The In Vitro Assessment for Antimicrobial Activity program provides capability in a broad range of in vitro assessments to evaluate promising candidate countermeasures for antimicrobial activity against microbial pathogens and vectors, including those derived from clinical specimens.
This is one of several programs provided by NIAID's Division of Microbiology and Infectious Diseases to support infectious disease product developers.
This contract program supports the development and refinement of animal models and animal replacement technologies and provides in vivo and complex human cell-based in vitro model preclinical testing services, ranging from screening and proof-of-concept to GLP efficacy studies.
Julie E. Ledgerwood, D.O.
View all research conducted at the Vaccine Research Center (VRC)
The goal of the Vaccine Production Program Laboratory (VPPL) is to efficiently translate candidate research vaccines into materials for proof-of-concept clinical trials and to enable advanced development and licensure by partners. The VPPL is responsible for process design and development, clinical good manufacturing practices (cGMP) manufacturing, pre-clinical safety testing, and regulatory activities for all VRC products. Since its inception in 2001, the VPPL has overseen the manufacture of over 68 bulk pharmaceutical compounds formulated into 40 different vaccine and therapeutic products. These products include candidate vaccines for HIV, influenza, filoviruses including Ebola and Marburg, and alphaviruses including chikungunya, SARS, and West Nile.
The laboratory includes development groups for stable cell line generation, cell culture (upstream), purification (downstream), formulation, and analytical testing (characterization and lot release). The focus is on developing state-of-the-art production methodology that will support both the cGMP manufacturing of material for VRC clinical trials and the effective transfer of successful candidates to partner organizations for eventual commercialization. Flexibility is essential as candidate formats include DNA plasmids, viral vectors, recombinant antibodies and other proteins, virus-like particles, and self-assembling nanoparticles.
Once a process is developed, it is transferred to the Vaccine Pilot Plant (VPP), located in Frederick, Maryland, for cGMP production. The VPP, completed in 2005, has four independent production trains in a facility of 126,900 square feet. Two trains operate at 100 liter scale, one train at 400 liter scale, and one train at 2,000 liter scale. There are also suites for inoculum preparation and for media/buffer preparation. The filling operations are qualified to perform small-scale lots up to 5,000 vials and large-scale lots up to 15,000 vials. The warehouse is sized to handle raw materials and supplies sufficient to maintain production operations with coordination and control through the adjacent dispensary. Quality control laboratories and a quality assurance department are responsible for oversight of cGMP manufacture including validation, compliance, lot release, and document control. The VPP is operated under contract by Leidos Biomedical Research, Inc. at the Frederick National Lab.
The Regulatory Science Group within the VPPL is responsible for setting regulatory strategy and for managing regulatory activities for all VRC products. The regulatory group has assembled more than twenty investigational new drugs (INDs) and master files since 2001. The group also leads the development and performance of good laboratory practices (GLP) safety studies for planned clinical products. The Translational Program Management Group of the VPPL is responsible for the cross-VRC coordination of VRC projects and programs. The VPPL establishes Collaborative Research and Development Agreements (CRADAs) with industry partners to bring innovative vaccine technologies in for development or to continue development of VRC candidates beyond Phase I or II clinical development.
The Vaccine Production Program (VPP) applies cutting-edge science and engineering approaches to plasmid design, cell expression, purification, formulation, and analytics. The multidisciplinary nature of the work requires expertise in a number of scientific fields, including molecular and cellular biology, biochemistry, biophysics, biomedical and chemical engineering, and traditional pharmaceutics. The broad portfolio of knowledge, coupled with a strong culture of collaboration and innovation, makes the VPP one of the most versatile product development organizations in infectious disease research. Notable achievements include the development of novel quadrivalent and hexavalent mosaic nanoparticle vaccines to fight influenza, as well as breakthroughs in production and stabilization of trimeric protein vaccine candidates for human immunodeficiency virus (HIV) and respiratory syncytial virus (RSV).
The Vaccine Production Program portfolio includes a diverse array of both traditional and novel vaccine technologies and biologics, including:
Structure of Influenza Mosaic Multi-Component Nanoparticle. Left: 3D model of hexa-valent nanoparticle. Center: 2D Cryo-EM image of purified nanoparticle. Right: 3D reconstruction of 2D Cryo-EM data.
The Vaccine Production Program (VPP) is a science-driven organization that exists as a unique bridge between government and the biopharmaceutical industry. Research at the VPP combines the rigor of academic research with cutting-edge development. Specialized teams of scientists and engineers work in close collaboration with each other, other labs and programs within the VRC and the NIH, as well as with academic and industry partners.
