Development Scientist: Master Seed, FMD Vaccine Virus Derivation
Resume:
KRISTEN M. LAMB, PH.D.
Phone: 762-***-**** (Cell)
E-mail: *.********.****@*****.***
**** ****** **. ***. *** College Station, TX 77864 KM Lamb CV – page 1 of 8
OBJECTIVE
Utilize my skills and experience for the advancement of biological research in human and/or animal health at a high-impact organization.
TECHNICAL SKILL SET
Virology Experience:
Viral titration, viral propagation, purification and characterization, serum neutralization (SN) assays
(fluorescence and syncytia-based detection methods), production of viral vector constructs, RNA Dependent RNA polymerase (RDRP) assay (luminescence-based assay, related to MOA of small molecule), viral plaque assay, proof-of-concept work for modified SN assays with fluorescently labeled virus, cell based assays for small molecule anti-viral capacity. FeLV ELISA assay execution for project team.
Viruses: Vaccine strain of Foot and Mouth disease virus, Hepatitis C, Venezuelan equine encephalitis virus (VEEV), Respiratory Syncytial Virus (RSV), Measles Virus (MeV), Canine distemper virus (CDV), Canine Parvovirus (CPV), Canine adenovirus type 2 (CAV2), Canine parainfluenza virus (CPi), Seneca Valley Fever Virus (SVA), Porcine Circovirus type 2. Molecular Biology Experience:
Plasmid and genomic DNA purification (cell culture, clinical tissue samples, E. coli, blood, feces), RNA purification (cell culture, from purified virus, clinical tissue samples), UV/Vis determination of DNA quantity and purity, PCR, qPCR, reverse transcription PCR, cloning, site directed mutagenesis, electrophoresis of DNA and RNA, DNA sequence analysis, DNA restriction digests, homologous recombination, oligonucleotide designs for PCR-based assays. Design/analysis tools used – Serial Cloner, Sequencher, Vector NTI, NCBI website tools, DNAStar. Cell Culture Experience:
Cell transfection, stable cell line formation. Cell lines: BHK (adherent and suspension variations), BHK- T7, Vero, Vero-hSLAM, Hep2, MDCK, Fibroblast, CRFK. Biochemistry Experience:
Protease activity assay (using Folin-Ciocalteau reagent), electrophoresis, SDS PAGE, Western Blotting, immunofluorescence assays, protein folding/unfolding, circular dichroism (CD), dynamic light scattering
(DLS), enzyme activity and inhibition assay and IC50 determination (spectrophometric). Modified activity assays for given proteins (procedures are used to this day in that lab). Executed, modified, and produced protocol for non-specific protease activity of unknown protease from organism. Protein Expression and Purification:
Construction of expression plasmids, transformation of E. coli with plasmids, and protein expression and purification (cation exchange, reverse phase (RP), size exclusion (SEC), affinity chromatography (nickel, cobalt, methotrexate) using AKTA FPLC, peristaltic pump, gravity flow, and other FPLC/HPLC systems). Protein expression in baculovirus system. Protein expression and construct designing also includes KRISTEN M. LAMB
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membrane-associated proteins. Expressed proteins in E. coli expression systems as well as in mammalian cells. Produced from scratch protocols for protein purification for given proteins (some novel) and modified purification procedures based on construct variation. Protein Characterization:
Protein concentration (Bradford and Lowry protein assays), Circular dichroism (CD), dynamic light scattering (DLS), active site analysis utilizing online programs (ex: DoGSiteScorer), protein folding prediction programs (ex: Phyre2 Server), Sybyl small molecule docking. Structural Biology Experience: Crystallography and Microscopy– Successfully crystallized a number of ternary structures of DHFR (Klebsiella pneumoniae, Vancomycin-resistant enterococcus, human), moved from cloning and protein purification to crystal production, freezing, X-ray crystallography data collection, scaling, refinement, and analysis for structure-based drug. Experience with PDB structure submission. Used these structures and others for small molecule docking into crystal structures using Sybyl.
Crystallography tools used – HKL2000, d*Trek, CCP4, Phenix, Coot, Pymol, MolProbity, Chimera, RCSB PDB Validation tools.
Electron Microscopy – cryo-tomography sample preparation (included but not limited to growth of cells infected with Measles virus on cryo-EM grids and then freezing them), cryo-tomography data reconstruction and analysis, negative stain EM, immune-labeled EM, thin-sectioning EM. Electron Microscopy tools used – IMOD, Amira.
Chemistry:
Column chromatography, extractions, distillations, experience in graduate level organic synthesis lab in the pharmacology department at UTMB
Other Related Experience and Qualification: BSL2/BSL3 environments, clearance for working on select agent (VEEV), scientific publication writing, laboratory protocol writing, experience in training others in laboratory techniques, experience leading junior scientists (2 years in depth), experience in X- ray facility management and training of individuals in data collection and initial data processing techniques.
