PubMed

Recent Publications

CX-5461 is a DNA G-quadruplex stabilizer with selective lethality in BRCA1/2 deficient tumours.

CX-5461 is a DNA G-quadruplex stabilizer with selective lethality in BRCA1/2 deficient tumours.

Nat Commun. 2017 Feb 17;8:14432

Authors: Xu H, Di Antonio M, McKinney S, Mathew V, Ho B, O'Neil NJ, Santos ND, Silvester J, Wei V, Garcia J, Kabeer F, Lai D, Soriano P, Banáth J, Chiu DS, Yap D, Le DD, Ye FB, Zhang A, Thu K, Soong J, Lin SC, Tsai AH, Osako T, Algara T, Saunders DN, Wong J, Xian J, Bally MB, Brenton JD, Brown GW, Shah SP, Cescon D, Mak TW, Caldas C, Stirling PC, Hieter P, Balasubramanian S, Aparicio S

Abstract
G-quadruplex DNAs form four-stranded helical structures and are proposed to play key roles in different cellular processes. Targeting G-quadruplex DNAs for cancer treatment is a very promising prospect. Here, we show that CX-5461 is a G-quadruplex stabilizer, with specific toxicity against BRCA deficiencies in cancer cells and polyclonal patient-derived xenograft models, including tumours resistant to PARP inhibition. Exposure to CX-5461, and its related drug CX-3543, blocks replication forks and induces ssDNA gaps or breaks. The BRCA and NHEJ pathways are required for the repair of CX-5461 and CX-3543-induced DNA damage and failure to do so leads to lethality. These data strengthen the concept of G4 targeting as a therapeutic approach, specifically for targeting HR and NHEJ deficient cancers and other tumours deficient for DNA damage repair. CX-5461 is now in advanced phase I clinical trial for patients with BRCA1/2 deficient tumours (Canadian trial, NCT02719977, opened May 2016).

PMID: 28211448 [PubMed - in process]



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Stem cells in the adult CNS revealed: examining their regulation by myelin basic protein.

Stem cells in the adult CNS revealed: examining their regulation by myelin basic protein.

Neural Regen Res. 2016 Dec;11(12):1916-1917

Authors: Xu W, Lakshman N, Morshead CM

PMID: 28197181 [PubMed - in process]



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LUMIER: A Discovery Tool for Mammalian Protein Interaction Networks.

LUMIER: A Discovery Tool for Mammalian Protein Interaction Networks.

Methods Mol Biol. 2017;1550:137-148

Authors: Barrios-Rodiles M, Ellis JD, Blencowe BJ, Wrana JL

Abstract
Protein-protein interactions (PPIs) play an essential role in all biological processes. In vivo, PPIs occur dynamically and depend on extracellular cues. To discover novel protein-protein interactions in mammalian cells, we developed a high-throughput automated technology called LUMIER (LUminescence-based Mammalian IntERactome). In this approach, we co-express a Luciferase (LUC)-tagged fusion protein along with a Flag-tagged protein in an efficiently transfectable cell line such as HEK-293T cells. The interaction between the two proteins is determined by co-immunoprecipitation using an anti-Flag antibody, and the presence of the LUC-tagged interactor in the complex is subsequently detected via its luciferase activity. LUMIER can easily detect transmembrane protein partners, interactions that are signaling- or splice isoform-dependent, as well as those that may occur only in the presence of posttranslational modifications. Using various collections of Flag-tagged proteins, we have generated protein interaction networks for several TGF-β family receptors, Wnt pathway members, and have systematically analyzed the effect of neural-specific alternative splicing on protein interaction networks. The results have provided important insights into the physiological and functional relevance of some of the novel interactions found. LUMIER is highly scalable and can be used for both low- and high-throughput strategies. LUMIER is thus a valuable tool for the identification and characterization of dynamically regulated PPIs in mammalian systems. Here, we describe a manual version of LUMIER in a 96-well format that can be easily implemented in any laboratory.

