Abstract A49: Cabozantinib attenuates CXCL8-driven macrophage-dependent migration and invasion of PTEN-deficient prostate cancer
Authors: Neisen Jessica L. , McCabe Nuala , Kennedy Richard D. , Waugh David JJ 
Published: May 2015

Background: The interaction between a tumor and the surrounding non-malignant stroma impacts on nearly every aspect of malignant progression. The heterogeneity of the tumor microenvironment (TME), however, makes mapping this interaction difficult. Each stromal population contributes to various functions and each tumor type has its own distinct pathway of interaction with different stromal populations. The majority of these interactions are mediated by soluble factors secreted into the TME. The loss of the tumor suppressor PTEN, an early event in prostate tumorigenesis, results in a subsequent increase in secretion of the pro-inflammatory CXCL8 chemokine into the TME (Maxwell et al 2013). The specific role of CXCL8 signaling within the prostate TME has not been fully classified and warrants further investigation.

Methods: PTENnull PC3 cells were cultured in the absence and presence of human macrophage-representative THP-1 monocytes. Tumor-derived CXCL8 signaling was attenuated using (i) CXCL8 neutralizing antibodies, (ii) CXCR1 and CXCR2 receptor-targeting small peptide inhibitors (PEPducins), and (iii) a CXCL8-targeted shRNA method to generate stable PC3-120p cells with reduced CXCL8 secretion – a control line termed PC3-NTp was co-developed. PC3 cell motility and invasion was assessed by real-time transwell invasion through MatrigelTM and wound closure assays. Phospho-receptor tyrosine kinase (RTK) arrays were used to identify signaling pathways downstream of CXCL8 signaling which are relevant to prostate tumor cross-talk with infiltrating monocytes.

Results: The presence of THP-1 cells more than doubled the rate of PC3 invasion through MatrigelTM (2.4-fold, p=0.0098) and increased wound closure after 6 h (50.63% v 25.65% for PC3 cells alone; p=0.0108). Macrophage-dependent enhancement of invasion and motility was attenuated following inhibition of CXCL8 signaling (i) by neutralizing antibody (90% decrease in normalized invasion and 43% decrease in wound closure, p&lt;0.001 and p=0.03 respectively relative to IgG) and (ii) PEPducin inhibition of CXCR1 and CXCR2 (38% reduction in wound closure, p=0.0002) compared to non-targeting peptides. Low CXCL8-secreting PC3-120p cells were unable to initiate THP-1-potentiated motility and invasion, however, this response was rescued by addition of exogenous human CXCL8. Exogenous CXCL8 was shown to up-regulate secretion of RTK-activating cytokines and growth factors from THP-1 cells. Addition of the RTK inhibitor cabozantinib (XL184) but not crizotinib abrogated THP-1-dependent invasion and wound healing response of PTEN-deficient PC3 cells. Differential analysis of RTKs regulated by both cabozantinib and CXCL8 neutralizing antibody in co-culture identify c-Kit, RET, VEGFR1 and VEGFR2 as potential downstream regulators of CXCL8-dependent monocyte-enhanced PC3 invasion.

Conclusions: Tumor associated macrophages (TAMs) are correlated with prostate cancer progression. Secretion of CXCL8 from PTEN-deficient prostate cancer cells activates macrophages, which then sustains cancer cell migration and invasion. Attenuating CXCL8 signaling within the TME and its downstream effects provides potential therapeutic targets to reduce metastasis in PTEN-deficient, CXCL8-high prostate cancer patients.

Citation Format: Jessica L. Neisen, Nuala McCabe, Richard D. Kennedy, David JJ Waugh. Cabozantinib attenuates CXCL8-driven macrophage-dependent migration and invasion of PTEN-deficient prostate cancer. [abstract]. In: Abstracts: AACR Special Conference on Cellular Heterogeneity in the Tumor Microenvironment; 2014 Feb 26-Mar 1; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(1 Suppl):Abstract nr A49. doi:10.1158/1538-7445.CHTME14-A49