Targeting the Cohesive Cluster Phenotype in Chordoma via β1 Integrin Increases Ionizing Radiation Efficacy

Abstract

Chordoma is a rare, radiation-resistant, skull-base and spinal tumor with high local recurrence containing mixed cell-adhesion phenotypes. We characterized DNA damage response (DDR) signaling (γH2AX, pKAP1, pATM) and survival response to ionizing radiation (IR) in human chordoma samples (42 resections, 23 patients) to test if blocking cell adhesion sensitizes U-CH1 tumor cells to IR. U-CH1 cells expressed brachyury, YAP, and laminin adhesion receptors (CD49c, CD49f, CD44), and approximately 15% to 20% of U-CH1 cells featured an α6 integrin-dependent (CD49f) cohesive cluster phenotype, which confers therapeutic resistance and aids metastasis. DDR to IR in U-CH1 cells was compared to normal prostate epithelial (PrEC) and tumor cells (DU145). Flow cytometry showed a dose- and time-dependent increase in γH2AX and pKAP1 expression in all cell lines. However, nearly 50% of U-CH1 cells exhibited nonresponsive phenotype to IR (measured by γH2AX and pKAP1) independent of cell cycle status. Immunofluorescence microscopy verified that only 15% of U-CH1 clustered cells were γH2AX or pKAP1 positive (versus 80% of nonclustered cells) 2 hours following 2-Gy IR. Conversely, both tumor cell lines were uniformly defective in pATM response. HYD1, a synthetic ECM ligand, inhibited DDR through an unresolved γH2AX response. β1 integrin-blocking antibody (AIIB2) decreased cell survival 50% itself and approximately doubled the IR-induced cell kill at all IR doses observed at 2 and 4 weeks posttreatment. These results suggest that a heterogeneity of DDR to IR exists within a chordoma population. Blocking integrin function alone and/or as an adjuvant to IR may eradicate chordomas containing the cohesive cluster phenotype.

Identification of therapeutic targets in chordoma through comprehensive genomic and transcriptomic analyses

“…These processes include ILK (integrin-linked kinase), integrin, and GTPase signaling, which are associated with the PI3K/Akt/mTor pathways; phospholipase C and calcium signaling pathways, which are associated with Fgf signaling; regulation of gene expression through the nuclear receptors FXR (farnesoid X receptor), TR (thyroid hormone receptor), LXR (liver X receptor), and RXR (retinoid X receptor); and what appear to be tumor microenvironment processes as defined by epithelial adherens junction signaling and tight junction signaling. Although ILK signaling has not been specifically reported in chordomas, obstructing β1 integrin function was suggested to be therapeutically relevant following radiation in chordomas (Harryman et al 2017). Separately, the importance of the microenvironment of chordomas has been previously demonstrated through in vitro analyses, whereby hypoxia combined with exposure to CCN2, a connective tissue growth factor, up-regulated notochord progenitor markers (Patel et al 2014).…”

Gene Editing of α6 Integrin Inhibits Muscle Invasive Networks and Increases Cell–Cell Biophysical Properties in Prostate Cancer

“…The existence of the cohesive cluster phenotype in prostate cancer specimens has been observed previously (12). Although the cohesive cluster phenotype can significantly slow tumor progression (43), it is dependent upon b1 integrin function and can provide resistance to chemotherapeutic and radiotherapeutic approaches by a variety of mechanisms (45,46). It remains to be determined if the cohesive cluster phenotype that blocks invasion here may result in tumors that are more resistant to chemotherapeutic agents.…”

Novel Regulation of Integrin Trafficking by Rab11-FIP5 in Aggressive Prostate Cancer

“…In this cancer type, invasion and metastasis is significantly dependent on α6 integrin. α6 integrin is prominent in prostate cancer invasion, as well as in bone and visceral metastasis as a cohesive cluster phenotype (48, 49) which is targetable. We identified various intracellular trafficking regulatory proteins that are significantly associated with the ratio of intracellular to cell surface localization of α6 integrin.…”