Richard A Slayden

Colorado State University | CSU · Department of Microbiology, Immunology & Pathology

Toxin-Antitoxin loci regulate adaptive responses to stresses associated with the host environment and drug exposure. Phylogenomic studies have shown that Mycobacterium tuberculosis encodes a naturally expanded type II toxin-antitoxin system, including ParDE/RelBE superfamily members. Type II toxins are presumably regulated exclusively through prote...

Background: NIAID has a programme for testing drug candidates against biodefense and emerging bacterial pathogens that uses defined strain panels consisting of standard laboratory reference strains and strains of clinical origin. Objectives: The current studies were performed to assess the activity of standard-of-care drugs, determine benchmark...

Despite significant research efforts, treatment options for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remain limited. This is due in part to a lack of therapeutics that increase host defense to the virus. Replication of SARS-CoV-2 in lung tissue is associated with marked infiltration of macrophages and activation of innate immune...

Twenty lupane type A-ring azepano-triterpenoids were synthesized from betulin and its related derivatives and their antitubercular activity against Mycobacterium tuberculosis, mono-resistant MTB strains, and nontuberculous strains Mycobacterium abscessus and Mycobacterium avium were investigated in the framework of AToMIc (Anti-mycobacterial Target...

Background Francisella tularensis is a highly virulent and contagious Gram-negative intracellular bacterium that causes the disease tularaemia in mammals and is classified as a Category A priority pathogen. Methods We utilized a systematic analysis of antibacterial potency, extent of dissemination by analysis of bacterial burden in a secondary vit...

Since its initial discovery in late 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the cause of COVID19, has spread worldwide and despite significant research efforts, treatment options remain limited. Replication of SARS-CoV-2 in lung is associated with marked infiltration of macrophages and activation of innate immune inflamm...

Objectives Francisella tularensis is a highly virulent and contagious gram-negative intracellular bacterium that causes the disease tularemia in mammals and is classified as a Category A priority pathogen. The high infectivity, difficulty of obtaining a durable cure of disseminated disease and the emergence of drug resistance warrants investigation...

Infection with Influenza A virus can lead to the development of encephalitis and subsequent neurological deficits ranging from headaches to neurodegeneration. Post-encephalitic parkinsonism has been reported in surviving patients of H1N1 infections, but not all cases of encephalitic H1N1 infection present with these neurological symptoms, suggestin...

Drug susceptibility profiles of NIAID Category A and B priority and emerging pathogens to standard of care drugs were assessed to determine susceptibility against clinical strains in addition to reference laboratory strains to establish a comparison resource for the performance of drug candidates, to define non-redundant minimal strain panels for i...

Filamenting temperature sensitive protein Z (FtsZ) is an essential bacterial cell division protein and a promising target for the development of new antibacterial therapeutics. As a part of our ongoing SAR studies on 2,5,6-trisubstituted benzimidazoles as antitubercular agents targeting Mtb-FtsZ, a new library of compounds with modifications at the...

Inflammatory activation of glial cells promotes loss of dopaminergic neurons in Parkinson's disease (PD). The transcription factor, Nuclear Factor-kappa B (NF-κB), regulates the expression of multiple neuroinflammatory cytokines and chemokines in activated glial cells that are damaging to neurons. Thus, inhibition of NF-κB signaling in glial cells...

There has been a significant reduction in annual tuberculosis incidence since the World Health Organization declared tuberculosis a global health threat. However, treatment of M. tuberculosis infections requires lengthy multidrug therapeutic regimens to achieve a durable cure. The development of new drugs that are active against resistant strains a...

We report the results of a symposium aimed at identifying validated biomarkers that can be used to complement clinical observations for diagnosis and prognosis of joint injury leading to equine osteoarthritis (OA). Biomarkers might also predict pre-fracture change that could lead to catastrophic bone failure in equine athletes. The workshop was att...

2-Alkyl-1,2-benzisoselenazol-3(2H)-ones, represented by the ebselen (1a), are being studied intensively for a range of medicinal applications. We describe both a new thermal and photoinduced copper-mediated cross-coupling between potassium selenocyanate (KSeCN) and N-substituted ortho-halobenzamides to form 2-alkyl-1,2-benzisoselenazol-3(2H)-ones c...

