Carbonic anhydrase 9

CA9
Available structures PDB Ortholog search: PDBe RCSB List of PDB id codes 2HKF, 3IAI, 5FL4, 5FL5, 5FL6
Identifiers
Aliases	CA9, CAIX, MN, carbonic anhydrase 9
External IDs	OMIM: 603179; MGI: 2447188; HomoloGene: 20325; GeneCards: CA9; OMA:CA9 - orthologs
Gene location (Human) Chr. Chromosome 9 (human)[1] Band 9p13.3 Start 35,673,928 bp[1] End 35,681,159 bp[1]
Gene location (Mouse) Chr. Chromosome 4 (mouse)[2] Band 4|4 A5 Start 43,506,966 bp[2] End 43,513,729 bp[2]
RNA expression pattern Bgee Human Mouse (ortholog) Top expressed in body of stomach gallbladder fundus cardia cerebellar hemisphere gonad right hemisphere of cerebellum right testis pylorus left testis Top expressed in epithelium of stomach mucous cell of stomach zygote pyloric antrum primary oocyte duodenum secondary oocyte large intestine jejunum colon More reference expression data BioGPS More reference expression data
Gene ontology Molecular function metal ion binding lyase activity zinc ion binding carbonate dehydratase activity protein binding carbonic anhydrase Cellular component nucleolus cell projection nucleus membrane basolateral plasma membrane microvillus membrane integral component of membrane plasma membrane Biological process response to hypoxia bicarbonate transport morphogenesis of an epithelium secretion response to testosterone regulation of transcription from RNA polymerase II promoter in response to hypoxia Sources:Amigo / QuickGO
Orthologs Species Human Mouse Entrez 768 230099 Ensembl ENSG00000107159 ENSMUSG00000028463 UniProt Q16790 Q8VHB5 RefSeq (mRNA) NM_001216 NM_139305 RefSeq (protein) NP_001207 NP_647466 Location (UCSC) Chr 9: 35.67 – 35.68 Mb Chr 4: 43.51 – 43.51 Mb PubMed search [3] [4]
Wikidata
View/Edit Human View/Edit Mouse

Carbonic anhydrase IX (CA9/CA IX) is an enzyme that in humans is encoded by the CA9 gene.^[5][6][7] It is one of the 14 carbonic anhydrase isoforms found in humans and is a transmembrane dimeric metalloenzyme with an extracellular active site that facilitates acid secretion in the gastrointestinal tract.^[8] CA IX is overexpressed in many types of cancer including clear cell renal cell carcinoma (RCC) as well as carcinomas of the cervix, breast and lung where it promotes tumor growth by enhancing tumor acidosis.^[9][10]

Function

Carbonic anhydrases (CAs) are a large family of zinc metalloenzymes that catalyze the reversible hydration of carbon dioxide. They participate in a variety of biological processes, including respiration, calcification, acid-base balance, bone resorption, and the formation of aqueous humor, cerebrospinal fluid, saliva, and gastric acid. They show extensive diversity in tissue distribution and in their subcellular localization.^[7]

CA IX is mainly expressed in the gastrointestinal tract where it facilitates acid secretion.^[11] The CA IX enzyme, along with the CA II enzyme, binds to Anion Exchanger 2 (AE2) which increases bicarbonate transport and maximizes the rate of acid secretion by gastric parietal cells.^[8]

Structure

CA IX is a transmembrane glycoprotein with an extracellular active site.^[9] The cytoplasmic tail of the enzyme contains three residues that may be phosphorylated (Thr-443, Ser-448, and Tyr-449) and participate in signal transduction.^[9][12] Phosphorylated tyrosine 449 can interact with PI3K which activates protein kinase B to affect cellular glucose metabolism.^[13]

Under physiological conditions, the enzyme exists as two nearly identical dimers.^[14] Both dimers are stabilized by two hydrogen bonds between Arg-137 and the Ala-127 carbonyl oxygen as well as many Van der Waals interactions.^[14] One dimer, however, has additional stabilization due to a disulfide bridge formed by two cysteine residues.^[14]

Two hydrogen bonds between Arg-137 and the Ala-127 carbonyl oxygen stabilize the CA IX dimer.

