Lentiviral Fluorescent Genetic Barcoding for Multiplex Fate Tracking of Leukemic Cells

Abstract

Tracking the behavior of leukemic samples both in vitro and in vivo plays an increasingly large role in efforts to better understand the leukemogenic processes and the effects of potential new therapies. Such work can be accelerated and made more efficient by methodologies enabling the characterization of leukemia samples in multiplex assays. We recently developed three sets of lentiviral fluorescent genetic barcoding (FGB) vectors that create 26, 14, and 6 unique immunophenotyping-compatible color codes from GFP-, yellow fluorescent protein (YFP)-, and monomeric kusabira orange 2 (mKO2)-derived fluorescent proteins. These vectors allow for labeling and tracking of individual color-coded cell populations in mixed samples by real-time flow cytometry. Using the prototypical Hoxa9/Meis1 murine model of acute myeloid leukemia, we describe the application of the 6xFGB vector system for assessing leukemic cell characteristics in multiplex assays. By transplanting color-coded cell mixes, we investigated the competitive growth behavior of individual color-coded populations, determined leukemia-initiating cell frequencies, and assessed the dose-dependent potential of cells exposed to the histone deacetylase inhibitor Entinostat for bone marrow homing. Thus, FGB provides a useful tool for the multiplex characterization of leukemia samples in a wide variety of applications with a concomitant reduction in workload, processing times, and mouse utilization.

Keywords

• fluorescent genetic barcoding
• lentiviral gene transfer
• acute myeloid leukemia
• Hoxa9
• Meis1
• H9M
• leukemic stem cell
• LSC