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Characterization of small spheres derived from various solid tumor cell lines: are they suitable targets for T cells?

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Abstract

T cell based immunotherapy has been investigated in a variety of malignancies and analyses have been mostly founded on in vitro data with tumor cell monolayers. However, three-dimensional (3D) culture models might mimic more closely the ‘in vivo’ conditions than 2D monolayers. Therefore, we analyzed the expression of tumor-associated antigens (TAA) and of molecules involved in antigen processing and presentation (APM) in tumor spheres, which served as an in vitro model for micrometastasis which might be enriched in tumor propagating cancer stem cells. For enrichment of sphere cells 12 human solid tumor cell lines were cultured in serum-free medium. Expression of a variety of TAA and APM were analyzed by RT-PCR and/or flow cytometry and compared to expression in corresponding adherent bulk cells grown in regular growth medium. Compared to adherent cells, spheres showed equal or higher mRNA expression levels of LMP2, LMP7 and MECL-1, of TAP1 and TAP2 transporters and, surprisingly, also of TAA including differentiation antigens. However, downregulation or loss of HLA-I and HLA-II molecules in spheres was observed in 8 of 10 and 1 of 2 cell lines, respectively, and was unresponsive to stimulation with IFN-γ. Although tumor spheres express TAA and molecules of intracellular antigen processing, they are defective in antigen presentation due to downregulation of HLA surface expression which may lead to immune evasion.

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Abbreviations

CTL:

Cytotoxic T lymphocyte

TAA:

Tumor-associated antigens

APM:

Molecules involved in antigen processing and presentation

SFM:

Serum-free growth medium

GM:

Serum containing growth medium

HLA:

Human leukocyte antigen

IMP:

Influenza matrix protein

WT1:

Wilm′s tumor suppressor gene 1

PAX2:

Paired box gene 2

CEA:

Carcinoembryonic Antigen

CK20:

Cytokeratin 20

HMBS:

Hydroxymethylbilane synthase

GAPDH:

Glyceraldehyde-3-phosphate dehydrogenase

TAP1/2:

Transporters 1/2

LMP2/7:

Large multifunctional peptidase 2/7

MECL-1:

Multicatalytic endopeptidase complex 1

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Acknowledgments

Supported by a grant of the Berliner Krebsgesellschaft, Hiege-Stiftung for Melanoma Research, EU Integrated Project Cancer Immunology and Immunotherapy. CRAR received funding from Wilhelm Sander-Stiftung.

Conflict of interest

The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. Department of Medicine III, Charité, Campus Benjamin Franklin, Hindenburgdamm 30, 12200, Berlin, Germany

    Antonia Busse, Anne Letsch, Alberto Fusi, Anika Nonnenmacher, David Stather, Sebastian Ochsenreither & Ulrich Keilholz

  2. Institute for Pathology & Comprehensive Cancer Center Charité, Charitéplatz 1, 10117, Berlin, Germany

    Christian RA Regenbrecht

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  1. Antonia Busse
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  2. Anne Letsch
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  3. Alberto Fusi
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Correspondence to Antonia Busse.

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Busse, A., Letsch, A., Fusi, A. et al. Characterization of small spheres derived from various solid tumor cell lines: are they suitable targets for T cells?. Clin Exp Metastasis 30, 781–791 (2013). https://doi.org/10.1007/s10585-013-9578-5

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  • DOI: https://doi.org/10.1007/s10585-013-9578-5

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