T lymphocytes from
tumor-draining lymph nodes (TDLN), after activation and expansion in vitro, can mediate regression of metastatic
tumor in animal models. We have shown that TDLNs are subject to
tumor-induced suppression that is
tumor specific, T-cell mediated, and dependent on the duration of
tumor growth, but the mechanism of this suppression remains largely unknown. Recently, in other model systems,
tumor-bearer T cells have been shown to have decreased expression of
T-cell receptor-zeta (TCR zeta), a key component in
antigen-driven activation pathways. We sought to investigate whether the suppression of TDLN reactivity that accompanies prolonged
tumor growth was associated with decreased expression of TCR zeta in fresh and in vitro activated lymph node lymphocytes. Mice bearing subcutaneous
tumor deposits of MCA 205 had TDLN cells harvested after various durations of
tumor growth, then activated in vitro with anti-CD3 for 2 days (activation phase), followed by expansion with
interleukin-2 (IL-2) (10 U/ml) for 3 days (expansion phase). Two-color flow cytometry was used to determine TCR zeta expression in fresh and activated TDLN cells. Antitumor reactivity was assessed by the ability of activated TDLN to mediate regression of lung
metastases. There was a time-dependent suppression of the antitumor reactivity of the activated TDLN; activated TDLN from mice bearing
tumors 14 days or less were able to mediate the regression of established lung
metastases, whereas activated TDLN from animals bearing
tumors 21 days or more were ineffective. In addition, TCR zeta expression on T lymphocytes from fresh and activated TDLN was also depressed in a time-dependent manner. Because
tumor-induced immunosuppression in our model is known to be T cell mediated, we examined whether the Th2
cytokine IL-4, when added in vitro during activation or expansion, could suppress antitumor reactivity and lead to a depression in TCR zeta expression of TDLN cells in a fashion similar to prolonged
tumor growth. The addition of 10 U/ml of
IL-4 in vitro had a marked suppressive effect on the antitumor activity of day 14 TDLN; the effect was most pronounced when
IL-4 was present during the expansion phase. Fluorescence-activated cell sorter analysis of day 14 TDLN exposed to
IL-4 in vitro demonstrated a marked decrease in TCR zeta expression, comparable to that seen in late
tumor-bearer TDLN. Thus, TDLN from late
tumor-bearers show a consistent decrease in TCR zeta expression that is associated with suppressed antitumor reactivity, and exposure to
IL-4 in vitro results in qualitatively and quantitatively similar changes. Our observations suggest a mechanism whereby Th2 cells could mediate immunosuppression by downregulating a critical component of the
T-cell-receptor signal transduction machinery.