GPR35 as a therapeutic target to halt breast cancer progression and osteolytic bone metastasis

Bone metastasis is a frequent and devastating complication of breast cancer (BC) and causes severe skeletal-related events, reduced quality of life, and poor overall survival. BC cells engage in bidirectional interactions with the bone microenvironment by modulating the expression of various chemokines and their receptors, thereby promoting metastatic colonization and niche formation. Numerous G protein-coupled receptors (GPCRs) are implicated in oncogenic processes. Among them, G protein-coupled receptor 35 (GPR35), the cognate receptor for CXCL17, has recently been identified as a key regulator of tumor progression and metastasis. Nevertheless, its precise role in cancer progression, metastatic spread, and modulation of the bone microenvironment remains poorly understood. GPR35 is overexpressed in BC patients, and its elevated expression is strongly associated with reduced overall survival. Consistently, our study also demonstrates elevated GPR35 expression in breast cancer bone-metastatic (BC-BoneMet) cell lines, further supporting its potential role in driving bone metastatic progression. To functionally validate these observations, we generated GPR35 knockout (KO) BC-BoneMet cell lines using the CRISPR/Cas9 system. GPR35 deletion significantly reduced cell migration, invasion, and colony-forming ability in vitro . In vivo studies showed that loss of GPR35 in breast tumor cells led to significantly reduced primary tumor growth in syngeneic mice, supporting a critical role for GPR35 in driving tumor progression. To evaluate the therapeutic benefits of targeting the GPR35, we employed a highly selective GPR35 inhibitor (GPR35i). In functional studies, GPR35i markedly reduced the proliferation, migration, and clonogenicity of BC-BoneMet cells even under CXCL17 stimulation. Furthermore, flow cytometry revealed that GPR35i significantly induces apoptosis and cell cycle delay in BC-BoneMet cells. Given that most BC bone metastases are osteolytic (bone-destructive) and osteoblasts regulate osteoclast functions, we also investigated the effects of the tumor secretome on bone cells. The secretome of GPR35-deficient tumor cells failed to promote bone cell differentiation. Similar effects were observed when the secretome of GPR35i-treated BC-BoneMet cells was applied to osteoblast and osteoclast differentiation assays. Conclusively, these findings demonstrate that GPR35 plays a key role in breast cancer bone metastasis by remodeling the bone microenvironment and supporting the formation of a metastatic niche. These findings highlight GPR35 as a promising therapeutic target for preventing osteolytic progression and reducing metastatic burden in bone.