Abstract
Malignant transformation of cells is frequently associated with abnormalities in human leukocyte antigen (HLA) expression. MHC class I loss or down-regulation in cancer cells is a major immune escape route used by a large variety of human tumours to evade antitumour immune responses mediated by cytotoxic T lymphocytes. The goal of our study was to explore HLA genotyping and phenotyping in a variety of melanoma tumour cell lines. A total of 91 melanoma cell lines were characterised for HLA class I and II genotype. In addition, 61 out of the 91 cell lines were also analysed for HLA class I and II cell surface molecule expression by flow cytometry. Unexpectedly, we found that 19.7% of the melanoma cell lines were homozygous for HLA class I genotypes, sometimes associated with HLA class II homozygosity (8.79%) and sometimes not (10.98%). The frequency of homozygosity was significantly higher compared with the control groups (1.6%). To identify the reasons underlying the high frequency of HLA homozygosity we searched for genomic deletions using eight pairs of highly polymorphic microsatellite markers covering the entire extended HLA complex on the short arm of chromosome 6. Our results were compatible with hemizygous deletions and suggest that loss of heterozygosity on chromosome arm 6p is a common feature in melanoma cell lines. In fact, although autologous normal DNA from the patients was not available and could not be tested, the retention in some cases of heterozygosity for a number of microsatellite markers would indicate a hemizygous deletion. In the rest of the cases, markers at 6p and 6q showed a single allele pattern indicating the probable loss of part or the whole of chromosome 6. These results led us to conclude that loss of heterozygosity in chromosome 6 is nonrandom and is possibly an immunologically relevant event in human malignant melanoma. Other well-established altered HLA class I phenotypes were also detected by flow cytometry that correspond to HLA class I total loss and HLA-ABC and/or specific HLA-B locus down-regulation.
Similar content being viewed by others
References
Arguello JR, Little AM, Pay AL, Gallardo D, Rojas I, Marsh SGE, Goldman JM, Madrigal JA (1998) Mutation detection and typing of polymorphic loci through double-strand conformation analysis. Nat Genet 18:192–194
Bauer S, Groh V, Wu J, Steinle A, Phillips JH, Lanier LL, Spies T (1999) Activation of NK cells and T cells by NKG2D, a receptor for stress-inducible MICA. Science 285:727–729
Cerwenka A, Baron JL, Lanier LL (2001) Ectopic expression of retinoic acid early inducible-1 gene (RAE-1) permits natural killer cell-mediated rejection of a MHC class I-bearing tumour in vivo. Proc Natl Acad Sci U S A 98:11521–11526
de Nooij-van Dalen AG, van Buuren-van Seggelen VH, Lohman PH, Giphart-Gassler M (1998) Chromosome loss with concomitant duplication and recombination both contribute most to loss of heterozygosity in vitro. Genes Chromosomes Cancer 21:30–38
Dean M, Carrington M, O’Brien SJ (2002) Balanced polymorphism selected by genetic versus infectious human disease. Annu Rev Genomics Hum Genet 3:263–292
Garcia-Lora A, Algarra I, Gaforio JJ, Ruiz-Cabello F, Garrido F (2001) Immunoselection by T lymphocytes generates repeated MHC class I-deficient metastatic tumour variants. Int J Cancer 91:109–119
Garrido F, Ruiz-Cabello F, Cabrera T, Perez-Villar JJ, Lopez-Botet M, Duggan-Keen M, Stern PL (1997) Implications for immunosurveillance of altered HLA class I phenotypes in human tumours. Immunol Today 18:89–95
Goldberg SF, Miele ME, Hatta N, Takata M, Paquette-Straub C, Freedman LP, Welch DR (2003) Melanoma metastasis suppression by chromosome 6: evidence for a pathway regulated by CRSP3 and TXNIP. Cancer Res 63:432–440
Groh V, Steinle A, Bauer S, Spies T (1998) Recognition of stress-induced MHC molecules by intestinal epithelial gammadelta T cells. Science 279:1737–1740
Groh V, Rhinehart R, Randolph-Habecker J, Topp MS, Riddell SR, Spies T (2001) Costimulation of CD8alphabeta T cells by NKG2D via engagement by MIC induced on virus-infected cells. Nat Immunol 2:255–260
