Abstract
The immunogenicity of the disialoganglioside, GD3, a melanoma-tumor-associated antigen, has been evaluated in non-human primates. Sera from four chimpanzees and two monkeys were evaluated for anti-GD3 antibody activity by solid-phase radioimmunoassay using GD3 and control gangliosides as targets. Serum from one monkey, immunized with cells from a melanoma cell line, was strongly reactive with GD3, having a titer of >2500. In contrast, serum from this animal was non-reactive with several other gangliosides including the structurally similar GM3. Anti-GD3 reactivity was also demonstrable, albeit in low titer, in the sera of an additional monkey and a chimpanzee. Each of these animals had likewise been immunized using cells from melanoma cell lines. On the basis of these observations, suggestive of a primate anti-GD3 antibody response, we initiated a series of immunizations of chimpanzee using purified GD3 bound to Salmonella minnesota, R595. IgG reactive with melanoma cells in the cell-binding assay was first detected in sera collected after 4 immunizations and increased in titer against each reactive melanoma cell line during the immunizations. Reactivity of this serum with melanoma cell lines demonstrated a direct correlation with the expression of GD3 by the respective cell line. Anti-GD3 reactivity was evident in solid-phase radioimmunoassay against purified GD3 beginning with serum collected after 11 immunizations. By comparison with its binding to the control ganglioside panel, this serum demonstrated strong specificity for GD3 (titer=640) while having only marginal reactivity with GM3 (titer=40). Immune serum from this animal was also able specifically to block subsequent binding of a murine IgM anti-GD3 antibody (DMab7) to target GD3 in solid-phase radioimmunoassay. Together, these observations suggest that GD3, in the form of a purified molecule bound to a bacterial matrix or as part of the intact melanoma cell membrane, can be immunogenic in non-human primates, and is able to elicit an antibody response of appropriate specificity.
Keywords: Melanoma, Melanoma Cell, Antibody Response, Melanoma Cell Line, Immune Serum
Footnotes
Supported in part by grant CA32672 from the National Cancer Institute, Veterans Administration Program 821 and by the Yerkes Regional Primate Center, Atlanta, Georgia. The Yerkes Center is fully accredited by the American Association for Accreditation of Laboratory Animal Care
References
- 1.Cheresh DA, Honski CJ, Staffilino LK, Jung G, Reisfeld RA. Disialoganglioside GD3 on human melanoma serves as a relevant target antigen for monoclonal antibody mediated tumor cytolysis. Proc Natl Acad Sci USA. 1984;82:5155–5159. doi: 10.1073/pnas.82.15.5155. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 2.Dippold WG, Lyoyd KO, Li LTC, Ikeda H, Oettgen HF, Old LJ. Definition of six antigenic systems with mouse monoclonal antibodies. Proc Natl Acad Sci USA. 1980;77:6114–6118. doi: 10.1073/pnas.77.10.6114. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3.Dippold WG, Knuth A, Buschenfelde K-HM. Inhibition of human melanoma cell growth in vitro by monoclonal anti-GD3 ganglioside antibody. Cancer Res. 1984;44:806–810. [PubMed] [Google Scholar]
- 4.Fredman P, Noren R, Mansson J-E, Svennerholm L. Concentration, composition and structure of gangliosides from the central nervous system of dogfish, Squalus acanthia, and cod, Gadus callarias . Biochim Biophys Acta. 1982;713:410–418. doi: 10.1016/0005-2760(82)90260-0. [DOI] [PubMed] [Google Scholar]
- 5.Gallanos C, Luderlitz O, Westphal O. Preparation of antisera against the lipid-A component of bacterial lipopolysaccharides. Eur J Biochem. 1971;24:116–122. doi: 10.1111/j.1432-1033.1971.tb19661.x. [DOI] [PubMed] [Google Scholar]
- 6.Graus F, Cordon-Condo C, Houghton AN, Melamed MR, Old LJ. Distribution of the ganglioside GD3 in the human nervous system detected by R24 mouse monoclonal antibody. Brain Res. 1984;324:190–194. doi: 10.1016/0006-8993(84)90642-5. [DOI] [PubMed] [Google Scholar]
