Acidic metabolic waste products accumulate in the tumor microenvironment because of high metabolic activity and insufficient perfusion. In tumors, the acidity of the interstitial space and the relatively well-maintained intracellular pH influence cancer and stromal cell function, their mutual interplay, and their interactions with the extracellular matrix. Tumor pH is spatially and temporally heterogeneous, and the fitness advantage of cancer cells adapted to extracellular acidity is likely particularly evident when they encounter less acidic tumor regions, for instance, during invasion. Through complex effects on genetic stability, epigenetics, cellular metabolism, proliferation, and survival, the compartmentalized pH microenvironment favors cancer development. Cellular selection exacerbates the malignant phenotype, which is further enhanced by acid-induced cell motility, extracellular matrix degradation, attenuated immune responses, and modified cellular and intercellular signaling. In this review, we discuss how the acidity of the tumor microenvironment influences each stage in cancer development, from dysplasia to full-blown metastatic disease.

The contribution of sodium-dependent bicarbonate transport to intracellular pH (pH _i ) regulation in vascular smooth muscle cells is controversial, partly because the molecular identity of the transporter(s) responsible has not been identified. Here, using the pH-sensitive fluorophore 2′,7′-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein (BCECF), we show that smooth muscle cells of intact mouse mesenteric, coronary, and cerebral small arteries all display a sodium- and bicarbonate-dependent pH _i recovery after an NH ₄ ⁺ -prepulse. The sodium-dependent bicarbonate flux was largely 4,4′-diisothiocyanatostilbene-2,2′-disulphonic acid (DIDS) sensitive (56% to 91%) and of a magnitude similar to the amiloride-sensitive flux. Additionally, steady-state pH _i was lower (0.2 to 0.4 pH units magnitude) in all 3 vascular beds when CO ₂ /bicarbonate was omitted. RT-PCR analyses showed that NBCn1 (slc4a7) is the only Na ⁺ -dependent bicarbonate transporter of the slc4 family detectable at the mRNA level in all 3 vascular beds investigated. Whole-mount immunolabeling and immunogold electron microscopy confirmed the presence of NBCn1 protein in the sarcolemma of mouse mesenteric small arterial smooth muscle cells. Intact mouse mesenteric small arteries were electropermeated to facilitate transfection with small interfering RNA targeting NBCn1, which resulted in an approximate 43% decrease in the ratio of NBCn1 to glyceraldehyde-3-phosphate dehydrogenase mRNA. After knock-down, we found a decreased steady-state pH _i (0.21±0.08 pH units) as well as a 68±10% decrease in the net Na ⁺ -dependent, amiloride-insensitive base influx after acid load. Finally, omission of CO ₂ /bicarbonate resulted in a decreased contractile response to norepinephrine after sustained exposure to the agonist, underlining the importance of CO ₂ /bicarbonate for vascular contractility. We conclude that NBCn1 mediates the Na ⁺ -dependent bicarbonate transport important for pH _i regulation in smooth muscle cells of mouse mesenteric, coronary, and cerebral small arteries.

A unifying feature of solid tumors is a markedly altered pH profile compared to normal tissues. This reflects that solid tumors, despite completely different origins, often share several phenotypic properties with implications for intra- and extracellular pH. These include: a metabolic shift in most cancer cells towards more acid-producing pathways, reflecting both oncogenic signaling and the development of hypoxia in poorly perfused regions of the tumors; the poorly perfused and often highly dense tumor microenvironment, reducing the diffusive flux of acid equivalents compared to that in normal tissues; and the markedly altered regulation of the expression and activity of pH-regulatory transport proteins in the cancer cells. While some of these properties of tumors have been well described in recent years, the great majority of the research in this clinically important area has focused on proton transport, in particular via the Na+/H+-exchanger 1 (SLC9A1, NHE1) and various H+ ATPases. We have, however, recently demonstrated that at least under some conditions, including in vitro models of HER2 positive breast cancer, and measurements obtained directly in freshly dissected human mammary tumors, bicarbonate transporters such as the electroneutral Na+,HCO3--cotransporter (SLC4A7, NBCn1), are upregulated and play central roles in pH regulation. In this review, we summarize and discuss the current knowledge regarding the regulation and roles of bicarbonate transport in cancer.

