Suppression of apoptosis by expression of antiapoptotic BCL2 family members is a hallmark of acute myeloblastic leukemia (AML). Induced myeloid leukemia cell differentiation protein (MCL1), an ...antiapoptotic BCL2 family member, is commonly upregulated in AML cells and is often a primary mode of resistance to treatment with the BCL2 inhibitor venetoclax. Here, we describe VU661013, a novel, potent, selective MCL1 inhibitor that destabilizes BIM/MCL1 association, leads to apoptosis in AML, and is active in venetoclax-resistant cells and patient-derived xenografts. In addition, VU661013 was safely combined with venetoclax for synergy in murine models of AML. Importantly, BH3 profiling of patient samples and drug-sensitivity testing
accurately predicted cellular responses to selective inhibitors of MCL1 or BCL2 and showed benefit of the combination. Taken together, these data suggest a strategy of rationally using BCL2 and MCL1 inhibitors in sequence or in combination in AML clinical trials. SIGNIFICANCE: Targeting antiapoptotic proteins in AML is a key therapeutic strategy, and MCL1 is a critical antiapoptotic oncoprotein. Armed with novel MCL1 inhibitors and the potent BCL2 inhibitor venetoclax, it may be possible to selectively induce apoptosis by combining or thoughtfully sequencing these inhibitors based on a rational evaluation of AML.
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Structural biology of the Bcl-2 family of proteins Petros, Andrew M.; Olejniczak, Edward T.; Fesik, Stephen W.
BBA - Molecular Cell Research,
03/2004, Letnik:
1644, Številka:
2
Book Review, Journal Article
Recenzirano
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The proteins of the Bcl-2 family are important regulators of programmed cell death. Structural studies of Bcl-2 family members have provided many important insights into their molecular mechanism of ...action and how members of this family interact with one another. To date, structural studies have been performed on six Bcl-2 family members encompassing both anti- (Bcl-x
L, Bcl-2, KSHV-Bcl-2, Bcl-w) and pro-apoptotic (Bax, Bid) members. They all show a remarkably similar fold despite an overall divergence in amino acid sequence and function (pro-apoptotic versus anti-apoptotic). The three-dimensional structures of Bcl-2 family members consist of two central, predominantly hydrophobic α-helices surrounded by six or seven amphipathic α-helices of varying lengths. A long, unstructured loop is present between the first two α-helices. The structures of the Bcl-2 proteins show a striking similarity to the overall fold of the pore-forming domains of bacterial toxins. This finding led to experiments which demonstrated that Bcl-x
L, Bcl-2, and Bax all form pores in artificial membranes. A prominent hydrophobic groove is present on the surface of the anti-apoptotic proteins. This groove is the binding site for peptides that mimic the BH3 region of various pro-apoptotic proteins such as Bak and Bad. Structures of Bcl-x
L in complex with these BH3 peptides showed that they bind as an amphipathic α-helix and make extensive hydrophobic contacts with the protein. These data have not only helped to elucidate the interactions important for hetero-dimerization of Bcl-2 family members but have also been used to guide the discovery of small molecules that block Bcl-x
L and Bcl-2 function. In the recently determined structure of the anti-apoptotic Bcl-w protein, the protein was also found to have a hydrophobic groove on its surface capable of binding BH3-containing proteins and peptides. However, in the native protein an additional carboxy-terminal α-helix interacts with the hydrophobic groove. This is reminiscent of how the carboxy-terminal α-helix of the pro-apoptotic protein Bax binds into its hydrophobic groove. This interaction may play a regulatory role and for Bax may explain why it is found predominately in the cytoplasm prior to activation. The hydrophobic groove of the pro-apoptotic protein, Bid protein, is neither as long nor as deep as that found in Bcl-x
L, Bcl-2, or Bax. In addition, Bid contains an extra α-helix, which is located between α1 and α2 with respect to Bcl-x
L, Bcl-2, and Bax. Although there are still many unanswered questions regarding the exact mechanism by which the Bcl-2 family of proteins modulates apoptosis, structural studies of these proteins have deepened our understanding of apoptosis on the molecular level.
Nuclear magnetic resonance (NMR) spectroscopy has evolved into a powerful tool for fragment-based drug discovery over the last two decades. While NMR has been traditionally used to elucidate the ...three-dimensional structures and dynamics of biomacromolecules and their interactions, it can also be a very valuable tool for the reliable identification of small molecules that bind to proteins and for hit-to-lead optimization. Here, we describe the use of NMR spectroscopy as a method for fragment-based drug discovery and how to most effectively utilize this approach for discovering novel therapeutics based on our experience.
Short interfering RNA (siRNA) is widely used for studying gene function and holds great promise as a tool for validating drug targets and treating disease. A critical assumption in these applications ...is that the effect of siRNA on cells is specific, i.e., limited to the specific knockdown of the target gene. In this article, we characterize the specificity of siRNA by applying gene expression profiling. Several siRNAs were designed against different regions of the same target gene for three different targets. Their effects on cells were compared by using DNA microarrays to generate gene expression signatures. When the siRNA design and transfection conditions were optimized, the signatures for different siRNAs against the same target were shown to correlate very closely, whereas the signatures for different genes revealed no correlation. These results indicate that siRNA is a highly specific tool for targeted gene knockdown, establishing siRNA-mediated gene silencing as a reliable approach for large-scale screening of gene function and drug target validation.
Drugging an undruggable pocket on KRAS Kesslera, Dirk; Gmachla, Michael; Mantoulidisa, Andreas ...