PMID: 28188528 [PubMed - in process]



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SMYD2 lysine methyltransferase regulates leukemia cell growth and regeneration after genotoxic stress.

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SMYD2 lysine methyltransferase regulates leukemia cell growth and regeneration after genotoxic stress.

Oncotarget. 2017 Feb 06;:

Authors: Zipin-Roitman A, Aqaqe N, Yassin M, Biechonski S, Amar M, van Delft MF, Gan OI, McDermott SP, Buzina A, Ketela T, Shlush L, Xie S, Voisin V, Moffat J, Minden MD, Dick JE, Milyavsky M

Abstract
The molecular determinants governing escape of Acute Myeloid Leukemia (AML) cells from DNA damaging therapy remain poorly defined and account for therapy failures. To isolate genes responsible for leukemia cells regeneration following multiple challenges with irradiation we performed a genome-wide shRNA screen. Some of the isolated hits are known players in the DNA damage response (e.g. p53, CHK2), whereas other, e.g. SMYD2 lysine methyltransferase (KMT), remains uncharacterized in the AML context. Here we report that SMYD2 knockdown confers relative resistance to human AML cells against multiple classes of DNA damaging agents. Induction of the transient quiescence state upon SMYD2 downregulation correlated with the resistance. We revealed that diminished SMYD2 expression resulted in the upregulation of the related methyltransferase SET7/9, suggesting compensatory relationships. Indeed, pharmacological targeting of SET7/9 with (R)-PFI2 inhibitor preferentially inhibited the growth of cells expressing low levels of SMYD2.Finally, decreased expression of SMYD2 in AML patients correlated with the reduced sensitivity to therapy and lower probability to achieve complete remission. We propose that the interplay between SMYD2 and SET7/9 levels shifts leukemia cells from growth to quiescence state that is associated with the higher resistance to DNA damaging agents and rationalize SET7/9 pharmacological targeting in AML.

PMID: 28187429 [PubMed - as supplied by publisher]



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Fc Engineering for Developing Therapeutic Bispecific Antibodies and Novel Scaffolds.

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Fc Engineering for Developing Therapeutic Bispecific Antibodies and Novel Scaffolds.

Front Immunol. 2017;8:38

Authors: Liu H, Saxena A, Sidhu SS, Wu D

Abstract
Therapeutic monoclonal antibodies have become molecules of choice to treat autoimmune disorders, inflammatory diseases, and cancer. Moreover, bispecific/multispecific antibodies that target more than one antigen or epitope on a target cell or recruit effector cells (T cell, natural killer cell, or macrophage cell) toward target cells have shown great potential to maximize the benefits of antibody therapy. In the past decade, many novel concepts to generate bispecific and multispecific antibodies have evolved successfully into a range of formats from full bispecific immunoglobulin gammas to antibody fragments. Impressively, antibody fragments such as bispecific T-cell engager, bispecific killer cell engager, trispecific killer cell engager, tandem diabody, and dual-affinity-retargeting are showing exciting results in terms of recruiting and activating self-immune effector cells to target and lyse tumor cells. Promisingly, crystallizable fragment (Fc) antigen-binding fragment and monomeric antibody or half antibody may be particularly advantageous to target solid tumors owing to their small size and thus good tissue penetration potential while, on the other hand, keeping Fc-related effector functions such as antibody-dependent cellular cytotoxicity, complement-dependent cytotoxicity, antibody-dependent cell-mediated phagocytosis, and extended serum half-life via interaction with neonatal Fc receptor. This review, therefore, focuses on the progress of Fc engineering in generating bispecific molecules and on the use of small antibody fragment as scaffolds for therapeutic development.

PMID: 28184223 [PubMed - in process]



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MARK4 inhibits Hippo signaling to promote proliferation and migration of breast cancer cells.

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MARK4 inhibits Hippo signaling to promote proliferation and migration of breast cancer cells.

