The disruption of epigenetic pathways are key driving mechanisms that contribute to myriad human diseases. Recentadvances in chromatin biology and high-throughput genetic sequencing have discovered that such disruptions underlie asubstantial number of cancers. While the affected epigenetic pathways that contribute to cancer pathophysiology are quitediverse, a common theme among them is that aberrant regulation of epigenetic enzymes can lead to dysregulated expressionof key disease driver genes. For example, aberrant DNA methylation, histone H3 methylation and/or histone H3deacetylation can repress genes, and lead to a more deleterious disease phenotype. Indeed, broad-acting pan-epigeneticinhibitors have shown initial promise clinically by grossly disrupting disease signaling pathways by altering the expressionprofile of key genes. However, a fundamental issue related to the mechanism of these traditional epigenetic inhibitors centerson their potential to affect thousands of genes simultaneously (both on- and off-target). Thus, while treatment with histonedeacetylase (HDAC) inhibitors causes the desired effect of activation of specific target genes, this activation also comeswith off-target activation of potentially hundreds to thousands of additional genes. Recently, technological advancementson the CRISPR/Cas9 system have been developed to allow for induction of site-specific chromatin modifications that canmodulate gene expression. Our technological advancements build on these approaches and allow for the possibility ofdeveloping targeted, epigenetically based gene therapeutics with real potential for clinical translation. Here, we outline an approach that leverages a site-specific dCas9-FKBP fusion protein, coupled with a syntheticbifunctional chemical epigenetic modifier (CEM). The CEM consists of three modular components: (1) FK506 (which bindsFKBP); (2) a short inert chemical polyethylene glycol (PEG) linker; and (3) a chemical entity that interacts with hostepigenetic machinery. Ultimately, the dCas9-FKBP CEM has the ability to target any locus in the genome, and "activate"epigenetic activity to modulate gene expression. We propose to implement this strategy to a clinically relevant target: TP53.Our proposed technology seeks to target endogenous histone acetylation enzymes to the TP53 locus to reverse its epigeneticrepression, and thus increase the sensitivity of tumor cells to chemotherapeutic agents. As a first step, we propose to evaluateour technology in preclinical models of colorectal cancer. Colorectal cancer is the third most commonly diagnosed cancerand third cancer most common cause of cancer-related death among both men and women. Additionally, colorectal canceris known to be driven by both mutated and epigenetically silenced TP53, and there are available preclinical model systemsthat recapitulate was is observed clinically. Long-term our novel platform could represent an innovative and curativetreatment for many epigenetically driven human cancers. NARRATIVE
While the pathways underlying cancer is quite diverse, a common theme among them is the inappropriate
repression of tumor suppressor genes or activation of oncogenes (i.e., abnormal epigenetic regulation).
Technologies that can specifically restore the normal state of these otherwise normal genes would provide a
new way to treat cancer. Our proposed technology will make a novel contribution to cancer research by offering
a new personalized epigenome-based approach therapeutic that treats cancer through a novel mechanism at the
root of the gene mis-regulation. men ; men's ; Methylation ; Biological Models ; Biologic Models ; Model System ; Mus ; Mice ; Mice Mammals ; Murine ; Parents ; Polyethylene Glycols ; Macrogols ; Polyethylene Oxide ; Polyethyleneoxide ; Polyoxyethylenes ; Repression ; Role ; social role ; Signal Pathway ; Stains ; Staining method ; Technology ; Up-Regulation ; Upregulation ; Woman ; Fluorescein ; Generations ; Tumor Suppressor Genes ; Anti-Oncogenes ; Antioncogenes ; Cancer Suppressor Genes ; Emerogenes ; Onco-Suppressor Genes ; Oncogenes-Tumor Suppressors ; Recessive Oncogenes ; Tumor Suppressing Genes ; oncosuppressor gene ; TP53 gene ; Antioncogene Protein p53 ; Cellular Tumor Antigen P53 ; Oncoprotein p53 ; P53 ; Phosphoprotein P53 ; Phosphoprotein pp53 ; Protein TP53 ; TP53 ; TRP53 ; Tumor Protein p53 ; Tumor Protein p53 Gene ; p53 Antigen ; p53 Genes ; p53 Tumor Suppressor ; protein p53 ; Tacrolimus Binding Proteins ; FK-506-Binding Protein ; FK506 Binding Proteins ; FKBP ; FKBP Rotamase ; Guide RNA ; gRNA ; Fluorescein-5-isothiocyanate ; 5-Isothiocyanatofluorescein ; FITC ; Mediating ; Apoptosis ; Apoptosis Pathway ; Programmed Cell Death ; Chimeric Proteins ; Chimera Protein ; Fusion Protein ; base ; improved ; Site ; Clinical ; Isothiocyanates ; Chemicals ; Plant Roots ; root ; CDKN1A gene ; CDK-Interacting Protein 1 ; CDKN1 ; CDKN1A ; CIP1 ; Cyclin-Dependent Kinase Inhibitor 1A ; WAF1 ; Wildtype p53-Activated Fragment 1 ; p21 gene ; p21 protein ; Functional disorder ; Dysfunction ; Physiopathology ; pathophysiology ; Therapeutic ; Genetic ; Pansy ; Violet ; Viola ; Hour ; System ; Heterograft ; Heterologous Transplantation ; Xenograft ; Xenotransplantation ; xeno-transplant ; xeno-transplantation ; Xenograft procedure ; Tumor Cell ; neoplastic cell ; novel ; Colon Cancer ; Colonic Carcinoma ; cancer in the colon ; Colon Carcinoma ; FK 506 ; FK506 ; chemotherapeutic agent ; BAX ; BCL2-Associated X Protein Gene ; BCL2L4 ; BAX gene ; Regulation ; Cancer Treatment ; Malignant Neoplasm Therapy ; Malignant Neoplasm Treatment ; anti-cancer therapy ; anticancer therapy ; cancer-directed therapy ; cancer therapy ; cancer diagnosis ; Cell Cycle Arrest ; chromatin modification ; Histone Acetylation ; Molecular Interaction ; Binding ; HCT 116 Cells ; HCT- 116 ; HCT-116 ; HCT116 ; HCT116 Cells ; ANX5 ; ANX5 Gene ; ANXA5 ; Anchorin CII ; Anchorin CII Gene ; Annexin A5 Gene ; Annexin A5 Protein ; Annexin V ; CBP-I ; Calphobindin I ; ENX2 ; ENX2 Gene ; Endonexin II ; Endonexin II Gene ; Lipocortin V Gene ; Lipocortin-V ; PAP-I ; PP4 Gene ; Placental Anticoagulant Protein I ; Placental Protein 4 ; Thromboplastin Inhibitor ; VAC-Alpha ; Vascular Anticoagulant-Alpha ; annexin A5 ; ANXA5 gene ; Aberrant DNA Methylation ; Deacetylation ; HDAC Agent ; HDAC inhibitor ; Histone deacetylase inhibition ; Histone Deacetylase Inhibitor ; Mutate ; Preclinical Models ; Pre-Clinical Model ; Quantitative RTPCR ; qRTPCR ; Quantitative Reverse Transcriptase PCR ; in vivo ; mRNA Expression ; Cancer Etiology ; Cancer Cause ; Epigenetic Process ; Epigenetic ; Epigenetic Change ; Epigenetic Mechanism ; Oncogene Activation ; Colorectal Cancer ; Colo-rectal Cancer ; Pathway interactions ; pathway ; vector ; disease phenotype ; anticancer research ; anti-cancer research ; cancer research ; Coupled ; innovation ; innovate ; innovative ; therapeutic gene ; gene therapeutics ; gene-based therapeutic ; gene-based therapeutics ; genes therapeutic ; genes therapeutics ; clinically relevant ; clinical relevance ; human disease ; tumor ; SW480 ; SW-480 ; epigenome ; epigenetic regulation ; CRISPR/Cas technology ; CRISPR method ; CRISPR methodology ; CRISPR technique ; CRISPR technology ; CRISPR-CAS-9 ; CRISPR-based method ; CRISPR-based technique ; CRISPR-based technology ; CRISPR-based tool ; CRISPR/Cas method ; CRISPR/Cas9 ; CRISPR/Cas9 technology ; Cas nuclease technology ; Clustered Regularly Interspaced Short Palindromic Repeats method ; Clustered Regularly Interspaced Short Palindromic Repeats methodology ; Clustered Regularly Interspaced Short Palindromic Repeats technique ; Clustered Regularly Interspaced Short Palindromic Repeats technology ; curative treatments ; curative intervention ; curative therapeutic ; curative therapy ; clinical translation ; recruit ; genomic locus ; gene locus ; genetic locus ; epigenetic silencing ; epigenetic gene silencing ; Affect ; inhibitor/antagonist ; inhibitor ; Automobile Driving ; driving ; Biology ; Western Blotting ; Western Immunoblotting ; protein blotting ; Malignant Neoplasms ; Cancers ; Malignant Tumor ; malignancy ; neoplasm/cancer ; Cell Cycle ; Cell Division Cycle ; Cells ; Cell Body ; Chromatin ; Cessation of life ; Death ; Disease ; Disorder ; Engineering ; Enzymes ; Enzyme Gene ; Flow Cytometry ; Flow Cytofluorometries ; Flow Cytofluorometry ; Flow Microfluorimetry ; Flow Microfluorometry ; flow cytophotometry ; Gene Activation ; Gene Expression ; Patient Care ; Patient Care Delivery ; Genes ; Genome ; Histone Deacetylase ; HDAC ; HDAC Proteins ; Histone H3 ; Human ; Modern Man ; In Vitro ; Lead ; Pb element ; heavy metal Pb ; heavy metal lead ;
