CC-90009, a novel cereblon E3 ligase modulator targets acute myeloid leukemia blasts and leukemia stem cells

A number of clinically validated drugs have been developed by repurposing the CUL4-DDB1-CRBN-RBX1 (CRL4CRBN) E3 ubiquitin ligase complex with molecular glue degraders to eliminate disease-driving proteins. Here, we present the identification of a first-in-class GSPT1-selective cereblon E3 ligase modulator, CC-90009. Biochemical, structural and molecular characterization demonstrates that CC-90009 co-opts the CRL4CRBN to selectively target GSPT1 for ubiquitination and proteasomal degradation. Depletion of GSPT1 by CC-90009 rapidly induces AML apoptosis, reducing leukemia engraftment and leukemia stem cells (LSC) in large scale primary patient xenografting of 35 independent AML samples, including those with adverse risk features. Using a genome-wide CRISPR-Cas9 screen for effectors of CC-90009 response, we uncovered the ILF2 and ILF3 heterodimeric complex as a novel regulator of cereblon expression. Knockout of ILF2/ILF3 decreases the production of full-length cereblon protein via modulating CRBN mRNA alternative splicing, leading to diminished response to CC-90009. The screen also revealed that the mTOR signaling and the integrated stress response (ISR) specifically regulate the response to CC-90009 in contrast to other cereblon modulators. Hyperactivation of the mTOR pathway by inactivation of TSC1 and TSC2 protected against the growth inhibitory effect of CC-90009 by reducing CC-90009 induced binding of GSPT1 to cereblon and subsequent GSPT1 degradation. On the other hand, GSPT1 degradation promoted the activation of the GCN1/GCN2/ATF4 pathway and subsequent apoptosis in AML cells. Collectively, CC-90009 activity is mediated by multiple layers of signaling networks and pathways within AML blasts and LSC, whose elucidation gives insight into further assessment of CC-90009's clinical utility.

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Elucidating the Mechanism of Action of CC-90009, a Novel Cereblon E3 Ligase Modulator, in AML Via Genome-Wide CRISPR Screen

Blood ◽

10.1182/blood-2019-125492 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 405-405 ◽

Cited By ~ 2

Author(s):

Gang Lu ◽

Christine Surka ◽

Chin-Chun Lu ◽

In Sock Jang ◽

Kai Wang ◽

...

Keyword(s):

Cell Lines ◽

Mechanism Of Action ◽

E3 Ligase ◽

Inhibitory Effect ◽

Growth Inhibitory Effect ◽

Employment Equity ◽

Growth Inhibitory ◽

Genome Wide ◽

A Genome ◽

Equity Ownership

CC-90009 is a novel cereblon E3 ligase modulator (CELMoD) currently under investigation in a phase I clinical study in relapsed or refractory acute myeloid leukemia (R/R AML) (CC-90009-AML-001; NCT02848001). CC-90009 coopts the CUL4-DDB1-CRBN-RBX1 (CRL4CRBN) E3 ubiquitin ligase complex to target the translation termination factor G1 to S phase transition 1 (GSPT1) for ubiquitination and proteasomal degradation, resulting in rapid induction of apoptosis and growth inhibition in AML cell lines and primary patient blasts. To further elucidate the mechanism of action of CC-90009 in AML, we performed a genome-wide CRISPR/Cas9 screen to identify gene(s) whose knockout abrogate(s) the response to CC-90009 in a sensitive AML cell line. In addition to well-established key regulatory proteins required for the activity of all known cereblon modulators, which include components of the CRL4CRBN complex, E2 ubiquitin conjugating enzymes UBE2G1 and UBE2D3, and members of the neddylation and deneddylation machinery, interestingly, the screen identified the ILF2 and ILF3 heterodimeric complex as a novel regulator of cereblon expression. Knockout of ILF2/ILF3 decreased the production of full-length CRBN transcript via modulating alternative splicing of CRBN mRNA, leading to significant downregulation of cereblon expression and hence diminished response to CC-90009. The screen also revealed that mTOR signaling and the integrated stress response (ISR) specifically regulate the response to CC-90009 in contrast to other cereblon modulators. Since CC-90009 inhibits protein translation, it is reasonable to expect interactions with regulators of this pathway. Hyperactivation of the mTOR pathway by inactivation of TSC1 and TSC2 protected against the growth inhibitory effect of CC-90009 , at least in part by reducing CC-90009 induced binding of GSPT1 to cereblon and subsequent GSPT1 degradation. On the other hand, GSPT1 degradation promoted the activation of the GCN1/GCN2/ATF4 pathway and subsequent apoptosis in AML cells. Loss of GCN2 significantly attenuated the growth inhibitory effect of CC-90009, and this effect can be rescued with GCN2 wild-type but not enzymatically-dead mutants. Collectively, the antitumor activity of CC-90009, a first-in-class GSPT1 degrader, in AML cell lines is mediated by multiple layers of signaling networks and machinery, the elucidation of which reveals the underlying mechanism by which CC-90009 exerts its anti-AML activity and informs on the pathways for further study of CC-90009's clinical utility. Disclosures Lu: Celgene Corporation: Employment, Equity Ownership. Surka:Celgene: Employment, Equity Ownership. Lu:Celgene Corporation: Employment, Equity Ownership. Jang:Celgene: Employment, Equity Ownership. Wang:Celgene: Employment, Equity Ownership. Rolfe:Celgene: Employment, Equity Ownership.