EMBO Rep. 2017 Feb 09;:

Authors: Heidary Arash E, Shiban A, Song S, Attisano L

Abstract
The Hippo pathway is a critical regulator of tissue size, and aberrations in pathway regulation lead to cancer. MST1/2 and LATS1/2 kinases comprise the core of the pathway that, in association with adaptor proteins SAV and MOB, functions in a sequential manner to phosphorylate and inhibit the transcription factors YAP and TAZ. Here we identify mammalian MARK family members as activators of YAP/TAZ. We show that depletion of MARK4 in MDA-MB-231 breast cancer cells results in the loss of nuclear YAP/TAZ and decreases the expression of YAP/TAZ targets. We demonstrate that MARK4 can bind to MST and SAV, leading to their phosphorylation, and that MARK4 expression attenuates the formation of a complex between MST/SAV and LATS, which depends on the kinase activity of MARK4. Abrogation of MARK4 expression using siRNAs and CRISPR/Cas9 gene editing attenuates the proliferation and migration of MDA-MB-231 cells. Our results show that MARK4 acts as a negative regulator of the Hippo kinase cassette to promote YAP/TAZ activity and that loss of MARK4 restrains the tumorigenic properties of breast cancer cells.

PMID: 28183853 [PubMed - as supplied by publisher]



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Integrated analysis of proteome, phosphotyrosine-proteome, tyrosine-kinome and tyrosine-phosphatome in acute myeloid leukemia.

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Integrated analysis of proteome, phosphotyrosine-proteome, tyrosine-kinome and tyrosine-phosphatome in acute myeloid leukemia.

Proteomics. 2017 Feb 07;:

Authors: Tong J, Helmy M, Cavalli FM, Jin L, St-Germain J, Karisch R, Taylor P, Minden MD, Taylor MD, Neel BG, Bader GD, Moran MF

Abstract
Reversible protein-tyrosine phosphorylation is catalyzed by the antagonistic actions of protein-tyrosine kinases (PTKs) and phosphatases (PTPs), and represents a major form of cell regulation. Acute myeloid leukemia (AML) is an aggressive hematological malignancy that results from the acquisition of multiple genetic alterations, which in some instances are associated with deregulated protein-phosphotyrosine (pY)-mediated signaling networks. However, although individual PTKs and PTPs have been linked to AML and other malignancies, analysis of protein-pY networks as a function of activated PTKs and PTPs has not been done. In this study, mass spectrometry was used to characterize AML proteomes, and phospho-proteome-subsets including pY proteins, PTKs, and PTPs. AML proteomes resolved into two groups related to high or low degrees of maturation according to French-American-British (FAB) classification, and reflecting differential expression of cell surface antigens. AML pY proteomes reflect canonical, spatially organized signaling networks, unrelated to maturation, with heterogeneous expression of activated receptor and non-receptor PTKs. We present the first integrated analysis of the pY-proteome, activated PTKs, and PTPs. Every PTP and most PTKs have both positive and negative associations with the pY-proteome. pY proteins resolve into groups with shared PTK and PTP correlations. These findings highlight the importance of pY turnover and the PTP phosphatome in shaping the pY-proteome in AML. This article is protected by copyright. All rights reserved.

PMID: 28176486 [PubMed - as supplied by publisher]



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Discovery and structure activity relationship of the first potent cryptosporidium FIKK kinase inhibitor.

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Discovery and structure activity relationship of the first potent cryptosporidium FIKK kinase inhibitor.

Bioorg Med Chem. 2017 Jan 24;:

Authors: Osman KT, Ye J, Shi Z, Toker C, Lovato D, Jumani RS, Zuercher W, Huston CD, Edwards AM, Lautens M, Santhakumar V, Hui R

Abstract
FIKKs are parasite-specific protein kinases with distinctive sequence motifs and their biological roles have not been completely elucidated. Here, we report the first potent Cryptosporidium FIKK (CpFIKK) inhibitor. We identified 4b as a potent (IC50=0.2nM) inhibitor of CpFIKK catalytic activity. In addition, we identified both CpCDPK1 selective as well as dually acting CpFIKK-CDPK1 inhibitors from the same structural class of compounds. We evaluated these CpFIKK inhibitors for inhibition of parasite growth in vitro. The observed effects on parasite growth did not correlate with CpFIKK inhibition, suggesting that CpFIKK may not be involved in parasite growth.

