Benhur Lee, MD
- PROFESSOR | Microbiology
Research Topics:AIDS/HIV, Antivirals, Biodefense, Cell Biology, Dendritic Cells, Drug Design and Discovery, Endothelial Cells, Gene Therapy, Glycobiology, Hematopoiesis, Immune Antagonism, Infectious Disease, Interferon, Interferon Antagonists, Interferon Resistance, Lymphocytes, Macrophage, Membranes, Nipah Virus, Nucleus, Protein Trafficking & Sorting, Retrovirus, Stem Cells, Systems Biology, Trafficking, Vaccine Development, Viruses and Virology
Professor Benhur Lee is a virologist with wide-ranging research interests who joined the Department of Microbiology at the Icahn School of Medicine at Mount Sinai in 2014. He holds the Ward-Coleman Chair in Microbiology. Prior to his recruitment to Mount Sinai, Dr. Lee spent 12 years as a member of the Department of Microbiology, Immunology & Molecular Genetics as well as the Department of Pathology and Laboratory Medicine at the David Geffen School of Medicine at UCLA (University of California, Los Angeles). He joined UCLA in 2001 as an Assistant Professor, and was promoted on an accelerated schedule twice: to Associate Professor in 2007, and to full Professor in 2011. Dr. Lee graduated from Yale University School of Medicine in 1995 with an M.D. and induction into the National Medical Honor Society (Alpha Omega Alpha), and did his clinical and post-doctoral training at the University of Pennsylvania Medical Center (1995-2001) where he also served as Chief Resident (1997). He worked on HIV fusion and entry during his post-doctoral years under Dr. Robert Doms (1997-2001).
Dr. Lee is board-certified in Clinical Pathology and was an attending physician on the Transfusion Medicine Service at UCLA. He was also a member of the UCLA AIDS Institute and the Center for AIDS Research. At UCLA, Dr. Lee was a member of the MD/PhD (MSTP) admissions committee, and served as the Affinity Group Leader and Graduate Advisor for students interested in Virology and Gene Therapy. He was Vice-Chair (Clinical and Translational efforts) of the training committee overseeing an NIH T32 training program in Virology & Gene Therapy, and was a standing member on the Embryonic Stem Cell Research Oversight Committee at UCLA. Dr. Lee hopes to leverage his prior academic experience and contribute to Mount Sinai in equally substantive ways.
The Lee Lab has a special interest in emerging viruses, with a focus on molecular viral-host interactions at the level enveloped virus entry and budding mechanisms. We currently use HIV and henipaviruses (a emerging genus of lethal paramyxoviruses) as our primary model systems. Dr. Lee believes in multidisciplinary approaches to science and has had graduate students and post-docs whose primary expertise ranges across fields such as biomathematics, biophysics, biochemistry, cell biology, and molecular virology. In addition, he also has active projects translating insights gained from basic studies on host-pathogen interactions into broad-spectrum anti-viral therapeutics, and has long-standing collaborations with colleagues in medical chemistry. The Lee Lab has access to BSL4 (UTMB-Galveston) facilities for high-containment work on lethal pathogens.
Currently, the Lee lab has two post-docs trained and certified to work at BSL4 containment facilities. Dr. Lee’s latest foray into translation science involves developing a highly efficient and robust reverse genetics system for paramyxoviruses, so as to facilitate development of live attenuated paramyxovirus based vectors and vaccines. In addition, he has research contracts for generating targeted lentiviral vectors for crossing the blood-brain-barrier; once again, using insights gained from basic studies into virus entry and tropism.
Multi-Disciplinary Training AreasBiophysics and Systems Pharmacology [BSP], Developmental and Stem Cell Biology [DSCB], Immunology [IMM], Microbiology [MIC]
BSc (Magna cum laude), Santa Clara University
MD, Yale University School of Medicine
Residency, Hospital of the University of Pennsylvania
NA, Hospital of the University of Pennsylvania
Dolph Adams Award 2006
Burroughs Wellcome Fund Career Development Award
Henipavirus & Paramyxoviridae
Henipaviruses are the newest genus of paramyxoviruses, and recent studies have shown that bats, on a global scale, are host to major genera of mammalian paramyxoviruses1. Paramyxoviruses are membrane enveloped, singled-stranded, negative-sense RNA viruses that include pathogens of global biomedical and agricultural import. Click here for a quick introduction to paramyxovirus phylogeny.
