Pharmacology

James Hammond

Dr James Hammond

Professor

Education:
BSc (Hons), Physiology & Pharmacology, University of Western Ontario, 1979
PhD, Pharmacology, University of Alberta, 1983

Teaching: PMCOL 337

Research:
Pharmacological, functional and molecular characterization of the membrane transport systems responsible for the cellular uptake and release of endogenous nucleosides and nucleobases and their anticancer/antiviral analogues.

Research Interests / Laboratory Techniques

Multiple subtypes of nucleoside/nucleobase transporters have been identified by both functional and molecular approaches. They vary in their sensitivities to inhibition by agents such as nitrobenzylthioinosine (NBMPR) and dipyridamole, and by their dependence on ion (sodium) gradients. Two broad gene families have been identified: 1) Equilibrative nucleoside transporters (SLC29A; ENT) that operate by facilitated diffusion and 2) Concentrative sodium/nucleoside co-transporters (SLC28A; CNT). Seven of these transporters have been cloned from a number of mammalian (including human) tissues (ENT1, ENT2, ENT3, ENT4, CNT1, CNT2, CNT3). We cloned the canine ENT1 and the mouse ENT1 and ENT2, as well as two novel splice variants of the mouse ENT1 transporter. The physiological and pharmacological significance of the multiple transporter subtypes/variants has yet to be established. It is expected that nucleoside transporter heterogeneity would impact significantly on the actions of many drugs that act through adenosine receptors or by modifying cellular nucleoside metabolism. This has particular relevance to the clinical activities of the cytotoxic nucleoside analogues used in cancer chemotherapy (e.g. cytosine arabinoside, gemcitabine, 2-chlorodeoxyadenosine), as well as drugs that modify vascular function via modulation of the adenosine signalling pathways.
In addition, a novel nucleobase transporter has recently been identified (SLC43A3) which may play an important role in mediating the cellular uptake and cytotoxicity of nucleobase analogue drugs such as the 6-thiopurines used in cancer therapy and for the treatment of intestinal bowel disorders such as Crohn’s Disease.


Current research:

1) The analysis of the protein structural elements involved in the interaction of drugs and substrates with the multiple subtypes and isoforms of equilibrative nucleoside and nucleobase transporters. Studies involve the generation of point mutants (site-directed mutagenesis) and their expression in transporter-deficient mammalian cell lines. Nucleoside transport activity of the native and recombinant proteins is measured directly by the cellular accumulation of radiolabelled substrates and indirectly using specific probes for the proteins associated with the transporter.

2) The role of protein phosphorylation in the activity and membrane turnover of the purine transporters, Investigation of mechanisms to modify transporter expression/activity by modifying regulatory pathways to enhance therapies with nucleoside/nucleobase analogue drugs. This work also includes examination of the impact of transporter dimerization and intracellular protein interactions on transporter function (Funded by NSERC).
3) Characterization of ENT4 as a drug target for the treatment of ischemia-reperfusion injury. ENT4 transports adenosine predominantly under acidic conditions. Therefore, ENT4 may be important for the regulation of adenosine in the vasculature in acidic environments such as those seen in ischemic tissues (Funded by the Heart and Stroke Foundation of Canada).
4) Analysis of the phenotypes associated with the genetic knockout of the SLC29A1 or SLC29A4-encoded transporters (ENT1, ENT4) in mice. We have identified a clear vascular phenotype, as well as a soft tissue aberrant mineralization phenotype. Studies are ongoing to understand the mechanisms underlying these biological changes and their relevance to human disease states (Funded by the CIHR and the Heart and Stroke Foundation of Canada)
5) Analysis of the role of variations in the expression of SLC43A3 in the therapeutic efficacy and side-effect profile of 6-mercaptopurine when used to treat acute lymphoblastic leukemia (Funded by Cancer Research Society of Canada).

Selected Recent Publications

Best KA, Bone DB, Vilas G, Gros R, Hammond JR (2018) Changes in aortic reactivity associated with the loss of equilibrative nucleoside transporter 1 (ENT1) in mice. PLoS ONE 13(11): e0207198. https://doi.org/10.1371/journal.pone.0207198

Ii H, Warraich S, Tenn N, Quinonez D, Holdsworth DW, Hammond JR, Dixon SJ and Séguin CA. (2016) Disruption of biomineralization pathways in spinal tissues of a mouse model of diffuse idiopathic skeletal hyperostosis. Bone 90:37-49. PMID: 27237608.

Hughes SJ, Cravetchi X, Vilas G and Hammond JR. (2015) Adenosine A1 receptor activation modulates human equilibrative nucleoside transporter 1 (hENT1) activity via PKC-mediated phosphorylation of serine-281. Cell Signal 27(5):1008-18. PMID: 25725289.

Bone DB, Antic M, Quinonez D, Hammond JR. (2015) Hypoxanthine uptake by skeletal muscle microvascular endothelial cells from equilibrative nucleoside transporter 1 (ENT1)-null mice: effect of oxidative stress. Microvasc Res 98:16-22. PMID: 25448155

Bone DB, Antic M, Vilas G and Hammond JR. (2014) Oxidative stress modulates nucleobase transport in microvascular endothelial cells. Microvasc Res 95C:68-75. PMID: 24976360.

Warraich S, Bone DB, Quinonez D, Ii H, Choi DS, Holdsworth DW, Drangova M, Dixon SJ, Seguin CA and Hammond JR. (2013) Loss of equilibrative nucleoside transporter 1 (ENT1) in mice leads to progressive ectopic mineralization of spinal tissues resembling diffuse idiopathic skeletal hyperostosis (DISH) in humans. J Bone Miner Res 28(5):1135-49. PMID: 23184610.

Park JS and Hammond JR. (2012) Cysteine Residues in the TM9-TM11 Region of the Human Equilibrative Nucleoside Transporter Subtype 1 Play an Important Role in Inhibitor Binding and Translocation Function. Mol Pharmacol 82(5):784-94. PMID: 22837314.

Park JS, Hughes SJ, Cunningham FK and Hammond JR. (2011) Identification of Cysteines Involved in the Effects of Methanethiosulfonate Reagents on Human Equilibrative Nucleoside Transporter 1. Mol Pharmacol 80(4):735-46. PMID: 21791574.

Bone DB, Choi DS, Coe IR and Hammond JR. (2010) Nucleoside/nucleobase transport and metabolism by microvascular endothelial cells isolated from ENT1 -/- mice. Am J Physiol Heart Circ Physiol 299 (3):H847-H856. PMID: 20543083.

Robillard KR, Bone DB, Park JS and Hammond JR. (2008) Characterization of mENT1Delta11, a novel alternative splice variant of the mouse equilibrative nucleoside transporter 1. Mol Pharmacol 74 (1):264-273. PMID: 18413666.

Bone DB and Hammond JR. (2007) Nucleoside and nucleobase transporters of primary human cardiac microvascular endothelial cells: Characterization of a novel nucleobase transporter. Am J Physiol Heart Circ Physiol 293:H3325-H3332. PMID: 17921321.