Currently, there are few methods to visualize cell cycle progression in leukemia stem cells (LSC). Human LSC are characterized as quiescent self-renewing cells that drive leukemic transformation of myeloproliferative neoplasms and myelodysplastic syndromes. Additionally, current methods require sequential transduction and clonal selection. These methods and conditions are not feasible to study primary patient samples and LSC, which are generally limited in sample size. To alleviate this challenge, we generated a lentiviral biscistronic vector encoding FUCCI (fluorescent ubiquitination-based cell-cycle indicator) probes. Time-lapse confocal imaging of transduced 293 cells and SKNO-1 cells exhibited normal cell division and distinct nuclear staining of either green or red fluorescence. Further, analysis by flow cytometry of 293 cell confirms that S/G2/M and G1/G0 phases of cell cycle correlate to expression of fluorescent reporters expressed from bicistronic lentiviral vector and recapitulate FACS plots using traditional Ki67 and 7AAD staining. The development of this new reagent offers a new and more effective method to track cell cycle progression in limited mammalian cell populations. Future studies will utilize primary patient CD34+CD38- selected leukemic stem cell populations.

Citation Format: Gabriel Pineda, Florence Lambert-Fliszar, Gennarina L. Riso, Kathleen M. Kane, Catriona Jamieson. Characterization of cell-cycle progression using a novel bi-cistronic lentiviral FUCCI reporter. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-159. doi:10.1158/1538-7445.AM2014-LB-159