Acidic tissue microenvironment contributes to tumor progression via multiple effects including the activation of angiogenic factors and proteases, reduced cell-cell adhesion, increased migration and invasion, etc. In addition, intratumoral acidosis can influence the uptake of anticancer drugs and modulate the response of tumors to conventional therapy. Acidification of the tumor microenvironment often develops due to hypoxia-triggered oncogenic metabolism, which leads to the extensive production of lactate, protons and carbon dioxide. In order to avoid intracellular accumulation of the acidic metabolic products, which is incompatible with the survival and proliferation, tumor cells activate molecular machinery that regulates pH by driving transmembrane inside-out and outside-in ion fluxes. Carbonic anhydrase IX (CA IX) is a hypoxia-induced catalytic component of the bicarbonate import arm of this machinery. Through its catalytic activity, CA IX directly participates in many acidosis-induced features of tumor phenotype as demonstrated by manipulating its expression and/or by in vitro mutagenesis. CA IX can function as a survival factor protecting tumor cells from hypoxia and acidosis, as a pro-migratory factor facilitating cell movement and invasion, as a signaling molecule transducing extracellular signals to intracellular pathways (including major signaling and metabolic cascades) and converting intracellular signals to extracellular effects on adhesion, proteolysis and other processes. These functional implications of CA IX in cancer are supported by numerous clinical studies demonstrating the association of CA IX with various clinical correlates and markers of aggressive tumor behavior. Although our understanding of the many faces of CA IX is still incomplete, existing knowledge supports the view that CA IX is a biologically and clinically relevant molecule, exploitable in anticancer strategies aimed at targeting adaptive responses to hypoxia and/or acidosis.

Solid tumors, including breast cancer, are characterized by the hypoxic microenvironment, extracellular acidosis, and chemoresistance. Hypoxia marker, carbonic anhydrase IX (CAIX), is a pH regulator providing a selective survival advantage to cancer cells through intracellular neutralization while facilitating tumor invasion by extracellular acidification. The expression of CAIX in breast cancer patients is associated with poor prognosis and metastases. Importantly, CAIX-positive hypoxic tumor regions are enriched in cancer stem cells (CSCs). Here we investigated the biological effects of CA9-silencing in breast cancer cell lines. We found that CAIX-downregulation in hypoxia led to increased levels of let-7 (lethal-7) family members. Simultaneously with the increase of let-7 miRNAs in CAIX-suppressed cells, LIN28 protein levels decreased, along with downstream metabolic pathways: pyruvate dehydrogenase kinase 1 (PDK1) and phosphorylation of its substrate, pyruvate dehydrogenase (PDH) at Ser-232, causing attenuation of glycolysis. In addition to perturbed glycolysis, CAIX-knockouts, in correlation with decreased LIN28 (as CSC reprogramming factor), also exhibit reduction of the further CSC-associated markers NANOG (Homeobox protein NANOG) and ALDH1 (Aldehyde dehydrogenase isoform 1). Oppositely, overexpression of CAIX leads to the enhancement of LIN28, ALDH1, and NANOG. In conclusion, CAIX-driven regulation of the LIN28/let-7 axis augments glycolytic metabolism and enhances stem cell markers expression during CAIX-mediated adaptation to hypoxia and acidosis in carcinogenesis.

Hypoxia is a common phenomenon that occurs in most solid tumors. Regardless of tumor origin, the evolution of a hypoxia-adapted phenotype is critical for invasive cancer development. Pancreatic ductal adenocarcinoma is also characterized by hypoxia, desmoplasia, and the presence of necrosis, predicting poor outcome. Carbonic anhydrase IX (CAIX) is one of the most strict hypoxia regulated genes which plays a key role in the adaptation of cancer cells to hypoxia and acidosis. Here, we summarize clinical data showing that CAIX expression is associated with tumor necrosis, vascularization, expression of Frizzled-1, mucins, or proteins involved in glycolysis, and inevitably, poor prognosis of pancreatic cancer patients. We also describe the transcriptional regulation of CAIX in relation to signaling pathways activated in pancreatic cancers. A large part deals with the preclinical evidence supporting the relevance of CAIX in processes leading to the aggressive behavior of pancreatic tumors. Furthermore, we focus on CAIX occurrence in pre-cancerous lesions, and for the first time, we describe CAIX expression within intraductal papillary mucinous neoplasia. Our review concludes with a detailed account of clinical trials implicating that treatment consisting of conventionally used therapies combined with CAIX targeting could result in an improved anti-cancer response in pancreatic cancer patients.

In contrast to human carbonic anhydrase IX (hCA IX) that has been extensively studied with respect to its molecular and functional properties as well as regulation and expression, the mouse ortholog has been investigated primarily in relation to tissue distribution and characterization of CA IX-deficient mice. Thus, no data describing transcriptional regulation and functional properties of the mouse CA IX (mCA IX) have been published so far, despite its evident potential as a biomarker/target in pre-clinical animal models of tumor hypoxia. Here, we investigated for the first time, the transcriptional regulation of the Car9 gene with a detailed description of its promoter. Moreover, we performed a functional analysis of the mCA IX protein focused on pH regulation, cell–cell adhesion, and migration. Finally, we revealed an absence of a soluble extracellular form of mCA IX and provided the first experimental evidence of mCA IX presence in exosomes. In conclusion, though the protein characteristics of hCA IX and mCA IX are highly similar, and the transcription of both genes is predominantly governed by hypoxia, some attributes of transcriptional regulation are specific for either human or mouse and as such, could result in different tissue expression and data interpretation.

Oxygen is absolutely essential for correct functioning of living organisms and alterations in its concentration lead to serious consequences. In tumor tissues, oxygen plays an important role in energy production and modulation of red- ox balance. Insufficient oxygen supply within tissues results in hypoxia that is a characteristic feature of the tumor microenvironment. Hypoxia- inducible transcriptional factor represents a key executor of a cellular and molecular response to hypoxia and can activate the expression of more than hundred genes involved in various essential cellular processes. From the clinical point of view, phenotypic alterations caused by hypoxia are serious. Tumor hypoxia has been associated with resistance to therapy, disease progression and recurrence as well as increased mortality. Therefore, intratumoral hypoxia represents a clinically relevant problem, and its detection within tumors is very important for patient stratification for a suitable treatment. Currently available strategies directed towards the detection of hypoxic regions within tumor tissue suffer from numerous limitations e. g. invasiveness, inaccessibility of tumor tissue, low sensibility, inaccurate interpretation etc. On the other hand, the use of an intrinsic endogenous hypoxic marker, which can be detected through immunohistochemistry, is relatively simple, routinely available, and reproducible and can be performed on both prospective and retrospective samples. These include carbonic anhydrase IX (CA IX), one of the most strongly hypoxia-induced proteins and a prominent indicator of chronic hypoxia. Moreover, hypoxia-induced proteins (including CA IX) are also potential targets of anticancer therapy, and their practical application is a subject of intense research.