One face of the dimer contains proteoglycan (PG) domains-a feature that is unique from other CA enzymes- and the opposite face contains the C-termini which help the enzyme attach to the cell membrane.^[15] CA IX contains an N-linked glycosylation site bearing mannose-type glycan structures on Asn-309 as well as an O-linked glycosylation site on Thr-78.^[16]

CA IX dimer with its four distinct pairs of domains labeled: the proteoglycan domain (PG), catalytic domain (blue), transmembrane segment (TM) and the intracellular tail (IC).

Regulation

Expression of CA IX is primarily regulated at the transcriptional level.^[17] The promoter region of the CA9 gene contains an HRE (hypoxia responsive element) where HIF-1 can bind, which allows hypoxic conditions to increase CA IX expression.^[17] Expression can also be regulated post-translationally by metalloproteinases which cause shedding of the enzyme's ectodomain.^[18] Unlike other CA isozymes, CA IX is not inhibited by high lactate concentrations.^[19] However, it is inhibited by bicarbonate.^[19]

Clinical significance

CA IX is a transmembrane protein and is a tumor-associated carbonic anhydrase isoenzyme. It is over-expressed in VHL mutated clear cell renal cell carcinoma (ccRCC) and hypoxic solid tumors, but is low-expressed in normal kidney and most other normal tissues. It may be involved in cell proliferation and transformation. This gene is mapped to 9p13-p12.^[7]

CA IX is a cellular biomarker of hypoxia. Furthermore, recent studies examining the association between CA IX levels and various clinicopathological outcomes suggest that CA IX expression may also be a valuable prognostic indicator for overall survival^[20] although this association has been questioned.^[21]

CA IX shows high expression in carcinomas of the uterine cervix, kidney, oesophagus, lung, breast, colon, brain, and vulva compared to expression in few noncancerous tissues.^[22][11] Its overexpression in cancerous tissues compared to normal ones is due to hypoxic conditions in the tumor microenvironment caused by abnormal vasculature and subsequent transcriptional activation by HIF-1 binding.^[17] In clear cell renal carcinomas, CA IX shows high expression under normoxia due to a mutation in the VHL gene that normally negatively regulates HIF-1.^[22] Because of its overexpression in many types of cancer and low expression in normal tissues, CAIX has become a useful target for clear cell RCC and breast cancer tumor imaging in mice.^[23][24]

CA IX plays a very significant role in tumor acidification as it has very high catalytic activity with the highest rate of proton transfer of the known CAs.^[16] The enzyme converts carbon dioxide outside of the tumor into bicarbonate and protons, contributing to extracellular acidosis and promoting tumor growth by regulating the pH of the cytosol.^[10]

As a drug target

Because of its low expression in normal tissues and overexpression in many cancer tissues, CA IX has also become a desirable drug target. Girentuximab, an antibody that binds to CA IX, failed to improve disease-free as well as overall survival of patients with clear cell RCC in Phase III clinical trials.^[25]

However, a number of small molecules have been used to inhibit CA IX. The main classes of these inhibitors are inorganic anions, sulfonamides, phenols, and coumarins.^[15] Anions and sulfonamides inhibit CA IX by coordinating the zinc ion within CA IX while phenols bind to the zinc-coordinated water molecule.^[15] Coumarins serve as mechanism-based inhibitors that are hydrolyzed by the enzyme to form a cis-2-hydroxycinnamic acid derivative that then binds to the active site.^[26]

