Showing papers by "Yves Pommier published in 2018"

ONC201 kills breast cancer cells in vitro by targeting mitochondria.

Abstract: // Yoshimi Endo Greer 1 , Natalie Porat-Shliom 2 , Kunio Nagashima 3 , Christina Stuelten 2 , Dan Crooks 4 , Vishal N. Koparde 5 , Samuel F. Gilbert 1 , Celia Islam 1 , Ashley Ubaldini 1 , Yun Ji 6 , Luca Gattinoni 6 , Ferri Soheilian 3 , Xiantao Wang 7 , Markus Hafner 7 , Jyoti Shetty 8 , Bao Tran 8 , Parthav Jailwala 5 , Maggie Cam 5 , Martin Lang 4 , Donna Voeller 1 , William C. Reinhold 9 , Vinodh Rajapakse 9 , Yves Pommier 9 , Roberto Weigert 2 , W. Marston Linehan 4 and Stanley Lipkowitz 1 1 Women’s Malignancies Branch, Center for Cancer Research (CCR), National Cancer Institute (NCI), National Institutes of Health (NIH), Bethesda, MD, USA 2 Laboratory of Cellular and Molecular Biology, CCR, NCI, NIH, Bethesda, MD, USA 3 Electron Microscope Laboratory, Leidos Biomedical Research, Inc. Frederick National Laboratory for Cancer Research (FNLCR), Frederick, MD, USA 4 Urologic Oncology Branch, CCR, NCI, NIH, Bethesda, MD, USA 5 CCR Collaborative Bioinformatics Resource, Leidos Biomedical Research, Inc., FNLCR, Frederick, MD, USA 6 Experimental Transplantation and Immunology Branch, CCR, NCI, NIH, Bethesda, MD, USA 7 RNA Molecular Biology Group, Laboratory of Muscle Stem Cells and Gene Regulation, National Institute of Arthritis and Musculoskeletal and Skin Diseases, NIH, Bethesda, MD, USA 8 CCR Sequencing Facility, Leidos Biomedical Research, Inc., FNLCR, Frederick, MD, USA 9 Developmental Therapeutics Branch, CCR, NCI, NIH, Bethesda, MD, USA Correspondence to: Stanley Lipkowitz, email: lipkowis@mail.nih.gov Keywords: ONC201; breast cancer; mitochondria Received: November 03, 2017 Accepted: March 06, 2018 Published: April 06, 2018 ABSTRACT We report a novel mechanism of action of ONC201 as a mitochondria-targeting drug in cancer cells. ONC201 was originally identified as a small molecule that induces transcription of TNF-related apoptosis-inducing ligand (TRAIL) and subsequently kills cancer cells by activating TRAIL death receptors. In this study, we examined ONC201 toxicity on multiple human breast and endometrial cancer cell lines. ONC201 attenuated cell viability in all cancer cell lines tested. Unexpectedly, ONC201 toxicity was not dependent on either TRAIL receptors nor caspases. Time-lapse live cell imaging revealed that ONC201 induces cell membrane ballooning followed by rupture, distinct from the morphology of cells undergoing apoptosis. Further investigation found that ONC201 induces phosphorylation of AMP-dependent kinase and ATP loss. Cytotoxicity and ATP depletion were significantly enhanced in the absence of glucose, suggesting that ONC201 targets mitochondrial respiration. Further analysis indicated that ONC201 indirectly inhibits mitochondrial respiration. Confocal and electron microscopic analysis demonstrated that ONC201 triggers mitochondrial structural damage and functional impairment. Moreover, ONC201 decreased mitochondrial DNA (mtDNA). RNAseq analysis revealed that ONC201 suppresses expression of multiple mtDNA-encoded genes and nuclear-encoded mitochondrial genes involved in oxidative phosphorylation and other mitochondrial functions. Importantly, fumarate hydratase deficient cancer cells and multiple cancer cell lines with reduced amounts of mtDNA were resistant to ONC201. These results indicate that cells not dependent on mitochondrial respiration are ONC201-resistant. Our data demonstrate that ONC201 kills cancer cells by disrupting mitochondrial function and further suggests that cancer cells that are dependent on glycolysis will be resistant to ONC201.

