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Publications (23)

  • Matthew Tcheng · Ismael Samudio · Eric A. Lee · [...] · Paul A. Spagnuolo
    Article · Aug 2016 · Leukemia and Lymphoma
  • L. Angka · P.A. Spagnuolo
    [Show abstract] [Hide abstract] ABSTRACT: PURPOSE: Concomitant use of nutraceuticals with chemotherapy is very common. Cancer patients self-medicate to relieve the side effects associated with chemotherapy, improve disease outcome and to regain control of their medical care. However, there is limited empirical evidence on potential drug-nutraceutical interactions and their resulting effect on chemotherapy efficacy. METHOD: To investigate drug-nutraceutical interactions we created and screened a library of commonly used nutraceuticals for their modulatory effects on the activity of cytarabine and daunorubicin, two primary chemotherapeutics used to treat acute myeloid leukemia (AML). Combination screening was performed in 3 AML cell lines (OCI-AML2, KG1a and U937) using the MTS viability assay. Lead compounds were validated using with the Annexin V/ Propidium iodide assay and CalcuSyn drug combination software. RESULTS: We identified zinc as a nutraceutical that enhanced AML chemotherapy efficacy with combination index (CI) values of 0.649, 0.632 and 0.615 at EC 25, 50 and 75, respectively; CI values <0.9, >1.1 or between 0.9-1.1 denote statistical synergy, antagonism or additivity, respectively. In contrast, we show that echinacea hindered AML chemotherapy efficacy by significantly reducing the ability of cytarabine to induce cell death. CONCLUSION: Given the positive and negative effects of nutraceuticals, patients undergoing chemotherapy must consult with their oncologist before consuming over-thecounter supplements. © 2015, Canadian Society for Pharmaceutical Sciences. All rights reserved.
    Article · Aug 2015
  • Eric A Lee · Leonard Angka · Sarah-Grace Rota · [...] · Paul A Spagnuolo
    [Show abstract] [Hide abstract] ABSTRACT: Treatment regimens for acute myeloid leukemia (AML) continue to offer weak clinical outcomes. Through a high-throughput cell-based screen, we identified avocatin B, a lipid derived from avocado fruit, as a novel compound with cytotoxic activity in AML. Avocatin B reduced human primary AML cell viability without effect on normal peripheral blood stem cells. Functional stem cell assays demonstrated selectivity toward AML progenitor and stem cells without effects on normal hematopoietic stem cells. Mechanistic investigations indicated that cytotoxicity relied on mitochondrial localization, as cells lacking functional mitochondria or CPT1, the enzyme that facilitates mitochondria lipid transport, were insensitive to avocatin B. Furthermore, avocatin B inhibited fatty acid oxidation and decreased NADPH levels, resulting in ROS-dependent leukemia cell death characterized by the release of mitochondrial proteins, apoptosis-inducing factor, and cytochrome c. This study reveals a novel strategy for selective leukemia cell eradication based on a specific difference in mitochondrial function. Cancer Res; 75(12); 2478-88. ©2015 AACR. ©2015 American Association for Cancer Research.
    Article · Jun 2015 · Cancer Research
  • [Show abstract] [Hide abstract] ABSTRACT: Mitochondria contain multiple copies of their own 16.6 kb circular genome. To explore the impact of mitochondrial DNA (mtDNA) damage on mitochondrial (mt) function and viability of AML cells, we screened a panel of DNA damaging chemotherapeutic agents to identify drugs that could damage mtDNA. We identified bleomycin as an agent that damaged mtDNA in AML cells at concentrations that induced cell death. Bleomycin also induced mtDNA damage in primary AML samples. Consistent with the observed mtDNA damage, bleomycin reduced mt mass and basal oxygen consumption in AML cells. We also demonstrated that the observed mtDNA damage was functionally important for bleomycin-induced cell death. Finally, bleomycin delayed tumor growth in xenograft mouse models of AML and anti-leukemic concentrations of the drug induced mtDNA damage in AML cells preferentially over normal lung tissue. Taken together, mtDNA-targeted therapy may be an effective strategy to target AML cells and bleomycin could be useful in the treatment of this disease.
    Article · Mar 2015 · Apoptosis
  • L. Angka · P.A. Spagnuolo
    [Show abstract] [Hide abstract] ABSTRACT: Nutraceuticals are food-derived bioactive compounds with anti-cancer treatment potential. From a vitamin A metabolite (All-trans-retinoic acid) for acute promyelocytic leukemia to an edible mushroom extract (maitake) in Phase 2 clinical trial for myelodysplastic syndromes, nutraceuticals are found at all stages of clinical development for treatment of various hematological malignancies. Here, we report the advances in nutraceutical discovery and therapeutic relevance for various hematological malignancies.
