| 1. |
- Rehling, Daniel, et al.
(author)
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Crystal structures of NUDT15 variants enabled by a potent inhibitor reveal the structural basis for thiopurine sensitivity
- 2021
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In: Journal of Biological Chemistry. - : Elsevier BV. - 0021-9258 .- 1083-351X. ; 296
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Journal article (peer-reviewed)abstract
- The enzyme NUDT15 efficiently hydrolyzes the active metabolites of thiopurine drugs, which are routinely used for treating cancer and inflammatory diseases. Loss-of-function variants in NUDT15 are strongly associated with thiopurine intolerance, such as leukopenia, and preemptive NUDT15 genotyping has been clinically implemented to personalize thiopurine dosing. However, understanding the molecular consequences of these variants has been difficult, as no structural information was available for NUDT15 proteins encoded by clinically actionable pharmacogenetic variants because of their inherent instability. Recently, the small molecule NUDT15 inhibitor TH1760 has been shown to sensitize cells to thiopurines, through enhanced accumulation of 6-thio-guanine in DNA. Building upon this, we herein report the development of the potent and specific NUDT15 inhibitor, TH7755. TH7755 demonstrates a greatly improved cellular target engagement and 6-thioguanine potentiation compared with TH1760, while showing no cytotoxicity on its own. This potent inhibitor also stabilized NUDT15, enabling analysis by X-ray crystallography. We have determined high-resolution structures of the clinically relevant NUDT15 variants Arg139Cys, Arg139His, Val18Ile, and V18_V19insGlyVal. These structures provide clear insights into the structural basis for the thiopurine intolerance phenotype observed in patients carrying these pharmacogenetic variants. These findings will aid in predicting the effects of new NUDT15 sequence variations yet to be discovered in the clinic.
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| 2. |
- Visnes, Torkild, et al.
(author)
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Small-molecule inhibitor of OGG1 suppresses proinflammatory gene expression and inflammation
- 2018
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In: Science. - : American Association for the Advancement of Science (AAAS). - 0036-8075 .- 1095-9203. ; 362:6416, s. 834-
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Journal article (peer-reviewed)abstract
- The onset of inflammation is associated with reactive oxygen species and oxidative damage to macromolecules like 7,8-dihydro-8-oxoguanine (8-oxoG) in DNA. Because 8-oxoguanine DNA glycosylase 1 (OGG1) binds 8-oxoG and because Ogg1-deficient mice are resistant to acute and systemic inflammation, we hypothesized that OGG1 inhibition may represent a strategy for the prevention and treatment of inflammation. We developed TH5487, a selective active-site inhibitor of OGG1, which hampers OGG1 binding to and repair of 8-oxoG and which is well tolerated by mice. TH5487 prevents tumor necrosis factor-alpha-induced OGG1-DNA interactions at guanine-rich promoters of proinflammatory genes. This, in turn, decreases DNA occupancy of nuclear factor kappa B and proinflammatory gene expression, resulting in decreased immune cell recruitment to mouse lungs. Thus, we present a proof of concept that targeting oxidative DNA repair can alleviate inflammatory conditions in vivo.
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| 3. |
- Marklund, Maja, et al.
(author)
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Spatio-temporal analysis of prostate tumors in situ suggests pre-existence of treatment-resistant clones
- 2022
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In: Nature Communications. - : Springer Nature. - 2041-1723. ; 13:1
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Journal article (peer-reviewed)abstract
- The molecular mechanisms underlying lethal castration-resistant prostate cancer remain poorly understood, with intratumoral heterogeneity a likely contributing factor. To examine the temporal aspects of resistance, we analyze tumor heterogeneity in needle biopsies collected before and after treatment with androgen deprivation therapy. By doing so, we are able to couple clinical responsiveness and morphological information such as Gleason score to transcriptome-wide data. Our data-driven analysis of transcriptomes identifies several distinct intratumoral cell populations, characterized by their unique gene expression profiles. Certain cell populations present before treatment exhibit gene expression profiles that match those of resistant tumor cell clusters, present after treatment. We confirm that these clusters are resistant by the localization of active androgen receptors to the nuclei in cancer cells post-treatment. Our data also demonstrates that most stromal cells adjacent to resistant clusters do not express the androgen receptor, and we identify differentially expressed genes for these cells. Altogether, this study shows the potential to increase the power in predicting resistant tumors. Spatial heterogeneity in prostate cancer can contribute to its resistance to androgen deprivation therapy (ADT). Here, the authors analyse prostate cancer samples before and after ADT using Spatial Transcriptomics, and find heterogeneous pre-treatment tumour cell populations and stromal cells that are associated with resistance.
