High aldehyde dehydrogenase and expression of cancer stem cell markers selects for breast cancer cells with enhanced malignant and metastatic ability

TLDR

Cancer stem cells have been identified in leukemia and solid tumors, but their role in metastasis remains poorly understood due to technical challenges with primary human cells. The study aimed to develop pre‑clinical models to investigate whether stem‑like breast cancer cells drive metastatic behavior. Four human breast‑cancer lines were profiled for CD44/CD24, CD133, and ALDH activity, and ALDH‑high/CD44+ subpopulations were isolated by FACS and tested for proliferation, colony formation, adhesion, migration, invasion in vitro and tumorigenicity/metastasis in NOD/SCID/IL2γR null mice. ALDH‑high/CD44+CD24− and ALDH‑high/CD44+CD133+ cells exhibited markedly higher growth, colony formation, adhesion, migration, invasion, tumorigenicity, and metastasis compared to ALDH‑low counterparts, indicating that these stem‑like populations likely mediate breast‑cancer metastasis.

Abstract

Cancer stem cells (CSCs) have recently been identified in leukaemia and solid tumours; however, the role of CSCs in metastasis remains poorly understood. This dearth of knowledge about CSCs and metastasis is due largely to technical challenges associated with the use of primary human cancer cells in pre-clinical models of metastasis. Therefore, the objective of this study was to develop suitable pre-clinical model systems for studying stem-like cells in breast cancer metastasis, and to test the hypothesis that stem-like cells play a key role in metastatic behaviour. We assessed four different human breast cancer cell lines (MDA-MB-435, MDA-MB-231, MDA-MB-468, MCF-7) for expression of prospective CSC markers CD44/CD24 and CD133, and for functional activity of aldehyde dehydrogenase (ALDH), an enzyme involved in stem cell self-protection. We then used fluorescence-activated cell sorting and functional assays to characterize differences in malignant/metastatic behaviour in vitro (proliferation, colony-forming ability, adhesion, migration, invasion) and in vivo (tumorigenicity and metastasis). Sub-populations of cells demonstrating stem-cell-like characteristics (high expression of CSC markers and/or high ALDH) were identified in all cell lines except MCF-7. When isolated and compared to ALDH(low)CD44(low/-) cells, ALDH(hi)CD44(+)CD24(-) (MDA-MB-231) and ALDH(hi)CD44(+)CD133(+) (MDA-MB-468) cells demonstrated increased growth (P < 0.05), colony formation (P < 0.05), adhesion (P < 0.001), migration (P < 0.001) and invasion (P < 0.001). Furthermore, following tail vein or mammary fat pad injection of NOD/SCID/IL2gamma receptor null mice, ALDH(hi)CD44(+)CD24(-) and ALDH(hi)CD44(+)CD133(+) cells showed enhanced tumorigenicity and metastasis relative to ALDH(low)CD44(low/-) cells (P < 0.05). These novel results suggest that stem-like ALDH(hi)CD44(+)CD24(-) and ALDH(hi)CD44(+)CD133(+) cells may be important mediators of breast cancer metastasis.