Overcoming Resistance to DNA-Targeted Agents by Epigenetic Activation of Schlafen 11 (SLFN11) Expression with Class I Histone Deacetylase Inhibitors.

Abstract: Purpose: Schlafen 11 (SLFN11), a putative DNA/RNA helicase is a dominant genomic determinant of response to DNA-damaging agents and is frequently not expressed in cancer cells. Whether histone deacetylase (HDAC) inhibitors can be used to release SLFN11 and sensitize SLFN11-inactivated cancers to DNA-targeted agents is tested here.Experimental Design:SLFN11 expression was examined in The Cancer Genome Atlas (TCGA), in cancer cell line databases and in patients treated with romidepsin. Isogenic cells overexpressing or genetically inactivated for SLFN11 were used to investigate the effect of HDAC inhibitors on SLFN11 expression and sensitivity to DNA-damaging agents.Results:SLFN11 expression is suppressed in a broad fraction of common cancers and cancer cell lines. In cancer cells not expressing SLFN11, transfection of SLFN11 sensitized the cells to camptothecin, topotecan, hydroxyurea, and cisplatin but not to paclitaxel. SLFN11 mRNA and protein levels were strongly induced by class I (romidepsin, entinostat), but not class II (roclinostat) HDAC inhibitors in a broad panel of cancer cells. SLFN11 expression was also enhanced in peripheral blood mononuclear cells of patients with circulating cutaneous T-cell lymphoma treated with romidepsin. Consistent with the epigenetic regulation of SLFN11, camptothecin and class I HDAC inhibitors were synergistic in many of the cell lines tested.Conclusions: This study reports the prevalent epigenetic regulation of SLFN11 and the dominant stimulatory effect of HDAC inhibitors on SLFN11 expression. Our results provide a rationale for combining class I HDAC inhibitors and DNA-damaging agents to overcome epigenetic inactivation of SLFN11-mediated resistance to DNA-targeted agents. Clin Cancer Res; 24(8); 1944-53. ©2018 AACR.

PRMT5-mediated arginine methylation of TDP1 for the repair of topoisomerase I covalent complexes.

Abstract: Human tyrosyl-DNA phosphodiesterases (TDP) hydrolyze the phosphodiester bond between DNA and the catalytic tyrosine of Top1 to excise topoisomerase I cleavage complexes (Top1cc) that are trapped by camptothecin (CPT) and by genotoxic DNA alterations. Here we show that the protein arginine methyltransferase PRMT5 enhances the repair of Top1cc by direct binding to TDP1 and arginine dimethylation of TDP1 at residues R361 and R586. Top1-induced replication-mediated DNA damage induces TDP1 arginine methylation, enhancing its 3'- phosphodiesterase activity. TDP1 arginine methylation also increases XRCC1 association with TDP1 in response to CPT, and the recruitment of XRCC1 to Top1cc DNA damage foci. PRMT5 knockdown cells exhibit defective TDP1 activity with marked elevation in replication-coupled CPT-induced DNA damage and lethality. Finally, methylation of R361 and R586 stimulate TDP1 repair function and promote cell survival in response to CPT. Together, our findings provide evidence for the importance of PRMT5 for the post-translational regulation of TDP1 and repair of Top1cc.

Discovery, Synthesis, and Evaluation of Oxynitidine Derivatives as Dual Inhibitors of DNA Topoisomerase IB (TOP1) and Tyrosyl-DNA Phosphodiesterase 1 (TDP1), and Potential Antitumor Agents

Abstract: Tyrosyl–DNA phosphodiesterase 1 (TDP1) is a recently discovered enzyme repairing DNA lesions resulting from stalled topoisomerase IB (TOP1)–DNA covalent complex. Inhibiting TDP1 in conjunction with TOP1 inhibitors can boost the action of the latter. Herein, we report the discovery of the natural product oxynitidine scaffold as a novel chemotype for the development of TOP1 and TDP1 inhibitors. Three kinds of analogues, benzophenanthridinone, dihydrobenzophenanthridine, and benzophenanthridine derivatives, were synthesized and evaluated for both TOP1 and TDP1 inhibition and cytotoxicity. Analogue 19a showed high TOP1 inhibition (+++) and induced the formation of cellular TOP1cc and DNA damage, resulting in cancer cells apoptosis at nanomolar concentration range. In vivo studies indicated that 19a exhibits antitumor efficiency in HCT116 xenograft model. 41a exhibited additional TDP1 inhibition with IC50 value of 7 μM and synergistic effect with camptothecin in MCF-7 cells. This work will facilitate future ef...