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| 4. |
- Llona-Minguez, Sabin, et al.
(author)
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Discovery of the First Potent and Selective Inhibitors of Human dCTP Pyrophosphatase 1
- 2016
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In: Journal of Medicinal Chemistry. - : American Chemical Society (ACS). - 0022-2623 .- 1520-4804. ; 59:3, s. 1140-1148
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Journal article (peer-reviewed)abstract
- The dCTPase pyrophosphatase 1 (dCTPase) regulates the intracellular nucleotide pool through hydrolytic degradation of canonical and noncanonical nucleotide triphosphates (dNTPs). dCTPase is highly expressed in multiple carcinomas and is associated with cancer cell sternness. Here we report on the development of the first potent and selective dCTPase inhibitors that enhance the cytotoxic effect of cytidine analogues in leukemia cells. Boronate 30 displays a promising in vitro ADME profile, including plasma and mouse microsomal half-lives, aqueous solubility, cell permeability and CYP inhibition, deeming it a suitable compound for in vivo studies.
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| 5. |
- Llona-Minguez, Sabin, et al.
(author)
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Identification of Triazolothiadiazoles as Potent Inhibitors of the dCTP Pyrophosphatase 1
- 2017
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In: Journal of Medicinal Chemistry. - : American Chemical Society (ACS). - 0022-2623 .- 1520-4804. ; 60:5, s. 2148-2154
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Journal article (peer-reviewed)abstract
- The dCTP pyrophosphatase 1 (dCTPase) is involved in the regulation of the cellular dNTP pool and has been linked to cancer progression. Here we report on the discovery of a series of 3,6-disubstituted triazolothiadiazoles as potent dCTPase inhibitors. Compounds 16 and 18 display good correlation between enzymatic inhibition and target engagement, together with efficacy in a cellular synergy model, deeming them as a promising starting point for hit -to-lead development.
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| 6. |
- Rudling, Axel, et al.
(author)
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Fragment-Based Discovery and Optimization of Enzyme Inhibitors by Docking of Commercial Chemical Space
- 2017
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In: Journal of Medicinal Chemistry. - : American Chemical Society (ACS). - 0022-2623 .- 1520-4804. ; 60:19, s. 8160-8169
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Journal article (peer-reviewed)abstract
- Fragment-based lead discovery has emerged as a leading drug development strategy for novel therapeutic targets. Although fragment-based drug discovery benefits immensely from access to atomic-resolution information, structure-based virtual screening has rarely been used to drive fragment discovery and optimization. Here, molecular docking of 0.3 million fragments to a crystal structure of cancer target MTH1 was performed. Twenty-two predicted fragment ligands, for which analogs could be acquired commercially, were experimentally evaluated. Five fragments inhibited MTH1 with IC50 values ranging from 6 to 79 mu M. Structure-based optimization guided by predicted binding modes and analogs from commercial chemical libraries yielded nanomolar inhibitors. Subsequently solved crystal structures confirmed binding modes predicted by docking for three scaffolds. Structure-guided exploration of commercial chemical space using molecular docking gives access to fragment libraries that are several orders of magnitude larger than those screened experimentally and can enable efficient optimization of hits to potent leads.
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| 7. |
- Gubanova, Evgenia, et al.
(author)
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SMG-1 suppresses CDK2 and tumor growth by regulating both the p53 and Cdc25A signaling pathways
- 2013
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In: Cell Cycle. - : Informa UK Limited. - 1538-4101 .- 1551-4005. ; 12:24, s. 3770-3780
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Journal article (peer-reviewed)abstract
- The DNA damage response is coordinated by phosphatidylinositol 3-kinase-related kinases, ATM, ATR, and DNA-PK. SMG-1 is the least studied stress-responsive member of this family. Here, we show that SMG-1 regulates the G 1/S checkpoint through both a p53-dependent, and a p53-independent pathway. We identify Cdc25A as a new SMG-1 substrate, and show that cells depleted of SMG-1 exhibit prolonged Cdc25A stability, failing to inactivate CDK2 in response to radiation. Given an increased tumor growth following depletion of SMG-1, our data demonstrate a novel role for SMG-1 in regulating Cdc25A and suppressing oncogenic CDK2 driven proliferation, confirming SMG-1 as a tumor suppressor.
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| 8. |
- Ogris, Christoph, et al.