References

1. GRCh38: Ensembl release 89: ENSG00000107159 – Ensembl, May 2017
2. GRCm38: Ensembl release 89: ENSMUSG00000028463 – Ensembl, May 2017
3. "Human PubMed Reference:". National Center for Biotechnology Information, U.S. National Library of Medicine.
4. "Mouse PubMed Reference:". National Center for Biotechnology Information, U.S. National Library of Medicine.
5. Opavský R, Pastoreková S, Zelník V, Gibadulinová A, Stanbridge EJ, Závada J, Kettmann R, Pastorek J (May 1996). "Human MN/CA9 gene, a novel member of the carbonic anhydrase family: structure and exon to protein domain relationships". Genomics. 33 (3): 480–7. doi:10.1006/geno.1996.0223. PMID 8661007.
6. Nakagawa Y, Uemura H, Hirao Y, Yoshida K, Saga S, Yoshikawa K (October 1998). "Radiation hybrid mapping of the human MN/CA9 locus to chromosome band 9p12-p13". Genomics. 53 (1): 118–9. doi:10.1006/geno.1998.5483. PMID 9787087.
7. "Entrez Gene: CA9 carbonic anhydrase IX".
8. Morgan PE, Pastoreková S, Stuart-Tilley AK, Alper SL, Casey JR (August 2007). "Interactions of transmembrane carbonic anhydrase, CAIX, with bicarbonate transporters". American Journal of Physiology. Cell Physiology. 293 (2): C738-48. doi:10.1152/ajpcell.00157.2007. PMID 17652430.
9. Frost, Susan C.; McKenna, Robert (Oct 2013). Carbonic Anhydrase: Mechanism, Regulation, Links to Disease, and Industrial Applications. Springer Science & Business Media. ISBN 9789400773592.
10. Chiche J, Brahimi-Horn MC, Pouysségur J (April 2010). "Tumour hypoxia induces a metabolic shift causing acidosis: a common feature in cancer". Journal of Cellular and Molecular Medicine. 14 (4): 771–94. doi:10.1111/j.1582-4934.2009.00994.x. PMC 3823111. PMID 20015196.
11. "Tissue expression of CA9 - Summary - The Human Protein Atlas". www.proteinatlas.org. Retrieved 2019-03-14.
12. Ditte P, Dequiedt F, Svastova E, Hulikova A, Ohradanova-Repic A, Zatovicova M, Csaderova L, Kopacek J, Supuran CT, Pastorekova S, Pastorek J (December 2011). "Phosphorylation of carbonic anhydrase IX controls its ability to mediate extracellular acidification in hypoxic tumors". Cancer Research. 71 (24): 7558–67. doi:10.1158/0008-5472.CAN-11-2520. PMID 22037869.
13. Dorai T, Sawczuk IS, Pastorek J, Wiernik PH, Dutcher JP (December 2005). "The role of carbonic anhydrase IX overexpression in kidney cancer". European Journal of Cancer. 41 (18): 2935–47. doi:10.1016/j.ejca.2005.09.011. PMID 16310354.
14. Alterio V, Hilvo M, Di Fiore A, Supuran CT, Pan P, Parkkila S, Scaloni A, Pastorek J, Pastorekova S, Pedone C, Scozzafava A, Monti SM, De Simone G (September 2009). "Crystal structure of the catalytic domain of the tumor-associated human carbonic anhydrase IX". Proceedings of the National Academy of Sciences of the United States of America. 106 (38): 16233–8. Bibcode:2009PNAS..10616233A. doi:10.1073/pnas.0908301106. PMC 2752527. PMID 19805286.
15. De Simone G, Supuran CT (February 2010). "Carbonic anhydrase IX: Biochemical and crystallographic characterization of a novel antitumor target". Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics. 1804 (2): 404–9. doi:10.1016/j.bbapap.2009.07.027. PMID 19679200.
16. Hilvo M, Baranauskiene L, Salzano AM, Scaloni A, Matulis D, Innocenti A, Scozzafava A, Monti SM, Di Fiore A, De Simone G, Lindfors M, Jänis J, Valjakka J, Pastoreková S, Pastorek J, Kulomaa MS, Nordlund HR, Supuran CT, Parkkila S (October 2008). "Biochemical characterization of CA IX, one of the most active carbonic anhydrase isozymes". The Journal of Biological Chemistry. 283 (41): 27799–809. doi:10.1074/jbc.M800938200. PMID 18703501.