NCI Comparative Oncology Program Testing of Non-Camptothecin Indenoisoquinoline Topoisomerase I Inhibitors in Naturally Occurring Canine Lymphoma.

Abstract: Purpose: Only one chemical class of topoisomerase I (TOP1) inhibitors is FDA approved, the camptothecins with irinotecan and topotecan widely used. Because of their limitations (chemical instability, drug efflux-mediated resistance, and diarrhea), novel TOP1 inhibitors are warranted. Indenoisoquinoline non-camptothecin topoisomerase I (TOP1) inhibitors overcome chemical instability and drug resistance that limit camptothecin use. Three indenoisoquinolines, LMP400 (indotecan), LMP776 (indimitecan), and LMP744, were examined in a phase I study for lymphoma-bearing dogs to evaluate differential efficacy, pharmacodynamics, toxicology, and pharmacokinetics. Experimental Design: Eighty-four client-owned dogs with lymphomas were enrolled in dose-escalation cohorts for each indenoisoquinoline, with an expansion phase for LMP744. Efficacy, tolerability, pharmacokinetics, and target engagement were determined. Results: The MTDs were 17.5 mg/m2 for LMP 776 and 100 mg/m2 for LMP744; bone marrow toxicity was dose-limiting; up to 65 mg/m2 LMP400 was well-tolerated and MTD was not reached. None of the drugs induced notable diarrhea. Sustained tumor accumulation was observed for LMP744; γH2AX induction was demonstrated in tumors 2 and 6 hours after treatment; a decrease in TOP1 protein was observed in most lymphoma samples across all compounds and dose levels, which is consistent with the fact that tumor response was also observed at low doses LMP744. Objective responses were documented for all indenoisoquinolines; efficacy (13/19 dogs) was greatest for LMP744. Conclusions: These results demonstrate proof-of-mechanism for indenoisoquinoline TOP1 inhibitors supporting their further clinical development. They also highlight the value of the NCI Comparative Oncology Program (https://ccr.cancer.gov/Comparative-Oncology-Program) for evaluating novel therapies in immunocompetent pets with cancers.

Characterization and structure-activity relationships of indenoisoquinoline-derived topoisomerase I inhibitors in unsilencing the dormant Ube3a gene associated with Angelman syndrome

Abstract: Angelman syndrome (AS) is a severe neurodevelopmental disorder lacking effective therapies. AS is caused by mutations in ubiquitin protein ligase E3A (UBE3A), which is genomically imprinted such that only the maternally inherited copy is expressed in neurons. We previously demonstrated that topoisomerase I (Top1) inhibitors could successfully reactivate the dormant paternal allele of Ube3a in neurons of a mouse model of AS. We also previously showed that one such Top1 inhibitor, topotecan, could unsilence paternal UBE3A in induced pluripotent stem cell-derived neurons from individuals with AS. Although topotecan has been well-studied and is FDA-approved for cancer therapy, its limited CNS bioavailability will likely restrict the therapeutic use of topotecan in AS. The goal of this study was to identify additional Top1 inhibitors with similar efficacy as topotecan, with the expectation that these could be tested in the future for safety and CNS bioavailability to assess their potential as AS therapeutics. We tested 13 indenoisoquinoline-derived Top1 inhibitors to identify compounds that unsilence the paternal allele of Ube3a in mouse neurons. Primary cortical neurons were isolated from embryonic day 14.5 (E14.5) mice with a Ube3a-YFP fluorescent tag on the paternal allele (Ube3am+/pYFP mice) or mice that lack the maternal Ube3a allele and hence model AS (Ube3am−/p+ mice). Neurons were cultured for 7 days, treated with drug for 72 h, and examined for paternal UBE3A protein expression by Western blot or fluorescence immunostaining. Dose responses of the compounds were determined across a log range of drug treatments, and cytotoxicity was tested using a luciferase-based assay. All 13 indenoisoquinoline-derived Top1 inhibitors unsilenced paternal Ube3a. Several compounds exhibited favorable paternal Ube3a unsilencing properties, similar to topotecan, and of these, indotecan (LMP400) was the most effective based on estimated Emax (maximum response of unsilencing paternal Ube3a) and EC50 (half maximal effective concentration). We provide pharmacological profiles of indenoisoquinoline-derived Top1 inhibitors as paternal Ube3a unsilencers. All 13 tested compounds were effective at unsilencing paternal Ube3a, although with variable efficacy and potency. Indotecan (LMP400) demonstrated a better pharmacological profile of Ube3a unsilencing compared to our previous lead compound, topotecan. Taken together, indotecan and its structural analogues are potential AS therapeutics whose translational potential in AS treatment should be further assessed.