- 7.He X, Svennerholm L, Fredman P, Wickstrand CJ, Bigner DD (1988) (in press)
- 8.Hersey P. Review of melanoma antigens recognized by monoclonal antibodies (Mabs): Their functional significance and applications in diagnosis and treatment of melanoma. Pathology. 1985;17:346–354. doi: 10.3109/00313028509063778. [DOI] [PubMed] [Google Scholar]
- 9.Honsik CJ, Jung G, Reisfeld RA. LYmphokine activated killer cells targeted by monoclonal antibodies to the disialogangliosides GD2 and GD3 specifically lyse human tumor cells of neuroectodermal origin. Proc Natl Acad Sci USA. 1986;83:7893–7897. doi: 10.1073/pnas.83.20.7893. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 10.Houghton AN, Mintzer D, Cordon-Cardo C, Welt S, Fliegel B, Vadhan S, Carswell E, Melamed MR, Oettgen HO, Old LJ. Mouse monoclonal IgG3 antibody detecting GD3 ganglioside: a phase 1 trial in patients with malignant melanoma. Proc Natl Acad Sci USA. 1985;82:1242–1246. doi: 10.1073/pnas.82.4.1242. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 11.Hunter RW, Greenwood FC. Preparation of iodine-131 labelled human growth hormone of high specific activity. Nature. 1962;194:495–496. doi: 10.1038/194495a0. [DOI] [PubMed] [Google Scholar]
- 12.Mutzner PA, Stuhlmiller GM, Seigler HF. Characterization of melanoma cell membrane tumor associated antigens using xeno-, allo- and autoserum. I. Immunofluorescence. J Surg Oncol. 1980;14:367–377. doi: 10.1002/jso.2930140411. [DOI] [PubMed] [Google Scholar]
- 13.Real FX, Houghton AN, Albino AP, Cordon-Cardo C, Melamed MR, Oettgen HO, Old LJ. Surface antigens of melanomas and melanocytes defined by mouse monoclonal antibodies: specificity analysis and comparison of antigen expression in cultured cells and tissues. Cancer Res. 1985;45:4401–4411. [PubMed] [Google Scholar]
- 14.Seigler HF, Mohanakumar T, Metzgar RS, Stuhlmiller GM. Human melanoma and leukemia associated antigens defined by non-human primate antisera. Fed Proc. 1975;34:1642–1646. [PubMed] [Google Scholar]
- 15.Seigler HF, Cox E, Mutzner PA, Shepard L, Nicholson E, Shingleton WW. Specific active immunotherapy for melanoma. Ann Surg. 1979;190:366–372. doi: 10.1097/00000658-197909000-00012. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16.Seigler HF, Wallack MK, Vervaert CE, Bash JA, Roberson KM, Stuhlmiller GM (1988) Melanoma patient antibody responses to melanoma tumor associated antigens defined by murine monoclonal antibodies. J Biol Response Mod (in press) [PubMed]
- 17.Singer KH, Hashimoto K, Stuhlmiller GM, Lazarus GS. Antibody-induced release of cellular proteinases: Loss of adhesion of human melanoma cells after binding of anti-melanoma antibody. J Immunol. 1982;128:1321–1326. [PubMed] [Google Scholar]
- 18.Stuhlmiller GM, Seigler HF. Characterization of a chimpanzee anti-human melanoma antiserum. Cancer Res. 1975;35:2132–2137. [PubMed] [Google Scholar]
- 19.Stuhlmiller GM, Mohanakumar T, Metzgar RS, Seigler HF. The use of non-human primates for production of antisera to human tumor associated antigens. J Immunol Methods. 1976;12:267–274. doi: 10.1016/0022-1759(76)90048-x. [DOI] [PubMed] [Google Scholar]
- 20.Stuhlmiller GM, Borowitz MJ, Seigler HF. D6.1, a murine monoclonal antimelanoma antibody from congenitally athymic nude mice immunized with partially purified melanoma tumor-associated antigen. Hybridoma. 1984;3:333–346. doi: 10.1089/hyb.1984.3.333. [DOI] [PubMed] [Google Scholar]
- 21.Stuhlmiller GM, Darrow TL, Haupt DM, Seigler HF. Immune response of chimpanzee to purified melanoma 250 kilodalton tumor-associated antigen. Cancer Immunol Immunother. 1987;25:193–200. doi: 10.1007/BF00199147. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 22.Svennerholm L, Fredman P. A procedure for quantitative isolation of brain gangliosides. Biochim Biophys Acta. 1980;617:97–109. doi: 10.1016/0005-2760(80)90227-1. [DOI] [PubMed] [Google Scholar]
- 23.Wickstrand CJ, Bigner DD. Expression of fetal brain antigens by human tumors of neuroectodermal origin as defined by monoclonal antibodies. Cancer Res. 1982;42:267–275. [PubMed] [Google Scholar]