Abstract Background: Most prostate cancer in vitro models are derived from metastasized disease and thus are not well suited to study the early events in prostate carcinogenesis. Aims: Here, we used the immortalized non-malignant prostate epithelial cell line RWPE-1 to create a sequential in vitro model of prostate epithelial cell transformation. RWPE-1 cells were first transduced to overexpress androgen receptor (AR), secondly transduced to overexpress TMPRSS2-ERG fusion and thirdly CRISPR’d to delete either one or both of the PTEN alleles. Functional and transcriptomic characterization of the cell line series allows better understanding of the early prostate transformation as well as provides a useful tool set to further study additional early events in prostate cancer. Results: AR expressing RWPE-1 cells maintain normal prostate epithelial phenotype where the presence of androgens acts as differentiation stimuli leading to growth reduction through cell cycle arrest. TMPRRS2-ERG expression together with AR activation results into even more pronounced growth retardation, suggesting that in this model TMPRRS2-ERG alone does not have the transformative potential. Adding heterozygous PTEN deletion increases the growth and migration potential of these cells in the presence of androgens, and cells with PTEN deletion together with TMPRSS2-ERG migrate faster than only PTEN deleted cells, highlighting the synergistic role of TMPRSS2-ERG and PTEN in prostate cancer progression. At the level of gene expression, the growth suppressing effect of AR seems to encompass wide range of biosynthetic processes in all lines of the model. However, increased inflammatory signaling and KRAS pathway activity is seen in the PTEN deleted cells in comparison to the TMPRSS-ERG only expressing cells, which could explain the differences observed in growth in presence of androgens. Additional studies are needed e. g. to assess the tumorigenicity of these cells. Conclusion: The established sequential in vitro model of prostate epithelial transformation can serve as a useful model in future research of early prostate cancer events. Citation Format: Hanna E. Rauhala, Konsta Kukkonen, Tapio Visakorpi. Novel sequential in vitro model to study prostate epithelial transformation abstract. In: Proceedings of the AACR Special Conference in Cancer Research: Innovations in Prostate Cancer Research and Treatment; 2026 Jan 20-22; Philadelphia PA. Philadelphia (PA): AACR; Cancer Res 2026;86 (2Suppl): Abstract nr B063.
Rauhala et al. (2026) studied this question.