Novel Fluoroindenoisoquinoline Non-Camptothecin Topoisomerase I Inhibitors

Abstract: Contrary to other anticancer targets, topoisomerase I (TOP1) is targeted by only one chemical class of FDA-approved drugs: topotecan and irinotecan, the derivatives of the plant alkaloid, camptothecin. The indenoisoquinolines LMP400, LMP744, and LMP776 are novel noncamptothecin TOP1 inhibitors in clinical trial, which overcome the limitations of camptothecins. To further improve metabolic stability, their methoxy groups have been replaced by fluorine, as in the fluoroindenoisoquinolines NSC 781517 (LMP517), NSC 779135 (LMP135), and NSC 779134 (LMP134). We tested the induction and stability of TOP1 cleavage complexes (TOP1cc), and the induction and persistence of DNA damage measured by histone H2AX phosphorylation (γH2AX) compared with their parent compounds LMP744 and LMP776 in leukemia CCRF-CEM and colon carcinoma HCT116 cells. The fluoroindenoisoquinolines induced TOP1cc and γH2AX at nanomolar concentrations, and at higher levels than the parent indenoisoquinolines. The fluoroindenoisoquinoline LMP135 showed greater antitumor activity than topotecan in small-cell lung cancer cell H82 xenografts. It was also more potent than topotecan in the NCI-60 cancer cell line panel. Bioinformatics tools (http://discover.nci.nih.gov/cellminercdb) were used to investigate the following: (i) the correlations of fluoroindenoisoquinolines activity with other drugs, and (ii) genomic determinants of response in the NCI-60. The activity of the fluoroindenoisoquinolines was mostly correlated with camptothecin derivatives and the parent indenoisoquinolines, consistent with TOP1 targeting. Genomic analyses and activity assays in CCRF-CEM SLFN11-deleted cells showed that SLFN11 expression is a dominant determinant of response to LMP135. This study shows the potential value of the fluoroindenoisoquinolines for further development as novel anticancer agents targeting TOP1. Mol Cancer Ther; 17(8); 1694-704. ©2018 AACR.

Systematic Analysis of Compounds Specifically Targeting Telomeres and Telomerase for Clinical Implications in Cancer Therapy.

