Expression of hexokinase II and Glut-1 in untreated human breast cancer

Abstract

Expressions of HKII and Glut-1 were studied in untreated primary human breast cancers by immunohistochemistry. 79% of the breast cancers were HKII-positive and 61% were Glut-1-positive. Average positive malignant cell areas were 66 ± 41% for HKII and 29 ± 36% for Glut-1. HKII staining was cytoplasmic, suggesting mitochondrial localization with no variations in staining intensities. Glut-1 staining was heterogeneous, cytoplasmic and membranous and varied with histology and tumor stage. Cells expressing HKII did not always express Glut-1 and vice versa. Increased FDG-uptake appeared to be associated with increased Glut-1 expression (P = 0.021), but not with HKII expression (p = 0.6).

FDG uptake in breast cancer tissue appears to be associated with the extent of immunodetectable expression of Glut-1, but not that of HKII, and FDG uptake may differ between individual tumors depending on tumor stage and histology.

Introduction

The high rates of glucose uptake and metabolism often displayed by malignant tumors are reported to be associated with increased expression of glucose transporter 1 (Glut-1) and increased activity of glycolytic enzymes. The Glut-1 mediated glucose influx and the increased phosphorylation rates enable cancer cells to maintain their high growth rates and metabolic activity. These characteristics of malignant tumors have been widely utilized to diagnose and monitor response to treatment of cancers by imaging their uptake of ¹⁸F-labeled glucose analog 2-fluoro-2-deoxy-D-glucose (FDG) with PET. It is currently accepted that increased influx of the glucose analog, higher rates of phosphorylation and diminished rates of dephosphorylation of the intracellular phosphorylated sugar result in intracellular accumulation of the polar FDG-6-phosphate (FDG-P) in cancer cells.

Hexose enters cancer cells by facilitated diffusion mediated primarily by Glut-1 and is then converted to hexose-6-phosphate by hexokinase (HK). There are four isoenzymes of HK in mammalian tissues designated as type-I, II, III, and IV (the latter is usually referred to as glucokinase). HKI, II and III are similar in their molecular masses (approximately 100 kDa), have high glucose affinity and are susceptible to product inhibition by glucose-6-phosphate. HKI and HKII are associated with mitochondria while HKIII is localized at the periphery of the nucleus. Glucokinase is primarily located in the liver and pancreatic beta cells, has a molecular mass of 50 kDa, has lower affinity to glucose and is not susceptible to inhibition by glucose-6-phosphate [31].

As the first glycolytic enzyme, HK can play a key role in sugar metabolism of malignant cells and it has been suggested that HKs, HKII in particular, regulate glucose metabolism in cancer cells [16]. Indeed, it has been shown that HK specific activity is higher in tissue homogenates of malignant tumors as compared to benign tumors or normal tissue [22]. In human breast cancers, a 13-fold increase in HK specific activity as compared to normal breast [11], as well as intratumoral and intertumoral heterogeneity in specific activity of HK [12], were found. Gudnason et al [10] have reported increased expression of HKI and appearance of HKII in human malignant breast tumors. In addition, there are data suggesting that phosphorylation may be the rate-limiting step for FDG-uptake in human breast cancers in vivo [28]. The measured levels of HK specific activity in tumor homogenates, as well as the rates of FDG-uptake can be modified by factors other than increased expression or enzymatic activity (e.g. tumor cellularity, tumor necrosis or the extent of non-malignant elements). Hence, they may not accurately reflect the levels of HK isoenzymes in the cancer cells per se.

The intratumoral expression of glucose transporters in cells of human breast cancers has been described previously [6], [33] but studies describing the expression of hexokinases in cancer cells in tissues of human breast or other cancers are lacking. In this communication we report the results of an immunohistochemical study of the intratumoral expression of HKII in untreated primary breast cancer in tissue sections from breast cancer patients and in normal breast tissues from patients undergoing breast reduction. The relationships between the expression of HKII and Glut-1, as well as possible association with FDG uptake in human breast cancer, were assessed.