Identification of the JNK-Active Triple-Negative Breast Cancer Cluster Associated With an Immunosuppressive Tumor Microenvironment

Background: Although an immunosuppressive tumor microenvironment (TME) is key for tumor progression, the molecular characteristics associated with the immunosuppressive TME remain unknown in triple-negative breast cancer (TNBC). Our previous functional proteomic study of TNBC tumors identified that C-JUN N-terminal kinase (JNK) pathway-related molecules were enriched in a cluster associated with the inflammatory pathway. However, the role of the JNK pathway in the TNBC TME is still unclear. Methods: Transcriptomic analysis was conducted using The Cancer Genome Atlas datasets. The effect ofJNK-IN-8, a covalent pan-JNK inhibitor, on TNBC tumor growth, lung metastasis, and the TME was measured in TNBC syngeneic mouse models (n = 13 per group). Tumor (n =43) or serum (n =46) samples from TNBC patients were analyzed using multiplex immunohistochemistry or Luminex assay. All statistical tests were 2-sided. Results: CIBERSORT analysis revealed that TNBC patients with high phosphorylated JNK level (n = 47) had more regulatory T cell (Treg) infiltration than those with a low phosphorylated JNK level (n =47) (P = .02). Inhibition of JNK signaling statistically significantly reduced tumor growth (P < .001) and tumor-infiltrating Tregs (P = .02) while increasing the infiltration of CD8(+) T cells in TNBC mouse models through the reduction of C-C motif ligand 2 (CCL2). Tumor-associated macrophages were the predominant cells secreting CCL2, and inhibition of JNK signaling reduced CCL2 secretion of human primary macrophages. Moreover, in patients with TNBC (n = 43), those with high levels of CCL2(+) tumor-associated macrophages had more Treg and less CD8(+) T cell infiltration (P = .04), and the serum CCL2 level was associated with poor overall survival (hazard ratio = 2.65, 95% confidence interval = 1.29 to 5.44, P = .008) in TNBC patients (n = 46). Conclusions: The JNK/C-JUN/CCL2 axis contributes to TNBC aggressiveness via forming an immunosuppressive TME and can offer novel therapeutic strategies for TNBC.

Automated summary

The study identified that high phosphorylated JNK level in TNBC is associated with increased Treg infiltration. Inhibition of JNK signaling led to reduced tumor growth and Treg infiltration, and increased CD8(+) T cell infiltration, possibly through suppression of CCL2 secretion. This suggests that targeting the JNK/C-JUN/CCL2 axis may offer new therapeutic strategies for combating the aggressiveness of TNBC.