There is growing awareness of the link between drug-target residence time and in vivo drug activity, and there are increasing efforts to determine the molecular factors that control the lifetime of a drug-target complex. Rational alterations in drug-target residence time require knowledge of both the ground and transition states on the inhibition r...

A critical goal of lead compound selection and optimization is to maximize target engagement whilst minimizing off-target binding. Since target engagement is a function of both the thermodynamics and kinetics of drug-target interactions, it follows that the structures of both the ground states and transition states on the binding reaction coordinat...

Much is known about the mode of action of drugs and resistance mechanisms under laboratory growth conditions, but research on the bacterial transcriptional response to drug pressure in vivo or efficacious mode of action and transient resistance mechanisms of clinically employed drugs is limited. Accordingly, to assess active alternative metabolism...

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Melioidosis is caused by the facultative intracellular bacterium Burkholderia pseudomallei and is potentially fatal. Despite a growing global burden and high fatality rate, little is known about the disease. Recent studies demonstrate that cyclooxygenase-2 (COX-2) inhibition is an effective post-exposure therapeutic for pulmonary melioidosis, which...

Confirmation of NF-ĸB activity with infection of treatment. A representative western blot image of the p65 activity with infection and treatment after 90 minutes of infection. Infection of RAW 264.7 with Bp82 resulted in an increase in p65 expression, and pre-treatment with TA was able to significantly reduce this expression. Data is representative...

Dose Response. RAW 264.7 cells were pretreated with 10 μM, 20 μM, 50 μM, 100 μM, 150 μM and 200 μM TA and 0.01% DMSO for 30 minutes and infected with Bp82 at an MOI of 5. Cell viability was determined via optical absorbance at 570nm utilizing the crystal violet method. Each treatment was normalized to uninfected/untreated control for each time poin...

Histogram confirming unstained control. This is a representative image of the histogram shifts that occur when comparing the uninfected/unstained, uninfected/stained, and Bp82 infected/stained. (TIF)

Flow cytometry gating of intact cells. RAW 264.7 cells were gated on a linear forward-scatter vs. logarithmic side-scatter plot. (TIF)

Eliminating doublets from analyzed population. To eliminate doublets from the population analyzed for MCF a pulse-width vs logarithmic mean intensity plot was used to gate on the singlet population. This is a representative image for COX-2 analysis. This population was used to determine the MCF. (TIF)

Confirmation of COX-2 expression with infection and treatment. A representative immunofluorescence image with infection and treatment after 6 hours of infection. On the left, DAPI was used for the nuclear counter stain (blue).Infection of RAW 264.7 with Bp82 resulted in induction of COX-2 (red) and pre-treatment with TA significantly reduced COX-2...

Clinical diagnosis of acute infectious diseases during the early stages of infection is critical to administering the appropriate treatment to improve the disease outcome. We present a data driven analysis of the human cellular response to respiratory viruses including influenza , respiratory syncytia virus, and human rhinovirus, and compared this...

Previously, structure-based drug design was used to develop substituted diphenyl ethers with potency against the Mycobacterium tuberculosis (Mtb) enoyl-ACP reductase (InhA), however, the highly lipophilic centroid compound, SB-PT004, lacked sufficient efficacy in the acute murine Mtb infection model. A next generation series of compounds were desig...

β-Ketoacyl-ACP synthases (KAS) are key enzymes involved in the type II bacterial fatty acid biosynthesis (FASII) pathway and are putative targets for antibacterial discovery. Several natural product KAS inhibitors have previously been reported, including thiolactomycin (TLM), which is produced by Nocardia spp. Here we describe the synthesis and cha...

Tuberculosis (TB) and its drug resistant forms kills more people than any other infectious disease. This fact emphasizes the need to identify new drugs to treat TB. 2-Aminothiophenes (2AT) have been reported to inhibit Pks13, a validated anti-TB drug target. We synthesized a library of 42 2AT compounds. Among these, compound 33 showed remarkable po...