The specialized teams of scientists and engineers that work in close collaboration at the VPP are depicted in this image
The Cell Line Development group is responsible for generating cell lines that produce high and consistent levels of a target vaccine or therapeutic protein. This includes expression vector construction, transfection of the production host cell line, and generation of stable cell pools, followed by single-cell isolation, scale-up, and evaluation for most vaccine products. The final cell bank is ultimately used to support current good manufacturing practice (cGMP) manufacturing of the clinical trial material.
Left: High throughput cell growth monitoring. Center: Cell suspension. Right: Automated platform for cell culture, plating, and sampling
Research in Upstream Process Development centers on the identification and optimization of process parameters to support and control the growth of a mammalian or microbial cell line in a production bioreactor. The resulting process must ensure a robust and scalable method to deliver high-yielding expression of the recombinant vaccine or other protein of interest. Development begins with automated microbioreactors (15 to 250 mL) and progressively increases from 3 L bench-scale bioreactors up to a pilot scale of 50 L. The finalized processes are then transferred externally to enable larger scale manufacturing of clinical material under cGMP regulations.
Left: Automated micro-bioreactors evaluating growth conditions. Right: Cell culture production at 50 L scale.
Downstream Process Development employs diverse purification techniques to produce high-quality recombinant products from mammalian or microbial culture harvests. The unit operations include bulk filtration (removal of cells, cell debris, or other particulate matter), specialized chromatography steps (to separate soluble impurities like host cell proteins and DNA), and specialized filtration and/or buffer exchange steps. Ultimately, the process is transferred externally for cGMP execution at scales supporting up to 2000 L cell culture harvests. At this manufacturing scale, a high-yielding process is capable of delivering up to kilogram yields of clinical product.
Left: Small scale purification system. Center: High throughput parallel ultrafiltration system. Right: Mid-scale gel chromatography column.
The goal of analytical method development is to design and qualify analytical methods in areas such as specific binding activity, physicochemical attributes, impurity levels, etc. These methods are applied to the characterization and evaluation of product quality, defined by its biological activity (potency), identity, purity, and stability. The Analytical Development group is also responsible for transfer of these methods to the Quality Control laboratory of a cGMP manufacturing site.
Left: High throughput chemiluminescence platform. Center: Ultra Performance Liquid Chromatography systems. Right: Automated Western Blot system.
The Formulation Development group is responsible for developing formulations that ensure the quality of the vaccine candidates through cGMP manufacturing and clinical trials. Factors such as physical stress, temperature, light, and atmospheric oxygen can affect the physical and chemical stability of the vaccine. The Formulation Development group utilizes high-throughput methods focusing on molecular conformational, colloidal, and thermodynamic properties to identify optimal stabilizing solution conditions and additives.
Left: High throughput Background Membrane Imaging particle counter. Center: Loading a micro-volume Fluorescence cuvette. Right: Automated ultrafiltration liquid handler.
Scientific Operations facilitates the scientific mission of the VPP through the management of essential support systems that ensure efficient day-to-day operations. These systems include procurement, management of critical reagents and materials, safety, document retention, shipping/receiving, and coordination with NIH/NIAID groups and collaborators, including clinical sites worldwide.
Left: Shipping biologics for transport. Center: Aliquoting critical reagents. Right: Transferring cell banks to liquid nitrogen storage
Yang Y, Bastani N, Lagler SK, Harris D, Nagy A, Chen P, Patel A, Li Y, Gowetski DB, Lei QP. Development and application of an analytical approach to assess an antibody's potential for disulfide reduction. Biotechnol Prog. 2022 Mar;38(2):e3229.
Loomis RJ, DiPiazza AT, Falcone S, Ruckwardt TJ, Morabito KM, Abiona OM, Chang LA, Caringal RT, Presnyak V, Narayanan E, Tsybovsky Y, Nair D, Hutchinson GB, Stewart-Jones GBE, Kueltzo LA, Himansu S, Mascola JR, Carfi A, Graham BS. Chimeric Fusion (F) and Attachment (G) Glycoprotein Antigen Delivery by mRNA as a Candidate Nipah Vaccinev. Front Immunol. 2021 Dec 8;12:772864.
Cibelli NL, Arias GF, Figur ML, Khayat SS, Leach KM, Loukinov I, Gulla KC, Gowetski DB. Advances in Purification of SARS-CoV-2 Spike Ectodomain Protein Using High-Throughput Screening and Non-Affinity Methods. Res Sq [Preprint]. 2021 Aug 20:rs.3.rs-778537.
