Enhancing the Responsiveness of Immune Checkpoint Inhibitors in Breast Cancer Through PTHrP Blockade

초록

Immune checkpoint inhibitors (ICIs), such as anti-PD-1 and -CTLA-4 antibodies, are eective in many types of cancers, yet their efficacy on breast cancer is comparatively modest due to the immunouppressive ‘cold’ tumor microenvironment (TME). We have previously shown that parathyroid hormone-related peptide (PTHrP) from breast cancer contribute to the mobilization of myeloid-derived suppressor cells (MDSC) from the bone marrow via activation of osteoblasts, contributing to the immunosuppressive TME of breaset cancer. Accordingly, we hypothesized that blockade of PTHrP enhances the efficacy of ICI. We employed the CRISPR/Cas9 system to generate PTHrP knockout cell lines from 4T1 murine triple-negative breast cancer cells. After genotyping and Pthlh mRNA levels, we selected both heterozygous and homozygous PTHrP knockout 4T1 cell lines, which maintained the morphological characteristics and proliferation rates of the parental lines in vitro. In a subcutaneous tumor model, PTHrP knockout in 4T1 cells, previously non-responsive to PD-1 inhibitors, demonstrated a marked reduction in tumor growth following anti-PD-1 antibody treatment. Notably, tumor reduction was less significant in heterozygous knockdown cells compared to those with homozygous deletions. Furthermore, while the anti-PD-1 antibody did not significantly increase CD8-positive T cell counts in the TME of the 4T1 breast tumor model, the PTHrP knockout led to a decreased MDSC count and an increase in CD8-positive T cells within the TME. In conclusion, our study demonstrates that PTHrP plays a critical role in diminishing ICI efficacy in breast cancer by facilitating MDSC migration into the TME. Consequently, we propose a new therapeutic strategy involving integrating PTHrP neutralization with other chemotherapeutic agents in combination therapy, aiming to achieve greater treatment benefits for PTHrP-expressing cold tumors non-responsive to ICIs.