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Memorial Sloan-Kettering Cancer Center

10 ARTICLES PUBLISHED IN JoVE

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Biology

Detecting Somatic Genetic Alterations in Tumor Specimens by Exon Capture and Massively Parallel Sequencing
Helen H Won 1, Sasinya N Scott 1, A. Rose Brannon 1, Ronak H Shah 1, Michael F Berger 1,2
1Department of Pathology, Memorial Sloan-Kettering Cancer Center, 2Human Oncology and Pathogenesis Program, Memorial Sloan-Kettering Cancer Center

We describe the preparation of barcoded DNA libraries and subsequent hybridization-based exon capture for detection of key cancer-associated mutations in clinical tumor specimens by massively parallel "next generation" sequencing. Targeted exon sequencing offers the benefits of high throughput, low cost, and deep sequence coverage, thus yielding high sensitivity for detecting low frequency mutations.

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Medicine

Isolation of Cancer Stem Cells From Human Prostate Cancer Samples
Samuel J. Vidal 1, S. Aidan Quinn 1, Janis de la Iglesia-Vicente 1, Dennis M. Bonal 1, Veronica Rodriguez-Bravo 2, Adolfo Firpo-Betancourt 1, Carlos Cordon-Cardo 1, Josep Domingo-Domenech 1
1Department of Pathology, Icahn School of Medicine at Mount Sinai, 2Molecular Biology Program, Memorial Sloan-Kettering Cancer Center

The isolation of cancer stem cells (CSCs) directly from human tissues is requisite for their biological characterization. This manuscript describes a methodology for the isolation of prostate CSCs from human tissues, while also providing tips on troubleshooting difficult steps.

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Neuroscience

Feeder-free Derivation of Neural Crest Progenitor Cells from Human Pluripotent Stem Cells
Nadja Zeltner 1, Fabien G. Lafaille 2, Faranak Fattahi 1, Lorenz Studer 1
1Developmental Biology, Center for Stem Cell Biology, Sloan-Kettering Institute for Cancer Research, 2St. Giles Laboratory of Human Genetics of Infectious Diseases, The Rockefeller University

Neural crest (NC) cells derived from human pluripotent stem cells (hPSC) have great potential for modeling human development and disease and for cell replacement therapies. Here, a feeder-free adaptation of the currently widely used in vitro differentiation protocol for the derivation of NC cells from hPSCs is presented.

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Medicine

Femoral Bone Marrow Aspiration in Live Mice
Young Rock Chung 1, Eunhee Kim 1, Omar Abdel-Wahab 1
1Human Oncology and Pathogenesis Program, Department of Medicine, Memorial Sloan-Kettering Cancer Center

This protocol describes a procedure for serial sampling of femoral bone marrow (BM) without requiring the sacrifice of mice. This procedure facilitates longitudinal studies of the BM composition of mice over time and provides serial access to cells within the BM for ex vivo and transplantation studies.

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Medicine

Generation of Prostate Cancer Patient Derived Xenograft Models from Circulating Tumor Cells
Estrelania S. Williams 1, Veronica Rodriguez-Bravo 3, Uma Chippada-Venkata 2, Janis De Ia Iglesia-Vicente 1, Yixuan Gong 2, Matthew Galsky 2, William Oh 2, Carlos Cordon-Cardo 1, Josep Domingo-Domenech 1
1Department of Pathology, Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, 2Department of Hematology/Oncology, Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, 3Molecular Biology Program, Memorial Sloan-Kettering Cancer Center

This manuscript details a method used to generate prostate cancer patient derived xenografts (PDXs) from circulating tumor cells (CTCs). The generation of PDX models from CTCs provides an alternative experimental model to study prostate cancer; the most commonly diagnosed tumor and a frequent cause of death from cancer in men.

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Developmental Biology

Feeder-free Derivation of Melanocytes from Human Pluripotent Stem Cells
Scott J. Callahan 1,2, Yvonne Mica 3, Lorenz Studer 1
1The Center for Stem Cell Biology, Developmental Biology Program, Memorial Sloan-Kettering Cancer Center, 2Cancer Biology and Genetics Program, Gerstner Sloan-Kettering Graduate School, Sloan-Kettering Institute for Cancer Research, 3Thermo Fisher Scientific

This work describes an in vitro differentiation protocol to produce pigmented, mature melanocytes from human pluripotent stem cells via a neural crest and melanoblast intermediate stage using a feeder-free, 25 day protocol.

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Cancer Research

Generation of Prostate Cancer Cell Models of Resistance to the Anti-mitotic Agent Docetaxel
Lisa Mohr 1, Marc Carceles-Cordon 1, Jungreem Woo 1, Carlos Cordon-Cardo 1, Josep Domingo-Domenech 1, Veronica Rodriguez-Bravo 1,2
1Department of Pathology, Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, 2Department of Oncological Sciences, Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai

Resistance to cancer therapies contributes to disease progression and death. Determining the mechanistic underpinnings of resistance is crucial for improving therapeutic response. This manuscript details the protocol to generate taxane-resistant cell models of prostate cancer (PC) to help dissecting the pathways involved in progression to Docetaxel resistance in PC patients.

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Immunology and Infection

Spatial and Temporal Control of T Cell Activation Using a Photoactivatable Agonist
Elisa Sanchez 1,2, Morgan Huse 1
1Immunology Program, Memorial Sloan-Kettering Cancer Center, 2Weill-Cornell Graduate School of Medical Sciences

This protocol describes an imaging-based method to activate T lymphocytes using photoactivatable peptide-MHC, enabling precise spatiotemporal control of T cell activation.

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Cancer Research

Isolation and Characterization of Tumor-initiating Cells from Sarcoma Patient-derived Xenografts
Dan Han 1, Veronica Rodriguez-Bravo 1, Sudeh Izadmehr 1, Josep Domingo-Domenech 1, Carlos Cordon-Cardo 1
1Department of Pathology, Icahn School of Medicine at Mount Sinai

We describe a detailed protocol for the isolation of tumor-initiating cells from human sarcoma patient-derived xenografts by fluorescence-activated cell sorting, using human leukocyte antigen-1 (HLA-1) as a negative marker, and for the further validation and characterization of these HLA-1-negative tumor-initiating cells.

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Cancer Research

Identification, Histological Characterization, and Dissection of Mouse Prostate Lobes for In Vitro 3D Spheroid Culture Models
Disharee Nath 1,2, Julie R. White 3,4, Gennady Bratslavsky 1, Leszek Kotula 1,2
1Department of Urology, SUNY Upstate Medical University, 2Department of Biochemistry and Molecular Biology, SUNY Upstate Medical University, 3Laboratory of Comparative Pathology, Memorial Sloan-Kettering Cancer Center, 4Boulder BioPATH, Inc

Genetically engineered mice are useful models for investigating prostate cancer mechanisms. Here we present a protocol to identify and dissect prostate lobes from a mouse urogenital system, differentiate them based on histology, and isolate and culture the primary prostate cells in vitro as spheroids for downstream analyses.

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