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Pathway to Cancer Therapy
Don-Hong Wang
其他書名
From the Analysis of Viral Protein Kinase Pathway to the Development of Novel High-throughput Screening Assay
出版University of California, Davis, 2010
ISBN11245094969781124509495
URLhttp://books.google.com.hk/books?id=_XKuAQAACAAJ&hl=&source=gbs_api
註釋The aims of this doctoral research were to understand the mechanism of oncogenic viruses in cancer development and to identify chemical compounds that have potential therapeutic values. Cancer is a complex disease caused by a disruption in cellular homeostasis, leading to unregulated cell growth and differentiation of cancer stem cells which in turn invade other organ tissues through the cardiovascular or lymphatic systems. This unregulated cell growth can occur in cells that are infected by certain viruses that take over the host's cellular machinery. Kaposi's sarcoma-associated herpesvirus (KSHV), also identified as human herpesvirus 8, is an oncogenic virus that is associated with primary effusion lymphoma, B-cell lymphomas, multicentric Castleman's disease, and AIDS-associated malignancies. Open reading frame 36 (Orf36), designated as viral protein kinase (vPK), encodes a serine/threonine kinase that is required for optimal viral replication. My studies have demonstrated the association of vPK with three different cellular proteins that play key roles in viral transcription: KAP-1, RNA polymerase II (RNAPII), and SR proteins. I found vPK associates with and phosphorylates KAP-1 which subsequently releases the viral DNA from the heterochromatin repressors. vPK then modulates the activity of RNAP II to initiate the transcription mechanism within the viral genome. Finally, vPK recruits SR proteins to the transcription site for the regulation of viral gene expression, pre-mRNA splicing and mRNA export. These findings strongly suggest that vPK is an important regulator for KSHV replication, contributing to uncontrolled cell growth. In addition to the basic study of cellular mechanisms in cancer cells as described above, I participated in designing the methodology of targeted drug discovery. This novel multiplix assay was developed with both one-bead one-compound library and phage display library to uncover novel chemical compounds that can target biological proteins. Using this novel assay, we identified ninety-one different chemical compounds associated with the proteome of cancer cells. Analysis of these ninety-one small molecule compounds revealed that these compounds associate with several high profile oncogenic targets. Notably, I found that compound C15-1 targets lysine (K)-specific demethyase 2B (KDM2B), a lysine-specific demethylase protein with oncogenic properties. Through immunoprecipitation, compound C15-1 was confirmed to associate with full length KDM2B. BLASTx analysis revealed that this compound interacts with the JmjC catalytic domain of KDM2B to modulate the demethylation activity. Western blot analysis showed that this compound decreases the demethylation activity which results in an increase in the methyl ligands on histone H3 dimethylated lysine K36 (H3K36M̳e̳2). From these data, we hypothesize that compound C15-1 inhibits the demethylation activity of KDM2B. Furthermore, the biological data of compound C15-1 demonstrate the effectiveness of this high-throughput multiplix assay in identifying novel biological and chemical entities for potential therapeutic interventions.