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Thiosemicarbazone-Pt(II) Complex Causes a Growth Inhibitory Effect on Human Mesenchymal Stem Cells

Medicinal Chemistry ◽

10.2174/1573406411666150514101026 ◽

2015 ◽

Vol 11 (7) ◽

pp. 670-675 ◽

Cited By ~ 3

Author(s):

Josefa Garcia-Ruiz ◽

Ana Matesanz Garcia ◽

Ana Souza ◽

Pilar Castelo

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Human Mesenchymal Stem Cells ◽

Inhibitory Effect ◽

Growth Inhibitory Effect ◽

Growth Inhibitory

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A genome-wide RNAi screen in mouse embryonic stem cells identifies Mp1 as a key mediator of differentiation

The Journal of Cell Biology ◽

10.1083/jcb1956oia9 ◽

2011 ◽

Vol 195 (6) ◽

pp. i9-i9 ◽

Cited By ~ 1

Author(s):

Bart A. Westerman ◽

A. Koen Braat ◽

Nicole Taub ◽

Marko Potman ◽

Joseph H.A. Vissers ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Embryonic Stem ◽

Mouse Embryonic Stem Cells ◽

Rnai Screen ◽

Genome Wide ◽

A Genome

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Phenotypic Screen and Transcriptomics Approach Complement Each Other in Functional Genomics of Defensive Stink Gland Physiology

10.21203/rs.3.rs-1117784/v1 ◽

2021 ◽

Author(s):

Sabrina Lehmann ◽

Bibi Atika ◽

Daniela Grossmann ◽

Christian Schmitt-Engel ◽

Nadi Strohlein ◽

...

Keyword(s):

Functional Genomics ◽

Large Scale ◽

Reverse Genetics ◽

Expression Profiles ◽

Forward Genetics ◽

Large Set ◽

Knock Down ◽

Genome Wide ◽

Phenotypic Screen ◽

A Genome

Abstract Background Functional genomics uses unbiased systematic genome-wide gene disruption or analyzes natural variations such as gene expression profiles of different tissues from multicellular organisms to link gene functions to particular phenotypes. Functional genomics approaches are of particular importance to identify large sets of genes that are specifically important for a particular biological process beyond known candidate genes, or when the process has not been studied with genetic methods before. Results Here, we present a large set of genes whose disruption interferes with the function of the odoriferous defensive stink glands of the red flour beetle Tribolium castaneum. This gene set is the result of a large-scale systematic phenotypic screen using a reverse genetics strategy based on RNA interference applied in a genome-wide forward genetics manner. In this first-pass screen, 130 genes were identified, of which 69 genes could be confirmed to cause knock-down gland phenotypes, which vary from necrotic tissue and irregular reservoir size to irregular color or separation of the secreted gland compounds. The knock-down of 13 genes caused specifically a strong reduction of para-benzoquinones, suggesting a specific function in the synthesis of these toxic compounds. Only 14 of the 69 confirmed gland genes are differentially overexpressed in stink gland tissue and thus could have been detected in a transcriptome-based analysis. Moreover, of the 29 previously transcriptomics-identified genes causing a gland phenotype, only one gene was recognized by this phenotypic screen despite the fact that 13 of them were covered by the screen. Conclusion Our results indicate the importance of combining diverse and independent methodologies to identify genes necessary for the function of a certain biological tissue, as the different approaches do not deliver redundant results but rather complement each other. The presented phenotypic screen together with a transcriptomics approach are now providing a set of close to hundred genes important for odoriferous defensive stink gland physiology in beetles.