Although henipavirus spillover events are currently limited to Southeast Asia and Australia, a recent global survey of almost 5,000 bat specimens revealed at least 23 distinct viral clades within the henipavirus genus, with the known Nipah and Hendra viruses representing only two of those clades1. The phylogenetic diversity of henipaviruses is thus vastly greater than what was thought previously. Henipaviruses also exhibit a much broader species tropism than other paramyxoviruses, largely due to the highly conserved protein receptors that these viruses use. In the various taxonomic schemes proposed for the transitional dynamics of zoonotic pathogens2,3 all these features justifiably place henipavirus at or close to the penultimate stage for sustained transmission in human outbreaks.
Our laboratory4, and others5, independently discovered EphrinB2 as the major receptor for Nipah and Hendra virus entry. We also identified EphrinB3 as an alternative receptor for henipavirus entry6. EphrinB2 and B3 belong to a class of highly conserved receptor tyrosine kinases (RTKs) that are involved in germ layer differentiation, tissue boundary formation, and other critical developmental processes such as angiogenesis, neurogenesis and axonal guidance7.
Paramyxovirus entry and fusion is a molecular choreography requiring cognate interactions between protein partners, the fusion (F) and attachment (HN/H/G) envelope glycoproteins, in which motion and form must both, be carefully regulated. Please see "Modes of paramyxovirus fusion: a Henipavirus perspective" for a recent review on paramyxovirus fusion from this lab8.
Our research on henipavirus entry have focused on:
• Elucidating the molecular determinants of attachment protein (G)-receptor specificity4,6,9-13
• Identifying the parameters that modulate fusion (F) protein activity14-16
• Dissecting the receptor binding triggered membrane fusion cascade16-20
• Developing quantitative and immunological tools to shed light on the kinetics and stoichiometry of the fusion process12,16,18,21
• Characterizing the prevalence of cross-reactive neutralizing antibodies from wild-life (mostly bats) and humans at high-risk for exposure in various locales (West Africa, South America) that have now reported the presence of henipavirus-related viruses
In the longer term, we are developing highly efficient reverse genetics rescue systems for single-stranded negative-sense RNA viruses (Mononegavirales) that are more robust (easy to optimize) and do not required the use of vaccinia-driven T7 polymerase (a standard method in the field). Protocols will be published on our lab's web-site, as and when they are published in the peer reviewed literature.
Emerging Infectious Diseases
Emerging and re-emerging infectious diseases pose a constant and present threat to global health and economy. Even excluding obvious threats such as HIV and pandemic flu, the last two decades have seen world-changing episodes like the spread of the West Nile Virus across North America, and the 2003 SARS pandemic. Estimates put the global macro-economic impact of SARS at US$30-$100 billion. Emerging zoonoses are a particular concern when human and environmental factors force the unintended overlap of otherwise naive ecological niches, which increase the chances for viruses to jump host species and to generate "spillover events" in non-natural animal or human hosts.
In most cases, zoonotic viruses, being ill adapted to the new animal or human host, are highly pathogenic, and infections lead to rapid, severe disease with high fatality. Ebola and Marburg viruses (filioviruses), and Hendra and Nipah viruses (henipaviruses) are the most lethal examples of recent zoonoses. An overview of our lab's research on henipaviruses is provided here. For a more detailed introduction, please refer to Henipavirus: Ecology, Molecular Biology and Pathogenesis (Benhur Lee and Paul Rota, Editors, Current Topics in Microbiology and Immunology.2012, Vol 358.)
In addition, arenaviruses, hantaviruses, dengue and chikungunya viruses may even be considered as diseases of global warming as climate change increases the geographical range of their animal or arthropod hosts. Given the increasing number of viral zoonoses, the traditional "one bug-one drug" paradigm is clearly inadequate as a model for antiviral drug development. A recent focus in our lab is developing broad-spectrum antiviral strategies that target either viral or host cell components that are essential for multiple classes of virus.
Understanding the mechanisms of spread, the nature of persistence of these viruses in their animal hosts, and their pathogenesis in humans, is critical for the development of any counteracting antiviral strategies and vaccines, and underscores the importance of multidisciplinary approaches for studying viral Emerging Infectious Diseases.
HIV-1 (Quantitative Modeling of Virus Entry Efficiency)
HIV uses CD4 and a coreceptor (CCR5 and/or CXCR4) for viral entry. The efficiency of HIV receptor/coreceptor mediated entry has important implications for viral pathogenesis and transmission. The advent of CCR5 inhibitors in clinical use also underscores the need for quantitative and predictive tools that can guide therapeutic management. Historically, measuring the efficiency of CD4/CCR5 mediated HIV entry has relied on surrogate, inadequate, or slow throughput experimental tools.
We developed the Affinofile receptor affinity profiling system that has provided a quantitative and higher throughput method to characterize viral entry efficiency as a function of CD4 and CCR5 expression levels.
For details on the Affinofle system, please see Johnston et al (2009) for the original description.Different groups have used the Affinofile system to reveal the distinct pathophysiological properties associated with Env entry phenotypes.