EXPERIENCE
Zoetis- College Station, TX 2023-Present
Development Scientist (FIXED TERM)
RD IV
Derive RNA from plasmid DNA containing vaccine strain of virus genome and utilize that for transfection of suspension cell line for virus rescue. The ultimate goal is to rescue and grow up the vaccine strain of virus for master and then working seed. Successfully rescued vaccine strain of FMD directly into suspension cells without initially deriving it in adherent cells first. KRISTEN M. LAMB
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Boehringer-Ingelheim- Saint Joseph, MO/Athens, GA 2021-2023 Assay Development (R&D, GI)
Research Scientist IV
Worked to identify discriminating sequences between highly related strains using Bioinformatics. Optimized, gap assessment, and readiness experiments for transfer of ELISA assay for international collaborators. Optimization of WES/JES-based assays for combination vaccine potency evaluation assay development. Purified of monoclonal antibodies used in assay. Executed multi-stage virus inactivation kinetics studies for production. Worked to determine limit of detection of assays. Regularly worked in parallel with different sites within the organization as well as different functional areas to achieve common milestones. Routinely worked with quality control, including even picking up non-release assays and contributing to internal assay optimizations within their department. Newport Labs, Boehringer-Ingelheim- Worthington, Minnesota 2018-2021 Research and Development
Research Scientist II
PI to protein-based vaccines Worthington-branch. Analyzed native protein sequences for consensus sequence determination for protein-based vaccine design against viral target. Purified protein and formulated protein for protein-based vaccines and associated ELISA development. Purified Fc-labelled protein for ELISA assay development. Processed and provided molecular samples for NGS or qPCR diagnostics for pathogen identification, and analyzed sequencing results for de novo sequencing of small genome organisms. The sequencing projects and protein-based vaccine designs included being trained on and working with DNAStar suite for DNA and Protein sequence alignments and analysis. Expressed protein in antibiotic-free E. coli system (only BSL-1 bacterium) and baculovirus. Formulated autogenous vaccines and aided in safety studies thereof. Bridged transfer of assays from one organizational site to my current one at the time and adapted the assay for resources available locally. Actively sought out and reached out to potential collaborators for new technologies. Liberty Biosecurity- Worcester, Massachusetts 2017-2017 Research and Development
Research Scientist
Independently ran and drove two biochemical efforts of the start-up company. The first was executing protease assays routinely on a novel protein. Part of this entailed working to determine divalent cation dependency of the unknown protease. The second front was protein purification and purification optimization of two proteins related to microbiological assays, which served as an integral part of another branch of the company. Part of this work also entailed working with, purifying, and determining activity parameters of a thermophile. This being said, I worked closely with the microbiology arm of the company to provide protein reagents at quality and quantity required for international export. Merial, Boehringer-Ingelheim- Athens, Georgia 2016-2016 Biological Research and Development
Scientist II (Contract)
Executed and optimized serum neutralization assays. Carried out ELISA assays. Worked towards sequencing in-house parvovirus strain and designing a fluorescently labeled strain thereof for the purposes of designing a more cost-efficient cell culture assay. Headed up efforts for designing method for more cost-efficient parainfluenza serum neutralization assay. Provided data for and interacted with project team members for multiple projects. Routinely isolated DNA/RNA from clinical samples including blood, cartilage, and tendon etc.) Ran qPCR and reverse transcription PCRs. Cloned full-length parvovirus into KRISTEN M. LAMB
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larger vector. Purified virus from clinical samples for further animal studies. Routinely produced formal assay results reports for multiple project teams and maintained database of results in Analytical Database and in-house reagents in Freezer Works system. Designed oligos for PCR-based assays. Georgia State University – Atlanta, Georgia 2015-2016 Center for Inflammation, Immunity, and Infection
Postdoctoral Associate
Manipulated and generated of full-length respiratory syncytial virus and canine distemper virus. Performed mutagenesis of fusion proteins of respiratory syncytial virus for generating a full-length recombinant virus. Rescued recombinant viruses in cell culture. Designed full-length recombinant respiratory syncytial virus strains. Used RDRP assays to aid in determination of mechanism of small molecule inhibition of viral infection. Carried out compound dose response assays in mammalian cells. Performed TCID50 assays for Measles and respiratory syncytial virus titering (using fluorescence, luminescence, or syncytia depending on the recombinant strain). Maintained and aided efforts for improvement of in-house database for reagents. Isolated RNA/DNA from infected mammalian cells for RT-PCR, and PCR analysis of infected stain. Routinely ran MIDI and MINI prep purifications of plasmids. Purified virus strains for use in HTP screening for anti-viral small molecule studies. Emory University – Atlanta, Georgia 2014-2015
Department of Pediatrics, Division of Infectious Diseases Postdoctoral associate
Conducted In vitro assembly of respiratory syncytial virus matrix protein lattice. Aimed to determine arrangement of Measles Virus (MeV) glycoproteins in mature virions. Performed cryo-tomography reconstructions of RSV-infected cells to understand protein arrangements and changes at early-stages of infection. Purified respiratory syncytial virus matrix protein from E. coli protein expression system and produced liposomes from Avanti lipid to understand protein behavior within context of viral membrane environment. Designed and executed reactions for respiratory syncytial virus matrix lattice assembly. Evaluated assembly reactions by negative staining and viewing on JEOL1400 microscope. Grew HeLa cells on quantifoil grids and infected them successfully with Measles virus strains Measles-HcHisTagFlag and Measles-GFP. Evaluated infection of HeLa cells with Measles-GFP virus via fluorescence. Cryo- plunged froze infected HeLa cells on grids for cryo-tomography data collection. Processed cryotomography data resulting from above mentioned Measles grids as well as from other projects in the lab using IMOD software. Processed cryo-tomography data to support projects of other laboratory members. Carried out TCID50 assays for Measles tittering. In this position, also contributed heavily to other projects beyond the ones I headed up.
