title: (67/68)Galmydar: A metalloprobe for monitoring breast cancer resistance protein (BCRP)-mediated functional transport activity.
abstract: BACKGROUND
  For stratification of chemotherapeutic choices, radiopharmaceuticals capable of imaging breast cancer resistance protein (BCRP/ABCG2)-mediated functional transport are desired. To accomplish this objective, Galmydar, a fluorescent and moderately hydrophobic Ga(III) cationic complex and its (67/68)Ga-radiolabeled counterparts were interrogated in HEK293 cells stably transfected with BCRP and their WT counterparts transfected with empty vector. Additionally, the sensitivity and specificity of (68)Ga-Galmydar to evaluate functional expression of BCRP at the blood-brain barrier (BBB) was investigated in gene-knockout mdr1a/1b(-/-) (double knockout, dKO) and mdr1a/1b(-/-)ABCG2(-/-) (triple knockout, tKO) mouse models.
  
  
  METHODS
  For radiotracer uptake assays and live cell fluorescence imaging, either (67)Ga-Galmydar or its unlabeled counterpart was incubated in HEK293 cells transfected with BCRP (HEK293/BCRP) and their WT counterparts at 37°C under a continuous flux of CO2 (5%) in the presence or absence of Ko143, a potent BCRP antagonist, and cellular uptake was measured to assess the sensitivity of Galmydar to probe BCRP-mediated functional transport activity in cellulo. For assessing the potential of Galmydar to enable diagnostic imaging of targeted tissues in vivo, the (67)Ga-radiolabeled counterpart was incubated in either human serum albumin or human serum at 37°C and the percentage of unbound (67)Ga-Galmydar was determined. To evaluate the sensitivity of (68)Ga-Galmydar for molecular imaging of BCRP-mediated efflux activity in vivo, microPET/CT brain imaging was performed in dKO and tKO mice and their age-matched WT counterparts, 60min post-intravenous injection.
  
  
  RESULTS
  (67)Ga-Galmydar shows uptake profiles in HEK293 cells inversely proportional to BCRP expression, and antagonist (Ko143) induced accumulation in HEK293/BCRP cells, thus indicating target sensitivity and specificity. Furthermore, employing the fluorescent characteristics of Galmydar, optical imaging in HEK293/BCRP cells shows an excellent correlation with the radiotracer cellular accumulation data. (67)Ga-Galmydar shows > 85% unbound fraction and presence of parental compound in human serum. Finally, microPET/CT imaging shows higher retention of (68)Ga-Galmydar in brains of dKO and tKO mice compared to their age-matched WT counterparts, 60min post-intravenous tail-vein injection.
  
  
  CONCLUSIONS
  Combined data indicate that Galmydar could provide a template scaffold for development of a PET tracer for imaging BCRP-mediated functional transport activity in vivo.
authors.affiliation: ICCE Institute, Molecular Imaging Center, Mallinckrodt Institute of Radiology, Washington University School of Medicine, St. Louis, MO 63110, USA.
authors.forename: Jothilingam
authors.initials: J
authors.lastname: Sivapackiam
authors.affiliation: ICCE Institute, Molecular Imaging Center, Mallinckrodt Institute of Radiology, Washington University School of Medicine, St. Louis, MO 63110, USA.
authors.forename: Scott E
authors.initials: SE
authors.lastname: Harpstrite
authors.affiliation: ICCE Institute, Molecular Imaging Center, Mallinckrodt Institute of Radiology, Washington University School of Medicine, St. Louis, MO 63110, USA.
authors.forename: Julie L
authors.initials: JL
authors.lastname: Prior
authors.affiliation: ICCE Institute, Molecular Imaging Center, Mallinckrodt Institute of Radiology, Washington University School of Medicine, St. Louis, MO 63110, USA; Students and Teachers As Research Scientists (STARS), Washington University School of Medicine, St. Louis, MO 63110, USA.
authors.forename: Stephen
authors.initials: S
authors.lastname: Mattingly
authors.affiliation: ICCE Institute, Molecular Imaging Center, Mallinckrodt Institute of Radiology, Washington University School of Medicine, St. Louis, MO 63110, USA; Students and Teachers As Research Scientists (STARS), Washington University School of Medicine, St. Louis, MO 63110, USA; Department of Neurology, Washington University School of Medicine, St. Louis, MO 63110, USA; Department of Biomedical Engineering, School of Engineering & Applied Science, Washington University, St. Louis, MO 63105, USA. Electronic address: sharmav@mir.wustl.edu.
authors.forename: Vijay
authors.initials: V
authors.lastname: Sharma
mesh_terms: D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2 / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D056831:Coordination Complexes / Q000378:metabolism*; D005456:Fluorescent Dyes / Q000378:metabolism*; D005710:Gallium Radioisotopes; D057809:HEK293 Cells; D006801:Humans; D008297:Male; D051379:Mice; D009363:Neoplasm Proteins / Q000378:metabolism*; D061848:Optical Imaging; D009942:Organometallic Compounds / Q000378:metabolism*; D049268:Positron-Emission Tomography; D011849:Radioactive Tracers; D014057:Tomography, X-Ray Computed
chemical_list: C000612242:((ENB-3-isopropoxy-PIDMP)Ga(III)); C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D056831:Coordination Complexes; D005456:Fluorescent Dyes; D005710:Gallium Radioisotopes; D009363:Neoplasm Proteins; D009942:Organometallic Compounds; D011849:Radioactive Tracers
keywords: BCRP; Gallium; Metalloprobe