Objective: IGF-I is one of several anabolic factors being investigated for the treatment of osteoarthritis. Due to the short biological half-life, extended administration is required for more robust cartilage healing. Here we create a self-complimentary adeno-associated virus gene therapy vector utilizing the transgene for IGF-I. Design: Various...

The increasing number of clinical strains resistant to one or more of the front-line TB drugs complicates the management of this disease. To develop next-generation benzimidazole-based FtsZ inhibitors with improved efficacy, we employed iterative optimization strategies based on whole bacteria potency, bactericidal activity, plasma and metabolic st...

Stress-induced molecular programs designed to stall division progression are nearly ubiquitous in bacteria, with one well-known example being the participation of the SulA septum inhibiting protein in the SOS DNA damage repair response. Mycobacteria similarly demonstrate stress-altered growth kinetics, however no such regulators have been found in...

Determining the molecular basis for target selectivity is of particular importance in drug discovery. The ideal antibiotic should be active against a broad spectrum of pathogenic organisms with a minimal effect on human targets. CG400549, a Staphylococcus-specific 2-pyridone compound that inhibits the enoyl-acyl carrier protein reductase (FabI), ha...

Trisubstituted benzimidazoles have demonstrated potency against Gram-positive and Gram-negative bacterial pathogens. Previously, a library of novel trisubstituted benzimidazoles was constructed for high throughput screening, and compounds were identified that exhibited potency against M. tuberculosis H37Rv and clinical isolates, and were not toxic...

The diaryl ethers are a novel class of antituberculosis drug candidates that inhibit InhA, the enoyl-ACP reductase involved in the fatty acid biosynthesis (FASII) pathway, and have antibacterial activity against both drug-sensitive and drug-resistant strains of Mycobacterium tuberculosis. In the present work, we demonstrate that two time-dependent...

Identification of a novel class of anti-Burkholderia compounds is key in addressing antimicrobial resistance to current therapies as well as naturally occurring resistance. The FabI enoyl-ACP reductase in Burkholderia is an underexploited target that presents an opportunity for development of a new class of inhibitors. A library of substituted diph...

We introduce Iterative Feature Removal (IFR) as an unbiased approach for selecting features with diagnostic capacity from large data sets. The algorithm is based on recently developed tools in machine learning that are driven by sparse feature selection goals. When applied to genomic data, our method is designed to identify genes that can provide d...

The bacterial fatty acid biosynthesis pathway is a validated target for the development of novel chemotherapeutics. However, since Burkholderia pseudomallei carries genes that encode both FabI and FabV enoyl-acyl carrier protein (ACP) reductase homologues, the enoyl-ACP reductase that is essential for in vivo growth needs to be defined so that the...

FtsZ, an essential protein for bacterial cell division, is a highly promising therapeutic target, especially for the discovery and development of new-generation anti-TB agents. Following up the identification of two lead 2,5,6-trisubstituted benzimidazoles, 1 and 2, targeting Mtb-FtsZ in our previous study, an extensive SAR study for optimization o...

Molecular programs employed by Mycobacterium tuberculosis (Mtb) for the establishment of non-replicating persistence (NRP) are poorly understood. In order to investigate mechanisms regulating entry into NRP, we asked how cell cycle regulation is linked to downstream adaptations that ultimately result in NRP. Based on previous reports and our recent...

Understanding the Ly49 gene family can be challenging in terms of nomenclature and genetic organization. The Ly49 gene family has two major gene nomenclature systems, Ly49 and Killer Cell Lectin-like Receptor subfamily A (klra). Mice from different strains have varying numbers of these genes with strain specific allelic variants, duplications, dele...

Francisella tularensis is classified as a category A priority pathogen and causes fatal disseminated disease in humans upon inhalation of less than 50 bacteria. Although drugs are available for treatment, they are not ideal because of toxicity and route of delivery, and in some cases patients relapse upon withdrawal. We have an ongoing program to d...

A local evanescent array coupled biosensor is used to detect spherical polystyrene nanoparticles with diameters of 40 nm and 200 nm, whose sizes and refractive index are similar to virus particles. The sensitivity is similar to 1%/particle for 200 nm particles and 0.04%/particle for 40 nm particles. Mie scattering in an evanescent field theory is u...

