title: In vivo labeling of B16 melanoma tumor xenograft with a thiol-reactive gadolinium based MRI contrast agent.
abstract: Murine melanoma B16 cells display on the extracellular side of the plasma membrane a large number of reactive protein thiols (exofacial protein thiols, EPTs). These EPTs can be chemically labeled with Gd-DO3A-PDP, a Gd(III)-based MRI contrast agent bearing a 2-pyridinedithio chemical function for the recognition of EPTs. Uptake of gadolinium up to 10(9) Gd atoms per cell can be achieved. The treatment of B16 cells ex vivo with a reducing agent such as tris(2-carboxyethyl)phosphine (TCEP) results in an increase by 850% of available EPTs and an increase by 45% of Gd uptake. Blocking EPTs with N-ethylmaleimide (NEM) caused a decrease by 84% of available EPTs and a decrease by 55% of Gd uptake. The amount of Gd taken up by B16 cells is therefore dependent upon the availability of EPTs, whose actual level in turn changes according to the extracellular redox microenvironment. Then Gd-DO3A-PDP has been assessed for the labeling of tumor cells in vivo on B16.F10 melanoma tumor-bearing mice. Gd-DO3A-PDP (or Gd-DO3A as the control) has been injected directly into the tumor region at a dose level of 0.1 μmol and the signal enhancement in MR images followed over time. The washout kinetics of Gd-DO3A-PDP from tumor is very slow if compared to that of control Gd-DO3A, and 48 h post injection, the gadolinium-enhancement is still clearly visible. Therefore, B16 cells can be labeled ex vivo as well as in vivo according to a common EPTs-dependent route, provided that high levels of the thiol reactive probe can be delivered to the tumor.
authors.affiliation: Institute for Biostructures and Bioimages, CNR, Molecular Biotechnology Center, University of Turin, Via Nizza 52, I-10126 Torino, Italy.
authors.forename: Valeria
authors.initials: V
authors.lastname: Menchise
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Digilio
authors.forename: Eliana
authors.initials: E
authors.lastname: Gianolio
authors.forename: Evelina
authors.initials: E
authors.lastname: Cittadino
authors.forename: Valeria
authors.initials: V
authors.lastname: Catanzaro
authors.forename: Carla
authors.initials: C
authors.lastname: Carrera
authors.forename: Silvio
authors.initials: S
authors.lastname: Aime
mesh_terms: D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D045744:Cell Line, Tumor; D003287:Contrast Media / Q000008:administration & dosage / Q000737:chemistry; D056831:Coordination Complexes / Q000008:administration & dosage / Q000737:chemistry; D005682:Gadolinium / Q000008:administration & dosage / Q000737:chemistry; D015552:Injections, Intralesional; D007700:Kinetics; D008024:Ligands; D057230:Limit of Detection; D008279:Magnetic Resonance Imaging; D008297:Male; D008546:Melanoma, Experimental / Q000175:diagnosis* / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008810:Mice, Inbred C57BL; D052066:N-Ethylmaleimide-Sensitive Proteins / Q000737:chemistry / Q000378:metabolism; D011725:Pyridines / Q000737:chemistry; D013439:Sulfhydryl Reagents / Q000008:administration & dosage / Q000737:chemistry / Q000494:pharmacology; D013440:Sulfides / Q000737:chemistry
chemical_list: C436500:2,6-pyridinedithiolate; D003287:Contrast Media; D056831:Coordination Complexes; D008024:Ligands; D011725:Pyridines; D013439:Sulfhydryl Reagents; D013440:Sulfides; D005682:Gadolinium; D052066:N-Ethylmaleimide-Sensitive Proteins