Background—: Disturbances in pH affect artery function, but the mechanistic background remains controversial. We investigated whether Na ⁺ ,HCO ₃ ⁻ cotransporter NBCn1, by regulating intracellular pH (pH _i ), influences artery function and blood pressure regulation. Methods and Results—: Knockout of NBCn1 in mice eliminated Na ⁺ ,HCO ₃ ⁻ cotransport and caused a lower steady-state pH _i in mesenteric artery smooth muscle and endothelial cells in situ evaluated by fluorescence microscopy. Using myography, arteries from NBCn1 knockout mice showed reduced acetylcholine-induced NO-mediated relaxations and lower rho-kinase-dependent norepinephrine-stimulated smooth muscle Ca ²⁺ sensitivity. Acetylcholine-stimulated NO levels (electrode measurements) and N-nitro-l-arginine methyl ester–sensitive l-arginine conversion (radioisotope measurements) were reduced in arteries from NBCn1 knockout mice, whereas relaxation to NO-donor S-nitroso-N-acetylpenicillamine, acetylcholine-induced endothelial Ca ²⁺ responses (fluorescence microscopy), and total and Ser-1177 phosphorylated endothelial NO-synthase expression (Western blot analyses) were unaffected. Reduced NO-mediated relaxations in arteries from NBCn1 knockout mice were not rescued by superoxide scavenging. Phosphorylation of myosin phosphatase targeting subunit at Thr-850 was reduced in arteries from NBCn1 knockout mice. Evaluated by an in vitro assay, rho-kinase activity was reduced at low pH. Without CO ₂ /HCO ₃ ⁻ , no differences in pH _i , contraction or relaxation were observed between arteries from NBCn1 knockout and wild-type mice. Based on radiotelemetry and tail-cuff measurements, NBCn1 knockout mice were mildly hypertensive at rest, displayed attenuated blood pressure responses to NO-synthase and rho-kinase inhibition and were resistant to developing hypertension during angiotensin-II infusion. Conclusions—: Intracellular acidification of smooth muscle and endothelial cells after knockout of NBCn1 inhibits NO-mediated and rho-kinase–dependent signaling in isolated arteries and perturbs blood pressure regulation.

Genome‐wide association studies recently linked the locus for Na⁺,HCO₃⁻‐cotransporter NBCn1 (SLC4A7) to breast cancer susceptibility, yet functional insights have been lacking. To determine whether NBCn1, by transporting HCO₃⁻ into cells, may dispose of acid produced during high metabolic activity, we studied the expression of NBCn1 and the functional impact of Na⁺,HCO₃⁻‐cotransport in human breast cancer. We found that the plasmalemmal density of NBCn1 was 20–30% higher in primary breast carcinomas and metastases compared to matched normal breast tissue. The increase in NBCn1 density was similar in magnitude to that observed for Na⁺/H⁺‐exchanger NHE1 (SLC9A1), a transporter previously implicated in cell migration, proliferation and malignancy. In primary breast carcinomas, the apparent molecular weight for NBCn1 was increased compared to normal tissue. Using pH‐sensitive fluorophores, we showed that Na⁺,HCO₃⁻‐cotransport is the predominant mechanism of acid extrusion and is inhibited 34 ± 9% by 200 μM 4,4′‐diisothiocyanatostilbene‐2,2′‐disulfonic acid in human primary breast carcinomas. At intracellular pH (pH_i) levels >6.6, CO₂/HCO₃⁻‐dependent mechanisms accounted for >90% of total net acid extrusion. Na⁺/H⁺‐exchange activity was prominent only at lower pH_i‐values. Furthermore, steady‐state pH_i was 0.35 ± 0.06 units lower in the absence than in the presence of CO₂/HCO₃⁻. In conclusion, expression of NBCn1 is upregulated in human primary breast carcinomas and metastases compared to normal breast tissue. Na⁺,HCO₃⁻‐cotransport is a major determinant of pH_i in breast cancer and the modest DIDS‐sensitivity is consistent with NBCn1 being predominantly responsible. Hence, our results suggest a major pathophysiological role for NBCn1 that may be clinically relevant.