    Article · Mar 2015
  • [Show abstract] [Hide abstract] ABSTRACT: Background Recurrence of colorectal cancer (CRC) may arise due to the persistence of drug-resistant and cancer-initiating cells that survive exposure to chemotherapy. Proteins responsible for this recurrence include the chemokine receptor CXCR4, which is known to enable CRC metastasis, as well as the cancer-initiating cell marker and peptidase CD26, which terminates activity of its chemokine CXCL12. Methods We evaluated the expression and function of CXCR4 and CD26 in colon cancer cell lines and xenografts following treatment with common chemotherapies using radioligand binding, flow cytometry, immunofluorescence, and enzymatic assays. Results 5-Fluorouracil, oxaliplatin and SN-38 (the active metabolite of irinotecan), as well as cisplatin, methotrexate and vinblastine, each caused decreases in cell-surface CXCR4 and concomitant increases in CD26 on HT-29, T84, HRT-18, SW480 and SW620 CRC cell lines. Flow cytometry indicated that the decline in CXCR4 was associated with a significant loss of CXCR4+/CD26- cells. Elevations in CD26 were paralleled by increases in both the intrinsic dipeptidyl peptidase activity of CD26 as well as its capacity to bind extracellular adenosine deaminase. Orthotopic HT-29 xenografts treated with standard CRC chemotherapeutics 5-fluorouracil, irinotecan, or oxaliplatin showed dramatic increases in CD26 compared to untreated tumors. Consistent with the loss of CXCR4 and gain in CD26, migratory responses to exogenous CXCL12 were eliminated in cells pretreated with cytotoxic agents, although cells retained basal motility. Analysis of cancer-initiating cell CD44 and CD133 subsets revealed drug-dependent responses of CD26/CD44/CD133 populations, suggesting that the benefits of combining standard chemotherapies 5-fluoruracil and oxaliplatin may be derived from their complementary elimination of cell populations. Conclusion Our results indicate that conventional anticancer agents may act to inhibit chemokine-mediated migration through eradication of CXCR4+ cells and attenuation of chemokine gradients through elevation of CD26 activity.
    Article · Feb 2015 · Clinical Cancer Research
  • Eric A. Lee · Leonard Angka · Sarah-Grace Rota · [...] · Paul A. Spagnuolo
    Conference Paper · Dec 2014
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    [Show abstract] [Hide abstract] ABSTRACT: Evaluate the physiochemical characterization of 16-3-16/pluronic-based systems either used with DOPE or not by size and zeta-potential measurements.  Optimize OVCAR-3 cell culture, transfection, sample collection, and detection methods.  Evaluate the relationship between the physicochemical properties of Pluronic delivery systems and their efficiency in vitro. 1. Particle Size and Zeta Potential: in the absence of DOPE, the particle size of all transfection complexes was generally below 200 nm, except for Pluronics F127 and L44. ZP was positive for 0.1 CMC and neutral for 1 CMC and 2 CMC. These results indicate that GS/Pluronic based systems undergo strong compaction and condensation with DNA molecules. 2. Transfection efficiency: F87 and F108 at ratio 1:5 obtained the highest transfection efficiency comparing to 1:2 and 1:5, but further below the positive control lipofectamine 2000®. 3. Cell Viability: cell Viability of ratio 1:5 was better than ratio 1:10 and the positive control lipofectamine 2000®. • Size and zeta Potential: particle size and zeta potential measurements of the transfection GS/Pluronic-based nanoparticles as well as the transfection GS/Pluronic-based lipid vesicles were measured using Malvern Zetasizer Nano ZS instrument. • Transfection efficiency and Cell viability: three concentrations of Pluronic (0.1 CMC, 1 CMC, and 2 CMC) were used in combination with 1.5 mM GS and DNA at three ratios (1:2, 1:5, and 1:10) of pVG.telRL DNA to GS. EGFP expression and PI staining were quantified using flow cytometry. 50,000 OVCAR-3 human ovarian cancer cells per sample were analyzed using aguava easyCyte™ Flow Cytometer
    Full-text Conference Paper · Nov 2014
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    [Show abstract] [Hide abstract] ABSTRACT: Conventional plasmid DNA vectors play a significant role in gene therapy, but they also have considerable limitations: they can elicit adverse immune responses because of bacterial sequences they contain for maintenance and amplification in prokaryotes, their bioavailability is compromised because of their large molecular size, and they may be genotoxic. We constructed an in vivo platform to produce ministring DNA-mini linear covalently closed DNA vectors-that are devoid of unwanted bacterial sequences and encode only the gene(s) of interest and necessary eukaryotic expression elements. Transfection of rapidly and slowly dividing human cells with ministring DNA coding for enhanced green fluorescent protein resulted in significantly improved transfection, bioavailability, and cytoplasmic kinetics compared with parental plasmid precursors and isogenic circular covalently closed DNA counterparts. Ministring DNA that integrated into the genome of human cells caused chromosomal disruption and apoptotic death of possibly oncogenic vector integrants; thus, they may be safer than plasmid and circular DNA vectors.
