The activation of
signal transducer and activator of transcription 3 (STAT3) has been identified as a key mediator that drives the fundamental components of
melanoma malignancy, including immune suppression in
melanoma patients. Increasing evidence also suggests that regulatory T cells (Tregs) are important in suppressing anti-
tumor immunity and play a dominant role in negating efficacious
immunotherapy approaches. We hypothesized that
WP1066, a novel inhibitor of STAT3 signaling, reverses immune suppression through the inhibition of Tregs and that this contributes to the antitumor activity of this agent against
melanoma brain metastases. We found that the mean percentage of peripheral blood mononuclear cells expressing phosphorylated STAT3 (p-STAT3) was significantly elevated in samples from patients with
melanoma brain metastases compared to healthy donors, 16.13 +/- 2.48% versus 4.17 +/- 1.79%. The p-STAT3 inhibitor
WP1066 enhanced CD3+ (which contained Tregs) but not CD8+ T cell cytotoxicity against human A375
melanoma cells, indicating that this p-STAT3 blockade agent did not directly activate CD8+ T cells. Furthermore, the p-STAT3 inhibitor did not enhance the cytotoxicity of CD3+CD25- T cells (from which Tregs were excluded), indicating that the enhanced cytotoxicity of
WP1066 is secondary to its inhibition of Tregs. This was confirmed by demonstrating that
WP1066 inhibited FoxP3+ Treg induction in a dose-dependent manner. Moreover, CD3+ T cells exhibited markedly enhanced levels of phosphorylated ZAP-70, a critical proximal signal in T cell activation, after exposure to
WP1066. Similar effects were not observed in Treg-depleted CD3+CD25- T cell populations, confirming that the T cell activation by WP compounds is secondary to their inhibition of the Tregs. These results suggest that
WP1066 enhances T cell cytotoxicity against
melanoma through inhibition of Tregs.