PMID: 28162900 [PubMed - as supplied by publisher]



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Preclinical evaluation of taxane-binding peptide-modified polymeric micelles loaded with docetaxel in an orthotopic breast cancer mouse model.

Preclinical evaluation of taxane-binding peptide-modified polymeric micelles loaded with docetaxel in an orthotopic breast cancer mouse model.

Biomaterials. 2017 Jan 26;123:39-47

Authors: Logie J, Ganesh AN, Aman AM, Al-Awar RS, Shoichet MS

Abstract
We developed a novel taxane-binding peptide (TBP) modified, biodegradable polymeric micelle that overcomes limitations of drug loading and poor serum stability typically seen with particle delivery, leading to enhanced pharmacokinetics and tumor distribution of docetaxel (DTX). The use of the taxane-binding peptide to increase docetaxel loading is particularly compelling as it takes advantage of a known intracellular binding mechanism in a new way. Docetaxel is a potent chemotherapeutic with a therapeutic index often limited by the toxicity of the excipients that are necessary to enhance its solubility for intravenous delivery. Our polymeric micelle has terminal furan groups that enable facile antibody Fab conjugation by Diels-Alder chemistry for targeted delivery. Compared to the conventional ethanolic polysorbate 80 formulation (Free DTX), our nanoparticle (NP DTX) formulation exhibited a two-fold increase in exposure and tumor accumulation. Notably, the reduced toxicity of the NP DTX formulation increased the therapeutic index and allowed for higher dosing regimens, with a maximum tolerated dose (MTD) 1.6-fold higher than that of the Free DTX formulation, which is significant and similar to enhancements observed in clinical products for docetaxel and other drugs. These improved properties led to enhanced mouse survival in an orthotopic model of breast cancer; however, the targeted formulation of Fab-NP DTX did not further improve efficacy. Together, these results clearly demonstrate the benefits of the TBP-modified polymeric micelles as promising carriers for docetaxel.

PMID: 28161682 [PubMed - as supplied by publisher]



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Multilayered Control of Alternative Splicing Regulatory Networks by Transcription Factors.

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Multilayered Control of Alternative Splicing Regulatory Networks by Transcription Factors.

Mol Cell. 2017 Feb 02;65(3):539-553.e7

Authors: Han H, Braunschweig U, Gonatopoulos-Pournatzis T, Weatheritt RJ, Hirsch CL, Ha KC, Radovani E, Nabeel-Shah S, Sterne-Weiler T, Wang J, O'Hanlon D, Pan Q, Ray D, Zheng H, Vizeacoumar F, Datti A, Magomedova L, Cummins CL, Hughes TR, Greenblatt JF, Wrana JL, Moffat J, Blencowe BJ

Abstract
Networks of coordinated alternative splicing (AS) events play critical roles in development and disease. However, a comprehensive knowledge of the factors that regulate these networks is lacking. We describe a high-throughput system for systematically linking trans-acting factors to endogenous RNA regulatory events. Using this system, we identify hundreds of factors associated with diverse regulatory layers that positively or negatively control AS events linked to cell fate. Remarkably, more than one-third of the regulators are transcription factors. Further analyses of the zinc finger protein Zfp871 and BTB/POZ domain transcription factor Nacc1, which regulate neural and stem cell AS programs, respectively, reveal roles in controlling the expression of specific splicing regulators. Surprisingly, these proteins also appear to regulate target AS programs via binding RNA. Our results thus uncover a large "missing cache" of splicing regulators among annotated transcription factors, some of which dually regulate AS through direct and indirect mechanisms.

PMID: 28157508 [PubMed - in process]



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