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Project description:BRD4, a member of the BET family of histone readers, binds to acetylated lysine of histone H3 and promotes assembly of super-enhancer complexes that drive expression of key oncogenes in acute myeloid leukemia (AML) and other cancers. ARV-825 is a proteolysis-targeting chimera (PROTAC) that targets BRD4 for CRBN-mediated ubiquitination and degradation. We treated the AML cell line OCI-AML3, as well as primary tumor cells from a case of AML, with ARV-825 in vitro. At low concentrations, ARV-825 caused profound and sustained reduction in BRD4 protein levels. This caused reduction in transcription of key genes including MYC, anti-apoptotic BCL2 and BCLXL, PIM1, and CD44. Downstream effects included loss of CXCR4 surface expression and mitochondrial respiration; increased reactive oxygen species; toxicity to AML cells in vitro; and efficacy vs. OCI-AML3 cells in a mouse model of AML.
2019-10-01 | GSE101380 | GEO
Project description:Effect of PROTAC-mediated BRD4 degradation in acute myeloid leukemia
| PRJNA382703 | ENA
Project description:BackgroundProteolysis targeting chimeras (PROTACs) are heterobifunctional small molecules that utilize the ubiquitin-proteasome system to selectively degrade target proteins. This innovative technology has shown remarkable efficacy and specificity in degrading oncogenic proteins and has progressed through various stages of preclinical and clinical development for hematologic malignancies, including adult acute myeloid leukemia (AML). However, the application of PROTACs in pediatric AML remains largely unexplored.MethodsIn this study, we show the potent effect of GSPT1 degradation against AML cells induced by either a GSPT1-selective cereblon modulator CC-90009 or by an off-target effect caused by a CDK6-PROTAC named GU3341.ResultsBoth in vitro and ex vivo experiments revealed that GSPT1 degradation significantly inhibited tumor growth, induced cell cycle arrest, and triggered apoptosis in two pediatric AML subtypes characterized by RUNX1::RUNX1T1 and FUS::ERG fusion genes. Furthermore, the degradation of GSPT1 impaired the expression of RUNX1::RUNX1T1 and its cooperating transcription factors RUNX1 and ERG. Similarly, GSPT1 degradation also reduced FUS::ERG fusion transcript levels in AML cells harboring the translocation t(16;24)(p11:q22).ConclusionsThese findings suggest a new role of GSPT1 in regulating leukemic transcriptional networks and open a new therapeutic strategy to target leukemic fusion genes in pediatric AML patients.
| S-EPMC11763475 | biostudies-literature
Project description:AURKA is a potential kinase target in various malignancies. The kinase-independent oncogenic functions partially disclose the inadequate efficacy of the kinase inhibitor in a Phase III clinical trial. Simultaneously targeting the catalytic and noncatalytic functions of AURKA may be a feasible approach. Here, a set of AURKA proteolysis targeting chimeras (PROTACs) are developed. The CRBN-based dAurA383 preferentially degrades the highly abundant mitotic AURKA, while cIAP-based dAurA450 degrades the lowly abundant interphase AURKA in acute myeloid leukemia (AML) cells. The proteomic and transcriptomic analyses indicate that dAurA383 triggers the "mitotic cell cycle" and "stem cell" processes, while dAurA450 inhibits the "MYC/E2F targets" and "stem cell" processes. dAurA383 and dAurA450 are combined as a PROTAC cocktail. The cocktail effectively degrades AURKA, relieves the hook effect, and synergistically inhibits AML stem cells. Furthermore, the PROTAC cocktail induces AML regression in a xenograft mouse model and primary patient blasts. These findings establish the PROTAC cocktail as a promising spatial-temporal drug administration strategy to sequentially eliminate the multifaceted functions of oncoproteins, relieve the hook effect, and prevent cancer stem cell-mediated drug resistance.
