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Autoimmune expression of a cytoplasmic protein p60 in mice bearing metastasizing SV40‐transformed tumors
Author(s) -
Manneck Hans E.,
Pfaff Martin,
Anderer F. Alfred
Publication year - 1988
Publication title -
international journal of cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.475
H-Index - 234
eISSN - 1097-0215
pISSN - 0020-7136
DOI - 10.1002/ijc.2910420620
Subject(s) - immunofluorescence , antigen , biology , antibody , epitope , immune system , cytoplasm , microbiology and biotechnology , cell culture , cancer research , immunology , genetics
In STU mice bearing metastasizing SV40‐transformed 51A‐232B‐M tumors, an immune response against a cellular 60kDa protein (p60) developed in about 50% of the tumor‐bearing animals, in addition to the response against SV40 large T‐antigen and cellular protein p53. The anti‐p60 auto‐immune response could be observed as early as 11 days after tumor challenge and was strictly linked with metastatic spread but was not a prerequisite for metastasis. Anti‐p60 antibodies could not be detected in sera of animals bearing metastasizing Rous‐sarcoma virus‐transformed or methylcholanthrene‐induced tumors, or in sera from human cancer patients with clinically confirmed metastatic spread. The anti‐p60 auto‐antibodies showed a broad cross‐reactivity against components of similar size in a great number of cell lines of various species and in normal mouse tissue. The p60 auto‐antigen is a cytoplasmic protein which is neither phosphorylated nor glycosylated in vivo. Immunoblotting performed with fresh cell lysates under non‐reducing conditions using tumor‐bearer sera revealed a diffuse p60 double band, but under reducing conditions only one sharp p60 band was observed. The reaction of p60 with anti‐p60 auto‐antibodies could be completely blocked by pre‐treatment of fresh cell lysates with N‐ethylmaleimide or p‐chloromercuriphenyl sulfonate, or by oxidation in air prior to immunoblotting, indicating that the anti‐p60 autoimmune response was directed against an epitope sensitive to SH‐group‐blocking reagents. Immunofluorescence studies with tumor‐bearer sera showed only a very weak cytoplasmic fluorescence, possibly due to the nature of the p60 SH‐groups in situ being masked. Immunoprecipitates with monoclonal antibodies against SV40 large T‐antigen and p53 obtained from fresh cell lysates of SV40‐transformed tumor cells contained no associated p60 auto‐antigen. The p60 auto‐antigen was purified from tumor cell homogenates with an enrichment factor of about 2,000; its iso‐electric point is at Ph 6.8. Determination of the biological half‐life of p60 yielded a value of about 28 hr. The p60 auto‐antibodies in pools of tumor‐bearer sera taken at day 40 after tumor challenge all belonged to the IgG 1 subclass.