Bruce A Sullenger

Duke University, Durham, North Carolina, United States

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Publications (142)1199.95 Total impact

  • Erin E Soule · Kristin M Bompiani · Rebecca S Woodruff · Bruce A Sullenger ·
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    ABSTRACT: Potent and rapid-onset anticoagulation is required for several clinical settings, including cardiopulmonary bypass surgery. In addition, because anticoagulation is associated with increased bleeding following surgery, the ability to rapidly reverse such robust anticoagulation is also important. Previously, we observed that no single aptamer was as potent as heparin for anticoagulating blood. However, we discovered that combinations of two aptamers were as potent as heparin. Herein, we sought to combine two individual anticoagulant aptamers into a single bivalent RNA molecule in an effort to generate a single molecule that retained the potent anticoagulant activity of the combination of individual aptamers. We created four bivalent aptamers that can inhibit Factor X/Xa and prothrombin/thrombin and anticoagulate plasma, as well as the combination of individual aptamers. Detailed characterization of the shortest bivalent aptamer indicates that each aptamer retains full binding and functional activity when presented in the bivalent context. Finally, reversal of this bivalent aptamer with a single antidote was explored, and anticoagulant activity could be rapidly turned off in a dose-dependent manner. These studies demonstrate that bivalent anticoagulant aptamers represent a novel and potent approach to actively and reversibly control coagulation.
    11/2015; DOI:10.1089/nat.2015.0565
  • Rebecca S Woodruff · Bruce A Sullenger ·
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    ABSTRACT: As a novel class of therapeutics, aptamers, or nucleic acid ligands, have garnered clinical interest because of the ease of isolating a highly specific aptamer against a wide range of targets, their chemical flexibility and synthesis, and their inherent ability to have their inhibitory ability reversed. The following review details the development and molecular mechanisms of aptamers targeting specific proteases in the coagulation cascade. The ability of these anticoagulant aptamers to bind to and inhibit exosite function rather than binding within the active site highlights the importance of exosites in blocking protein function. As both exosite inhibitors and reversible agents, the use of aptamers is a promising strategy for future therapeutics. © 2015 American Heart Association, Inc.
    Arteriosclerosis Thrombosis and Vascular Biology 08/2015; 35(10). DOI:10.1161/ATVBAHA.115.300131 · 6.00 Impact Factor
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    ABSTRACT: Cardiovascular disease is the leading cause of death in the United States. Heart failure is a common, costly, and potentially fatal condition that is inadequately managed by pharmaceuticals. Cardiac repair therapies are promising alternative options. A potential cardiac repair therapy involves reprogramming human fibroblasts toward an induced cardiac progenitor-like state. We developed a clinically useful and safer reprogramming method by nonintegrative delivery of a cocktail of cardiac transcription factor-encoding mRNAs into autologous human dermal fibroblasts obtained from skin biopsies. Using this method, adult and neonatal dermal fibroblasts were reprogrammed into cardiac progenitor cells (CPCs) that expressed c-kit, Isl-1, and Nkx2.5. Furthermore, these reprogrammed CPCs differentiated into cardiomyocytes (CMs) in vitro as judged by increased expression of cardiac troponin T, α-sarcomeric actinin, RyR2, and SERCA2 and displayed enhanced caffeine-sensitive calcium release. The ability to reprogram patient-derived dermal fibroblasts into c-kit(+) CPCs and differentiate them into functional CMs provides clinicians with a potential new source of CPCs for cardiac repair from a renewable source and an alternative therapy in the treatment of heart failure.
