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Spheroids of FAP-Positive Cell Lines as a Model for Screening Drugs That Affect FAP Expression
Victor V Pleshkan1,2, Marina V Zinovyeva1, Dina V Antonova1
1Gene Immunooncotherapy Group, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, 117997 Moscow, Russia.
Abstract:
Fibroblast activation protein has a unique expression profile that manifests mainly in wounds and tumors, which anticipates it as an encouraging and selective target for anticancer therapy. However, research of the therapeutic potential of FAP is limited both by legal restraints when working in vivo and by the difficulty of obtaining standardized primary cultures of FAP-positive cancer-associated fibroblasts due to their high heterogeneity. We found that 3D spheroids of FAP-positive cell lines could serve as robust and convenient models of FAP expression, in contrast to monolayers. By exposing such spheroids to various factors and compounds, it is possible to study changes in FAP expression, which are easily detected by confocal microscopy. FAP expression increases under the influence of the TGFβ, does not depend on pH, and decreases during hypoxia and starvation. We believe that the proposed model could be used to organize large-scale high-throughput screening of drugs that target FAP expression.
Insights
Three-dimensional spheroids of cancer-associated fibroblast cell lines offer a robust model for studying Fibroblast Activation Protein (FAP) expression. This 3D spheroid model facilitates drug screening for targeting FAP in cancer therapy.
Area of Science:
- Oncology
- Cell Biology
- Biochemistry
Background:
- Fibroblast Activation Protein (FAP) is a promising therapeutic target due to its expression in tumor microenvironments.
- Research on FAP's therapeutic potential is hindered by in vivo restrictions and challenges in standardizing heterogeneous cancer-associated fibroblast cultures.
Purpose of the Study:
- To develop a reliable and convenient model for studying Fibroblast Activation Protein (FAP) expression.
- To investigate factors influencing FAP expression in a controlled in vitro setting.
- To establish a platform for high-throughput drug screening targeting FAP.
Main Methods:
- Utilized 3D spheroids derived from FAP-positive cell lines as an in vitro model.
- Exposed spheroids to various factors including TGFβ, pH changes, hypoxia, and starvation.
- Quantified FAP expression changes using confocal microscopy.
Main Results:
- 3D spheroids demonstrated robust and reproducible FAP expression, outperforming monolayer cultures.
- FAP expression was upregulated by TGFβ.
- FAP expression was independent of pH but downregulated by hypoxia and starvation.
Conclusions:
- 3D spheroids provide a practical and effective model for studying FAP expression dynamics.
- The proposed model can be instrumental in large-scale high-throughput screening of drugs targeting FAP.
- This approach may overcome limitations associated with in vivo studies and heterogeneous primary cell cultures.
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