Overall Research Theme
The general theme that unites the studies in our laboratory is Molecular Viral-Host Interactions. We have a special interest in enveloped virus entry and budding mechanisms, with an increasing focus on viruses that cause Emerging Infectious Diseases. We study highly pathogenic viruses, and use the Henipavirus and Human Immunodeficiency Virus (HIV-1) as primary model systems to represent the pathogenesis of acute and chronic viruses, respectively.
• Acutely pathogenic viruses, by definition, strongly subvert the host's innate immunity, and thus propagate and transmit themselves before the host's adaptive immune system can come into play.
• Chronic viruses that persist in the host result in "smouldering" infections. They can actively replicate in the host over long periods of time, and have evolved mechanisms to counteract or evade both the innate and adaptive immune systems.
Henipavirus is a new genus of paramyxovirus discovered around the turn of the 20th century. Nipah (NiV) and Hendra (HeV) virus are zoonotic viruses--transmitted to humans from their natural bat reservoir--that cause fatal encephalitis in 40-75% of infected patients. In many cases, the estimated time from infection to death is less than 2 weeks, with symptomatic periods lasting for 4-7 days. HIV-1, the causative agent of AIDS, is also a zoonotic virus (transmitted from its natural chimpanzee reservoir) that exploded on the world scene in the early 1980s. HIV/AIDS remains a global pandemic, although our increasing understanding of HIV/IDS pathogenesis makes this a particularly exciting and fruitful time for basic research to be translated into therapeutic, preventive, and vaccine strategies.
How viruses survive, proliferate and transmit within and between hosts is a testament to the past and present evolutionary battles between host and pathogen. The guiding objective of our research is to obtain and translate basic knowledge about viral entry and replication processes to therapeutic or interventional anti-viral strategies. We are particularly interested in developing novel methods, reagents, and therapeutics to address long-standing and intractable problems in our fields of interest. For example, a recent focus of our lab is developing broad-spectrum antiviral strategies that target either viral or host cell components that are essential for multiple classes of virus.EphrinB2 is the primary high affinity receptor that mediates NiV and HeV entry. EphrinB2 belong to a class of highly conserved receptor tyrosine kinases (RTKs) that are involved in germ layer differentiation, tissue boundary formation, and other critical developmental processes such as angiogenesis, neurogenesis and axonal guidance. The unique properties associated with NiV Env interactions with its entry receptor (EphrinB2), and the fortuitous property of EphrinB2 being expressed in highly relevant populations of various stem cells, has also resulted in exciting collaborations with stem cell biologists and gene therapists to exploit this high affinity interaction for therapeutic and interrogative purposes.
Targeting EphrinB2+ stem cells and interrogating the EphrinB2-Eph axis in pluripotent stem cell fate and differentiation. EphrinB2 is present on murine embryonic stem cells (ESC), hematopoietic stem cells (HSC), and neural stem cells (NSC), and has been previously described as a putative molecular marker of "stemness"1. In collaboration with stem cell biologists, we have developed an armamentarium of tools based on the picomolar affinity of NiV-G for EphrinB2 to interrogate the role of ephrinB2 in human pluripotent stem cell fate and differentiation. In addition, in collaboration with gene therapists, we are developing Nipah virus Env pseudotyped lentiviral vectors that can cross the blood brain barrier and target the CNS. Our initial efforts are encouraging (see K Palomares et al, 2012)2 and show that NiV Env pseudotypes can not only target specific populations of human ESC, HSC, and NSC in vitro, but when administered intravenously in vivo, can bypass the liver sink, a critical barrier in targeted gene delivery. Click here for a more detailed description of our human pluripotent stem cell work that is funded by the California Institute of Regenerative Medicine
Broad Spectrum Antiviral Strategies
Lipid targeted compounds that inhibit virus-cell membrane fusion, targeting the virus membrane. Advances in antiviral therapeutics have allowed for effective management of specific viral infections, most notably human immunodeficiency virus (HIV). Yet, the one-bug-one-drug paradigm of drug discovery is insufficient to meet the looming threat of emerging and re-emerging viral pathogens that endangers global human and livestock health. This underscores the need for broad-spectrum antivirals that act on multiple viruses based on some commonality in their viral life cycle, rather than on specific viral proteins.