University of Connecticut – Storrs, Connecticut 2011-2014 Department of Pharmacy
Postdoctoral fellow
Performed X-ray Crystallography of DHFR from three species. Expressed and purified Vancomycin- resistant enterococcus DHFR, human, and Klebsiella DHFR. Produced crystals of all three varieties. Produced multiple high-resolution diffraction X-ray crystallization data sets. Analyzed ternary crystal structures of DHFR from three species for the purposes of understanding structural variations between these and other species to guide designs of inhibitors for each. This includes characterizing and analyzing molecular differences between these. Performed Sybyl small-molecule docking as well as active site KRISTEN M. LAMB
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binding characterization using online tools for the purposes of understanding molecular binding site of proteins and biophysical characteristics affecting inhibitor binding. Successfully completed first structure of Klebsiella pneumonaie DHFR (ternary structure with cofactor and inhibitor). Performed IC50 assays with human, Vancomycin-resistant enterococcus, and Klebsiella pneumoniae DHFR inhibitors to aid in further inhibitor designs. This is for the purpose of understanding the activity of the proteins, and how they vary under the influence of small molecules with changes in side-chain or ring design. Managed and trained persons on the in-house X-ray beamline. Instructed, demonstrated, and advised two students’ biochemical and X-ray crystallography techniques for PharmD student projects. Served as guest lecturer for course, “Structure and Function of Biological Macromolecules.” As guest lecturer, described general X-ray crystallography procedures, beginning at setting up crystallization trials. Reviewed X-ray facility instrumentation and initial computational methods for solving structures. Served as assistant to ECE Professors’ Training Courses (2 sessions). During this, I reviewed practical methods in X-ray crystallography.
University of Texas Medical Branch – Galveston, Texas 2003-2010 Department of Biochemistry and Molecular Biology
Graduate Research
Produced cryo-EM single-particle reconstructions of pre- and post-entry nucleocapsids. Conducted pseudo-atomic modeling with crystal structure of capsid protein with a series of Venezuelan equine encephalitis virus mature nucleocapsid cryo-EM reconstructions at decreasing resolutions. Investigated Hepatitis C core protein construct’s ability to fold and participate in nucleocapsid-like-particle assembly. Expressed several protein constructs of Hepatitis C virus core protein. Sought to form uniform, homogeneous nucleocapsid-like-particles of Hepatitis C for structural studies, including cryo-EM. Used negative staining to evaluate Hepatitis C nucleocapsid-like-particles. University of West Georgia – Carrollton, Georgia 2000-2003 Department of Chemistry
Undergraduate Research/Organic Laboratory TA/General Chemistry Workshop Leader (Formal group study session leader)
Performed extractions on Cunninghamella elegans with nitro-polyaromatic hydrocarbons (nitro-PAHs) to evaluate whether the fungus could successfully biodegraded the nitro-PAHs. Made reagents for lab, helped students perform experiments, and further explained the chemistry to students. Lead discussions and problem-solving session in general chemistry.
COMPUTER EXPERTISE
• Linux, Macintosh, and Windows Platforms.
• MS Office (Word, Excel, Access, PowerPoint).
• Sybyl for small molecule docking into protein structures
• Phenix, CCP4, Coot, d*Trek, HKL2000 for crystallography
• Adobe PhotoShop, Acrobat, Image J, Quicktime.