    Stem Cells and Development 07/2015; DOI:10.1089/scd.2015.0073 · 3.73 Impact Factor
  • Jaewoo Lee · Youngju Lee · Bruce A. Sullenger ·

    Cancer Research 10/2014; 74(19 Supplement):2639-2639. DOI:10.1158/1538-7445.AM2014-2639 · 9.33 Impact Factor
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    ABSTRACT: Background: Pathologic cutaneous scarring affects over 40 million people worldwide and costs billions of dollars annually. Understanding mechanisms of fibroblast activation and granulation tissue contraction is the first step toward preventing pathologic scarring. The authors hypothesize that nucleic acids increase fibroblast activation and cause granulation tissue contraction and that sequestration of nucleic acids by application of a nucleic acid scavenger dendrimer, polyamidoamine third-generation dendrimer, will decrease pathologic scarring. Methods: In vitro experiments were performed to assess the effect of nucleic acids on pathologic scar-associated fibroblast activity. The effect of nucleic acids on cytokine production and migration on mouse fibroblasts was evaluated. Immunofluorescence microscopy was used to determine the effect of nucleic acids on the differentiation of human primary fibroblasts into myofibroblasts. Using a murine model, the effect of polyamidoamine third-generation dendrimer on granulation tissue contraction was evaluated by gross and histologic parameters. Results: Mouse fibroblasts stimulated with nucleic acids had increased cytokine production (i.e., transforming growth factor-β, monocyte chemotactic protein 1, interleukin-10, tumor necrosis factor-α, and interferon-γ), migration, and differentiation into myofibroblasts. Polyamidoamine third-generation dendrimer blocked cytokine production, migration, and differentiation into myofibroblasts. Using a murine model of granulation tissue contraction, polyamidoamine third-generation dendrimer decreased wound contraction and angiogenesis. Collagen deposition in polyamidoamine third-generation dendrimer-treated tissues was aligned more randomly and whorl-like compared with control tissue. Conclusions: The data demonstrate that nucleic acid-stimulated fibroblast activation and granulation tissue contraction are blocked by polyamidoamine third-generation dendrimer. Sequestration of pathogen-associated molecular patterns may be an approach for preventing pathologic scarring.
    Plastic &amp Reconstructive Surgery 09/2014; 134(3):420e-433e. DOI:10.1097/PRS.0000000000000471 · 2.99 Impact Factor
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    ABSTRACT: Coordinated enzymatic reactions regulate blood clot generation. To explore the contributions of various coagulation enzymes in this process, we utilized a panel of aptamers against factors VIIa, IXa, Xa, and prothrombin. Each aptamer dose-dependently inhibited clot formation, yet none was able to completely impede this process in highly procoagulant settings. However, several combinations of two aptamers synergistically impaired clot formation. One extremely potent aptamer combination was able to maintain human blood fluidity even during extracorporeal circulation, a highly procoagulant setting encountered during cardiopulmonary bypass surgery. Moreover, this aptamer cocktail could be rapidly reversed with antidotes to restore normal hemostasis, indicating that even highly potent aptamer combinations can be rapidly controlled. These studies highlight the potential utility of using sets of aptamers to probe the functions of proteins in molecular pathways for research and therapeutic ends.
    Chemistry & Biology 07/2014; 21(8). DOI:10.1016/j.chembiol.2014.05.016 · 6.65 Impact Factor
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    ABSTRACT: β-arrestins, ubiquitous cellular scaffolding proteins that act as signaling mediators of numerous critical cellular pathways, are attractive therapeutic targets because they promote tumorigenesis in several tumor models. However, targeting scaffolding proteins with traditional small molecule drugs has been challenging. Inhibition of β-arrestin 2 with a novel aptamer impedes multiple oncogenic signaling pathways simultaneously. Additionally, delivery of the β-arrestin 2-targeting aptamer into leukemia cells through coupling to a recently described cancer cell-specific delivery aptamer, inhibits multiple β-arrestin-mediated signaling pathways known to be required for chronic myelogenous leukemia (CML) disease progression, and impairs tumorigenic growth in CML patient samples. The ability to target scaffolding proteins such as β-arrestin 2 with RNA aptamers may prove beneficial as a therapeutic strategy. An RNA aptamer inhibits β-arrestin 2 activity.Inhibiting β-arrestin 2 impedes multiple tumorigenic pathways simultaneously.The therapeutic aptamer is delivered to cancer cells using a cell-specific DNA aptamer.Targeting β-arrestin 2 inhibits tumor progression in CML models and patient samples.