(author)
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A novel method for crosstalk analysis of biological networks : improving accuracy of pathway annotation
- 2017
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In: Nucleic Acids Research. - : Oxford University Press (OUP). - 0305-1048 .- 1362-4962. ; 45:2
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Journal article (peer-reviewed)abstract
- Analyzing gene expression patterns is a mainstay to gain functional insights of biological systems. A plethora of tools exist to identify significant enrichment of pathways for a set of differentially expressed genes. Most tools analyze gene overlap between gene sets and are therefore severely hampered by the current state of pathway annotation, yet at the same time they run a high risk of false assignments. A way to improve both true positive and false positive rates (FPRs) is to use a functional association network and instead look for enrichment of network connections between gene sets. We present a new network crosstalk analysis method BinoX that determines the statistical significance of network link enrichment or depletion between gene sets, using the binomial distribution. This is a much more appropriate statistical model than previous methods have employed, and as a result BinoX yields substantially better true positive and FPRs than was possible before. A number of benchmarks were performed to assess the accuracy of BinoX and competing methods. We demonstrate examples of how BinoX finds many biologically meaningful pathway annotations for gene sets from cancer and other diseases, which are not found by other methods.
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| 9. |
- Green, Alanna C., et al.
(author)
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Formate overflow drives toxic folate trapping in MTHFD1 inhibited cancer cells
- 2023
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In: Nature Metabolism. - : Springer Nature. - 2522-5812. ; 5:4, s. 642-659
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Journal article (peer-reviewed)abstract
- Cancer cells fuel their increased need for nucleotide supply by upregulating one-carbon (1C) metabolism, including the enzymes methylenetetrahydrofolate dehydrogenase–cyclohydrolase 1 and 2 (MTHFD1 and MTHFD2). TH9619 is a potent inhibitor of dehydrogenase and cyclohydrolase activities in both MTHFD1 and MTHFD2, and selectively kills cancer cells. Here, we reveal that, in cells, TH9619 targets nuclear MTHFD2 but does not inhibit mitochondrial MTHFD2. Hence, overflow of formate from mitochondria continues in the presence of TH9619. TH9619 inhibits the activity of MTHFD1 occurring downstream of mitochondrial formate release, leading to the accumulation of 10-formyl-tetrahydrofolate, which we term a ‘folate trap’. This results in thymidylate depletion and death of MTHFD2-expressing cancer cells. This previously uncharacterized folate trapping mechanism is exacerbated by physiological hypoxanthine levels that block the de novo purine synthesis pathway, and additionally prevent 10-formyl-tetrahydrofolate consumption for purine synthesis. The folate trapping mechanism described here for TH9619 differs from other MTHFD1/2 inhibitors and antifolates. Thus, our findings uncover an approach to attack cancer and reveal a regulatory mechanism in 1C metabolism.
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| 10. |
- Somaiah, Navita, et al.
(author)
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Homologous recombination mediates cellular resistance and fraction size sensitivity to radiation therapy
- 2013
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In: Radiotherapy and Oncology. - : Elsevier BV. - 0167-8140 .- 1879-0887. ; 108:1, s. 155-161
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Journal article (peer-reviewed)abstract
- Purpose: Cellular sensitivity to radiotherapy total dose and fraction size is strongly influenced by DNA double strand break (DSB) repair. Here, we investigate response to radiotherapy fraction size using CHO cell lines deficient in specific DNA repair pathways in response to radiation induced DNA double strand breaks (DSB). Experimental design: We irradiated CHO cell lines, AA8 (WT), irs1SF (XRCO-), V3-3 (DNA-PKcs-) and EM9 (XRCC1-) with 16 Gy in 1 Gy daily fractions over 3 weeks or 16 Gy in 4 Gy daily fractions over 4 days, and studied clonogenic survival, DNA DSB repair kinetics (RAD51 and 53BP1 foci staining) and cell cycle profiles (flow cytometry). Results: In response to fractionated radiotherapy, wild-type and DNA repair defective cells accumulated in late S/G2 phase. In cells proficient in homologous recombination (HR), accumulation in S/G2 resulted in reduced sensitivity to fraction size and increased cellular resistance (clonogenic survival). Sensitivity to fraction size was also lost in NHEJ-defective V3-3 cells, which likely rely on functional HR. By contrast, HR-defective irs1SF cells, with functional NHEJ, remained equally sensitive to fractionation throughout the 3-week treatment. Conclusions: The high fidelity of HR, which is independent of induced DNA damage level, is postulated to explain the low fractionation sensitivity and cellular resistance of cells in S/G2 phase. In conclusion, our results suggest that HR mediates resistance to fractionated radiotherapy, an observation that may help future efforts to improve radiotherapy outcome.
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