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Graduate Center of the City University of New York

6 ARTICLES PUBLISHED IN JoVE

Bioengineering

Harnessing the Bioorthogonal Inverse Electron Demand Diels-Alder Cycloaddition for Pretargeted PET Imaging

Thomas Reiner^*1, Jason S. Lewis¹, Brian M. Zeglis^*1

¹Department of Radiology, Memorial Sloan Kettering Cancer Center

The bioorthogonal inverse electron demand Diels-Alder cycloaddition has been harnessed to create an effective and modular pretargeted PET imaging strategy for cancer. In this protocol, the steps of this methodology are described in the context of a model system employing the colorectal cancer targeted antibody huA33 and a ⁶⁴Cu-labeled radioligand.

JoVE Core

The Bioconjugation and Radiosynthesis of ⁸⁹Zr-DFO-labeled Antibodies

Brian M. Zeglis¹, Jason S. Lewis¹

¹Department of Radiology, Memorial Sloan Kettering Cancer Center

Due to its multi-day radioactive half-life and favorable decay properties, the positron-emitting radiometal ⁸⁹Zr is extremely well-suited for use in antibody-based radiopharmaceuticals for PET imaging. In this protocol, the bioconjugation, radiosynthesis, and preclinical application of ⁸⁹Zr-labeled antibodies will be described.

Bioengineering

A Novel Technique for Generating and Observing Chemiluminescence in a Biological Setting

Gabriel E. Büchel^1,2, Brandon Carney^1,3, Jun Tang¹, Brian M. Zeglis^1,3, Jörg Eppinger², Thomas Reiner^1,4

¹Department of Radiology, Memorial Sloan Kettering Cancer Center, ²KAUST Catalysis Center, King Abdullah University of Science and Technology, ³Department of Chemistry, Hunter College, and PhD Program in Chemistry, Graduate Center of City University of New York, ⁴Department of Radiology, Weill Cornell Medical College

This protocol describes a new intraoperative imaging technique that uses a ruthenium complex as a source of chemiluminescent light emission, thereby producing high signal-to-noise ratios during in vivo imaging. Intraoperative imaging is an expanding field that could revolutionize the way that surgical procedures are performed.

Chemistry

Pretargeted Radioimmunotherapy Based on the Inverse Electron Demand Diels-Alder Reaction

Rosemery Membreno^1,2, Brendon E. Cook^1,2,3, Brian M. Zeglis^1,2,3,4

¹Department of Chemistry, Hunter College of the City University of New York, ²Ph.D. Program in Chemistry, Graduate Center of the City University of New York, ³Department of Radiology, Memorial Sloan Kettering Cancer Center, ⁴Department of Radiology, Weill Cornell Medical College

This protocol describes the synthesis and characterization of a trans-cyclooctene (TCO)-modified antibody and a ¹⁷⁷Lu-labeled tetrazine (Tz) radioligand for pretargeted radioimmunotherapy (PRIT). In addition, it details the use of these two constructs for in vivo biodistribution and longitudinal therapy studies in a murine model of colorectal cancer.

Chemistry

Synthesis and Bioconjugation of Thiol-Reactive Reagents for the Creation of Site-Selectively Modified Immunoconjugates

Maria Davydova¹, Guillaume Dewaele Le Roi^1,2, Pierre Adumeau¹, Brian M. Zeglis^1,2,3,4

In this protocol, we will describe the synthesis of PODS, a phenyoxadiazolyl methyl sulfone-based reagent for the site-selective attachment of cargos to the thiols of biomolecules, particularly antibodies. In addition, we will describe the synthesis and characterization of a PODS-bearing bifunctional chelator and its conjugation to a model antibody.

Genetics

Characterizing Histone Post-translational Modification Alterations in Yeast Neurodegenerative Proteinopathy Models

Seth A. Bennett^1,2, Samantha N. Cobos^1,3, Marcella Meykler¹, Michel Fallah¹, Navin Rana¹, Karen Chen¹, Mariana P. Torrente^1,4

¹Chemistry Department, Brooklyn College, ²Ph.D. Program in Biochemistry, Graduate Center of the City University of New York, ³Ph.D. Program in Chemistry, Graduate Center of the City University of New York, ⁴Ph.D. Programs in Chemistry, Biochemistry, and Biology, Graduate Center of the City University of New York

This protocol outlines experimental procedures to characterize genome-wide changes in the levels of histone post-translational modifications (PTM) occurring in connection with the overexpression of proteins associated with ALS and Parkinson's disease in Saccharomyces cerevisiae models. After SDS-PAGE separation, individual histone PTM levels are detected with modification-specific antibodies via Western blotting.