Differential Proteomic Responses Of Luminal-A And Basal-Like Breast Cancer Cell Lines During Growth Inhibition Induced By Co-Culture With Agarose Encapsulated Murine Renal Adenocarcinoma (Renca) Cells

Abstract The existence of a tumor growth regulatory network that is conserved across tumor types and species has been hypothesized. Our evidence for this regulatory network is derived from studies demonstrating tumor growth inhibition by agarose encapsulated cancer cells (cancer macrobeads). The ability of encapsulated, thus growth restricted cells, to inhibit freely growing cancer cells has been shown when the encapsulated cells are murine (RENCA) or human (e.g., J82). Clinical trials are underway (NCT01053013, NCT02046174). We have shown that RENCA macrobeads release >10 known tumor inhibitory proteins targeting several signaling pathways including Akt/PI3. To better understand the mechanism(s) of growth inhibition, we used a systems biology approach to identify protein profiles and interactions from macrobead-treated human breast carcinoma cell lines that are either mildly aggressive (MCF7) or highly aggressive (MDA-MB231 [MDA]). Target cells were co-cultured with RENCA macrobeads or left untreated for 5 days. Lysates of target cells were prepared in Laemmli buffer, frozen, and sent for MS/MS. Samples were run in duplicate, protein intensities normalized, and the treated/untreated ratio log10 transformed to get a normal distribution. Protein profiles were analyzed using Key Pathway Analysis (KPA) and Metacore software. Growth inhibition by RENCA macrobeads was confirmed (MCF7 25%; MDA 57%). KPA of MCF7 proteins showed upregulation of ubiquitin pathways involved in degradation of misfolded proteins. Stress induced apoptosis and DNA damage (which correlates with the growth inhibition) were the top 2 pathways upregulated using Pathway and Process Enrichment analysis. This is in line with a strong epigenetic gene deregulation (Metacore). Transcription Factor and Network analyses show upregulation of CREB, A2MR and ATF3 networks. Also, Regulated Network analysis suggests a role for A2MR or RAGE in the inhibition of proliferation and increased apoptosis. Macrobead-treated MDA cells showed no significant changes using KPA. Pathway, Network and Process enrichment (Metacore) showed 5 of the top 10 upregulated processes were associated with cytoskeleton remodeling. Cell cycle and protein folding processes were also upregulated. Transcription factors associated with this response were similar to that of MCF7. RAGE again appears in many of the pathways as indicated by Network and Regulated Network analysis. The protein response to RENCA macrobeads differs for these 2 cell lines. Proteins related to apoptosis are upregulated in the MCF7 cells whereas MDA have a preference for cytoskeleton remodeling. These data support the hypothesis that distinctive tumors may respond differently to RENCA macrobead exposure at both an epigenetic and protein level, nonetheless resulting in growth inhibition. Citation Format: Melissa A. Laramore, Peter James, Prithy C. Martis, Atira Dudley, Lawrence S. Gazda, Carl A. Borrebaeck, Barry H. Smith. Differential proteomic responses of luminal-A and basal-like breast cancer cell lines during growth inhibition induced by co-culture with agarose encapsulated murine renal adenocarcinoma (RENCA) cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5552. doi:10.1158/1538-7445.AM2017-5552