Squamous cell carcinoma cell aggregates escape suspension-induced, p53-mediated anoikis: fibronectin and integrin alphav mediate survival signals through focal adhesion kinase.
Resistance to anoikis, or apoptosis triggered by detachment from the extracellular matrix (ECM), lengthens the survival of malignant cells, facilitating reattachment and colonization of secondary sites. To examine the molecular mechanisms underlying resistance to anoikis in human oral squamous cell carcinoma (SCC) cells, we cultured human squamous carcinoma (HSC-3) cells in suspension on plates coated with poly-2-hydroxyethyl methacrylate, which blocks access to the ECM. Cells in suspension that formed multicellular aggregates had significantly lower levels of apoptosis than single cells. Aggregates, but not single cells, had high levels of fibronectin. Preincubation with a cyclic arginine-glycine-aspartic acid peptide or fibronectin-blocking antibody significantly increased anoikis. Single cells had markedly lower expression of the integrin alpha(v) receptor than aggregates. Blocking alpha(v) function with a blocking antibody or by transfection with an antisense oligonucleotide increased apoptosis and inhibited aggregation. In single cells but not aggregates, phosphorylation of the integrin-associated focal adhesion kinase (FAK) at tyrosine 397 was reduced, and p53 levels were increased. Apoptosis was increased by blocking FAK with an antisense oligonucleotide and reduced by blocking p53. These findings show that SCC cells escape suspension-induced anoikis by forming multicellular aggregates that avail themselves of fibronectin survival signals mediated by integrin alpha(v). Single cells in suspension that do not form aggregates undergo anoikis because of decreased FAK phosphorylation and increased p53 levels. Thus, SCC cells appear to use neighboring cells and the ECM molecule FN to promote the metastatic phenotype.
Department of Stomatology, School of Dentistry, and Howard Hughes Medical Institute, School of Medicine, University of California, San Francisco, California 94143, USA.
Yan
Y
Zhang
Hai
H
Lu
Paul
P
Dazin
Yvonne
Y
Kapila
D000818:Animals; D023102:Anoikis / Q000502:physiology*; D000970:Antineoplastic Agents / Q000378:metabolism; D002294:Carcinoma, Squamous Cell / Q000378:metabolism* / Q000473:pathology*; D002449:Cell Aggregation; D018929:Cell Culture Techniques / Q000379:methods; D045744:Cell Line, Tumor; D002470:Cell Survival; D005109:Extracellular Matrix / Q000737:chemistry / Q000378:metabolism; D005353:Fibronectins / Q000235:genetics / Q000378:metabolism; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D006801:Humans; D039564:Integrin alphaV / Q000235:genetics / Q000378:metabolism; D009062:Mouth Neoplasms / Q000378:metabolism / Q000473:pathology; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000378:metabolism; D009842:Oligopeptides / Q000378:metabolism; D010641:Phenotype; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*
D000970:Antineoplastic Agents; D005353:Fibronectins; D039564:Integrin alphaV; D016376:Oligonucleotides, Antisense; D009842:Oligopeptides; D016159:Tumor Suppressor Protein p53; C047981:arginyl-glycyl-aspartic acid; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human