Identification of an alternatively spliced isoform of carbonic anhydrase XII in diffusely infiltrating astrocytic gliomas

Abstract

Carbonic anhydrase XII (CA XII) is a transmembrane enzyme that is associated with neoplastic growth. CA XII has been proposed to be involved in acidification of the extracellular milieu, creating an appropriate microenvironment for rapid tumor growth. Because RNA sequence databases have indicated that two isoforms of CA XII might exist in human tissues, and because alternatively spliced protein forms have been linked to aggressive behavior of cancer cells, we designed a study to evaluate the presence of the two forms of CA XII in diffuse astrocytomas, a tumor type known for its aggressive and often noncurable behavior. Reverse transcription PCR of tumor samples surprisingly revealed that CA XII present in diffuse astrocytomas is mainly encoded by a shorter mRNA variant. We further showed by Western blotting that anti-CA XII antibody recognized both isoforms in the glioblastoma cell lines, and we then evaluated the expression of CA XII in astrocytomas using immunohistochemistry and correlated the results with various clinicopathological and molecular factors. Of 370 diffusely infiltrating astrocytomas, 363 cases (98%) showed immunoreactions for CA XII. Importantly, CA XII expression correlated with poorer patient prognosis in univariate (p = 0.010, log-rank test) and multivariate survival analyses (p = 0.039, Cox analysis). From these results, we conclude that CA XII is commonly expressed in diffuse astrocytomas and that it might be used as a biomarker of poor prognosis. The absence of 11 amino acids in the shorter isoform, which seems to be common in astrocytomas, may affect the normal quaternary structure and biological function of CA XII.

Carbonic anhydrase VII–a potential prognostic marker in gliomas

“…This result makes CA VII another CA isozyme with a significant correlation to the patient survival. Indeed, the previous studies have demonstrated that CA II, CA IX and CA XII all show similar trends as prognostic markers in diffuse astrocytomas [6,7,25,39]. The presence of several CA isozymes in astrocytomas may reflect the rapid turnover of acid metabolic products in highly malignant tumor tissues.…”

“…There are numerous reports showing that CA IX acts not only as a marker for particular tumors, but its presence also correlates to prognosis in several tumor categories, such as brain tumors, sarcomas, and renal, lung, rectal, bladder, oral, breast, and cervical carcinomas [7,[27][28][29][30][31][32][33][34]. Although studies on CA II and CA XII have been mainly focused on normal tissues, there are a few previous reports, showing their presence in cancer cells and especially in brain tumors where they have shown significant association with prognosis [6,7,25]. CA VII is a cytosolic isozyme which was first demonstrated in the brain tissue [35], where it has been linked to the regulation of GABAergic neuronal transmission in hippocampal neurons [36].…”

“…EGFR amplification was detected in astrocytic tumors using chromogenic in situ hybridization (CISH) [24]. The immunostaining results for CA IX and CA XII have been described earlier [7,25].…”

New insights into the physiological role of carbonic anhydrase IX in tumour pH regulation

“…Growth of tumour xenografts was also retarded by albumin-binding acetazolamide derivatives that target extracellular CA isoforms (Ahlskog et al, 2009). These dramatic effects illustrate the importance of extracellular CA activity and may explain the correlation between CAIX/CAXII expression in tumour tissue samples and poor patient prognosis (Saarnio et al, 1998;Liao and Stanbridge, 2000;Chia et al, 2001;Koukourakis et al, 2001;Loncaster et al, 2001;Potter and Harris, 2003;Haapasalo et al, 2006;Haapasalo et al, 2008). These results make a case for clinical trials for studying the responsiveness of tumours in patients to anti-CAIX/anti-CAXII therapies based on small molecule inhibitors (Supuran, 2008;Guler et al, 2010) or selective antibodies with inhibitory effects (Xu et al, 2010).…”

Synthesis and inhibition potency of novel ureido benzenesulfonamides incorporating GABA as tumor-associated carbonic anhydrase IX and XII inhibitors

“…Among the various biological functions, CAs are involved in pH buffering of the intra-and extra-cellular spaces [4][5][6][7] . Mainly two carbonic anhydrase isoforms hCA IX [8][9][10] and CA XII 11,12 , possessing extracellular active site, are over-expressed in most of hypoxic tumors, such as gliomas/ ependymomas, mesotheliomas, carcinomas of the bladder, cervix, kidneys, lungs, breast, and other dysplasia resistant to classic chemo-and radiotherapy 7,13,14 . Indeed, in response to hypoxia event, such as the one occuring in the tumor area, the activated hypoxia inducible factor-1 (HIF-1) causes an over-expression of hCA IX 6 , which increases the production of protons in the district involved.…”