17. Tafreshi NK, Lloyd MC, Bui MM, Gillies RJ, Morse DL (2014). "Carbonic Anhydrase IX as an Imaging and Therapeutic Target for Tumors and Metastases". Carbonic Anhydrase: Mechanism, Regulation, Links to Disease, and Industrial Applications. Subcellular Biochemistry. Vol. 75. Springer Netherlands. pp. 221–54. doi:10.1007/978-94-007-7359-2_12. ISBN 9789400773585. PMC 4282494. PMID 24146382.
18. Zatovicova M, Sedlakova O, Svastova E, Ohradanova A, Ciampor F, Arribas J, Pastorek J, Pastorekova S (November 2005). "Ectodomain shedding of the hypoxia-induced carbonic anhydrase IX is a metalloprotease-dependent process regulated by TACE/ADAM17". British Journal of Cancer. 93 (11): 1267–76. doi:10.1038/sj.bjc.6602861. PMC 2361518. PMID 16278664.
19. Innocenti A, Vullo D, Scozzafava A, Supuran CT (February 2005). "Carbonic anhydrase inhibitors. Inhibition of isozymes I, II, IV, V, and IX with anions isosteric and isoelectronic with sulfate, nitrate, and carbonate". Bioorganic & Medicinal Chemistry Letters. 15 (3): 567–71. doi:10.1016/j.bmcl.2004.11.056. PMID 15664814.
20. Kirkpatrick JP, Rabbani ZN, Bentley RC, Hardee ME, Karol S, Meyer J, Oosterwijk E, Havrilesky L, Secord AA, Vujaskovic Z, Dewhirst MW, Jones EL (February 2008). "Elevated CAIX Expression is Associated with an Increased Risk of Distant Failure in Early-Stage Cervical Cancer". Biomarker Insights. 3: 45–55. doi:10.4137/bmi.s570. PMC 2688355. PMID 19578493.
21. Li J, Zhang G, Wang X, Li XF (2015). "Is carbonic anhydrase IX a validated target for molecular imaging of cancer and hypoxia?". Future Oncology. 11 (10): 1531–41. doi:10.2217/fon.15.11. PMC 4976829. PMID 25963430.
22. Pastorekova S, Ratcliffe PJ, Pastorek J (June 2008). "Molecular mechanisms of carbonic anhydrase IX-mediated pH regulation under hypoxia". BJU International. 101 Suppl 4 (s4): 8–15. doi:10.1111/j.1464-410X.2008.07642.x. PMID 18430116. S2CID 8780292.
23. Stillebroer AB, Franssen GM, Mulders PF, Oyen WJ, van Dongen GA, Laverman P, Oosterwijk E, Boerman OC (September 2013). "ImmunoPET imaging of renal cell carcinoma with (124)I- and (89)Zr-labeled anti-CAIX monoclonal antibody cG250 in mice". Cancer Biotherapy & Radiopharmaceuticals. 28 (7): 510–5. doi:10.1089/cbr.2013.1487. PMC 3741422. PMID 23697926.
24. Kijanka MM, van Brussel AS, van der Wall E, Mali WP, van Diest PJ, van Bergen En Henegouwen PM, Oliveira S (December 2016). "Optical imaging of pre-invasive breast cancer with a combination of VHHs targeting CAIX and HER2 increases contrast and facilitates tumour characterization" (PDF). EJNMMI Research. 6 (1): 14. doi:10.1186/s13550-016-0166-y. PMC 4747965. PMID 26860296.
25. Chamie K, Donin NM, Klöpfer P, Bevan P, Fall B, Wilhelm O, Störkel S, Said J, Gambla M, Hawkins RE, Jankilevich G, Kapoor A, Kopyltsov E, Staehler M, Taari K, Wainstein AJ, Pantuck AJ, Belldegrun AS (July 2017). "Adjuvant Weekly Girentuximab Following Nephrectomy for High-Risk Renal Cell Carcinoma: The ARISER Randomized Clinical Trial". JAMA Oncology. 3 (7): 913–920. doi:10.1001/jamaoncol.2016.4419. PMC 5824229. PMID 27787547.
26. Maresca A, Temperini C, Vu H, Pham NB, Poulsen SA, Scozzafava A, Quinn RJ, Supuran CT (March 2009). "Non-zinc mediated inhibition of carbonic anhydrases: coumarins are a new class of suicide inhibitors". Journal of the American Chemical Society. 131 (8): 3057–62. doi:10.1021/ja809683v. hdl:2158/594272. PMID 19206230. S2CID 207136680.