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A New Genome-to-Genome Comparison Approach for Large-Scale Revisiting of Current Microbial Taxonomy

Microorganisms ◽

10.3390/microorganisms7060161 ◽

2019 ◽

Vol 7 (6) ◽

pp. 161 ◽

Cited By ~ 1

Author(s):

Ming-Hsin Tsai ◽

Yen-Yi Liu ◽

Von-Wun Soo ◽

Chih-Chieh Chen

Keyword(s):

Microbial Diversity ◽

Large Scale ◽

Gene Selection ◽

Marker Gene ◽

Genome Comparison ◽

Marker Genes ◽

Species Classification ◽

Genome Wide ◽

A Genome ◽

Comparison Approach

Microbial diversity has always presented taxonomic challenges. With the popularity of next-generation sequencing technology, more unculturable bacteria have been sequenced, facilitating the discovery of additional new species and complicated current microbial classification. The major challenge is to assign appropriate taxonomic names. Hence, assessing the consistency between taxonomy and genomic relatedness is critical. We proposed and applied a genome comparison approach to a large-scale survey to investigate the distribution of genomic differences among microorganisms. The approach applies a genome-wide criterion, homologous coverage ratio (HCR), for describing the homology between species. The survey included 7861 microbial genomes that excluded plasmids, and 1220 pairs of genera exhibited ambiguous classification. In this study, we also compared the performance of HCR and average nucleotide identity (ANI). The results indicated that HCR and ANI analyses yield comparable results, but a few examples suggested that HCR has a superior clustering effect. In addition, we used the Genome Taxonomy Database (GTDB), the gold standard for taxonomy, to validate our analysis. The GTDB offers 120 ubiquitous single-copy proteins as marker genes for species classification. We determined that the analysis of the GTDB still results in classification boundary blur between some genera and that the marker gene-based approach has limitations. Although the choice of marker genes has been quite rigorous, the bias of marker gene selection remains unavoidable. Therefore, methods based on genomic alignment should be considered for use for species classification in order to avoid the bias of marker gene selection. On the basis of our observations of microbial diversity, microbial classification should be re-examined using genome-wide comparisons.

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A Comprehensive Survey on the Terpene Synthase Gene Family Provides New Insight into Its Evolutionary Patterns

Genome Biology and Evolution ◽

10.1093/gbe/evz142 ◽

2019 ◽

Vol 11 (8) ◽

pp. 2078-2098 ◽

Cited By ~ 8

Author(s):

Shu-Ye Jiang ◽

Jingjing Jin ◽

Rajani Sarojam ◽

Srinivasan Ramachandran

Keyword(s):

Gene Family ◽

Large Scale ◽

Family Members ◽

Terpene Synthase ◽

Limited Information ◽

Terpene Synthases ◽

Genome Wide ◽

A Genome ◽

Family Expansion ◽

Insight Into

Abstract Terpenes are organic compounds and play important roles in plant growth and development as well as in mediating interactions of plants with the environment. Terpene synthases (TPSs) are the key enzymes responsible for the biosynthesis of terpenes. Although some species were employed for the genome-wide identification and characterization of the TPS family, limited information is available regarding the evolution, expansion, and retention mechanisms occurring in this gene family. We performed a genome-wide identification of the TPS family members in 50 sequenced genomes. Additionally, we also characterized the TPS family from aromatic spearmint and basil plants using RNA-Seq data. No TPSs were identified in algae genomes but the remaining plant species encoded various numbers of the family members ranging from 2 to 79 full-length TPSs. Some species showed lineage-specific expansion of certain subfamilies, which might have contributed toward species or ecotype divergence or environmental adaptation. A large-scale family expansion was observed mainly in dicot and monocot plants, which was accompanied by frequent domain loss. Both tandem and segmental duplication significantly contributed toward family expansion and expression divergence and played important roles in the survival of these expanded genes. Our data provide new insight into the TPS family expansion and evolution and suggest that TPSs might have originated from isoprenyl diphosphate synthase genes.

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Large-scale GWAS reveals insights into the genetic architecture of same-sex sexual behavior

Science ◽

10.1126/science.aat7693 ◽

2019 ◽

Vol 365 (6456) ◽

pp. eaat7693 ◽

Cited By ~ 53

Author(s):

Andrea Ganna ◽

Karin J. H. Verweij ◽

Michel G. Nivard ◽

Robert Maier ◽

Robbee Wedow ◽

...

Keyword(s):

Sexual Behavior ◽

Genetic Architecture ◽

Large Scale ◽

Genome Wide Association Study ◽

Same Sex ◽

Genome Wide ◽

A Genome ◽

Number Of Sexual Partners ◽

Opposite Sex ◽

Males And Females

Twin and family studies have shown that same-sex sexual behavior is partly genetically influenced, but previous searches for specific genes involved have been underpowered. We performed a genome-wide association study (GWAS) on 477,522 individuals, revealing five loci significantly associated with same-sex sexual behavior. In aggregate, all tested genetic variants accounted for 8 to 25% of variation in same-sex sexual behavior, only partially overlapped between males and females, and do not allow meaningful prediction of an individual’s sexual behavior. Comparing these GWAS results with those for the proportion of same-sex to total number of sexual partners among nonheterosexuals suggests that there is no single continuum from opposite-sex to same-sex sexual behavior. Overall, our findings provide insights into the genetics underlying same-sex sexual behavior and underscore the complexity of sexuality.

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