Abstract 3335: Self-renewal, tumorigenicity and metastatic potential of CD44+ cells in ER-negative lines and dissociated primary human breast cancers is increased in CD24+ subsets
Authors: Azzam Diana J. , Drews-Elger Katherine , Zhao Dekuang , Chendong Pan , Wander Seth A. , El-Ashry Dorraya , Slingerland Joyce M. 
Published: June 2012

Primary human breast cancer subpopulations with a CD44+ CD24NEG/LOW ESA+ phenotype exhibit self-renewal and generate xenograft tumors with as few as 100 cells in immunodeficient mice. While CD44 expression is strongly associated with self renewal in several cancers, the relative contributions of CD24 negative versus low subpopulations to stemness is poorly defined. Early reports have not distinguished negative from low CD24 expression in human lines and primary tumors, and both, together with CD44+ status have been linked to breast cancer stem cell phenotypes. In contrast, certain reports have associated CD24 expression with tumor progression and metastasis. Here, we show increased mitogenic kinase activation, expression of ESC and EMT genes in the CD24LOW subpopulation of CD44+ cells in both triple negative MDA-MB-231 breast cancer cells and primary dissociated breast cancers (DTs) compared to the CD24NEG cells. Moreover, tumorigenicity was increased and metastasis arose exclusively from orthotopic xeno-implantation of the CD44+CD24LOW cells. MDA-MB-231 and two different primary DTs were live sorted into CD44+CD24NEG and CD44+CD24LOW enriched subpopulations. Cells expressing other putative stem cell markers, ALDH1 and ESA were almost entirely CD44+CD24LOW. CD44+CD24LOWcells showed higher PI3-kinase/Akt, MAPK, and Src activites compared to CD44+ CD24NEG cells. While both subpopulations formed soft agar colonies and/or mammospheres, those arising from CD44+CD24LOW were greater in number and size than from CD44+CD24NEG cells. When cultured in 2D after sorting, CD44+ CD24LOW cells gave rise to both CD44+ CD24LOW and CD44+ CD24NEG progeny while CD44+ CD24NEG yielded only CD44+ CD24NEG progeny. CD44+CD24LOW expressed higher levels of ESC genes and EMT-associated genes than CD44+CD24NEG cells and uniquely expressed stem cell-associated miRNAs. On orthotopic injection into BalbC nude mice, both subpopulations initiated tumors with as few as 100 cells, however tumors arising from CD44+CD24LOW cells had a shorter latency and grew more rapidly than those arising from CD44+CD24NEG cells. In vivo imaging of luciferase positive cells showed CD44+CD24LOW cells yield metastasis while CD44+CD24NEG cells did not.

Our data demonstrate, for the first time, distinct biological properties between the CD44+CD24LOW and CD44+CD24NEG enriched subpopulations of MDA-MB-231 and primary human breast cancer DTs. Low surface expression of CD24 is associated with ALDH1 and ESA+ status, increased self-renewal as shown by mammosphere and colony formation, tumorigenicity and metastatic potential of CD44+ cells. The ability to isolate and characterize such TSC-enriched subpopulations will be necessary for development of therapeutic strategies that preferentially target breast cancer stem cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3335. doi:10.1158/1538-7445.AM2011-3335