    PLoS ONE 04/2014; 9(4):e93441. DOI:10.1371/journal.pone.0093441 · 3.23 Impact Factor

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    Partha Ray · Bruce A Sullenger · Rebekah R White ·
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    ABSTRACT: Posttranslational modifications on proteins can serve as useful biomarkers for disease. However, their discovery and detection in biological fluids is challenging. Aptamers are oligonucleotide ligands that demonstrate high affinity toward their target proteins and can discriminate closely related proteins with superb specificity. Previously, we generated a cyclophilin B aptamer (M9-5) that could discriminate sera from pancreatic cancer patients and healthy volunteers with high specificity and sensitivity. In our present work we further characterize the aptamer and the target protein, cyclophilin B, and demonstrate that the aptamer could discriminate between cyclophilin B expressed in human cells versus bacteria. Using mass-spectrometric analysis, we discovered post-translational modifications on cyclophilin B that might be responsible for the M9-5 selectivity. The ability to distinguish between forms of the same protein with differing post-translational modifications is an important advantage of aptamers as tools for identification and detection of biomarkers.
    10/2013; 23(6). DOI:10.1089/nat.2013.0439
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    Eda K Holl · Kara L Shumansky · George Pitoc · Elizabeth Ramsburg · Bruce A Sullenger ·
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    ABSTRACT: Toll-like receptor (TLR) family members, 3, 7 and 9 are key components in initiation and progression of autoimmune disorders such as systemic lupus erythematosus (SLE). These TLRs are often referred to as nucleic acid-sensing TLRs based on their ability to recognize DNAs or RNAs produced by pathogens or damaged cells. During autoimmune disease progression these receptors recognize self nucleic acids as well as self nucleic acid-containing complexes and contribute to inflammatory cytokine production and subsequent enhancement of serum autoantibody levels. We have recently discovered that nucleic-acid scavenging polymers (NASPs) can neutralize the proinflammatory effects of nucleic acids. Here, we begin to explore what effects such NASPs have on normal immune function. We show that such NASPs can inhibit TLR activation without affecting nucleic acid-independent T cell activation. Moreover, we observe that stimulation of immune cells by encapsulated nucleic acids, such as those found in viral particles, is unaffected by NASPs. Thus NASPs only limit the activation of the immune system by accessible extra-cellular nucleic acid and do not engender non-specific immune suppression. These important findings suggest that NASPs represent a new approach toward anti-inflammatory drug development as these agents can potentially be utilized to block overt autoimmune disorders and inflammation while allowing normal immune responses to occur.
    PLoS ONE 07/2013; 8(7):e69413. DOI:10.1371/journal.pone.0069413 · 3.23 Impact Factor
  • R S Woodruff · Y Xu · J Layzer · W Wu · M L Ogletreee · B A Sullenger ·
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    ABSTRACT: Background: Exposure of the plasma protein factor XII (FXII) to an anionic surface generates activated FXII that not only triggers the intrinsic pathway of blood coagulation through the activation of FXI but also mediates various vascular responses through activation of the plasma contact system. While deficiencies of FXII are not associated with excessive bleeding, thrombosis models in factor-deficient animals have suggested that this protein contributes to stable thrombus formation. Therefore, FXII has emerged as an attractive therapeutic target to treat or prevent pathological thrombosis formation without increasing the risk for hemorrhage. Objectives: Using an in vitro directed evolution and chemical biology approach, we sought to isolate a nuclease-resistant RNA aptamer that binds specifically to FXII and directly inhibits FXII coagulant function. Methods and results: We describe the isolation and characterization of a high-affinity RNA aptamer targeting FXII/activated FXII (FXIIa) that dose dependently prolongs fibrin clot formation and thrombin generation in clinical coagulation assays. This aptamer functions as a potent anticoagulant by inhibiting the autoactivation of FXII, as well as inhibiting intrinsic pathway activation (FXI activation). However, the aptamer does not affect the FXIIa-mediated activation of the proinflammatory kallikrein-kinin system (plasma kallikrein activation). Conclusions: We have generated a specific and potent FXII/FXIIa aptamer anticoagulant that offers targeted inhibition of discrete macromolecular interactions involved in the activation of the intrinsic pathway of blood coagulation.