Gaudinski MR, Berkowitz NM, Idris AH, Coates EE, Holman LA, Mendoza F, Gordon IJ, Plummer SH, Trofymenko O, Hu Z, Campos Chagas A, O'Connell S, Basappa M, Douek N, Narpala SR, Barry CR, Widge AT, Hicks R, Awan SF, Wu RL, Hickman S, Wycuff D, Stein JA, Case C, Evans BP, Carlton K, Gall JG, Vazquez S, Flach B, Chen GL, Francica JR, Flynn BJ, Kisalu NK, Capparelli EV, McDermott A, Mascola JR, Ledgerwood JE, Seder RA; VRC 612 Study Team. A Monoclonal Antibody for Malaria Prevention. N Engl J Med. 2021 Aug 26;385(9):803-814.
Gulla K, Cibelli N, Cooper JW, Fuller HC, Schneiderman Z, Witter S, Zhang Y, Changela A, Geng H, Hatcher C, Narpala S, Tsybovsky Y, Zhang B, Vrc Production Program, McDermott AB, Kwong PD, Gowetski DB. A non-affinity purification process for GMP production of prefusion-closed HIV-1 envelope trimers from clades A and C for clinical evaluation. Vaccine. 2021 Jun 8;39(25):3379-3387.
Chen P, Chen M, Menon A, Hussain AI, Carey E, Lee C, Horwitz J, O'Connell S, Cooper JW, Schwartz R, Gowetski DB. Development of a High Yielding Bioprocess for a Pre-fusion RSV Subunit Vaccine. J Biotechnol. 2021 Jan 10;325:261-270.
View all research conducted at the Vaccine Research Center (VRC)
Respiratory syncytial virus (RSV) infects the lungs and breathing passages, and, in the United States, nearly all children have been infected with RSV by age two. In healthy people, symptoms of RSV infection are usually mild and resolve within a week. However, RSV can cause serious illness or death in vulnerable individuals, including premature and very young infants, children with chronic lung disease or congenital heart disease, and people who are over age 65. In the U.S., RSV is the most common cause of bronchiolitis (inflammation of the small airways in the lungs) in children younger than one year old and causes approximately 58,000 hospitalizations among children under five annually. RSV infection is estimated to cause about 14,000 annual deaths in U.S. adults over age 65. Globally, RSV affects an estimated 64 million people and causes 160,000 deaths each year.
NIAID conducts and supports basic research on RSV to improve understanding of the virus and how it causes disease, as well as factors in animals and humans that affect susceptibility to RSV infection. Research is also underway to develop vaccines to prevent RSV.
To learn about risk factors for RSV and current prevention and treatment strategies visit the MedlinePlus respiratory syncytial virus site.
For more than 50 years, NIAID’s commitment to RSV research has been unparalleled. NIAID researchers were the first to identify and characterize RSV and have provided fundamental knowledge that improves our understanding, treatment, and prevention of RSV disease. NIAID basic research has led to the only preventive treatment currently available for RSV and given us new techniques to manipulate the virus that have brought us closer to a safe and effective vaccine.
NIAID-funded basic and clinical studies helped establish the fundamental knowledge necessary for the private sector to develop protein vaccines. These vaccines are safe and effective at preventing severe RSV in some target populations.
The study found that infants who were not infected with RSV in the first year of life had a 26% lower risk of asthma at 5 years of age than those who were infected with RSV as infants.
Immunological evaluations in NHP models are essential for the advancement of vaccine research. In particular, vaccination and simian immunodeficiency virus (SIV) challenge of macaque species is the best animal model for evaluating candidate HIV vaccines in pre-clinical studies. As a result, the Nonhuman Primate Immunogenicity Core (NIC) was established to support the research efforts of investigators at the VRC.
The NIC manages basic, translational, and preclinical NHP studies. Basic research interests include lymphocyte trafficking, the generation and maintenance of memory responses in systemic and mucosal sites, and pathogenesis following SIV infection. Translational research studies are performed to optimize a variety of different platforms and vectors by studying the effects of different adjuvants, schedules, and delivery methods on immunogenicity. Finally, the data from preclinical testing of vaccines in NHP models is used to support moving clinical products forward to human testing and can be critical to regulatory filings.
