New fluorinated fructose analogs as selective probes of the hexose transporter protein GLUT5

Abstract

Facilitated hexose transporters (GLUTs) mediate the transport of hexoses and other substrates across the membranes of numerous cell types, and while some are expressed ubiquitously (e.g., GLUT1), others are more tissue specific (e.g., GLUT5). These properties have been exploited for the imaging of cancer cells by the use of hexose based probes, including fluorinated hexose derivatives for use with positron emission tomography (PET). However, design of new probes has been hampered by a limited understanding of how GLUT transporters interact with their substrates at the molecular level. Two fluorinated fructose surrogates designed for uptake by the GLUT5 transporter are described here: 3-deoxy-3-fluoro-D-fructose (3-FDF) and 1-deoxy-1-fluoro-2,5-anhydromannitol (1-FDAM). Synthesis (both cold and radiolabeled) and in vitro analysis of their transport characteristics in two breast cancer cell lines (EMT-6 and MCF-7) expressing GLUT5 are detailed. Both analogues are readily taken up into both cancer cell lines, with uptake mediated primarily by GLUT5. They also have low IC50 values, indicating a high affinity for the transporter, suggesting that the uptake of these probes would be unaffected by endogenously circulating fructose. Selective uptake by GLUT5 was also demonstrated in Xenopus oocytes. Finally, these results are the first demonstration that a hexose existing predominantly in the pyranose ring structure (3-FDF) is transported by GLUT5, strongly suggesting that this transporter can handle both furanose and pyranose forms of fructose.

d‐Fructose Modification Enhanced Internalization of Mixed Micelles in Breast Cancer Cells via GLUT5 Transporters

“…In general, the cellular uptake could be competitively inhibited by preincubation of free d ‐fructose indicating that PPF MM internalization was partially mediated by GLUT5. The internalization of PPN MM and PPF MM displayed no significant differences with and without free d ‐fructose in L929 cells ( p > 0.05, Figure B), suggesting that PPF MM internalization in L929 cells was not mediated through GLUT5, but likely through other transporters/receptors such as GLUT2 . Also, PPF MM showed higher cellular uptake on L929 cells than PPN MM (Figure S17, Supporting Information), which could be due to other pathways involved in the internalization of PPF MM.…”

“…Together, these results revealed that GLUT5 recognizes d ‐fructose located on the surface of PPF MM thus inducing GLUT5‐mediated internalization of PPF MM. However, the presence of free d ‐fructose cannot completely inhibit the internalization of PPF MM suggesting some other transporters/receptors such as GLUT2‐mediated pathway be involved in the internalization of PPF MM …”

“…Next, Zhao et al fabricated rod‐shaped fructose micelles of varying structures and size distributions which showed length‐dependent cellular uptake and movement behavior in vitro . Also, fluorinated fructose analogues have been developed for positron emission tomography‐imaging in cancer diagnosis of GLUT5‐overexpressing tumors . Carboxyl end‐functionalized poly(1‐ O ‐methacryloyl‐2,3:4,5‐di‐ O ‐isopropylidene‐β‐ d ‐fructopyranose) was prepared and grafted onto the surface of amine‐functionalized nanodiamonds to be applied to human breast cancer therapy .…”

Tunable GLUT–Hexose Binding and Transport via Modulation of Hexose C‐3 Hydrogen‐Bonding Capabilities

“…Although 4 showed no effect on GLUT5‐mediated transport of [ 14 C]‐ d ‐fructose (Figure ), it did inhibit [ 14 C]‐ d ‐glucose uptake into EMT‐6 cells (IC 50 =3.1±0.3 m m , Figure ). This represents the first example of an apparent target of a d ‐glucose‐specific transporter protein by a derivative of 2,5‐AM; compounds of this class have previously been shown to target d ‐fructose transporters such as GLUT5 . Inhibition of [ 14 C]‐ d ‐glucose transport by 3 and 5 was also examined.…”

“…Early structure–activity relationship studies demonstrated the necessity of secondary hydroxyl groups at the C‐3, C‐4 and C‐5 positions of d ‐fructose for GLUT5‐mediated transport . However, the hydroxyl group at C‐6 of d ‐fructose derivatives plays a negligible role in binding with GLUT5 (Figure ) . Consequently, GLUT5 targeting probes have been developed by the modification of the C‐6 position; however, these probes suffer from rapid efflux, presumably because of the absence of the C‐6 hydroxyl group, whose removal precludes metabolic trapping through phosphorylation by hexokinase …”