    Journal of Thrombosis and Haemostasis 05/2013; 11(7). DOI:10.1111/jth.12302 · 5.72 Impact Factor
  • J. E. Bond · E. Holl · M. A. Selim · B. Sullenger · H. Levinson ·

    Wound Repair and Regeneration 03/2013; 21(2):A17-A17. · 2.75 Impact Factor
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    Elizabeth D Pratico · Bruce A Sullenger · Smita K Nair ·
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    ABSTRACT: Induction of an effective immune response that can target and eliminate malignant cells or virus-infected cells requires the stimulation of antigen-specific effector T cells. A productive and long-lasting memory response requires 2 signals: a specific signal provided by antigen recognition through engagement of the T cell receptor and a secondary signal via engagement of costimulatory molecules (such as OX40) on these newly activated T cells. The OX40-OX40-ligand interaction is critical for the generation of productive effector and memory T cell functions. Thus agonistic antibodies that stimulate OX40 on activated T cells have been used as adjuvants to augment immune responses. We previously demonstrated that an aptamer modified to stimulate murine OX40 enhanced vaccine-mediated immune responses in a murine melanoma model. In this study, we describe the development of an agonistic aptamer that targets human OX40 (hOX40). This hOX40 aptamer was isolated using systematic evolution of ligands by exponential enrichment and binds the target purified protein with high affinity [dissociation constants (K(d))<10 nM]. Moreover, the hOX40 aptamer-streptavidin complex has an apparent binding affinity of ∼50 nM for hOX40 on activated T cells as determined by flow cytometry and specifically binds activated human T cells. A multivalent version of the aptamer, but not a mutant version of the aptamer, was able to stimulate OX40 on T cells and enhance cell proliferation and interferon-gamma production. Future studies will assess the therapeutic potential of hOX40 aptamers for ex vivo stimulation of antigen specific T cells in conjunction with dendritic cell-based vaccines for adoptive cellular therapy.
    10/2012; 23(1). DOI:10.1089/nat.2012.0388
  • Anandi Krishnan · Erwin A Vogler · Bruce A Sullenger · Richard C Becker ·
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    ABSTRACT: The anticoagulant properties of a novel RNA aptamer that binds FIXa depend collectively on the intensity of surface contact activation of human blood plasma, aptamer concentration, and its binding affinity for FIXa. Accordingly, anticoagulation efficiency of plasma containing any particular aptamer concentration is low when coagulation is strongly activated by hydrophilic surfaces compared to the anticoagulation efficiency in plasma that is weakly activated by hydrophobic surfaces. Anticoagulation efficiency is lower at hypothermic temperatures possibly because aptamer-FIXa binding decreases with decreasing temperatures. Experimental results demonstrating these trends are qualitatively interpreted in the context of a previously established model of anticoagulation efficiency of thrombin-binding DNA aptamers that exhibit anticoagulation properties similar to the FIXa aptamer. In principle, FIXa aptamer anticoagulants should be more efficient and therefore more clinically useful than thrombin-binding aptamers because aptamer binding to FIXa competes only with FX that is at much lower blood concentration than fibrinogen (FI) that competes with thrombin-binding aptamers. Our findings may have translatable relevance in the application of aptamer anticoagulants for clinical conditions in which blood is in direct contact with non-biological surfaces such as those encountered in cardiopulmonary bypass circuits.
    Journal of Thrombosis and Thrombolysis 10/2012; 35(1). DOI:10.1007/s11239-012-0778-7 · 2.17 Impact Factor
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    ABSTRACT: Gemcitabine is a nucleoside analog that is currently the best available single-agent chemotherapeutic drug for pancreatic cancer. However, efficacy is limited by our inability to deliver sufficient active metabolite into cancer cells without toxic effects on normal tissues. Targeted delivery of gemcitabine into cancer cells could maximize effectiveness and concurrently minimize toxic side effects by reducing uptake into normal cells. Most pancreatic cancers overexpress epidermal growth factor receptor (EGFR), a trans-membrane receptor tyrosine kinase. We utilized a nuclease resistant RNA aptamer that binds and is internalized by EGFR on pancreatic cancer cells to deliver gemcitabine-containing polymers into EGFR-expressing cells and inhibit cell proliferation in vitro. This approach to cell type–specific therapy can be adapted to other targets and to other types of therapeutic cargo.
