Skip to main content
PMC home page
Cancer Immunology, Immunotherapy : CII logoLink to Cancer Immunology, Immunotherapy : CII
. 1991 Sep;32(5):325–330. doi: 10.1007/BF01789051

Analysis of T cell receptor β and γ genes from peripheral blood, regional lymph node and tumor-infiltrating lymphocyte clones from melanoma patients

Mark R Albertini 1, Janice A Nicklas 3, Bettejayne F Chastenay 3, Timothy C Hunter 3, Richard J Albertini 3, Steven S Clark 1, Jacquelyn A Hank 1, Paul M Sondel 1,4,5
PMCID: PMC11038145  PMID: 1825620

Abstract

A total of 199 T cell clones from two melanoma patients were derived from progenitor T cells from recurrent melanoma, regional lymph nodes (either involved or uninvolved with malignancy) and peripheral blood by inoculating single cells directly into the wells of microtiter plates before in vitro expansion. The surface marker phenotype of most clones was CD4+CD8−, although some were CD4−CD8+. Genomic DNA prepared from all clones was analyzed by Southern blot hybridization using T cell receptor (TCR) β and γ gene probes, seeking clones with identical TCR gene rearrangement patterns as direct evidence for in vivo progenitor T cell clonal amplification. ProbingHindIII-digested DNA with TCRβ and TCRγ probes revealed several clones with identical TCR gene rearrangement patterns. These clones had subsequent probing ofBamHI-digested DNA with TCRβ and TCRγ probes, which showed all but 2 clones to have distinct rearrangement patterns. These analyses provide clear molecular evidence for in vivo polyclonal CD4+ T cell populations in each of several separate immune compartments in these patients.

Keywords: Melanoma, Regional Lymph Node, Cell Receptor, Cell Clone, Melanoma Patient

Footnotes

This investigation was supported by National Institutes of Health, National Research Service Award CA-08 397 from the National Cancer Institute as well as NIH CA-32 685, CA-30 688, DOE FG028 760 502 and American Cancer Society Grant ACS CH-237