Further reading

• Nishimori I, Onishi S (2001). "Carbonic anhydrase isozymes in the human pancreas". Digestive and Liver Disease. 33 (1): 68–74. doi:10.1016/s1590-8658(01)80138-9. PMID 11303978.
• Pastoreková S, Závadová Z, Kostál M, Babusíková O, Závada J (April 1992). "A novel quasi-viral agent, MaTu, is a two-component system". Virology. 187 (2): 620–6. doi:10.1016/0042-6822(92)90464-Z. PMID 1312272.
• Pastorek J, Pastoreková S, Callebaut I, Mornon JP, Zelník V, Opavský R, Zat'ovicová M, Liao S, Portetelle D, Stanbridge EJ (October 1994). "Cloning and characterization of MN, a human tumor-associated protein with a domain homologous to carbonic anhydrase and a putative helix-loop-helix DNA binding segment". Oncogene. 9 (10): 2877–88. PMID 8084592.
• Závada J, Závadová Z, Pastoreková S, Ciampor F, Pastorek J, Zelník V (May 1993). "Expression of MaTu-MN protein in human tumor cultures and in clinical specimens". International Journal of Cancer. 54 (2): 268–74. doi:10.1002/ijc.2910540218. PMID 8486430. S2CID 19669856.
• Pastoreková S, Parkkila S, Parkkila AK, Opavský R, Zelník V, Saarnio J, Pastorek J (February 1997). "Carbonic anhydrase IX, MN/CA IX: analysis of stomach complementary DNA sequence and expression in human and rat alimentary tracts". Gastroenterology. 112 (2): 398–408. doi:10.1053/gast.1997.v112.pm9024293. PMID 9024293.
• Saarnio J, Parkkila S, Parkkila AK, Waheed A, Casey MC, Zhou XY, Pastoreková S, Pastorek J, Karttunen T, Haukipuro K, Kairaluoma MI, Sly WS (April 1998). "Immunohistochemistry of carbonic anhydrase isozyme IX (MN/CA IX) in human gut reveals polarized expression in the epithelial cells with the highest proliferative capacity". The Journal of Histochemistry and Cytochemistry. 46 (4): 497–504. doi:10.1177/002215549804600409. PMID 9524195.
• Ivanov SV, Kuzmin I, Wei MH, Pack S, Geil L, Johnson BE, Stanbridge EJ, Lerman MI (October 1998). "Down-regulation of transmembrane carbonic anhydrases in renal cell carcinoma cell lines by wild-type von Hippel-Lindau transgenes". Proceedings of the National Academy of Sciences of the United States of America. 95 (21): 12596–601. Bibcode:1998PNAS...9512596I. doi:10.1073/pnas.95.21.12596. PMC 22876. PMID 9770531.
• Grabmaier K, Vissers JL, De Weijert MC, Oosterwijk-Wakka JC, Van Bokhoven A, Brakenhoff RH, Noessner E, Mulders PA, Merkx G, Figdor CG, Adema GJ, Oosterwijk E (March 2000). "Molecular cloning and immunogenicity of renal cell carcinoma-associated antigen G250". International Journal of Cancer. 85 (6): 865–70. doi:10.1002/(SICI)1097-0215(20000315)85:6<865::AID-IJC21>3.0.CO;2-Q. PMID 10709109. S2CID 40122146.
