Discipline: Biological Sciences
Subcategory: Cancer Research
Alexxa C. Cruz-Bonilla - University of Puerto Rico, Mayaguez
Hedgehog (Hh) signaling activity is highly upregulated in about 30-40% of tumor specimens and it has been associated with poor prognosis in breast tumors of the triple-negative sub-type. Hh signaling activity has been linked with the phenotype of tumor-associated macrophages (TAMs) in tumors, yet its oncogenic role within the context of triple-negative breast cancer (TNBC) is not well-understood. We hypothesize that TAMs support the oncogenic activity of Hh signaling in TNBC in a paracrine manner. Hh signaling activity was examined in monocytes, TAMs, and MDA-MB-231 (TNBC cell line) through qRT-PCR and Western Blot. The qRT-PCR results showed no significant difference in the levels of GLI1 and SMO genes in TAM monocultures +/- SHH ligand, indicating the absence of canonical Hh signaling activity. The SHH ligand was only expressed in tumor cells but not in TAMs. Tumor cell growth was examined in a co-culture system when increasing TNBC to a set TAM quantity, with a 2:1 and 1:1 ratio. These ratios were also observed under -/+ NVP-LDE225 (NVP) treatment; a Hh pathway inhibitor. Macrophage retention in the co-cultures was analyzed in comparison to the TAM monocultures, these demonstrated no statistical difference across all conditions. All experiment replicates of n=3 that were performed demonstrated replicability amongst all samples. Initial examination of growth rates indicates that NVP does not affect the TAM and TNBC monocultures; having a non-significant difference when analyzed through a One-Way Anova. Utilizing Dunnet’s multiple comparison test, the results showed a decreasing tendency in the tumor proliferation rates between the co-cultures treated -/+ NVP and when compared to the TNBC monocultures. In the absence of NVP, both 2:1 and 1:1 co-culture, the difference in proliferation rate was significantly higher (p<0.0001) than in the TNBC monocultures. Where the 1:1 co-culture -NVP conditions had a p<0.0001 and +NVP a p=0.0020, decreasing the proliferation rate when in treatment. Overall, results pointed to an increase in TNBC growth when in direct contact with TAMs and lower proliferation rates in co-cultures under NVP treatment. Utilizing the same parameters established for the co-culture system, a tri-culture model with a fibroblast cell line is currently being tested. Future work also includes replicating into the three-dimensional platform to better understand the role of cell-to-cell contact and the metabolic processes that occur in vivo.
Funder Acknowledgement(s): National Institutes of Health
Faculty Advisor: Prof. Maribella Domenech, email@example.com
Role: This research has been conducted solely by myself over a four-year span in unison with the Principle Investigator, Prof. Maribella Domenech.