| S-EPMC9353462 | biostudies-literature
Project description:Although the use of ATP-competitive tyrosine kinase inhibitors of oncoprotein BCR-ABL1 has enabled durable responses in patients with chronic myeloid leukemia (CML), issues of drug resistance and residual leukemic stem cells remain. To test whether the degradation of BCR-ABL1 kinase could offer improved response, we developed a series of proteolysis-targeting chimera (PROTAC) that allosterically target BCR-ABL1 protein and recruit the E3 ligase Von Hippel-Lindau, resulting in ubiquitination and subsequent degradation of the oncogenic fusion protein. In both human CML K562 cells and murine Ba/F3 cells expressing BCR-ABL1, lead compound GMB-475 induced rapid proteasomal degradation and inhibition of downstream biomarkers, such as STAT5, and showed increased sensitivity compared with diastereomeric controls lacking degradation activity. Notably, GMB-475 inhibited the proliferation of certain clinically relevant BCR-ABL1 kinase domain point mutants and further sensitized Ba/F3 BCR-ABL1 cells to inhibition by imatinib, while demonstrating no toxicity toward Ba/F3 parental cells. Reverse phase protein array analysis suggested additional differences in levels of phosphorylated SHP2, GAB2, and SHC associated with BCR-ABL1 degradation. Importantly, GMB-475 reduced viability and increased apoptosis in primary CML CD34+ cells, with no effect on healthy CD34+ cells at identical concentrations. GMB-475 degraded BCR-ABL1 and reduced cell viability in primary CML stem cells. Together, these findings suggest that combined BCR-ABL1 kinase inhibition and protein degradation may represent a strategy to address BCR-ABL1-dependent drug resistance, and warrant further investigation into the eradication of persistent leukemic stem cells, which rely on neither the presence nor the activity of the BCR-ABL1 protein for survival. SIGNIFICANCE: Small-molecule-induced degradation of BCR-ABL1 in CML provides an advantage over inhibition and provides insights into CML stem cell biology. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/79/18/4744/F1.large.jpg.
| S-EPMC6893872 | biostudies-literature
Project description:Acute myeloid leukemia (AML) is a highly cancerous and aggressive hematologic disease with elevated levels of drug resistance and relapse resulting in high mortality. Recently, bromodomains and extra-terminal (BET) protein inhibitors have been extensively researched in hematological tumors as potential anticancer agents. MZ1 is a novel BET inhibitor that mediates selective proteins degradation and suppression of tumor growth through proteolysis-targeting chimeras (PROTAC) technology. Accordingly, this study aimed to investigate the role and therapeutic potential of MZ1 in AML. In this study, we first identified that AML patients with high BRD4 expression had poor overall survival than those with low expression group. MZ1 inhibited AML cell growth and induced apoptosis and cycle arrest in vitro. MZ1 induced degradation of BRD4, BRD3 and BRD2 in AML cell strains. Additionally, MZ1 also initiated the cleavage of poly-ADP-ribose polymerase (PARP), which showed cytotoxic effects on NB4 (PML-RARa), K562 (BCR-ABL), Kasumi-1 (AML1-ETO), and MV4-11 (MLL-AF4) cell lines representing different molecular subtypes of AML. In AML mouse leukemia model, MZ1 significantly decreased leukemia cell growth and increased the mouse survival time. According to the RNA-sequencing analysis, MZ1 led to c-Myc and ANP32B genes significant downregulation in AML cell lines. Knockdown of ANP32B promoted AML cell apoptosis and inhibited cell growth. Overall, our data indicated that MZ1 had broad anti-cancer effects on AML cell lines with different molecular lesions, which might be exploited as a novel therapeutic strategy for AML patients.
| S-EPMC9543111 | biostudies-literature
Project description:Acute myeloid leukemia (AML) is often initiated by genetic alterations of machineries that regulate chromatin and transcription, thereby blocking cell differentiation. Such mechanisms may also render leukemia cells vulnerable to perturbations of transcriptional regulators, which includes small molecules targeting the coactivator protein BRD4. Numerous studies have validated BRD4 as a therapeutic target in diverse subtypes of AML; however, the vital function of BRD4 in this disease is only beginning to be understood. Here we discuss the recent progress in elucidating the transcriptional function of BRD4 in AML cells, with an emphasis on the desirable attributes, but also the inherent limitations, of targeting general coactivator proteins as cancer therapy.
