[What to do with the patient on serotherapy?].
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We assessed the in vivo anti-tumor effectiveness of monoclonal antibodies of different isotypes. Starting with a hybridoma cell secreting an IgG3 anti-Thy-1.1 antibody, we isolated three variant hybridoma cell lines secreting anti-Thy-1.1 antibody of the IgG1, IgG2a, and IgG2b isotypes. Each antibody displayed identical antigen binding properties, but differed in their ability to mediate in vitro lysis of Thy-1.1+ AKR/J SL2 lymphoma cells. In assays of complement dependent cytotoxicity, the relative activity of each antibody isotype was IgG2a = IgG2b greater than IgG3 greater than IgG1. In assays of antibody-dependent cell-mediated cytotoxicity when using non-immune spleen cells as effectors, the relative activities were IgG2a greater than or equal to IgG2b greater than IgG1 greater than IgG3. Infusion of equivalent amounts of each antibody (1.5 mg) in AKR/Cum (Thy-1.2+) mice inoculated subcutaneously with 3 X 10(5) AKR/J SL2 lymphoma cells resulted in significant inhibition of tumor growth only in mice treated with IgG2a antibody. However, the antibodies were cleared at different rates, with the IgG2a antibody having the slowest clearance. When antibody doses were adjusted to achieve equivalent serum levels 24 hr after infusion, all of the antibody isotypes exhibited at least some anti-tumor activity, although IgG2a antibody was again the most effective. These studies demonstrate that the difference in anti-tumor activity between antibodies of different isotypes may result from differences both in their serum clearance rate and their ability to interact with host effector mechanisms.
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We reported inhibition of growth of primary rat mammary carcinomas after infusions of tumor-bearer plasma absorbed with Protein A-Sepharose or inactivated CNBr Sepharose. Absorbed plasmas were depleted of the third component of complement (C3) (other complement components defined similarly) and C5 but not C1, C4, or C2. These results suggested that activation of the alternative pathway of complement might be involved in the observed antitumor effects. To test this concept sera were treated with ethylenedinitrilotetraacetic acid or [ethylenebis(oxyethylenenitrilo)]tetraacetic acid before absorption with Protein A-Sepharose. Ethylenedinitrilotetraacetic acid, by chelating calcium and magnesium, prevents activation of both the alternative and classical complement pathways. [Ethylenebis(oxyethylenenitrilo)]tetraacetic acid, by chelating calcium but not magnesium, permits activation of the alternative pathway but inhibits activation of the classical complement pathway. Sera in the presence or absence of chelating agent were absorbed with Protein A-Sepharose twice at room temperature. After absorption calcium was added to the sera. Rats were treated by i.v. injection of sera twice a week for 2 weeks. Measurements of tumor size were made weekly for 5-7 weeks and then tumor weight was determined. Groups were compared both for size of index and total tumors. The results can be summarized as follows: tumor-bearer sera before absorption did not inhibit the growth of rat primary mammary carcinomas; tumor-bearer sera after absorption with Protein A-Sepharose showed significant consumption of C3 and did inhibit tumor growth; tumor-bearer sera absorbed in the presence of ethylenedinitrilotetraacetic acid did not show a decrease in C3 functional activity and did not inhibit tumor growth; tumor-bearer sera absorbed in the presence of [ethylenebis(oxyethylenenitrilo)]tetraacetic acid did show a decrease in C3 functional activity and did inhibit tumor growth; sera from normal adult female rats after absorption with Protein A-Sepharose did inhibit tumor growth. The results are consistent with a role for the alternative pathway of complement in the inhibition of growth of rat primary mammary carcinomas observed after treatment with absorbed sera.
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