Synthesis, biological evaluation, and screening of 2,5-anhydro-D-mannitol derivatives towards selective targeting and imaging of GLUT5 transporter protein

• Author / Creator

  Rana, Natasha

• The solute carrier (SLC) family of membrane proteins is responsible for the internalization of a variety of essential small molecule compounds such as amino acids, monosaccharides, neurotransmitters, vitamins, as well as many inorganic ions. In recent years, these proteins have been recognized as potential therapeutic targets. Hexoses like glucose and fructose are energy sources and fuels for various metabolic processes inside the cell. Transport of hexoses through the cell membrane is facilitated through two distinct gene families (1) sodium-glucose transporters (SGLTs) and glucose transporters (GLUTs). GLUTs are passive transporters that use concentration and chemical gradient for the movement of hexoses across the cell membrane. To date, there are fourteen members (isoforms) of GLUTs, encoded by the human SLC2A genome family. Each GLUT varies in terms of which tissues it is found in, its substrate affinity and transport kinetics, and its sequence. In recent years, deregulation and alteration of energy metabolism have gained great attention due to their relation to various metabolic disorders, cancer, and diabetes. Overexpression of GLUTs in various cancers makes them an interesting and important biomarker to develop diagnostic and therapeutic probes for early detection and treatment. GLUT1 is a ubiquitous transporter that mediates the passage of D-glucose with high affinity. However, some cancers do not display overexpression of GLUT1, suggesting that D-glucose might not be the common energy source for all types of cancer or cancer cells switch their metabolic needs to other sugars like fructose, in glucose deficient environment. The principal fructose transporter, GLUT5, is responsible for absorbing dietary fructose and shows no facility for transporting D-glucose and other sugars. In addition, while normal breast tissues display limited expression of GLUT5, a significant percentage of breast tumor shows overexpression of this protein, making GLUT5 an intriguing target for imaging and detection. Fructose transport through GLUT5 occurs at an affinity (Ki) of 15mM. Interestingly, the C-2 deoxy analog of D-fructose (2,5-anhydro-D-mannitol; 2,5-AM) also displayed a similar affinity (Ki = 12.6 mM) to that of D-fructose. These observations inspired our decision to develop, evaluate and screen a library of 2,5-AM iii derivatives with the goal of developing diagnostic and therapeutic probes targeting GLUT5. Chapter 2 describes the preparation of a small library of 2,5-AM derivatives that were screened for inhibition of a previously reported PET tracer and potent GLUT5 substrate, 6-deoxy-6-[18F]fluoro-D-fructose ([18F]-6-FDF). Several of these derivatives were subjected to computational analysis involving docking and molecular dynamics simulations using the published GLUT5 three-dimensional structure. These calculations helped to identify the key interactions likely to be involved in the binding of 2,5-AM derivatives to GLUT5 and will inform the design of further generations of inhibitor libraries en route to potent molecular imaging probes targeting GLUT5. The most promising compounds from Chapter 2 were then used as a starting point for the design and synthesis of [18F]-containing PET radiotracers for potential use in medical imaging, and fluorescent probes applicable to confocal microscopy study or in situ optical detection of tumors (Chapter 3). Chapter 4 describes attempts to apply the fluorescent probe molecule 6-NBDF in a fluorescence-based assay to assess 2,5-AM derivatives for inhibition of its uptake by GLUT5. Importantly, this assay would avoid the need for a radioactive reporter molecule, as was used in Chapter 2. Conditions were developed for a robust assay that will be suitable for automation in order to screen large libraries of drug-like compounds or natural product extracts. Finally, Chapter 5 details possible future directions of this research, including different methods of forming GLUT5 targeting small molecules as NIR dye conjugates, drug conjugates, and dual probes.

• Subjects / Keywords

  • Chemistry
  • Solute carrier
  • Membrane proteins
  • Proteins
  • Cell membrane
  • Metabolic processes
  • Transporter proteins

• Graduation date

  Fall 2022

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/r3-z6v3-ws34

• License

  This thesis is made available by the University of Alberta Library with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.