    Full-text Article · May 2014 · Molecular Therapy - Nucleic Acids
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    [Show abstract] [Hide abstract] ABSTRACT: Genome-wide characterization of the in vivo cellular response to perturbation is fundamental to understanding how cells survive stress. Identifying the proteins and pathways perturbed by small molecules affects biology and medicine by revealing the mechanisms of drug action. We used a yeast chemogenomics platform that quantifies the requirement for each gene for resistance to a compound in vivo to profile 3250 small molecules in a systematic and unbiased manner. We identified 317 compounds that specifically perturb the function of 121 genes and characterized the mechanism of specific compounds. Global analysis revealed that the cellular response to small molecules is limited and described by a network of 45 major chemogenomic signatures. Our results provide a resource for the discovery of functional interactions among genes, chemicals, and biological processes.
    Full-text Article · Apr 2014 · Science
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    Leonard Angka · Eric A Lee · Sarah G Rota · [...] · Paul A Spagnuolo
    [Show abstract] [Hide abstract] ABSTRACT: To identify novel anti-cancer agents, we created and screened a unique nutraceutical library for activity against acute myeloid leukemia (AML) cells. From this screen, we determined that glucopsychosine was selectively toxic toward AML cell lines and primary AML patient samples with no effect toward normal hematopoietic cells. It delayed tumor growth and reduced tumor weights in mouse xenografts models without imparting toxicity. Glucopsychosine increased cytosolic calcium and induced apoptosis through calpain enzymes. Extracellular calcium was functionally important for glucopsychosine-induced AML cell death and surface calcium channel expression is altered in AML cells highlighting a unique mechanism of glucopsychosine's selectivity.
    Full-text Article · Mar 2014 · Cancer letters
  • [Show abstract] [Hide abstract] ABSTRACT: The signal transducer and activator of transcription (STAT) proteins represent a family of cytoplasmic transcription factors that regulate a pleiotropic range of biological processes. In particular, Stat3 protein has received particular attention as it regulates the expression of genes involved in a variety of malignant processes including proliferation, survival, migration and drug resistance. Multiple myeloma (MM) is an incurable haematologic malignancy that often exhibits abnormally high levels of Stat3 activity. Although current treatment strategies can improve the clinical management of MM, it remains uniformly incurable with a dismal median survival time post-treatment of 3-4 years. Thus, novel targeted therapeutics are critically needed to improve MM patient outcomes. We herein report the development of a series of small molecule Stat3 inhibitors with potent anti-MM activity in vitro. These compounds showed high-affinity binding to Stat3's SH2 domain, inhibited intracellular Stat3 phosphorylation, and induced apoptosis in MM cell lines at low micromolar concentrations.
    Article · Aug 2013 · Journal of Medicinal Chemistry
  • [Show abstract] [Hide abstract] ABSTRACT: A focused library of hetero-trisubstituted purines was developed for improving the cell penetrating and biological efficacy of a series of anti-Stat3 protein inhibitors. From this SAR study, lead agent 22e was identified as being a promising inhibitor of MM tumour cells (IC50's <5μM). Surprisingly, biophysical and biochemical characterization proved that 22e was not a Stat3 inhibitor. Initial screening against the kinome, prompted by the purine scaffold's history for targeting ATP binding pockets, suggests possible targeting of the JAK family kinases, as well for ABL1 (nonphosphorylated F317L) and AAK1.
    Article · May 2013 · Bioorganic & medicinal chemistry
  • P A Spagnuolo · R Hurren · M Gronda · [...] · A D Schimmer
    [Show abstract] [Hide abstract] ABSTRACT: Parthenolide is selectively toxic to leukemia cells; however, it also activates cell protective responses which may limit its clinical application. Therefore, we sought to identify agents that synergistically enhance parthenolide's cytotoxicity. Using a high-throughput combination drug screen, we identified the hypoglycemic, vildagliptin, which synergized with parthenolide to induce death of the leukemia stem cell line, TEX (combination index (CI) = 0.36 and 0.16, at EC 50 and 80, respectively; where CI < 1 denotes statistical synergy). The combination of parthenolide and vildagliptin reduced the viability and clonogenic growth of cells from acute myeloid leukemia patients and had limited effects on the viability of normal human peripheral blood stem cells. The basis for synergy was independent of vildagliptin's primary action as an inhibitor of dipeptidyl peptidase (DPP) IV. Rather, using chemical and genetic approaches we demonstrated that the synergy was due to inhibition of the related enzymes DPP8 and 9. In summary, these results highlight DPP8 and 9 inhibition as a novel chemosensitizing strategy in leukemia cells. Moreover, these results suggest that the combination of vildagliptin and parthenolide could be useful for the treatment of leukemia.Leukemia accepted article preview online, 15 January 2013; doi:10.1038/leu.2013.9.
