Olfactomedin III expression contributes to anoikis-resistance in clonal variants of a human lung squamous carcinoma cell line

• Published in: Experimental cell research 2012-03, Vol.318 (5), p.593-602
• Main Authors: Keenan, Joanne, Joyce, Helena, Aherne, Sinead, O'Dea, Shirley, Doolan, Padraig, Lynch, Vincent, Clynes, Martin
• Format: Article
• Language: English
• Subjects: Anoikis; Apoptosis; Autophagy; Carcinoma, Squamous Cell; Caspase 3 - metabolism; Cell culture; Cell Cycle; Cell Line, Tumor; Cell Survival; Cells; Extracellular Matrix Proteins - genetics; Extracellular Matrix Proteins - metabolism; Extracellular Matrix Proteins - secretion; Focal Adhesion Kinase 1 - metabolism; Gene Expression; Gene Expression Profiling; Gene Knockdown Techniques; Glycoproteins - genetics; Glycoproteins - metabolism; Glycoproteins - secretion; Humans; Lung cancer; Lung carcinoma; Lung Neoplasms; Olfactomedin; Olfactomedin 3; Oligonucleotide Array Sequence Analysis; Phagocytosis; Phosphorylation; Real-Time Polymerase Chain Reaction; Resistance to control; RNA Interference; siRNA; Tumor cell lines; Western blotting
• ISSN: 0014-4827; 1090-2422
• DOI: 10.1016/j.yexcr.2012.01.012

Three clonal subpopulations of DLKP, a poorly differentiated squamous lung carcinoma cell line, display striking differences in ability to survive in suspension (anoikis resistance). DLKP-SQ is anoikis resistant (7.5% anoikis at 24h). In contrast, DLKP-M and DLKP-I are sensitive to anoikis (49.2% and 42.6% respectively). DLKP-I shows increased apoptosis consistently over all time points tested while DLKP-M appear to slow down metabolically and perhaps delays onset of anoikis by undergoing autophagy. Expression microarray analysis identified pronounced differential expression of Olfactomedin 3 (OLFM3) between the clones. High expression of OLFM3 was confirmed at the RNA level by qRT-PCR in DLKP-SQ and at the protein level by Western blotting (within the cell and secreted). Little or no OLFM3 was detected in the other two clones (DLKP-M and DLKP-I). Following siRNA knockdown of OLFM3 in DLKP-SQ, anoikis was increased 2.8-fold to 21% which was intermediate between the anoikis levels in DLKP-SQ and DLKP-M or DLKP-I. This knockdown correlated with increased apoptosis in suspension but not in attached culture conditions. Addition of recombinant OLFM3 reduced anoikis in DLKP-I. This is the first instance of OLFM3 being linked with anoikis resistance in a human cancer cell line.