• Having managed and maintained an R-Axis IV in-house crystallography beamline, I am quick to familiarize myself with new instrumentation. This is also due to spending years in graduate school working on JEOL 2100 cryo-EM microscope.
KRISTEN M. LAMB
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EXTERNAL PUBLICATIONS
Lamb, Kristen, et al. Antimicrobial Agents and Chemotherapy. 2014 December; 58(12): 7484-7491.
“Novel structure of Klebsiella pneumoniae dihydrofolate reductase and implications for inhibition.” Lamb, K.M., et al., Biochemistry. 2013 October 15; 52(41): 7318-26. “Elucidating features that drive the design of selective antifolates using crystal structures of human dihydrofolate reductase.” Lamb, Kristen, et al., Virology. 2010 October 25; 406(2): 261-9. “Structure of a Venezuelan equine encephalitis virus assembly intermediate isolated from infected cells.” Lamb, Kristen, et al., Virology. 2010 October 25; 406(2): 261-9. “Structure of a Venezuelan equine encephalitis virus assembly intermediate isolated from infected cells.” G-Dayanandan, N., Paulsen, J.L., Viswanathan, K., Keshipeddy, S., Lombardo, M.N., Zhou, W., Lamb, K.M., Sochia, A.E., Alverson, J.B., Priestly, N.D., Wright, D.L., Anderson, A.C., J. Med. Chem. 2014 February 25; 57(6): 2643-2656. “Propargyl-linked antifolates are dual inhibitors of Candida albicans and Candida glabrata.”
Yan, D., Weisshaar, M., Lamb, K., Chung, H.K., Lin, M.Z., Plemper, R.K., Biochemistry. 2015 September 15; 54(36): 5589-5604. “Replication-Competent Influenza Virus and Respiratory Syncytial Virus Luciferase Reporter Strains Engineered for Co-infections Identify Antiviral Compounds in Combination Screens.”
Yi, H., Strauss, J.D., Ke, Z., Alonas, E., Dillard, R.S., Hampton, C.M., Lamb, K.M., Hammonds, J.E., Santangelo, P.J., Spearman, P.W., Wright, E.R., J. Histochem. Cytochem. 2015 October; 63(10): 780- 792. “Native Immunogold Labeling of Cell Surface Proteins and Viral Glycoproteins for Cryo-Electron Microscopy and Cryo-Electron Tomography Applications.” INTERNAL PRESENTATIONS
Oral Presentation: Towards a More Cost-efficient Canine Parainfluenza (CPi) Virus Serum Neutralization (SN) Assay. B&B Merial Meeting. December 15, 2016. Oral Presentation: Determining Glycoprotein Arrangement and Assembly Details in Measles Virus. Emory Children’s Center: Division of Infectious Disease Monthly Meeting. December 18, 2014. EXTERNAL PRESENTATIONS
Poster Presentation: Elucidating Features that Drive the Design of Selective Antifolates Using Human Dihydrofolate Reductase. 44
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Mid-Atlantic Macromolecular Crystallography Meeting in Durham, North Carolina, May 2013.
Poster Presentation: Elucidating Features that Drive the Design of Selective Antifolates Using Human Dihydrofolate Reductase. North Eastern Structure Symposium (NESS) in Hartford, Connecticut, May 2013.
KRISTEN M. LAMB
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Poster Presentation: Understanding Molecular Interactions Regulating Venezuelan Equine Encephalitis Virus Nucleocapsid Plasticity. Keck Annual Research Conference in Houston, Texas, November 2007. Poster Presentation: (multiple years) Understanding Molecular Interactions Regulating Venezuelan Equine Encephalitis Virus Nucleocapsid Plasticity. Sealy Center for Structural Biology Symposium. May 2006-2010.
EDUCATION
Ph.D.-Biomedical Sciences (Biochemistry and Molecular Biology) University of Texas Medical Branch, Fall 2010.
Dissertation: Understanding the Assembly of Simple ssRNA Virus Nucleocapsids. Impact: Increasing the understanding of early stage and assembly of mature virions could increase the opportunities for therapeutics exploiting these points. B.S.-Chemistry, University of West Georgia, Spring, 2003 HONORS
Keck Fellowship Recipient W. M. Keck Center for Virus Imaging 2005-2007 Pharmacology Scholarship UTMB Pharmacology Department 2003-2004 AFFILIATIONS
Biophysical Society, 2011 to 2017.
American Chemical Society, 2001 to 2010.
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REFERENCES
1. Debra Kendall, Ph.D.
Distinguished Professor, Pharmaceutical Sciences, University of Connecticut *****.*******@*****.***
2. Bradley Feilmeier, M.S.
Sr. Principal Business Analyst, Boehringer-Ingelheim *******.*********@******.***
3. Karen Lavery, Ph.D.
Sr. Manager, Metro Biotech
*****@************.***
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