    Article · Jan 2013 · Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K
  • [Show abstract] [Hide abstract] ABSTRACT: Despite efforts to understand and treat acute myeloid leukemia (AML), there remains a need for more comprehensive therapies to prevent AML-associated relapses. To identify new therapeutic strategies for AML, we screened a library of on- and off-patent drugs and identified the antimalarial agent mefloquine as a compound that selectively kills AML cells and AML stem cells in a panel of leukemia cell lines and in mice. Using a yeast genome-wide functional screen for mefloquine sensitizers, we identified genes associated with the yeast vacuole, the homolog of the mammalian lysosome. Consistent with this, we determined that mefloquine disrupts lysosomes, directly permeabilizes the lysosome membrane, and releases cathepsins into the cytosol. Knockdown of the lysosomal membrane proteins LAMP1 and LAMP2 resulted in decreased cell viability, as did treatment of AML cells with known lysosome disrupters. Highlighting a potential therapeutic rationale for this strategy, leukemic cells had significantly larger lysosomes compared with normal cells, and leukemia-initiating cells overexpressed lysosomal biogenesis genes. These results demonstrate that lysosomal disruption preferentially targets AML cells and AML progenitor cells, providing a rationale for testing lysosomal disruption as a novel therapeutic strategy for AML.
    Article · Dec 2012 · The Journal of clinical investigation
  • [Show abstract] [Hide abstract] ABSTRACT: Src Homology 2 (SH2) domains are the paradigm of phosphotyrosine (pY) protein recognition modules and mediate numerous cancer-promoting protein-protein complexes. Effective SH2 domain mimicry with pY-binding coordination complexes offers a promising route to new and selective disruptors of pY-mediated protein-protein interactions. We herein report the synthesis and in vitro characterization of a library of coordination complex SH2 domain proteomimetics. Compounds were designed to interact with phosphopeptides via a two-point interaction, principally with pY, and to make secondary interactions with pY+2/3, thereby achieving sequence-selective discrimination. Here, we report that lead mimetics demonstrated high target phosphopeptide affinity (K(a) ∼ 10(7) M(-1)) and selectivity. In addition, biological screening in various tumor cells for anticancer effects showed a high degree of variability in cytotoxicity among receptors, which supported the proposed two-point binding mode. Several receptors potently disrupted cancer cell viability in breast cancer, prostate cancer, and acute myeloid leukemia cell lines.
    Article · Jul 2012 · Inorganic Chemistry
  • Conference Paper · Nov 2011
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    [Show abstract] [Hide abstract] ABSTRACT: To identify FDA-approved agents targeting leukemic cells, we performed a chemical screen on two human leukemic cell lines and identified the antimicrobial tigecycline. A genome-wide screen in yeast identified mitochondrial translation inhibition as the mechanism of tigecycline-mediated lethality. Tigecycline selectively killed leukemia stem and progenitor cells compared to their normal counterparts and also showed antileukemic activity in mouse models of human leukemia. ShRNA-mediated knockdown of EF-Tu mitochondrial translation factor in leukemic cells reproduced the antileukemia activity of tigecycline. These effects were derivative of mitochondrial biogenesis that, together with an increased basal oxygen consumption, proved to be enhanced in AML versus normal hematopoietic cells and were also important for their difference in tigecycline sensitivity.
    Full-text Article · Nov 2011 · Cancer cell
  • Article · Nov 2011 · Molecular Cancer Therapeutics
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    Mahadeo A Sukhai · Paul A Spagnuolo · Scott Weir · [...] · Aaron D Schimmer
    [Show abstract] [Hide abstract] ABSTRACT: Advancing novel therapeutic agents for the treatment of malignancy into the marketplace is an increasingly costly and lengthy process. As such, new strategies for drug discovery are needed. Drug repurposing represents an opportunity to rapidly advance new therapeutic strategies into clinical trials at a relatively low cost. Known on-patent or off-patent drugs with unrecognized anticancer activity can be rapidly advanced into clinical testing for this new indication by leveraging their known pharmacology, pharmacokinetics, and toxicology. Using this approach, academic groups can participate in the drug discovery field and smaller biotechnology companies can "de-risk" early-stage drug discovery projects. Here, several scientific approaches used to identify drug repurposing opportunities are highlighted, with a focus on hematologic malignancies. In addition, a discussion of the regulatory issues that are unique to drug repurposing and how they impact developing old drugs for new indications is included. Finally, the mechanisms to enhance drug repurposing through increased collaborations between academia, industry, and nonprofit charitable organizations are discussed.
    Full-text Article · Jun 2011 · Blood