| S-EPMC5550326 | biostudies-other
Project description:Targeting protein degradation with Proteolysis-Targeting Chimeras (PROTACs) is an area of great current interest in drug discovery. Nevertheless, although the high effectiveness of PROTACs against a wide variety of targets has been established, most degraders reported to date display limited intrinsic tissue selectivity and do not discriminate between cells of different types. Here, we describe a strategy for selective protein degradation in a specific cell type. We report the design and synthesis of a trastuzumab-PROTAC conjugate (Ab-PROTAC 3) in which E3 ligase-directed degrader activity is caged with an antibody linker which can be hydrolyzed following antibody-PROTAC internalization, releasing the active PROTAC and inducing catalytic protein degradation. We show that 3 selectively targets bromodomain-containing protein 4 (BRD4) for degradation only in HER2 positive breast cancer cell lines, while sparing HER2 negative cells. Using live cell confocal microscopy, we show internalization and lysosomal trafficking of the conjugate specifically in HER2 positive cells, leading to the release of active PROTAC in quantities sufficient to induce potent BRD4 degradation. These studies demonstrate proof-of-concept for tissue-specific BRD4 degradation, overcoming limitations of PROTAC selectivity, with significant potential for application to novel targets.
| S-EPMC7309268 | biostudies-literature
Project description:BRD4, a member of the BET family of histone readers, binds to acetylated lysine of histone H3 and promotes assembly of super-enhancer complexes that drive expression of key oncogenes in acute myeloid leukemia (AML) and other cancers. ARV-825 is a proteolysis-targeting chimera (PROTAC) that targets BRD4 for CRBN-mediated ubiquitination and degradation. BM-MSCs are an important element of the bone marrow microenvironment of AML. To understand how targeting BRD4 in BM-MSCs may contribute to the overall effect on AML of targeting BRD4, we treated BM-MSCs from two normal donors with ARV-825 in vitro. Treatment of BM-MSC monocultures with ARV-825 for 24 hr caused extensive changes in gene expression, highly uniform between triplicates. Although the cultures from the two normal donors showed different profiles, their changes with ARV-825 were highly similar. These changes implicated effects on oxidative stress, osteogenic differentiation, retinoid metabolism, F-actin polymerization, CXCL12, and proliferation.
2019-10-01 | GSE101449 | GEO
Project description:BackgroundT-cell acute lymphoblastic leukemia (T-ALL) is an aggressive disease with a high risk of induction failure and poor outcomes, with relapse due to drug resistance. Recent studies show that bromodomains and extra-terminal (BET) protein inhibitors are promising anti-cancer agents. ARV-825, comprising a BET inhibitor conjugated with cereblon ligand, was recently developed to attenuate the growth of multiple tumors in vitro and in vivo. However, the functional and molecular mechanisms of ARV-825 in T-ALL remain unclear. This study aimed to investigate the therapeutic efficacy and potential mechanism of ARV-825 in T-ALL.MethodsExpression of the BRD4 were determined in pediatric T-ALL samples and differential gene expression after ARV-825 treatment was explored by RNA-seq and quantitative reverse transcription-polymerase chain reaction. T-ALL cell viability was measured by CCK8 assay after ARV-825 administration. Cell cycle was analyzed by propidium iodide (PI) staining and apoptosis was assessed by Annexin V/PI staining. BRD4, BRD3 and BRD2 proteins were detected by western blot in cells treated with ARV-825. The effect of ARV-825 on T-ALL cells was analyzed in vivo. The functional and molecular pathways involved in ARV-825 treatment of T-ALL were verified by western blot and chromatin immunoprecipitation (ChIP).ResultsBRD4 expression was higher in pediatric T-ALL samples compared with T-cells from healthy donors. High BRD4 expression indicated a poor outcome. ARV-825 suppressed cell proliferation in vitro by arresting the cell cycle and inducing apoptosis, with elevated poly-ADP ribose polymerase and cleaved caspase 3. BRD4, BRD3, and BRD2 were degraded in line with reduced cereblon expression in T-ALL cells. ARV-825 had a lower IC50 in T-ALL cells compared with JQ1, dBET1 and OTX015. ARV-825 perturbed the H3K27Ac-Myc pathway and reduced c-Myc protein levels in T-ALL cells according to RNA-seq and ChIP. In the T-ALL xenograft model, ARV-825 significantly reduced tumor growth and led to the dysregulation of Ki67 and cleaved caspase 3. Moreover, ARV-825 inhibited cell proliferation by depleting BET and c-Myc proteins in vitro and in vivo.ConclusionsBRD4 indicates a poor prognosis in T-ALL. The BRD4 degrader ARV-825 can effectively suppress the proliferation and promote apoptosis of T-ALL cells via BET protein depletion and c-Myc inhibition, thus providing a new strategy for the treatment of T-ALL.
| S-EPMC8061034 | biostudies-literature