References

  • 1.Acha-Orbea H, Mitchell D, Timmermann L, Wraith D, Tausch G, Waldor M, Zamvil S, McDevitt H, Steinman L. Limited heterogeneity of T-cell receptors from lymphocytes mediating autoimmune encephalomyelitis allows specific immune intervention. Cell. 1988;54:263–273. doi: 10.1016/0092-8674(88)90558-2. [DOI] [PubMed] [Google Scholar]
  • 2.Albertini RJ. Somatic gene mutations in vivo as indicated by the 6-thioguanine-resistant T-lymphocytes in human blood. Mutat Res. 1985;150:411–422. doi: 10.1016/0027-5107(85)90138-1. [DOI] [PubMed] [Google Scholar]
  • 3.Anichini A, Fossati G, Parmianio G. Clonal analysis of the cytolytic T-cell response to human tumors. Immunol Today. 1987;8:385–389. doi: 10.1016/0167-5699(87)90215-5. [DOI] [PubMed] [Google Scholar]
  • 4.Eynde VC, Hainaut P, Herin M, Knuth A, Lemoine C, Weynants P, Bruggen PVD, Fauchet R, Boon T. Presence on a human melanoma of multiple antigens recognized by autologous CTL. Int J Cancer. 1989;44:634–640. doi: 10.1002/ijc.2910440413. [DOI] [PubMed] [Google Scholar]
  • 5.Fishleder AJ, Finke JH, Tubbs R, Bukowski RM. Induction by interleukin-2 of oligoclonal expansion of cultured tumor-infiltrating lymphocytes. J Natl Cancer Inst. 1990;82:124–128. doi: 10.1093/jnci/82.2.124. [DOI] [PubMed] [Google Scholar]
  • 6.Fleischer B, Bogdahn U. Growth of antigen specific, HLA restricted T-lymphocyte clones from cerebrospinal fluid. Clin Exp Immunol. 1983;52:38–44. [PMC free article] [PubMed] [Google Scholar]
  • 7.Fleischer B, Marquardt P, Poser S, Kreth H. Phenotypic markers and functional characteristics of T-lymphocyte clones from cerebrospinal fluid in multiple sclerosis. J Neuroimmunol. 1984;7:151–162. doi: 10.1016/s0165-5728(84)80015-6. [DOI] [PubMed] [Google Scholar]
  • 8.Fliedner V, Qiao L, Whiteside T, Leyvraz S, Barras C, Miescher S. Clonogenic and functional potential of T-lymphocytes infiltrating human solid tumors. In: Truitt R, Gale R, Bortin M, editors. Cellular immunotherapy of cancer. New York: Liss; 1987. pp. 223–232. [PubMed] [Google Scholar]
  • 9.Galili U, Vanky F, Rodriguez L, Klein E. Activated T-lymphocytes within human solid tumors. Cancer Immunol Immunother. 1979;6:129–133. [Google Scholar]
  • 10.Hamaoka T, Fujiwara H. Phenotypically and functionally distinct T-cell subsets in antitumor responses. Immunol Today. 1987;8:267–269. doi: 10.1016/0167-5699(87)90186-1. [DOI] [PubMed] [Google Scholar]
  • 11.Hersey P, Bolhuis R. ‘Nonspecific’ MHC-unrestricted killer cells and their receptors. Immunol Today. 1987;8:233–239. doi: 10.1016/0167-5699(87)90173-3. [DOI] [PubMed] [Google Scholar]
  • 12.Itoh K, Tilden A, Balch C. Interleukin-2 activation of cytotoxic T-lymphocytes infiltrating into human metastatic melanomas. Cancer Res. 1986;46:3011–3017. [PubMed] [Google Scholar]
  • 13.Itoh K, Platsoucas C, Balch C. Autologous tumor-specific cytotoxic T-lymphocytes in the infiltrate of human metastatic melanoma. J Exp Med. 1988;168:1419–1441. doi: 10.1084/jem.168.4.1419. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14.Knuth A, Wolfel T, Klehmann E, Boon T, Buschenfelde K. Cytolytic T-cell clones against an autologous melanoma: specificity study and definition of three antigens by immunoselection. Proc Natl Acad Sci USA. 1989;86:2804–2808. doi: 10.1073/pnas.86.8.2804. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 15.Lanier L, Phillips J. Evidence for three types of human cytotoxic lymphocytes. Immunol Today. 1986;7:132–134. doi: 10.1016/0167-5699(86)90076-9. [DOI] [PubMed] [Google Scholar]
  • 16.Lee SJ, Benjamin D, Weiner H, Seidman J, Duby A, Ang S, Hafler D. Clonality and T-cell receptor γ chains in immune compartments: examination of cerebrospinal fluid derived T-cell clones. In: Davis M, Kappler J, editors. The T-cell receptor. New York: Liss; 1988. pp. 87–97. [Google Scholar]
  • 17.Lefranc M, Forster A, Rabbitts T. Rearrangement of two distinct T-cell γ-chain variable-region genes in human DNA. Nature. 1986;319:420–422. doi: 10.1038/319420a0. [DOI] [PubMed] [Google Scholar]
  • 18.Marrack P, Kappler J. The antigen-specific, major histocompatibilitycomplex-restrictedreceptoron T-cells. AdvImmunol. 1986;38:1–30. doi: 10.1016/s0065-2776(08)60005-x. [DOI] [PubMed] [Google Scholar]
  • 19.Marrack P, Kappler J. The T-cell receptor. Science. 1987;238:1073–1078. doi: 10.1126/science.3317824. [DOI] [PubMed] [Google Scholar]
  • 20.Miescher S, Stoeck M, Qigo L, Barras C, Barrelet L, Fliedner V. Preferential clonogenic deficit of CD8-positive T-lymphocytes infiltrating human solid tumors. Cancer Res. 1988;48:6992–6998. [PubMed] [Google Scholar]