All studies that are conducted through the NIC are processed in a standardized manner using the same standard operating procedures (SOPs) for tissue preparation and T cell assays. For example, the NIC uses a qualified 19-color ICS panel and assay for the measurement of IFN-γ, IL-2, TNF, IL-4, IL-13, IL-17, IL-21, and CD40L from CD4 and CD8 T cells; memory T cell subsets and Tfh can also be identified using this panel. In addition, the NIC has qualified the Luminex assay for the measurement of 23 cytokines and chemokines from plasma or cell culture supernatant. To perform these operations, the NIC uses state-of-the-art technologies available at the VRC including 30-color FACS analyzers, 30-color sorters (one in BSL-3 containment), reagent manufacturing capabilities, and the Luminex system.
Other responsibilities of the NIC include the following: consulting on the design and implementation of NHP studies, maintaining a bank of NHP tissue samples for use by VRC investigators, designing new panels such as probe-binding B cell panels, and collating, analyzing, and coordinating data.
For more information on research conducted by Kathryn Foulds, Ph.D. visit the ImmunoTechnology Section.
Dr. Foulds received her M.S. in biotechnology in 1998 from Johns Hopkins University and her Ph.D. in cell and molecular biology in 2003 from the University of Pennsylvania. Her academic interests focused on immunology as well as molecular and microbiology. As a postdoctoral fellow in Dr. Robert Seder’s laboratory at the VRC, Dr. Foulds investigated the role of IL-10 and IFNγ in regulating the generation of memory T cells following vaccination or infection of mice. Dr. Foulds became the assistant director of flow cytometry for the Immune Tolerance Network in 2008, where she managed three remote flow cytometry cores that acquired data for 15 clinical trials. She analyzed the flow cytometry data for quality control as well as for interpreting study results. Dr. Foulds was also responsible for designing flow cytometry panels and coordinating research and development projects performed at the flow cores. Dr. Foulds accepted the position of co-chief, NIC, in 2008.
Potter EL, Gideon HP, Tkachev V, Fabozzi G, Chassiakos A, Petrovas C, Darrah PA, Lin PL, Foulds KE, Kean LS, Flynn JL, Roederer M. Measurement of leukocyte trafficking kinetics in macaques by serial intravascular staining. Sci Transl Med. 2021 Jan 13;13(576):eabb4582.
Corbett KS, Flynn B, Foulds KE, Francica JR, Boyoglu-Barnum S, Werner AP, Flach B, O'Connell S, Bock KW, Minai M, Nagata BM, Andersen H, Martinez DR, Noe AT, Douek N, Donaldson MM, Nji NN, Alvarado GS, Edwards DK, Flebbe DR, Lamb E, Doria-Rose NA, Lin BC, Louder MK, O'Dell S, Schmidt SD, Phung E, Chang LA, Yap C, Todd JM, Pessaint L, Van Ry A, Browne S, Greenhouse J, Putman-Taylor T, Strasbaugh A, Campbell TA, Cook A, Dodson A, Steingrebe K, Shi W, Zhang Y, Abiona OM, Wang L, Pegu A, Yang ES, Leung K, Zhou T, Teng IT, Widge A, Gordon I, Novik L, Gillespie RA, Loomis RJ, Moliva JI, Stewart-Jones G, Himansu S, Kong WP, Nason MC, Morabito KM, Ruckwardt TJ, Ledgerwood JE, Gaudinski MR, Kwong PD, Mascola JR, Carfi A, Lewis MG, Baric RS, McDermott A, Moore IN, Sullivan NJ, Roederer M, Seder RA, Graham BS. Evaluation of the mRNA-1273 Vaccine against SARS-CoV-2 in Nonhuman Primates. N Engl J Med. 2020 Oct 15;383(16):1544-1555.
Van Rompay KKA, Keesler RI, Ardeshir A, Watanabe J, Usachenko J, Singapuri A, Cruzen C, Bliss-Moreau E, Murphy AM, Yee JL, Webster H, Dennis M, Singh T, Heimsath H, Lemos D, Stuart J, Morabito KM, Foreman BM, Burgomaster KE, Noe AT, Dowd KA, Ball E, Woolard K, Presicce P, Kallapur SG, Permar SR, Foulds KE, Coffey LL, Pierson TC, Graham BS. DNA vaccination before conception protects Zika virus-exposed pregnant macaques against prolonged viremia and improves fetal outcomes. Sci Transl Med. 2019 Dec 18;11(523):eaay2736.
Iwamoto N, Mason RD, Song K, Gorman J, Welles HC, Arthos J, Cicala C, Min S, King HAD, Belli AJ, Reimann KA, Foulds KE, Kwong PD, Lifson JD, Keele BF, Roederer M. Blocking α4β7 integrin binding to SIV does not improve virologic control. Science. 2019 Sep 6;365(6457):1033-1036.
