Hypoxia activates the capacity of tumor‐associated carbonic anhydrase IX to acidify extracellular pH

Abstract

Acidic extracellular pH (pHe) is a typical attribute of a tumor microenviroment, which has an impact on cancer development and treatment outcome. It was believed to result from an accumulation of lactic acid excessively produced by glycolysis. However, metabolic profiles of glycolysis-impaired tumors have revealed that CO 2 is a significant source of acidity, thereby indicating a contribution of carbonic anhydrase (CA). The tumor-associated CA IX isoform is the best candidate, because its extracellular enzyme domain is highly active, expression is induced by hypoxia and correlates with poor prognosis. This study provides the first evidence for the role of CA IX in the control of pHe. We show that CA IX can acidify the pH of the culture medium in hypoxia but not in normoxia. This acidification can be perturbed by deletion of the enzyme active site and inhibited by CA IX-selective sulfonamides, which bind only to hypoxic cells containing CA IX. Our findings suggest that hypoxia regulates both expression and activity of CA IX in order to enhance the extracellular acidification, which may have important implications for tumor progression.

Alternative splicing variant of the hypoxia marker carbonic anhydrase IX expressed independently of hypoxia and tumour phenotype

“…Antimouse FITC-conjugated antibodies were from Vector Laboratories (Burlingame, CA, USA). Immunofluorescence was performed as before (Svastova et al, 2004). Antibodies against the mouse CA IX protein were described elsewhere (Gut et al, 2002).…”

“…The fluorescent CA inhibitor (FITC-CAI) was obtained by reaction of homosulphanilamide with fluorescein isothiocyanate and showed a K I value of 24 nM towards CA IX (Svastova et al, 2004;Cecchi et al, 2005). The inhibitor was dissolved in PBS with 20% DMSO at 100 mM concentration and diluted in a culture medium to a final 1 mM concentration just before addition to cells.…”

“…The inhibitor was dissolved in PBS with 20% DMSO at 100 mM concentration and diluted in a culture medium to a final 1 mM concentration just before addition to cells. The MDCK-CA IX cells (Svastova et al, 2004) were plated at a density of 400 000 cells per 3.5 cm dish in the medium containing the conditioned medium from MDCK-AS transfectants that secrete the human AS variant. Control cells were incubated in the absence of secreted AS.…”

“…Moreover, CA IX also appears to play an active role in tumour biology via modulation of cell adhesion and control of pH Svastova et al, 2004;Swietach et al, 2007). CA IX participates in bicarbonate transport metabolon and contributes to acidification of extracellular microenvironment in response to hypoxia (Svastova et al, 2004;Morgan et al, 2007).…”

Role of the HBx oncoprotein in carbonic anhydrase 9 induction

“…CA IX is one of the most hypoxia-responsive genes and was proposed to serve as an intrinsic marker of hypoxia [Potter and Harris, 2003]. It catalyzes the reversible conversion of CO 2 to bicarbonate ion and proton like other active carbonic anhydrases, and participates in ion transport and pH control [Svastova et al, 2004]. Additionally, CA IX also reduces E-cadherin-mediated adhesion by binding to β-catenin [Svastova et al, 2003].…”