title: In vivo imaging of drug-induced mitochondrial outer membrane permeabilization at single-cell resolution.
abstract: Observing drug responses in the tumor microenvironment in vivo can be technically challenging. As a result, cellular responses to molecularly targeted cancer drugs are often studied in cell culture, which does not accurately represent the behavior of cancer cells growing in vivo. Using high-resolution microscopy and fluorescently labeled genetic reporters for apoptosis, we developed an approach to visualize drug-induced cell death at single-cell resolution in vivo. Stable expression of the mitochondrial intermembrane protein IMS-RP was established in human breast and pancreatic cancer cells. Image analysis was then used to quantify release of IMS-RP into the cytoplasm upon apoptosis and irreversible mitochondrial permeabilization. Both breast and pancreatic cancer cells showed higher basal apoptotic rates in vivo than in culture. To study drug-induced apoptosis, we exposed tumor cells to navitoclax (ABT-263), an inhibitor of Bcl-2, Bcl-xL, and Bcl-w, both in vitro and in vivo. Although the tumors responded to Bcl-2 inhibition in vivo, inducing apoptosis in around 20% of cancer cells, the observed response was much higher in cell culture. Together, our findings show an imaging technique that can be used to directly visualize cell death within the tumor microenvironment in response to drug treatment.
authors.affiliation: Center for Systems Biology, Massachusetts General Hospital, Boston 02114, USA.
authors.forename: Sarah
authors.initials: S
authors.lastname: Earley
authors.forename: Claudio
authors.initials: C
authors.lastname: Vinegoni
authors.forename: Joshua
authors.initials: J
authors.lastname: Dunham
authors.forename: Rostic
authors.initials: R
authors.lastname: Gorbatov
authors.forename: Paolo Fumene
authors.initials: PF
authors.lastname: Feruglio
authors.forename: Ralph
authors.initials: R
authors.lastname: Weissleder
mesh_terms: D000814:Aniline Compounds / Q000494:pharmacology*; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis; D001943:Breast Neoplasms / Q000188:drug therapy / Q000503:physiopathology*; D045744:Cell Line, Tumor; D005260:Female; D049452:Green Fluorescent Proteins; D006801:Humans; D007091:Image Processing, Computer-Assisted; D051379:Mice; D008819:Mice, Nude; D051336:Mitochondrial Membranes / Q000187:drug effects / Q000502:physiology*; D024101:Mitochondrial Proteins / Q000378:metabolism; D057054:Molecular Imaging / Q000379:methods*; D010190:Pancreatic Neoplasms / Q000188:drug therapy / Q000503:physiopathology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000037:antagonists & inhibitors; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D059010:Single-Cell Analysis; D013449:Sulfonamides / Q000494:pharmacology*; D059016:Tumor Microenvironment
chemical_list: D000814:Aniline Compounds; D000970:Antineoplastic Agents; D024101:Mitochondrial Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D011993:Recombinant Fusion Proteins; D013449:Sulfonamides; D049452:Green Fluorescent Proteins; C528561:navitoclax