Guan XY, Zhang HE, Zhou H, Sham JS, Fung JM, Trent JM (2002) Characterization of a complex chromosome rearrangement involving 6q in a melanoma cell line by chromosome microdissection. Cancer Genet Cytogenet 134:65–70
Hoglund M, Gisselsson D, Hansen GB, White VA, Sall T, Mitelman F, Horsman D (2004) Dissecting karyotypic patterns in malignant melanomas: temporal clustering of losses and gains in melanoma karyotypic evolution. Int J Cancer 108:57–65
Jimenez P, Canton J, Collado A, Cabrera T, Serrano A, Real LM, Garcia A, Ruiz-Cabello F, Garrido F (1999) Chromosome loss is the most frequent mechanism contributing to HLA haplotype loss in human tumours. Int J Cancer 83:91–97
Jimenez P, Canton J, Concha A, Cabrera T, Fernandez M, Real LM, Garcia A, Serrano A, Garrido F, Ruiz-Cabello F (2000) Microsatellite instability analysis in tumours with different mechanisms for total loss of HLA expression. Cancer Immunol Immunother 48:684–690
Koopman LA, Corver WE, van der Slik AR, Giphart MJ, Fleuren GJ (2000) Multiple genetic alterations cause frequent and heterogeneous human histocompatibility leukocyte antigen class I loss in cervical cancer. J Exp Med 191:961–976
Lanier LL (2001) A renaissance for the tumour immunosurveillance hypothesis. Nat Med 7:1178–1180
Maleno I, Lopez-Nevot MA, Cabrera T, Salinero J, Garrido F (2002) Multiple mechanisms generate HLA class I altered phenotypes in laryngeal carcinomas: high frequency of HLA haplotype loss associated with loss of heterozygosity in chromosome region 6p21. Cancer Immunol Immunother 51:389–396
Mazurenko NN, Beliakov IS, Bliev AIU, Guo Z, Hu X, Vinokurova SV, Bidzhieva BA, Pavlova LS, Ponten J, Kiselev FL (2003) Cervical carcinoma progression-associated genetic alterations on chromosome 6. Mol Biol (Mosk) 37:472–481
Mendez R, Serrano A, Jager E, Maleno I, Ruiz-Cabello F, Knuth A, Garrido F (2001) Analysis of HLA class I expression in different metastases from two melanoma patients undergoing peptide immunotherapy. Tissue Antigens 57:508–519
Metzelaar-Blok JA, Jager MJ, Moghaddam PH, Van der Slik AR, Giphart MJ (1999) Frequent loss of heterozygosity on chromosome 6p in uveal melanoma. Hum Immunol 60:962–969
Paschen A, Mendez RM, Jimenez P, Sucker A, Ruiz-Cabello F, Song M, Garrido F, Schadendorf D (2003) Complete loss of HLA class I antigen expression on melanoma cells: a result of successive mutational events. Int J Cancer 103:759–767
Poetsch M, Dittberner T, Woenckhaus C (2003) Can different genetic changes characterize histogenetic subtypes and biologic behavior in sporadic malignant melanoma of the skin? Cell Mol Life Sci 60:1923–1932
Ramal LM, Feenstra M, van der Zwan AW, Collado A, Lopez-Nevot MA, Tilanus M, Garrido F (2000) Criteria to define HLA haplotype loss in human solid tumours. Tissue Antigens 55:443–448
Schreiber H, Wu, TH, Nachman J, Kast WM (2002) Immunodominance and tumour escape. Semin Cancer Biol 12:25–31
Torres MJ, Ruiz-Cabello F, Skoudy A, Berrozpe G, Jimenez P, Serrano A, Real FX, Garrido F (1996) Loss of an HLA haplotype in pancreas cancer tissue and its corresponding tumour derived cell line. Tissue Antigens 47:372-381
Vajdic CM, Hutchins AM, Kricker A, Aitken JF, Armstrong BK, Hayward NK, Armes JE (2003) Chromosomal gains and losses in ocular melanoma detected by comparative genomic hybridization in an Australian population-based study. Cancer Genet Cytogenet 144:12–17
Acknowledgements
We would like to thank Ms Carmen Amezcua, Ms Toñi and Ms Carmen Gonzalez for technical assistance, and K. Shashok for improving the use of English in the manuscript. This work was partially supported by the Fondo de Investigaciones Sanitarias (FIS), Consejeria de Salud, Junta de Andalucia, by the Plan Andaluz de Investigación (PAI), Spain, and by ESTDAB, project contract no. QLRI-CT-2001-01325.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Rodriguez, T., Méndez, R., Roberts, C.H. et al. High frequency of homozygosity of the HLA region in melanoma cell lines reveals a pattern compatible with extensive loss of heterozygosity. Cancer Immunol Immunother 54, 141–148 (2005). https://doi.org/10.1007/s00262-004-0561-5
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00262-004-0561-5