A high-throughput screen led to the discovery of 2-amino-4-oxo-4-phenylbutanoate inhibitors of the 1,4-dihydroxy-2-naphthoyl-CoA synthase (MenB) from the menaquinone biosynthesis pathway in Mycobacterium tuberculosis. However, these compounds are unstable in solution and eliminate to form the corresponding 4-oxo-4-phenylbut-2-enoates that then reac...

Proteins that are involved in regulation of cell division and cell cycle progression remain undefined in Mycobacterium tuberculosis. In addition, there is a growing appreciation that regulation of cell replication at the point of division is important in establishing a non-replicating persistent state. Accordingly, the objective of this study was t...

As an initial step in developing novel antibacterials against Burkholderia pseudomallei, we have characterized the FabI enoyl-ACP reductase homologues in the type II fatty acid biosynthesis pathway from this organism and performed an initial enzyme inhibition study. A BLAST analysis identified two FabI enoyl-ACP reductase homologues, bpmFabI-1 and...

Arenaviruses are responsible for acute hemorrhagic fevers worldwide and are recognized to pose significant threats to public health and biodefense. Small molecule compounds have recently been discovered that inhibit arenavirus entry and protect against lethal infection in animal models. These chemically distinct inhibitors act on the tripartite env...

Detection of virus-like nanoparticles via scattering using a chip-scale optical biosensor Appl. Phys. Lett. 101, 161111 (2012) Role of electropores on membrane blebbing—A model energy-based analysis J. Appl. Phys. 112, 064703 (2012) Local streamline generation by mechanical oscillation in a microfluidic chip for noncontact cell manipulations Appl....

Libraries of novel trisubstituted benzimidazoles were created through rational drug design. A good number of these benzimidazoles exhibited promising MIC values in the range of 0.5-6 μg/mL (2-15 μM) for their antibacterial activity against Mtb H37Rv strain. Moreover, five of the lead compounds also exhibited excellent activity against clinical Mtb...

Nanoparticles with diameter and refractive index similar to viruses were detected by using a local evanescent array coupled biosensor with an integrated photodetector array on a CMOS chip.

Menaquinone is an essential component of the electron transport chain in many pathogens and consequently enzymes in the menaquinone biosynthesis pathway are potential drug targets for the development of novel antibacterial agents. In order to identify leads that target MenB, the 1,4-dihydroxy-2-naphthoyl-CoA synthase from Mycobacterium tuberculosis...

Francisella tularensis is a category A select agent based on its infectivity and virulence but disease mechanisms in infection remain poorly understood. Murine pulmonary models of infection were therefore employed to assess and compare dissemination and pathology and to elucidate the host immune response to infection with the highly virulent Type A...

The emergence of multidrug-resistant Mycobacterium tuberculosis strains has made many of the currently available anti-tuberculosis (TB) drugs ineffective. Accordingly, there is a pressing need to identify new drug targets. Filamentous temperature-sensitive protein Z (FtsZ), a bacterial tubulin homologue, is an essential cell-division protein that p...

The first demonstration of a label-free LEAC biosensor immunoassay is reported. CMOS chips with integrated detection arrays detected tuberculosis related antibodies and 18kDa protein antigens. The limit of detection for thickness change was 120 pm.

A Label-free optical waveguide immunosensor is investigated both theoretically and experimentally. The local evanescent array coupled (LEAC) biosensor is based on a local evanescent field shift mechanism, which differs from those of other evanescent waveguide sensors. Antigens specifically bound by immobilized antibodies on the waveguide surface in...

A label-free optical waveguide immunosensor was designed, fabricated and tested. Different from other popular resonance-based biosensors, such as surface-plasmon-resonance (SPR) or ring/disk resonance biosensors, the local evanescent array coupled (LEAC) biosensor relies on a local evanescent field shift mechanism and can be readily manufactured us...

The National Institute of Allergy and Infectious Disease classifies Francisella tularensis as a Category A priority pathogen. Despite the availability of drugs for treating tularaemia, the mortality in naturally acquired cases can still approach 30%. In addition, the usefulness of existing drugs for treatment in response to exposure or for prophyla...