CA IX Stabilizes Intracellular pH to Maintain Metabolic Reprogramming and Proliferation in Hypoxia

Abstract

Tumor hypoxia represents a severe microenvironmental stress that is frequently associated with acidosis. Cancer cells respond to these stresses with changes in gene expression that promote survival at least in part through pH regulation and metabolic reprogramming. Hypoxia-induced carbonic anhydrase IX (CA IX) plays a critical adaptive role in response to hypoxic and acidic environments by catalytically hydrating extracellular CO 2 to produce bicarbonate for buffering intracellular pH (pHi). We used proteome-wide profiling to study the cellular response to transient CA IX knockdown in hypoxia and found a decrease in the levels of key glycolytic enzymes and lactate dehydrogenase A (LDHA). Interestingly, the activity of LDH was also decreased as demonstrated by native in-gel activity assay. These changes led to a significant reduction in glycolytic flux and extracellular lactate levels in cancer cells in vitro , contributing to a decrease in proliferation. Interestingly, addition of the alternative LDH substrate alpha-ketobutyrate restored LDHA activity, extracellular acidification, pHi, and cellular proliferation. These results indicate that in the absence of CA IX, reduction of pHi disrupts LDHA activity and hinders the cellular capacity to regenerate NAD + and secrete protons to the extracellular space. Hypoxia-induced CA IX therefore mediates adaptation to microenvironmental hypoxia and acidosis directly, by enzymatically converting extracellular CO 2 to bicarbonate, and indirectly, by maintaining glycolysis-permissive intracellular milieu.

<p>Experimental Carbonic Anhydrase Inhibitors for the Treatment of Hypoxic Tumors</p>

“…Two of the 15 known human (h) CA isoforms, hCA IX 58 and XII 59 are predominantly found in tumor cells and show a rather limited diffusion in normal cells. Both isoforms are multi-domain trans-membrane proteins with an extracellular CA domain, and were demonstrated to participate in the rather complex machinery of pH regulation, 57–63 which as mentioned above, is dysregulated in cancer cells due to the activation of HIF-1/2. It should also be noted that these two enzymes are just a part of the complex network of proteins/processes which regulate pH and metabolism in tumor cells, most of which were mentioned in the Introduction.…”

Hypoxia Signaling in Cancer: From Basics to Clinical Practice

“…The uptake and the release of extracellular metabolites and their transporters can also be inhibited in these metabolic alterations. Moreover, various metabolite transporter proteins–involving monocarboxylate transporters (MCTs), which contribute to lactate transport–can also be tested as a part of anti-angiogenic treatment combination [ 38 , 39 , 103 , 104 ].…”

The role of metabolic ecosystem in cancer progression — metabolic plasticity and mTOR hyperactivity in tumor tissues

“…The hypoxic or pseudohypoxic changes induce the Warburg effect; therefore, the concentration of glucose, lactate, and other metabolic intermediates (e.g., carbohydrates, ribose, specific lipids, FAs, acetate, and amino acids) can vary in cancer microenvironments. Increased lactate production and/or O 2 consumption resulting CO 2 release and their transport mechanisms (using MCTs and carbonic anhydrase IX (CAIX) transporters) also contribute to lowered pH in the tumor mass [54,55].…”