Su1729 Targeting Both Active and Quiescent Cancer (stem) Cells Using Combined Therapy

BACKGROUND: Recurrence of colorectal cancers after surgery and chemotherapy may be partly explained by the presence of chemoresistant cancer stem cells (CSC). Given the highly heterogeneous nature of CSCs it is possible that further characterization might identify a subset of CSC that eventually develop micro-metastases. Galectin-3 (Gal3) has been implicated in tumor progression, metastasis, and drug-resistance. We have previously reported that cell surface Gal3 renders colon cancer cells resistant to TNF-related Apoptosis-Inducing Ligand (TRAIL). Here, our aim was to determine how Gal3 is related to expression of stem cell markers and chemoresistance in different colon cancer cell populations. METHODS: We used the highly metastatic colon cancer cell line LiM6 and its TRAIL-resistant derivative LIM6-TR as cellular model. CSC populations were identified and isolated based on the expression of 5 established CSC-Markers CD24, CD44, CD166, EpCAM, and ALDH. Surface Gal3 in these CSCs was monitored by flow cytometry. Self-renewal, a functional CSC hallmark of CSC, was measured in sphere-formation assays. Response to TRAIL wasmeasured by APO-BRDU TUNEL assays. RESULTS: Only 15% of LIM6 and LIM6-TR cells were CSC as defined by the 5 markers (CD24+, CD44+, CD166+, EpCAM+, ALDH+). Surface Gal3 analysis identified two subpopulations within the CSC pool of LiM6 (54% Gal3+, 46% Gal3 negative). In contrast, CSC from LIM6-TR were uniformly Gal3+. Of note, not all Gal3+ colorectal cells can be attributed with CSC properties, since only 25-30% of Gal3+ CRCs express the remaining CSC markers simultaneously. Sphere assays demonstrated that the parental LIM6-TR had a 2-fold higher sphere forming ability than LIM6. The isolated Gal3+ CSCs formed 3.3-fold more spheres than parental Lim6 cells, while a 90% decrease in sphere formation was observed in the GAL3 negative CSC subpopulation. Depletion of Gal3 in LiM6-TR cells resulted in the loss of the Gal3+ CSC subpopulation, diminished sphere formation, and resensitization to TRAIL. Sphere formation of GAL+ CSCs in the presence of the Gal3 inhibitor LacNAc [5mM] led to a 60% reduction in sphere-formation. CONCLUSIONS: Our findings indicate that surface Gal3 is responsible for high sphere formation ability and TRAIL-resistance in a subpopulation of CSC. Further experiments will determine whether surface Gal3 in combination with other CSC markers can help to identify metastatic and chemoresistant CSCs in the circulation of CRC patients.