan-Jones
authors.forename: Pawel
authors.initials: P
authors.lastname: Swietach
authors.forename: Adrian L
authors.initials: AL
authors.lastname: Harris
mesh_terms: D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D000818:Animals; D061067:Antibodies, Monoclonal, Humanized / Q000494:pharmacology*; D000951:Antigens, Neoplasm / Q000235:genetics / Q000378:metabolism*; D000068258:Bevacizumab; D000071231:Carbonic Anhydrase IX; D002256:Carbonic Anhydrases / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects*; D005260:Female; D055785:Gene Knockdown Techniques; D005909:Glioblastoma / Q000378:metabolism; D045325:HCT116 Cells; D019073:HT29 Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D051379:Mice; D009336:Necrosis; D009368:Neoplasm Transplantation; D014162:Transfection; D014183:Transplantation, Heterologous
chemical_list: D020533:Angiogenesis Inhibitors; D061067:Antibodies, Monoclonal, Humanized; D000951:Antigens, Neoplasm; D000068258:Bevacizumab; C089226:CA9 protein, human; D000071231:Carbonic Anhydrase IX; D002256:Carbonic Anhydrases