• Kivelä AJ, Parkkila S, Saarnio J, Karttunen TJ, Kivelä J, Parkkila AK, Pastoreková S, Pastorek J, Waheed A, Sly WS, Rajaniemi H (September 2000). "Expression of transmembrane carbonic anhydrase isoenzymes IX and XII in normal human pancreas and pancreatic tumours". Histochemistry and Cell Biology. 114 (3): 197–204. doi:10.1007/s004180000181. PMID 11083462. S2CID 22170460.
• Wingo T, Tu C, Laipis PJ, Silverman DN (November 2001). "The catalytic properties of human carbonic anhydrase IX". Biochemical and Biophysical Research Communications. 288 (3): 666–9. doi:10.1006/bbrc.2001.5824. PMID 11676494.
• Kivela AJ, Saarnio J, Karttunen TJ, Kivelä J, Parkkila AK, Pastorekova S, Pastorek J, Waheed A, Sly WS, Parkkila TS, Rajaniemi H (October 2001). "Differential expression of cytoplasmic carbonic anhydrases, CA I and II, and membrane-associated isozymes, CA IX and XII, in normal mucosa of large intestine and in colorectal tumors". Digestive Diseases and Sciences. 46 (10): 2179–86. doi:10.1023/A:1011910931210. PMID 11680594. S2CID 40928937.
• Koukourakis MI, Giatromanolaki A, Sivridis E, Simopoulos K, Pastorek J, Wykoff CC, Gatter KC, Harris AL (November 2001). "Hypoxia-regulated carbonic anhydrase-9 (CA9) relates to poor vascularization and resistance of squamous cell head and neck cancer to chemoradiotherapy". Clinical Cancer Research. 7 (11): 3399–403. PMID 11705854.
• Grabmaier K, de Weijert M, Uemura H, Schalken J, Oosterwijk E (August 2002). "Renal cell carcinoma-associated G250 methylation and expression: in vivo and in vitro studies". Urology. 60 (2): 357–62. doi:10.1016/S0090-4295(02)01711-9. PMID 12137853.
• Kaluz S, Kaluzová M, Chrastina A, Olive PL, Pastoreková S, Pastorek J, Lerman MI, Stanbridge EJ (August 2002). "Lowered oxygen tension induces expression of the hypoxia marker MN/carbonic anhydrase IX in the absence of hypoxia-inducible factor 1 alpha stabilization: a role for phosphatidylinositol 3'-kinase". Cancer Research. 62 (15): 4469–77. PMID 12154057.
• Swinson DE, Jones JL, Richardson D, Wykoff C, Turley H, Pastorek J, Taub N, Harris AL, O'Byrne KJ (February 2003). "Carbonic anhydrase IX expression, a novel surrogate marker of tumor hypoxia, is associated with a poor prognosis in non-small-cell lung cancer". Journal of Clinical Oncology. 21 (3): 473–82. doi:10.1200/JCO.2003.11.132. PMID 12560438.
• Bui MH, Seligson D, Han KR, Pantuck AJ, Dorey FJ, Huang Y, Horvath S, Leibovich BC, Chopra S, Liao SY, Stanbridge E, Lerman MI, Palotie A, Figlin RA, Belldegrun AS (February 2003). "Carbonic anhydrase IX is an independent predictor of survival in advanced renal clear cell carcinoma: implications for prognosis and therapy". Clinical Cancer Research. 9 (2): 802–11. PMID 12576453.