[Frontiers in Bioscience E3, 1456-1466, June 1, 2011]

CD155 is involved in NK-cell mediated lysis of human hepatoblastoma in vitro

Matthias Pfeiffer1, Guido Seitz2, Peter Ruck3, Claudia Mueller4, Alexander Steinle5, Peter Lang1, Rupert Handgretinger1, Joerg Fuchs2, Steven Warmann2

1Department of Pediatric Oncology, University Children's Hospital Tuebingen, Hoppe-Seyler-Str.3, 72076 Tuebingen, Germany 2Department of Pediatric Surgery, University Children's Hospital Tuebingen, Hoppe-Seyler-Str.3, 72076 Tuebingen, Germany, 3Institute of Pathology, 71229 Leonberg, Germany, 4Department II of Internal Medicine, Section of Transplantation Immunology and Immunohematology, University Hospital, Otfried-Mueller Strasse 10, D-72076 Tuebingen, Germany, 5Department of Immunology, Institute for Cell Biology, University of Tuebingen, Auf der Morgenstelle 15, 72076 Tuebingen


1. Abstract
2. Introduction
3. Materials and Methods
3.1. Patients
3.2. Reagents and antibodies
3.3. Tumor cells and culture conditions
3.4. RT-PCR analysis
3.5. Flow cytometry
3.6. Sequencing of HLA class I genes in HB cells
3.7. Isolation of NK cells
3.8. Cytotoxicity assay
3.9. Cytological analyses and immunofluorescence
3.10. Statistical Analyses
4. Results
4.1. HB cells express NK cell-activating ligands.
4.2. HB cell lines show surface expression of CD155.
4.3. Quantification of HLA expression levels on HB cell lines
4.4. NK cell mediated cytotoxicity against HB cells
4.5. Anti CD155 monoclonal antibody (mAb) inhibits NK cell mediated cytotoxicity
4.6. Cytological changes in HB cells incubated with NK cells
5. Discussion
6. Acknowledgements
7. References


NK cells are involved in the lysis of different solid tumors and leukemias. NK-activity is thereby regulated by activating and inhibitory receptors. Until now, nothing is known about the NK-activity against hepatoblastoma and the involved receptors. We tested NK cells for cytotoxicity against HB in vitro. Expression levels of activating NK ligands were analysed on 13 primary HB samples as well as on 3 HB cell lines. ALL HB cell lines showed low HLA-class-I-expression. CD155 expression was strong on primary HB samples and cell lines. NKG2D-ligands (MICA/B, ULBP1-3) were heterogeneous expressed in primary samples and cell cultures. There were no differences between the various histological subtypes. NK cells showed strong cytotoxicity in vitro which was significantly increased through interleukin-2 and -15 stimulation (p<.0001). Blockade of CD155 resulted in decreased lysis rates. Our findings show that NK cells exert high activity against hepatoblastoma in vitro and that CD155 is involved in the NK mediated killing of HB. The inclusion of a NK-based immunotherapy into novel treatment strategies might be a promising alternative especially for advanced tumors.