A NEW INTERFERON-α ASSAY SYSTEM INCLUDING INDIRECT ANTITUMOR EFFECT

Basic and Clinical Study for Renal Cell Carcinoma

Abstract

In this report, the clinical value of a new interferon (INF) assay and measuring serum tumor necrosis factor (TNF) titer in renal cell carcinoma (RCC) patients is described. In order to find out ineffective cases, we established a new in vitro IFN assay system which was modified from human tumor clonogenic assay (HTCA) using an underlayer including monocytes (5×10⁴/dish) and lymphocytes (5×10⁵/dish) as the feeder cells obtained from human peripheral venous blood. This assay can evaluate both direct and indirect (immune mediated) antitumor effects. Various kinds of cytokines (TNFα, IL-1α, -1β) were measured simultaneously in the cultured supernatants in vitro as well as the serum value in vivo by sandwich immunoenzymometric assay to compare the relationship between antitumor effect and clinical course.
In the basic study, cultured ACHN cell line was used as the target cells. The inhibition of colony growth was observed in a dose relative manner. Continuous IFN-α exposure of 50 and 500IU/ml in the presence of feeder cells showed a significant reduction of colony growth in comparison with cultured plates containing the drug only. Among the cytokines in the supernatants after incubation for 24 hours with IFN-α (500IU/ml) including feeder cells, only TNFα showed significant elevation.
In the clinical study, 31 RCC patients were tried for sensitivity testing using modified HTCA system. The sufficient colony growth was observed in 19 cases (61%). In 25 cases serum cytokines were compared with preoperative and postoperative values; twenty cases received post-operative INF-α administrations, and 5 cases did not. Seven of the 20 cases with IFN-α therapy had some evaluable lesions.
1) The rates of colony survivals were correlated with TNFα titers in the supernatants (r=-0.90, p<0.01).
2) The serum levels of TNFα rose in 15 of the 20 cases (75%) after IFN-α therapy. The elevated value was significantly higher than the value of pretreatment period (p<0.05, paired t-test).
3) There was a significant correlation between the TNFα titers in the supernatants (in vitro) and in the sera (in vivo) treated with IFN-α.
4) All of 3 cases with low serum TNFα titers during IFN therapy showed progressive disease. But in 4 cases with high serum TNFα titers metastatic lesions did not change in more than one year.
These results indicate that antiproliferative effects in vitro assay system were corresponded to TNFα, and that the serum TNFα titers in vivo also had some relation to IFN-α therapy and clinical course. We conclude that this new assay system can predict the indirect antitumor effect of IFN-α, and that serum levels of TNFα titers are useful to monitor the immune mediated actions of IFN in RCC patients.