    10/2012; 22(5-5):295-305. DOI:10.1089/nat.2012.0353
  • E. Holl · B. Sullenger ·
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    ABSTRACT: Introduction Toll-like receptors (TLRs) play a critical role in innate and adaptive immune responses by responding to pathogenic nucleic acids. Moreover, they have recently been shown to play a role in the pathogenesis of autoimmune disorders such as systemic lupus erythematosus (SLE) due to their ability to recognize self-antigens. There has been growing evidence suggesting that blocking of nucleic acid-sending TLRs such as TLR7 and 9 results in ameliorated disease. Although effective in blocking autoimmune disease progression, TLR inhibitors are not ideal given that they interfere with normal responses to pathogens. Our goal is to create compounds that bind nucleic acids prior to their entry into the cell and engagement of TLRs. Here we determined the role of nucleic acid-binding cationic polymers in neutralizing the proinflammatory effects of nucleic acids on a variety of immune cells implicated in autoimmune disease development. Methods B cells and dendritic cells (DCs) of wild type and lupus prone mice were exposed to TLR7 and 9 synthetic agonists. Supernatants were subsequently assessed for nucleic acid-driven proinflammatory cytokine release, plasma cell differentiation and antibody production. Cells were also assessed for their ability to respond to encapsulated viral particles in the presence of nucleic-acid binding cationic polymers. Results Nucleic acid-binding polymers inhibit TLR activation and subsequent cytokine production by both dendritic cells (DCs) and B cells of wild type and lupus prone mice. They also block nucleic acid-driven plasma cell differentiation and antibody production. Additionally, they modulate co-stimulatory cell surface marker expression (CD80, CD86) in stimulated DCs and B cells. Stimulation of immune cells by encapsulated viral particles is unaffected in the presence of polymers. Conclusion These findings provide a new avenue in drug development as nucleic acid-binding agents can potentially be utilized to block overt autoimmune disorders while allowing normal immune responses to occur.
    Cytokine 09/2012; 59(3):499. DOI:10.1016/j.cyto.2012.06.033 · 2.66 Impact Factor
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    ABSTRACT: Development of effective, yet safe, antithrombotic agents has been challenging because such agents increase the propensity of patients to bleed. Recently, naturally occurring polyphosphates such as extracellular DNA, RNA, and inorganic polyphosphates have been shown to activate blood coagulation. In this report, we evaluate the anticoagulant and antithrombotic activity of nucleic acid-binding polymers in vitro and in vivo. Such polymers bind to DNA, RNA, and inorganic polyphosphate molecules with high affinity and inhibit RNA- and polyphosphate-induced clotting and the activation of the intrinsic pathway of coagulation in vitro. Moreover, [NH(2)(CH(2))(2)NH(2)](G = 3);dendri PAMAM(NH(2))(32) (PAMAM G-3) prevents thrombosis following carotid artery injury and pulmonary thromboembolism in mice without significantly increasing blood loss from surgically challenged animals. These studies indicate that nucleic acid-binding polymers are able to scavenge effectively prothrombotic nucleic acids and other polyphosphates in vivo and represent a new and potentially safer class of antithrombotic agents.
