Abstract 5070: CD44 alternative splice variants are associated with prostate cancer cell identity and migration

Abstract

Abstract Patient tumors can demonstrate tremendous cell-to-cell genetic heterogeneity while cultured cancer cell lines are often more homogeneous. This fundamental difference can undermine in vitro findings and their application to the more complex in vivo tumor environment. However, it should not be assumed that cultured cells are homogeneous even in well established and widely used cell lines. In my study of CD44 alternative splicing in the prostate cancer cell line PC3, I encountered varying CD44 alternative splicing expression profiles dependent on the origin of the PC3 cell line. CD44 is a cell surface glycoprotein that binds to components of the extracellular matrix, including hyaluronan, and is primarily involved in cell-cell and cell-matrix interactions. CD44 contains 10 variable exons that when combined in particular combinations significantly alter/guide CD44 downstream activities. In the context of cancer biology, alternative splicing of CD44 can control the epithelial to mesenchymal transition (EMT) of cancer cells and is associated with metastasis in many types cancers such as breast and prostate. To determine if cellular heterogeneity was the source of variable CD44 alternative splicing profiles, I isolated PC3 single cell clones. From these clones I identified two cell populations based on colony morphology: a compact colony subset with cells of a epithelial phenotype (PC3-EL), and a diffuse colony subset with cells of a mesenchymal phenotype (PC3-ML). I then performed reverse transcription-PCR (rt-PCR) on cDNA generated from these two populations with primers that specifically amplify the CD44 variable region to generate a fingerprint of CD44 alternative splicing. I found that PC3 cells with an epithelial phenotype express multiple CD44 variant exons (CD44v), including the epithelium-associated CD44v1,v8-10 variant (CD44E). However, phenotypically mesenchymal PC3 cells show a complete loss of CD44 splice variants and primarily express standard CD44v1 (CD44S). These differences in CD44 mRNA alternative spicing are manifested at the protein level as phenotypically epithelial PC3 cells express predominantly high molecular weight CD44 variants. I then monitored the association of CD44v expression on cell migration and determined that PC3 cells that express CD44E are non-motile, while PC3 cells that predominantly express CD44S are highly motile. Acquisition of CD44S over CD44E can signal EMT, an early and critical step in cancer metastasis. These data indicate that the CD44 alternative splice fingerprint may provide a predictive biomarker for EMT and the acquisition of other early pro-metastatic features in prostate cancer cells. Citation Format: David J. Bond, John D. Lewis. CD44 alternative splice variants are associated with prostate cancer cell identity and migration. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5070.