Abstract: The targeting of telomerase and telomere maintenance mechanisms represents a promising therapeutic approach for various types of cancer. In this work, we designed a new protocol to screen for and rank the efficacy of compounds specifically targeting telomeres and telomerase. This approach used two isogenic cell lines containing a circular human artificial chromosome (HAC, lacking telomeres) and a linear HAC (containing telomeres) marked with the EGFP transgene; compounds that target telomerase or telomeres should preferentially induce loss of the linear HAC but not the circular HAC. Our assay allowed quantification of chromosome loss by routine flow cytometry. We applied this dual-HAC assay to rank a set of known and newly developed compounds, including G-quadruplex (G4) ligands. Among the latter group, two compounds, Cu-ttpy and Pt-ttpy, induced a high rate of linear HAC loss with no significant effect on the mitotic stability of a circular HAC. Analysis of the mitotic phenotypes induced by these drugs revealed an elevated rate of chromatin bridges in late mitosis and cytokinesis as well as UFB (ultrafine bridges). Chromosome loss after Pt-ttpy or Cu-ttpy treatment correlated with the induction of telomere-associated DNA damage. Overall, this platform enables identification and ranking of compounds that greatly increase chromosome mis-segregation rates as a result of telomere dysfunction and may expedite the development of new therapeutic strategies for cancer treatment.Significance: An assay provides a unique opportunity to screen thousands of chemical compounds for their ability to inactivate replication of telomeric ends in cancer cells and holds potential to lay the foundation for the discovery of new treatments for cancer. Cancer Res; 78(21); 6282-96. ©2018 AACR.

Novel clinical indenoisoquinoline topoisomerase I inhibitors: a twist around the camptothecins.

Abstract: Topotecan and irinotecan are both derivatives of the plant alkaloid camptothecin [1]. They are widely used for the treatment of solid tumors including colon, lung and ovarian cancers as well as pediatric tumors. Since their FDA approval almost 20 years ago, they have remained the only marketed topoisomerase I (TOP1) inhibitors [2]. This situation is unique in the anticancer armamentarium and is primarily related to the fact that pharmaceutical companies turned to the discovery of protein kinase inhibitors (often referred to as “targeted therapies”) shortly before the clinical approval of topotecan and irinotecan. Yet, camptothecins can be nominated “targeted therapies” as they only target TOP1, and crystal structures demonstrate that only the active natural stereoisomers (20S) of the camptothecins (see top right of the Figure 1) bind to the TOP1-DNA interface (scheme in the middle of the Figure 1), stacking with the base pairs that flank the DNA cleavage site generated by TOP1 and forming a network of hydrogen bonds with amino acid residues of the TOP1 catalytic pocket [3]. This finding led to the concept of interfacial inhibition, a molecular mechanism of action for a wide variety of natural products including TOP2 inhibitors, rapamycin, tubulin inhibitors and anesthetic drugs [3]. Binding of a single molecule of a camptothecin or indenoisoquinoline [4] traps the TOP1-DNA complex by blocking the religation of the broken DNA and the release of TOP1, which remains covalently attached to the 3’-end of the break that TOP1 made to relax DNA. As a result, camptothecins and indenoisoquinolines act by producing TOP1 cleavage complexes (TOP1cc). Editorial

Mitochondrial tyrosyl-DNA phosphodiesterase 2 and its TDP2S short isoform.

Abstract: Tyrosyl-DNA phosphodiesterase 2 (TDP2) repairs abortive topoisomerase II cleavage complexes Here, we identify a novel short isoform of TDP2 (TDP2S) expressed from an alternative transcription start site TDP2S contains a mitochondrial targeting sequence, contributing to its enrichment in the mitochondria and cytosol, while full-length TDP2 contains a nuclear localization signal and the ubiquitin-associated domain in the N-terminus Our study reveals that both TDP2 isoforms are present and active in the mitochondria Comparison of isogenic wild-type (WT) and TDP2 knockout (TDP2-/-/-) DT40 cells shows that TDP2-/-/- cells are hypersensitive to mitochondrial-targeted doxorubicin (mtDox), and that complementing TDP2-/-/- cells with human TDP2 restores resistance to mtDox Furthermore, mtDox selectively depletes mitochondrial DNA in TDP2-/-/- cells Using CRISPR-engineered human cells expressing only the TDP2S isoform, we show that TDP2S also protects human cells against mtDox Finally, lack of TDP2 in the mitochondria reduces the mitochondria transcription levels in two different human cell lines In addition to identifying a novel TDP2S isoform, our report demonstrates the presence and importance of both TDP2 isoforms in the mitochondria

TDP1 suppresses mis-joining of radiomimetic DNA double-strand breaks and cooperates with Artemis to promote optimal nonhomologous end joining.