    Proceedings of the National Academy of Sciences 07/2012; 109(32):12938-43. DOI:10.1073/pnas.1204928109 · 9.67 Impact Factor
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    Nancy A Stearns · Jaewoo Lee · Kam W Leong · Bruce A Sullenger · David S Pisetsky ·
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    ABSTRACT: Antibodies to DNA (anti-DNA) are the serological hallmark of systemic lupus erythematosus (SLE) and can mediate disease pathogenesis by the formation of immune complexes. Since blocking immune complex formation can attenuate disease manifestations, the effects of nucleic acid binding polymers (NABPs) on anti-DNA binding in vitro were investigated. The compounds tested included polyamidoamine dendrimer, 1,4-diaminobutane core, generation 3.0 (PAMAM-G3), hexadimethrine bromide, and a β-cylodextrin-containing polycation. As shown with plasma from patients with SLE, NABPs can inhibit anti-DNA antibody binding in ELISA assays. The inhibition was specific since the NABPs did not affect binding to tetanus toxoid or the Sm protein, another lupus autoantigen. Furthermore, the polymers could displace antibody from preformed complexes. Together, these results indicate that NABPs can inhibit the formation of immune complexes and may represent a new approach to treatment.
    PLoS ONE 07/2012; 7(7):e40862. DOI:10.1371/journal.pone.0040862 · 3.23 Impact Factor
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    ABSTRACT: To reduce the adverse effects of cancer therapies and increase their efficacy, new delivery agents that specifically target cancer cells are needed. We and others have shown that aptamers can selectively deliver therapeutic oligonucleotides to the endosome and cytoplasm of cancer cells that express a particular cell surface receptor. Identifying a single aptamer that can internalize into many different cancer cell-types would increase the utility of aptamer-mediated delivery of therapeutic agents. We investigated the ability of the nucleolin aptamer (AS1411) to internalize into multiple cancer cell types and observed that it internalizes into a wide variety of cancer cells and migrates to the nucleus. To determine if the aptamer could be utilized to deliver therapeutic oligonucleotides to modulate events in the nucleus, we evaluated the ability of the aptamer to deliver splice-switching oligonucleotides. We observed that aptamer-splice-switching oligonucleotide chimeras can alter splicing in the nuclei of treated cells and are effective at lower doses than the splice switching oligonucleotides alone. Our results suggest that aptamers can be utilized to deliver oligonucleotides to the nucleus of a wide variety of cancer cells to modulate nuclear events such as RNA splicing.
    06/2012; 22(3):187-95. DOI:10.1089/nat.2012.0347
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    ABSTRACT: Most cases of pancreatic cancer are not diagnosed until they are no longer curable with surgery. Therefore, it is critical to develop a sensitive, preferably noninvasive, method for detecting the disease at an earlier stage. In order to identify biomarkers for pancreatic cancer, we devised an in vitro positive/negative selection strategy to identify RNA ligands (aptamers) that could detect structural differences between the secretomes of pancreatic cancer and non-cancerous cells. Using this molecular recognition approach, we identified an aptamer (M9-5) that differentially bound conditioned media from cancerous and non-cancerous human pancreatic cell lines. This aptamer further discriminated between the sera of pancreatic cancer patients and healthy volunteers with high sensitivity and specificity. We utilized biochemical purification methods and mass-spectrometric analysis to identify the M9-5 target as cyclophilin B (CypB). This molecular recognition-based strategy simultaneously identified CypB as a serum biomarker and generated a new reagent to recognize it in body fluids. Moreover, this approach should be generalizable to other diseases and complementary to traditional approaches that focus on differences in expression level between samples. Finally, we suggest that the aptamer we identified has the potential to serve as a tool for the early detection of pancreatic cancer.
    The Journal of clinical investigation 04/2012; 122(5):1734-41. DOI:10.1172/JCI62385 · 13.22 Impact Factor

Publication Stats

7k Citations
1,199.95 Total Impact Points


  • 1998-2014
    • Duke University
      • Department of Surgery
      Durham, North Carolina, United States
  • 1995-2013
    • Duke University Medical Center
      • • Department of Surgery
      • • Division of Surgical Sciences
      Durham, North Carolina, United States
  • 2007-2011
    • Translational Genomics Research Institute
      Phoenix, Arizona, United States
  • 1994
    • Howard Hughes Medical Institute
      Ashburn, Virginia, United States
  • 1990-1992
    • Memorial Sloan-Kettering Cancer Center
      • Division of Molecular Biology
      New York City, NY, United States