Abstract
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is considered as a promising anti-cancer therapeutic. However, many cancers have been found to be or to become inherently resistant to TRAIL. A combination of epigenetic modifiers, such as histone deacetylase inhibitors (HDACi’s), with TRAIL was effective to overcome TRAIL resistance in some cancers. Broad spectrum HDACi’s, however, show considerable toxicity constraining clinical use. Since overexpression of class I histone deacetylase (HDAC) has been found in colon tumors relative to normal mucosa, we have focused on small spectrum HDACi’s. We have now tested agonistic receptor-specific TRAIL variants rhTRAIL 4C7 and DHER in combination with several class I specific HDACi’s on TRAIL-resistant colon cancer cells DLD-1 and WiDr. Our data show that TRAIL-mediated apoptosis is largely improved in WiDr cells by pre-incubation with Entinostat-a HDAC1, 2, and 3 inhibitor- and in DLD-1 cells by RGFP966-a HDAC3-specific inhibitor- or PCI34051-a HDAC8-specific inhibitor. We are the first to report that using RGFP966 or PCI34051 in combination with rhTRAIL 4C7 or DHER represents an effective cancer therapy. The intricate relation of HDACs and TRAIL-induced apoptosis was confirmed in cells by knockdown of HDAC1, 2, or 3 gene expression, which showed more early apoptotic cells upon adding rhTRAIL 4C7 or DHER. We observed that RGFP966 and PCI34051 increased DR4 expression after incubation on DLD-1 cells, while RGFP966 induced more DR5 expression on WiDr cells, indicating a different role for DR4 or DR5 in these combinations. At last, we show that combined treatment of RGFP966 with TRAIL variants (rhTRAIL 4C7/DHER) increases apoptosis on 3D tumor spheroid models.
Highlights
Cancer occurs when cells divide in an uncontrolled fashion and escape the strict mechanisms of cell death
Our results show that RGFP966 improves Tumor necrosis factor-related apoptosis inducing ligand (TRAIL)-induced apoptosis via both DR4 and DR5 receptors and its antitumor effect in combination with rhTRAIL 4C7 or DHER is close to the effect by SAHA combined with TRAIL variants
To study the role of the respective histone deacetylase (HDAC) we performed cell viability assays testing the sensitivities of DLD-1 and WiDr cells to various histone deacetylase inhibitors (HDACi’s)
Summary
Cancer occurs when cells divide in an uncontrolled fashion and escape the strict mechanisms of cell death. A number of HDAC-selective inhibitors are under investigation for use in oncology, including (1) RGFP966, a HDAC3-specific inhibitor, decreases the growth of prostate cancer models [29]; (2) PCI34051, a HDAC8-specific inhibitor, induces apoptosis of T-cell malignancies [30]; and (3) Tubacin, a HDAC6-specific inhibitor, suppresses proliferation of acute lymphoblastic leukemia cells [31]. We chose RGFP966 or PCI34051 to be used in combination with apoptosis-inducing ligands in colon cancer Both rhTRAIL 4C7 and DHER have shown superior apoptosis-inducing effects in colon tumor cells compared to rhTRAIL, but a number of cell lines to a lesser (DLD-1) or higher (WiDr) extend show resistance [32,33]. WiDr using different HDACi’s with partially overlapping specificities We combined these inhibitors with two TRAIL variants, DR4-specific rhTRAIL 4C7 and DR5-specific rhTRAIL DHER, to further unravel the antitumor effects. In trying to understand mechanism, we monitored the surface expression of DR4 and DR5 and we analyzed cell cycle changes in HDACi’s-treated cells
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