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  • Itch also known as astrophin interacting protein AIP is

    2019-08-13

    Itch, also known as astrophin-1 interacting protein-4 (AIP4), is a HECT domain-E3 ligase. It was discovered in 1998 in mouse coat color genetic studies which displayed the disruption caused by the Itch gene inducing a lethal autoimmune inflammatory condition. The substrates of Itch are transmembrane proteins, signaling molecules and transcription factors, for example, CXCR4, c-FLIP and p63. For past years, Itch has been thought to play an important role in the regulation of programmed cell death pathway which is associated with tumor development [9], [10], [11]. However, although many studies have studied the relationship between Itch and tumorigenesis, the mechanism has not yet been confirmed. To date, a few compounds (6-8) have been reported to be HECT domain-E3 ligase inhibitors (Fig. 2) [12], [13], [14]. Interestingly, among these reported compounds, only compound 8 was identified as effective Itch inhibitor in a high throughput screening using Itch auto-ubiquitination as an indicator of Itch activity [14]. Compound 6 on the other hand, was found to targeting smurf2, Nedd4 and WWP1, which are all associated with the HECT domain-E3 ligase [12]. Compound 7 has been shown to directly inhibit Nedd4-1 ubiquitination directly and to inhibit proliferation of human melanoma cells in vitro[13]. Review of the literature suggests that structures with a rigid core may be necessary in this field [10], [11], [12] and that designed structures should contain two parts: a rigid core structure and a side chain at least four carbon atoms in length. As a result, we decided to use naphthoquinone bearing one chlorine blasticidin as the core structure. Benzylamine, used as the linker generates a side chain of adequate length and the amide at the para position can connect with various heterocycles (compound 9) to generate a series of potential E3 ligase inhibitors. A number of designed target compounds (10a-10q) have been synthesized and evaluated for their antiproliferative effects (Fig. 3).
    Results and discussion
    Conclusion In this paper, we describe the synthesis of several naphthoquinone-based compounds (10a-10q). Among these synthetic compounds, compound 10e acted as an Itch inhibitor, exhibiting significant inhibitory activity in several cancer cell lines and with no antiproliferative activity in normal cell lines. In addition, this compound has demonstrated antitumor efficacy in human RPMI-8226 multiple myeloma xenograft model. Compound 10e also causes a decrease of Itch levels in the RPMI-8226 multiple myeloma cell line. Consequently, compound 10e can serve as a lead compound in an investigation of the connection between Itch or other E3 ligases and may provide a new strategy for understanding the HECT domain-E3 ligase inhibitors.
    Experimental section
    Acknowledgment This research were supported by the Ministry of Science and Technology, Taiwan (grant no. ).
    Terrestrial plant seedlings adopt a developmental strategy termed skotomorphogenesis when grown in the subterranean dark environment. The seedlings display long hypocotyls, curved apical hooks, and closed cotyledons. This characterized morphogenesis helps the seedlings grow rapidly toward the soil surface and protects the fragile meristem against mechanical injuries , , . On their emergence above the soil surface, light induces the seedlings to undergo a dramatic developmental transition to photomorphogenesis, which involves inhibition of hypocotyl elongation, unfolding of the apical hook, and cotyledon opening , , . COP1 was originally identified through genetic screens in , whose recessive mutants display constitutive photomorphogenesis in darkness . The COP1 protein contains three protein–protein interaction domains: a RING-finger region at its N terminus, a central coiled-coil domain, and seven WD40 repeats at its C terminus . A major breakthrough in addressing COP1’s molecular actions was the discovery that COP1 is an E3 ubiquitin ligase that mediates the proteolysis of various photomorphogenesis positive transcription factors , . Extensive studies have shown that COP1 acts as a functional E3 ligase in the free form or COP1–SUPPRESSOR of phyA-105 (SPA)–CULLIN (CUL) 4 complex. Within the complex, the core-component SPA proteins directly interact with COP1 and enhance the COP1 E3 ligase activity , . In the dark, COP1 directly interacts with the positive photomorphogenesis transcription factors, such as ELONGATED HYPOCOTYL 5 (HY5) and HY5 HOMOLOG (HYH), for protein ubiquitination and degradation via the 26S proteasome system , (). By contrast, COP1 has been shown to markedly stabilize the photomorphogenesis-suppressing transcription factors, such as PHYTOCHROME-INTERACTING FACTORs (PIFs) and ETHYLENE-INSENSITIVE3/EIN3-Like 1 (EIN3/EIL1) , , . Thus, COP1 plays a central role in the repression of photomorphogenesis. However, the mechanism of how COP1 stabilizes the negative transcription factors is largely unknown.