We recently described a small molecular compound that inhibited the entry of all lipid-enveloped viruses tested (ME Wolf et al, PNAS, 2010). LJ001 is a membrane-binding compound with broad-spectrum antiviral activity in vitro. LJ001 acts on the virus, and not the cell, inhibiting enveloped virus infection at the level of entry. LJ001 is non-cytotoxic at antiviral concentrations, yet had the remarkable property of inhibiting all enveloped viruses tested, including those of global biomedical and biosecurity importance such as HIV, HCV, Influenza, Ebola, henipaviruses, bunyaviruses, arenaviruses, and poxviruses. LJ001 is not virolytic, does not act as a "detergent", and LJ001-treated virions are still able to bind to their receptors. LJ001 was lipophilic, and could bind to both viral and cellular membranes. Yet, it inhibited virus-cell but not cell-cell fusion. This puzzling dichotomy was illuminated when studies with lipid biosynthesis inhibitors indicated that LJ001 was indeed cytotoxic when the ability of a cell to repair and turnover its membranes is compromised. Thus, we posited that the antiviral activity of LJ001 relies on exploiting the physiological difference between inert viral membranes and biogenic cellular membranes with reparative capabilities.
Our latest studies have identified the molecular target of LJ001 and its precise molecular mechanism of action. LJ001 acts as a membrane-binding photosensitizer that induces singlet oxygen mediated modifications of specific phospholipid components of viral membranes. This results in changes in the biophysical properties of viral membranes that are not conducive for the membrane curvature dynamics required for productive viral-cell fusion. Importantly, these biophysical changes are not found in biogenic cellular membranes treated with antiviral concentrations of the compound, likely due to multiple endogenous mechanisms that protect lipids against oxidative damage.
Structure Activity Relations (SAR) studies based on this mechanistic understanding led to novel second-generation compounds with markedly enhanced antiviral potencies as a result of improved photochemical and photophysical properties. This is a multi-disciplinary trans-national project requiring expertise ranging from molecular and animal virology, to membrane biophysics and lipidomics, to medicinal and photo- chemistry.
Current projects that will extend these studies include:
• Developing new strategies for improved control and activation of these antiviral compounds in vivo.
• Using the unique properties of these photosensitizers for more in-depth study of the viral-fusion process e.g. capturing and visualizing fusion intermediates using cryo-electron tomography studies of virus-cell fusion trapped just before membrane merger, or dissecting the fine mechanics of class I-III fusion by regulating the time of light-induced inactivation.
Bioactive Proteasome Inhibitors Our basic studies on Nipah virus budding led us to discover that ubiquitin-regulated nuclear-cytoplasmic trafficking of the Nipah virus matrix (NiV-M) protein is critical for matrix budding function. Inhibiting ubiquitination of NiV-M by proteasome inhibitors such as Bortozemib, which is FDA-approved for oncologic indications, led to nuclear retention of NiV-M, and abrogation of NiV-M budding. Live NiV budding and replication is exquisitely sensitive to Bortozemib with an in vitro IC50 (~2.7 nM) that is about 100-fold lower than the peak plasma concentration (~300 nM) that can be achieved in patients.
The ubiquitin proteasome pathway (UPP) is implicated in the lifecycle of multiple viruses. The UPP regulates a wide array of protein function and cellular processes and many viruses are known to manipulate the host cell UPP to enable replication, egress and immune evasion. Many proteasome inhibitors (PSM Inbs) have a negative impact on viral infections in vitro, but the involvement of the UPP in multiple cellular functions, coupled with the lack of potency, specificity or in vivo stability of the first generation PSM Inbs, discouraged the consideration that PSM INbs could be developed safely as an antiviral therapeutic. In 2003, the FDA approval of the first PSM Inb, Bortezomib, for the treatment of multiple myeloma, provided proof-of-principle that PSM Inbs can be developed with acceptable toxicology profile and good pharmacokinetics/ bioavailability. This has sparked the development of many second generation PSM Inbs with improved potency, selectivity, and bioavailability - several of which are already in Phase I-III trials for oncologic applications.
Excitingly, we have now obtained in vitro preliminary data showing that Bortezomib can also inhibit, to varying degrees, the replication of multiple Category A-C pathogens: Filoviridae (Ebola), Paramyxoviridae (Nipah), Bunyaviridae (Rift Valley fever), Flaviviridae (Russian-Spring-Summer encephalitis), and Arenaviridae (Junin). These data indicate that multiple viral families, though clearly not all, are susceptible to proteasome inhibition, and suggest that the anti-viral efficacy of Bortezomib and/or 2G-PSM Inbs should be evaluated against a broader spectrum of viruses.
This project can be fairly described as discovery driven science. Our immediate primary goal is to empirically evaluate and re-purpose these bioavailable PSM Inbs as potential broad-spectrum antivirals for infections caused by acutely pathogenic viral agents such as those classified as NIAID Category A-C pathogens. Based on the results, we will then elucidate the mechanisms underlying the differential efficacy of the various proteasome inhibitors against distinct viral families.