Association of the epithelial-to-mesenchymal transition phenotype with responsiveness to the p21-activated kinase inhibitor, PF-3758309, in colon cancer models

Abstract

The p21-activated kinase (PAK) family of serine/threonine kinases, which are overexpressed in several cancer types, are critical mediators of cell survival, motility, mitosis, transcription, and translation. In the study presented here, we utilized a panel of colorectal cancer (CRC) cell lines to identify potential biomarkers of sensitivity or resistance that may be used to individualize therapy to the PAK inhibitor PF-03758309. We observed a wide range of proliferative responses in the CRC cell lines exposed to PF-03758309, this response was recapitulated in other phenotypic assays such as anchorage-independent growth, three-dimensional (3D) tumor spheroid formation, and migration. Interestingly, we observed that cells most sensitive to PF-03758309 exhibited up-regulation of genes associated with a mesenchymal phenotype (CALD1, VIM, ZEB1) and cells more resistant had an up-regulation of genes associated with an epithelial phenotype (CLDN2, CDH1, CLDN3, CDH17) allowing us to derive an epithelial-to-mesenchymal transition (EMT) gene signature for this agent. We assessed the functional role of EMT-associated genes in mediating responsiveness to PF-3758309, by targeting known genes and transcriptional regulators of EMT. We observed that suppression of genes associated with the mesenchymal phenotype conferred resistance to PF-3758309, in vitro and in vivo. These results indicate that PAK inhibition is associated with a unique response phenotype in CRC and that further studies should be conducted to facilitate both patient selection and rational combination strategies with these agents.

Tumor P-Glycoprotein Correlates with Efficacy of PF-3758309 in in vitro and in vivo Models of Colorectal Cancer

“…Through studies conducted prior to and in conjunction with this work (see companion article (Pitts et al, 2013), we identified additional markers associated with sensitivity to PF-309 in models of CRC and found that cells and tumors with an epithelial genotype (high CLDN2 , CDH1 , RAB25 , CLDN3 , CDH17 ) are more resistant and those with a mesenchymal genotype (high CALD1 , VIM , ANK2 , ZEB1 ) display sensitivity. Indeed, GEO tumors possess an epithelial genotype, requiring a higher concentration of PF-309 to achieve effect, while the SW480 tumors possess a mesenchymal genotype, requiring a lower concentration of PF-309 to achieve effect (Figures 5, 7, and 8).…”

“…We evaluated the sensitivity of CRC cell lines to PF-309 treatment in a panel of 27 lines. Cells were exposed to PF-309 at a range of concentrations for 72 h and cell content measured by SRB assay (see Pitts et al, 2013). From the 27 lines tested, we selected lines that had approximately 100-fold difference in IC50 and examined the ABCB1 and ABCG2 gene expression profiles (Figure 1).…”

“…For determining the epithelial to mesenchymal signature, epithelial (E; CLDN2 , CDH1 , RAB25 , CLDN3 , and CDH17 ), and mesenchymal (M) genes ( CALD1 , VIM , ANK2 , and ZEB1 ) derived from the PAK inhibitor (see companion article: (Pitts et al, 2013). The expressions of these genes were ranked normalized within cell line and then Z-normalization was performed on this rank-based matrix.…”

P21-Activated Kinase 4: A Druggable Target in the Elusive Oncogenic Kras Pathway?

“…A major drawback of this Pfizer drug is that it was a substrate of the multidrug transporter P-glycoprotein, which contributed to its low bioavailability in humans [12]. In addition, studies demonstrated that PF-03798309 was ineffective against mesenchymal tumor cell lines, implying that this agent has applicability against epithelial tumors only [13]. This certainly dampens the clinical impact of this drug, as most oncogenic KRAS-driven tumors show high plasticity and motility that mirror mesenchymal characteristics.…”

Targeting Cdc42 in cancer

“…For example, in xenograft studies, the administration of a PLD1 small-molecule inhibitor was shown to prevent in situ vascularization and tumor growth, phenocopying the lack of tumor growth observed in Pld −/− mice [77]. Similarly, the administration of PAK kinases small-molecule inhibitors in a variety of cancer models has also been shown to suppress tumor growth and oncogenic signaling [64, 65, 95, 97]. Finally, an ACK1 small-molecule inhibitor has been shown to suppress ACK1 activation and abrogate transcriptional activity of the androgen receptor in prostate cancer cells [78].…”