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Demonstration of a Correlation between Tumor Cell H-2 Antigen Content, Immunogenicity, and Tumorigenicity Using Lectin-resistant Tumor Variants¹

Cancer Research Division, Department of Pathology, Queen's University, Kingston, Ontario, K7L 3N6 Canada

Four wheat germ agglutinin-resistant (WGA^R) variants of the highly malignant murine tumor MDAY-D2 were examined for membrane alterations which might correlate with their decrease in tumorigenicity described previously. Injection s.c. of as many as 5 x 10⁶ cells of the WGA^R variants MDW1 or MDW3 was rejected by the normal syngeneic DBA/2 host, but they grew rapidly and progressively in athymic nude mice, suggesting that a vigorous T-cell-mediated immune response was responsible for rejection of the variants in the immunocompetent host. In contrast, an inoculum of as few as 10² MDW4, MDW5, or MDAY-D2 cells gave rise to progressively growing metastatic tumors in normal DBA/2 mice. Hence, the MDW1 and MDW3 variants appeared more immunogenic than did the MDW4, MDW5, or MDAY-D2 parent tumor.

Two lines of evidence suggested that all of the WGA^R variants and parental tumor in fact shared a common tumor-associated antigen; (a) rejection of MDW1 tumor cells by DBA/2 mice immunized them against a subsequent challenge of either MDAY-D2 or MDW4 cells; (b) antitumor cytotoxic T-lymphocyte (CTL) activity was detected in a 4-hr ⁵¹Cr release assay in spleens taken from tumor-bearing mice and restimulated in vitro with mitomycin C-treated tumor cells. MDAY-D2 and WGA^R variants were cross-reactive, both as immunogens at the restimulation stage and as CTL targets. Compared to the tumorigenic lines, MDW1 and MDW3 were more effective stimulators of tumor-specific CTL and were superior targets in the CTL assay. Similarly, stimulation of syngeneic DBA/2 spleen cells with trinitrophenol-modified tumor cells showed MDW1 and MDW3 were more effective stimulators of the trinitrophenyl-specific response and were more readily lysed by trinitrophenyl-directed CTL than were the other WGA^R variants or MDAY-D2.

Separation of microsomal membrane proteins on sodium dodecyl sulfate:polyacrylamide gel electrophoresis revealed an elevated level of two glycoproteins with apparent molecular weights of 43,000 and 47,000, present in the microsomes of the poorly tumorigenic variants MDW1 and MDW3. The glycoproteins were identified as H-2D and H-2K, respectively. Absorption of alloantiserum by whole tumor cells indicated that the WGA^R variants and MDAY-D2 had similar amounts of H-2 on their cell surface. In contrast, absorption tests, using tumor cell endoplasmic reticulum and plasma membrane fractions, as well as analysis of the membrane fractions by sodium dodecyl sulfate:polyacrylamide gel electrophoresis, showed an increased level of H-2 in the endoplasmic reticulum of the poorly tumorigenic MDW1 and MDW3 variants.

The results demonstrate an inverse correlation between the tumorigenicity and both the immunogenicity and cellular H-2 levels of the variant tumor cell lines.

¹ This work was supported by grants from the National Cancer Institute of Canada and the Medical Research Council of Canada.

² National Cancer Institute of Canada Research Scholar. To whom requests for reprints should be addressed.

Received 3/ 9/81. Accepted 6/25/81.