A Robust Discovery Platform for the Identification of Novel Mediators of Melanoma Metastasis

Arman Alberto Sorin Shadaloey, Alcida Karz, Rana S. Moubarak, Praveen Agrawal, Grace Levinson, Kevin Kleffman, Orlando Aristizabal, Iman Osman, Youssef Z. Wadghiri, Eva Hernando

Research output: Contribution to journalArticlepeer-review

Abstract

Metastasis is a complex process, requiring cells to overcome barriers that are only incompletely modeled by in vitro assays. A systematic workflow was established using robust, reproducible in vivo models and standardized methods to identify novel players in melanoma metastasis. This approach allows for data inference at specific experimental stages to precisely characterize a gene's role in metastasis. Models are established by introducing genetically modified melanoma cells via intracardiac, intradermal, or subcutaneous injections into mice, followed by monitoring with serial in vivo imaging. Once preestablished endpoints are reached, primary tumors and/ or metastases-bearing organs are harvested and processed for various analyses. Tumor cells can be sorted and subjected to any of several 'omics' platforms, including single-cell RNA sequencing. Organs undergo imaging and immunohistopathological analyses to quantify the overall burden of metastases and map their specific anatomic location. This optimized pipeline, including standardized protocols for engraftment, monitoring, tissue harvesting, processing, and analysis, can be adopted for patient-derived, short-term cultures and established human and murine cell lines of various solid cancer types.

Original languageEnglish (US)
Article numbere63186
JournalJournal of Visualized Experiments
Volume2022
Issue number181
DOIs
StatePublished - Mar 2022
Externally publishedYes

ASJC Scopus subject areas

  • General Neuroscience
  • General Chemical Engineering
  • General Biochemistry, Genetics and Molecular Biology
  • General Immunology and Microbiology

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