Abstract: The Artemis nuclease and tyrosyl-DNA phosphodiesterase (TDP1) are each capable of resolving protruding 3'-phosphoglycolate (PG) termini of DNA double-strand breaks (DSBs). Consequently, both a knockout of Artemis and a knockout/knockdown of TDP1 rendered cells sensitive to the radiomimetic agent neocarzinostatin (NCS), which induces 3'-PG-terminated DSBs. Unexpectedly, however, a knockdown or knockout of TDP1 in Artemis-null cells did not confer any greater sensitivity than either deficiency alone, indicating a strict epistasis between TDP1 and Artemis. Moreover, a deficiency in Artemis, but not TDP1, resulted in a fraction of unrepaired DSBs, which were assessed as 53BP1 foci. Conversely, a deficiency in TDP1, but not Artemis, resulted in a dramatic increase in dicentric chromosomes following NCS treatment. An inhibitor of DNA-dependent protein kinase, a key regulator of the classical nonhomologous end joining (C-NHEJ) pathway sensitized cells to NCS, but eliminated the sensitizing effects of both TDP1 and Artemis deficiencies. These results suggest that TDP1 and Artemis perform different functions in the repair of terminally blocked DSBs by the C-NHEJ pathway, and that whereas an Artemis deficiency prevents end joining of some DSBs, a TDP1 deficiency tends to promote DSB mis-joining.

Application of Sequential Palladium Catalysis for the Discovery of Janus Kinase Inhibitors in the Benzo[c]pyrrolo[2,3-h][1,6]naphthyridin-5-one (BPN) Series

Abstract: The present account describes the discovery and development of a new benzo[ c]pyrrolo[2,3- h][1,6]naphthyridin-5-one (BPN) JAK inhibitory chemotype that has produced selective JAK inhibitors. Sequential palladium chemistry was optimized for the rapid access to a focused library of derivatives to explore the structure-activity relationships of the new scaffold. Several compounds from the series displayed potencies in the low nanomolar range against the four members of the JAK family with various selectivity profiles. Compound 20a, with an azetidine amide side chain, showed the best selectivity for JAK1 kinase vs JAK2, JAK3, and TYK2, with low nanomolar potency (IC50 = 3.4 nM). On the other hand, BPNs 17b and 18 had good general activity against the JAK family with excellent kinome selectivity profiles. Many of the new BPNs inhibited JAK3-mediated STAT-5 phosphorylation, the production of inflammatory cytokines, and the proliferation of primary T cells. Moreover, BPN 17b showed very similar in vivo results to tofacitinib in a rheumatoid arthritis animal model.

Endogenous single-strand DNA breaks at RNA polymerase II promoters in Saccharomyces cerevisiae

Abstract: Molecular combing and gel electrophoretic studies revealed endogenous nicks with free 3'OH ends at ∼100 kb intervals in the genomic DNA (gDNA) of unperturbed and G1-synchronized Saccharomyces cerevisiae cells. Analysis of the distribution of endogenous nicks by Nick ChIP-chip indicated that these breaks accumulated at active RNA polymerase II (RNAP II) promoters, reminiscent of the promoter-proximal transient DNA breaks of higher eukaryotes. Similar periodicity of endogenous nicks was found within the ribosomal rDNA cluster, involving every ∼10th of the tandemly repeated 9.1 kb units of identical sequence. Nicks were mapped by Southern blotting to a few narrow regions within the affected units. Three of them were overlapping the RNAP II promoters, while the ARS-containing IGS2 region was spared of nicks. By using a highly sensitive reverse-Southwestern blot method to map free DNA ends with 3'OH, nicks were shown to be distinct from other known rDNA breaks and linked to the regulation of rDNA silencing. Nicks in rDNA and the rest of the genome were typically found at the ends of combed DNA molecules, occasionally together with R-loops, comprising a major pool of vulnerable sites that are connected with transcriptional regulation.