Proceedings of the National Academy of Sciences - PNAS,
08/2019, Letnik:
116, Številka:
32
Journal Article
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Odprti dostop
The 3 human RAS genes, KRAS, NRAS, and HRAS, encode 4 different RAS proteins which belong to the protein family of small GTPases that function as binary molecular switches involved in cell signaling. ...Activating mutations in RAS are among the most common oncogenic drivers in human cancers, with KRAS being the most frequently mutated oncogene. Although KRAS is an excellent drug discovery target for many cancers, and despite decades of research, no therapeutic agent directly targeting RAS has been clinically approved. Using structure-based drug design, we have discovered BI-2852 (1), a KRAS inhibitor that binds with nanomolar affinity to a pocket, thus far perceived to be “undruggable,” between switch I and II on RAS; 1 is mechanistically distinct from covalent KRASG12C inhibitors because it binds to a different pocket present in both the active and inactive forms of KRAS. In doing so, it blocks all GEF, GAP, and effector interactions with KRAS, leading to inhibition of downstream signaling and an antiproliferative effect in the low micromolar range in KRAS mutant cells. These findings clearly demonstrate that this so-called switch I/II pocket is indeed druggable and provide the scientific community with a chemical probe that simultaneously targets the active and inactive forms of KRAS.
Looking for fragments: A fragment‐based screen using NMR spectroscopy was applied to discover ligands that bind to the GTPase K‐Ras and modulate the activity of the nucleotide exchange factor Sos. ...Structural data on how these fragment‐derived hits bind to the guanosine diphosphate–K‐Ras complex (see picture) provides a starting point for the future discovery of drugs that target K‐Ras activation and signaling.
Overexpression of prosurvival proteins such as Bcl-2 and Bcl-XL has been correlated with tumorigenesis and resistance to chemotherapy, and thus, the development of antagonists of these proteins may ...provide a novel means for the treatment of cancer. We recently described the discovery of 1 (ABT-737), which binds Bcl-2, Bcl-XL, and Bcl-w with high affinity, shows robust antitumor activity in murine tumor xenograft models, but is not orally bioavailable. Herein, we report that targeted modifications at three key positions of 1 resulted in a 20-fold improvement in the pharmacokinetic/pharmacodynamic relationship (PK/PD) between oral exposure (AUC) and in vitro efficacy in human tumor cell lines (EC50). The resulting compound, 2 (ABT-263), is orally efficacious in an established xenograft model of human small cell lung cancer, inducing complete tumor regressions in all animals. Compound 2 is currently in multiple phase 1 clinical trials in patients with small cell lung cancer and hematological malignancies.
Cellular and genetic evidence suggest that inhibition of ATAD2 could be a useful strategy to treat several types of cancer. To discover small-molecule inhibitors of the bromodomain of ATAD2, we used ...a fragment-based approach. Fragment hits were identified using NMR spectroscopy, and ATAD2 was crystallized with three of the hits identified in the fragment screen.
The chromatin-associated protein WDR5 is a promising target for pharmacological inhibition in cancer. Drug discovery efforts center on the blockade of the “WIN site” of WDR5, a well-defined pocket ...that is amenable to small molecule inhibition. Various cancer contexts have been proposed to be targets for WIN site inhibitors, but a lack of understanding of WDR5 target genes and of the primary effects of WIN site inhibitors hampers their utility. Here, by the discovery of potent WIN site inhibitors, we demonstrate that the WIN site links WDR5 to chromatin at a small cohort of loci, including a specific subset of ribosome protein genes. WIN site inhibitors rapidly displace WDR5 from chromatin and decrease the expression of associated genes, causing translational inhibition, nucleolar stress, and p53 induction. Our studies define a mode by which WDR5 engages chromatin and forecast that WIN site blockade could have utility against multiple cancer types.
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•WDR5 is a regulator of ribosome protein gene transcription•The WIN site of WDR5 tethers it to chromatin•Potent small molecule inhibitors of the WIN site displace WDR5 from chromatin•WIN site inhibitors induce nucleolar stress and p53-dependent apoptosis
WDR5 is a chromatin-associated protein and promising anti-cancer target. Aho et al. show that WDR5 controls the expression of ribosome protein genes and describe how small molecule inhibitors of WDR5 displace it from chromatin, causing impeded translation, nucleolar stress, and induction of p53-dependent apoptosis in leukemia cells.
The oncoprotein transcription factor MYC is overexpressed in the majority of cancers. Key to its oncogenic activity is the ability of MYC to regulate gene expression patterns that drive and maintain ...the malignant state. MYC is also considered a validated anticancer target, but efforts to pharmacologically inhibit MYC have failed. The dependence of MYC on cofactors creates opportunities for therapeutic intervention, but for any cofactor this requires structural understanding of how the cofactor interacts with MYC, knowledge of the role it plays in MYC function, and demonstration that disrupting the cofactor interaction will cause existing cancers to regress. One cofactor for which structural information is available is WDR5, which interacts with MYC to facilitate its recruitment to chromatin. To explore whether disruption of the MYC–WDR5 interaction could potentially become a viable anticancer strategy, we developed a Burkitt’s lymphoma system that allows replacement of wild-type MYC for mutants that are defective for WDR5 binding or all known nuclear MYC functions. Using this system, we show that WDR5 recruits MYC to chromatin to control the expression of genes linked to biomass accumulation. We further show that disrupting the MYC–WDR5 interaction within the context of an existing cancer promotes rapid and comprehensive tumor regression in vivo. These observations connect WDR5 to a core tumorigenic function of MYC and establish that, if a therapeutic window can be established, MYC–WDR5 inhibitors could be developed as anticancer agents.
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