Arsenic Cellular Handling by Human Multidrug Resistance Protein 1 (MRP1/ABCC1) and MRP4 (ABCC4)

• Author / Creator

  Whitlock, Brayden D

• Arsenic is a proven human carcinogen to which hundreds of millions of people are chronically exposed at unsafe levels, primarily through contaminated drinking water. An environmentally ubiquitous metalloid, arsenic in different forms is currently being used therapeutically as an anti-cancer cytotoxic agent, and is in clinical trials to expand its uses. Understanding how the human body handles arsenic upon environmental and therapeutic exposures is thus crucial to both preventing and treating cancers. Arsenic is metabolized in cells to form methylated and other conjugated arsenicals. Multidrug resistance proteins, a group of membrane proteins belonging to the ATP-binding cassette transporter subfamily C (MRPs/ABCCs) have been demonstrated to transport these methyl and glutathione conjugated arsenicals, giving the proteins a potentially key role in the cellular efflux and eventual elimination of arsenic. Further, ABCC protein level correlates with cancer risk and chemotherapy outcomes. Studying the way ABCCs interact with arsenic can be done effectively in vitro. However, animal models are needed to understand their influence on toxicokinetics. In Chapter 3, this thesis examines species differences between mouse and human orthologues of ABCC4, a key urinary elimination pathway protein for arsenicals. First, the epitope of the M4I-10 anti-ABCC4 antibody is mapped to determine that it can be used for inter-species work. Mouse Abcc4 is shown to not confer resistance to arsenicals in HEK293 cells and not transport DMAV or MMA(GS)2 in HEK293 membrane vesicles, unlike its human counterpart ABCC4. However, mAbcc4 appears to be important in protecting mouse embryonic fibroblasts from arsenate, as MEFs with Abcc4 knocked out are more susceptible to arsenate toxicity. Understanding species differences allows for laboratory data developed in animal models to be more accurately and reliably translated to the clinic and to human populations exposed to arsenic at unsafe levels. Novel functions of ABCC1 are also reported and characterized in Chapters 4 and 5. In Chapter 4, ABCC1 is reported to both transport and confer resistance against the cytotoxicity of DMAV in HEK293 but not HeLa cells. This cell line difference is not attributable to phosphorylation of Y920/S921 residues, unlike the HEK/HeLa cell line differences for As(GS)3 transport. In Chapter 5, the protection from cytotoxicity of monomethylarsonic acid (MMAV) conferred by ABCC1 in HEK293 cells but not HeLa cells is reported. Whole cell accumulation of MMAV is characterized in HEK293 and HeLa cells stably transfected with ABCC1 and vector, showing that ABCC1 has a bigger impact in preventing accumulation of MMAV in HEK cells, consistent with the cytotoxicity data. Using cytotoxicity, cellular accumulation, and vesicle transport experiments, this thesis advances the knowledge of how ABCC1 and ABCC4 proteins are involved in handling arsenic.

• Subjects / Keywords

  • Multidrug resistance proteins
  • arsenic
  • cellular efflux
  • Membrane transport

• Graduation date

  Fall 2021

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/r3-erd5-y933

• License

  This thesis is made available by the University of Alberta Libraries 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.