Rapid proteotyping reveals cancer biology and drug response determinants in the NCI-60 cells

Abstract: We describe the rapid and reproducible acquisition of quantitative proteome maps for the NCI-60 cancer cell lines and their use to reveal cancer biology and drug response determinants. Proteome datasets for the 60 cell lines were acquired in duplicate within 30 working days using pressure cycling technology and SWATH mass spectrometry. We consistently quantified 3,171 proteotypic proteins annotated in the SwissProt database across all cell lines, generating a data matrix with 0.1% missing values, allowing analyses of protein complexes and pathway activities across all the cancer cells. Systematic and integrative analysis of the genetic variation, mRNA expression and proteomic data of the NCI-60 cancer cell lines uncovered complementarity between different types of molecular data in the prediction of the response to 240 drugs. We additionally identified novel proteomic drug response determinants for clinically relevant chemotherapeutic and targeted therapies. We anticipate that this study represents a significant advance toward the translational application of proteotypes, which reveal biological insights that are easily missed in the absence of proteomic data.

Novel screen for anti-cancer drugs that elevate chromosome instability (CIN) using human artificial chromosome (HAC).

Abstract: Human artificial chromosomes (HACs) bearing functional kinetochores have been exploited as promising systems for gene delivery and expression and in studies of different epigenetic modifications on kinetochore structure and function. The HAC-based technology has been also used to develop drug screening and assessment strategies to manipulate the CIN (chromosome instability) phenotype in cancer cells. More recently, we designed a new protocol for systematic analysis of compounds specifically targeting telomeres and telomerase. This approach used two isogenic cell lines containing a circular HAC (lacking telomeres) and a linear HAC (containing telomeres): compounds that target telomerase or telomeres should preferentially induce loss of the linear HAC but not the circular HAC. This platform enables identification and ranking of compounds that greatly increase chromosome mis-segregation rates as a result of telomere dysfunction and may expedite the development of new therapeutic strategies for cancer treatment.

Integrative analysis of pharmacogenomics in major cancer cell line databases using CellMinerCDB

Abstract: As precision medicine demands molecular determinants of drug response, CellMinerCDB provides (https://discover.nci.nih.gov/cellminercdb/) a web-based portal for multiple forms of pharmacological, molecular, and genomic analyses, unifying the richest cancer cell line datasets (NCI-60, NCI-SCLC, Sanger/MGH GDSC, and Broad CCLE/CTRP). CellMinerCDB enables genomic and pharmacological data queries for identifying pharmacogenomic determinants, drug signatures, and gene regulatory networks for researchers without requiring specialized bioinformatics support. It leverages overlaps of cell lines and tested drugs to allow assessment of data reproducibility. It builds on the complementarity and strength of each dataset. A panel of 41 drugs evaluated in parallel in the NCI-60 and GDSC is reported, supporting drug reproducibility across databases, repositioning of bisacodyl and acetalax for triple negative breast cancer, and identifying novel drug response determinants and genomic signatures for topoisomerase inhibitors and schweinfurthins in development. CellMinerCDB also allowed the identification of LIX1L as a novel mesenchymal gene regulating cellular migration and invasiveness.

HIV-1 Integrase-Targeted Short Peptides Derived from a Viral Protein R Sequence.

Abstract: HIV-1 integrase (IN) inhibitors represent a new class of highly effective anti-AIDS therapeutics. Current FDA-approved IN strand transfer inhibitors (INSTIs) share a common mechanism of action that involves chelation of catalytic divalent metal ions. However, the emergence of IN mutants having reduced sensitivity to these inhibitors underlies efforts to derive agents that antagonize IN function by alternate mechanisms. Integrase along with the 96-residue multifunctional accessory protein, viral protein R (Vpr), are both components of the HIV-1 pre-integration complex (PIC). Coordinated interactions within the PIC are important for viral replication. Herein, we report a 7-mer peptide based on the shortened Vpr (69–75) sequence containing a biotin group and a photo-reactive benzoylphenylalanyl residue, and which exhibits low micromolar IN inhibitory potency. Photo-crosslinking experiments have indicated that the peptide directly binds IN. The peptide does not interfere with IN-DNA interactions or induce higher-order, aberrant IN multimerization, suggesting a mode of action for the peptide that is distinct from clinically used INSTIs and developmental allosteric IN inhibitors. This compact Vpr-derived peptide may serve as a valuable pharmacological tool to identify a potential new pharmacologic site.