  • 21.Modlin R, Brenner M, Krangel M, Duby A, Bloom B. T-cell receptors of human suppressor cells. Nature. 1987;329:541–545. doi: 10.1038/329541a0. [DOI] [PubMed] [Google Scholar]
  • 22.Moller D, Komishi K, Kirby M, Balbi B, Crystal R. Bias toward use of a specific T-cell receptor β-chain variable region in a subgroup of individuals with sarcoidosis. J Clin Invest. 1988;82:1183–1191. doi: 10.1172/JCI113715. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 23.Mukherji B, Guha A, Chakraborty N, Sivanandham M, Nashed A, Sporn J, Ergin M. Clonal analysis of cytotoxic and regulatory T-cell responses against human melanoma. J Exp Med. 1989;169:1961–1979. doi: 10.1084/jem.169.6.1961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 24.Nicklas JA, Hunter T, Sullivan L, Berman J, O'Neill J, Albertini R. Molecular analysis of in vivo hprt mutations in human T-lymphocytes: I. Studies of low frequency ‘spontaneous’ mutants by Southern blots. Mutagenesis. 1987;2:341–347. doi: 10.1093/mutage/2.5.341. [DOI] [PubMed] [Google Scholar]
  • 25.Nicklas JA, Hunter T, O'Neill J, Albertini R. Molecular analyses of in vivo hprt mutations in human T-lymphocytes: III. Longitudinal study of hprt gene structural alterations and T-cell clonal origins. Mutation Res. 1989;215:147–160. doi: 10.1016/0027-5107(89)90178-4. [DOI] [PubMed] [Google Scholar]
  • 26.Nitta T, Oksenberg JR, Rao NA, Steinman L. Predominant expression of T-cell receptor Vα7 in tumor-infiltrating lymphocytes of uveal melanoma. Science. 1990;249:672–674. doi: 10.1126/science.2382141. [DOI] [PubMed] [Google Scholar]
  • 27.O'Neill JP, McGinniss M, Berman J, Sullivan L, Nicklas J, Albertini R. Refinement of a T lymphocyte cloning assay to quantify the in vivo thioguanine-resistant mutant frequency in humans. Mutagenesis. 1987;2:87–94. doi: 10.1093/mutage/2.2.87. [DOI] [PubMed] [Google Scholar]
  • 28.Padula SJ, Sjroi D, Lingenheld E, Love J, Jr, Chou C, Clark R. T-Cell receptor beta chain gene rearrangement shared by murine T-cell lines derived from a site of autoimmune inflammation. J Clin Invest. 1988;81:1810–1818. doi: 10.1172/JCI113524. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 29.Pardoll D, Kruisbeek A, Fowlkes B, Coligan J, Schwartz R. The unfolding story of T-cell receptor γ. FASEB J. 1987;1:103–109. doi: 10.1096/fasebj.1.2.2956146. [DOI] [PubMed] [Google Scholar]
  • 30.Poljak R. Hypothesis: a molecular model for the MHC-restricted recognition of antigens by the T-cell receptor. Ann Inst Pasteur Immunol. 1987;138:175–180. doi: 10.1016/s0769-2625(87)80069-7. [DOI] [PubMed] [Google Scholar]
  • 31.Reinherz E (1987) T-Cell receptors: who needs more? Nature 660–663 [DOI] [PubMed]
  • 32.Rosenberg SA, Packard B, Aebersold P, Solomon D, Topalian S, Toy S, Simon P, Lotze M, Yang J, Seipp C, Simpson C, Carter C, Bock S, Schwartzentruber D, Wei J, White D. Use of tumorinfiltrating lymphocytes and interleukin-2 in the immunotherapy of patients with metastatic melanoma. N Engl J Med. 1988;319:1676–1680. doi: 10.1056/NEJM198812223192527. [DOI] [PubMed] [Google Scholar]
  • 33.Royer D, Reinherz E. T-lymphocytes: ontogeny, function, and relevance to clinical disorders. N Engl J Med. 1987;317:1136–1142. doi: 10.1056/NEJM198710293171807. [DOI] [PubMed] [Google Scholar]
  • 34.Sondel PM, Hank J, Kohler P, Chen B, Sosman J. Lymphokine activated killing as treatment for human cancer: clinical extrapolations from laboratory studies with interleukin-2 expanded leukocytes. In: Mitchell M, editor. Immunity to cancer; II. New York: Liss; 1989. pp. 151–160. [PubMed] [Google Scholar]
  • 35.Stamenkovic I, Stegagno M, Wright K, Krane S, Amento E, Colvin R, Duquesnoy R, Kurnick J. Clonal dominance among T-lymphocyte infiltrates in arthritis. Proc Natl Acad Sci USA. 1988;85:1179–1183. doi: 10.1073/pnas.85.4.1179. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 36.Thompson D, Sutherland M, Scanzano R, Shenouda G. Recognition of a purifiedM r-40,000 organ specific immunogen of human lung cancer by class II-restricted T-cells. JNCI. 1987;79:653–661. [PubMed] [Google Scholar]
  • 37.Yanagi Y, Yoshikai Y, Leggett K, Clark S, Alekgander I, Mak T. A human T-cell-specific cDNA clone encodes a protein having extensive homology to immunoglobulin chains. Nature. 1984;308:145–149. doi: 10.1038/308145a0. [DOI] [PubMed] [Google Scholar]

Articles from Cancer Immunology, Immunotherapy : CII are provided here courtesy of Springer

RESOURCES