Kong R, Duan H, Sheng Z, Xu K, Acharya P, Chen X, Cheng C, Dingens AS, Gorman J, Sastry M, Shen CH, Zhang B, Zhou T, Chuang GY, Chao CW, Gu Y, Jafari AJ, Louder MK, O'Dell S, Rowshan AP, Viox EG, Wang Y, Choi CW, Corcoran MM, Corrigan AR, Dandey VP, Eng ET, Geng H, Foulds KE, Guo Y, Kwon YD, Lin B, Liu K, Mason RD, Nason MC, Ohr TY, Ou L, Rawi R, Sarfo EK, Schön A, Todd JP, Wang S, Wei H, Wu W; NISC Comparative Sequencing Program, Mullikin JC, Bailer RT, Doria-Rose NA, Karlsson Hedestam GB, Scorpio DG, Overbaugh J, Bloom JD, Carragher B, Potter CS, Shapiro L, Kwong PD, Mascola JR. Antibody Lineages with Vaccine-Induced Antigen-Binding Hotspots Develop Broad HIV Neutralization. Cell. 2019 Jul 25;178(3):567-584.e19.
Donaldson MM, Kao SF, Foulds KE. OMIP-052: An 18-Color Panel for Measuring Th1, Th2, Th17, and Tfh Responses in Rhesus Macaques. Cytometry A. 2019 Mar;95(3):261-263.
Immunological evaluations in NHP models are essential for the advancement of vaccine research. In particular, vaccination and simian immunodeficiency virus (SIV) challenge of macaque species is the best animal model for evaluating candidate HIV vaccines in pre-clinical studies. As a result, the Nonhuman Primate Immunogenicity Core (NIC) was established to support the research efforts of investigators at the VRC.
The NIC manages basic, translational, and preclinical NHP studies. Basic research interests include lymphocyte trafficking, the generation and maintenance of memory responses in systemic and mucosal sites, and pathogenesis following SIV infection. Translational research studies are performed to optimize a variety of different platforms and vectors by studying the effects of different adjuvants, schedules, and delivery methods on immunogenicity. Finally, the data from preclinical testing of vaccines in NHP models is used to support moving clinical products forward to human testing and can be critical to regulatory filings.
All studies that are conducted through the NIC are processed in a standardized manner using the same standard operating procedures (SOPs) for tissue preparation and T cell assays. For example, the NIC uses a qualified 19-color ICS panel and assay for the measurement of IFN-γ, IL-2, TNF, IL-4, IL-13, IL-17, IL-21, and CD40L from CD4 and CD8 T cells; memory T cell subsets and Tfh can also be identified using this panel. In addition, the NIC has qualified the Luminex assay for the measurement of 23 cytokines and chemokines from plasma or cell culture supernatant. To perform these operations, the NIC uses state-of-the-art technologies available at the VRC including 30-color FACS analyzers, 30-color sorters (one in BSL-3 containment), reagent manufacturing capabilities, and the Luminex system.
Other responsibilities of the NIC include the following: consulting on the design and implementation of NHP studies, maintaining a bank of NHP tissue samples for use by VRC investigators, designing new panels such as probe-binding B cell panels, and collating, analyzing, and coordinating data.
For more information on research conducted by Kathryn Foulds, Ph.D. visit the ImmunoTechnology Section.
Potter EL, Gideon HP, Tkachev V, Fabozzi G, Chassiakos A, Petrovas C, Darrah PA, Lin PL, Foulds KE, Kean LS, Flynn JL, Roederer M. Measurement of leukocyte trafficking kinetics in macaques by serial intravascular staining. Sci Transl Med. 2021 Jan 13;13(576):eabb4582.
Corbett KS, Flynn B, Foulds KE, Francica JR, Boyoglu-Barnum S, Werner AP, Flach B, O'Connell S, Bock KW, Minai M, Nagata BM, Andersen H, Martinez DR, Noe AT, Douek N, Donaldson MM, Nji NN, Alvarado GS, Edwards DK, Flebbe DR, Lamb E, Doria-Rose NA, Lin BC, Louder MK, O'Dell S, Schmidt SD, Phung E, Chang LA, Yap C, Todd JM, Pessaint L, Van Ry A, Browne S, Greenhouse J, Putman-Taylor T, Strasbaugh A, Campbell TA, Cook A, Dodson A, Steingrebe K, Shi W, Zhang Y, Abiona OM, Wang L, Pegu A, Yang ES, Leung K, Zhou T, Teng IT, Widge A, Gordon I, Novik L, Gillespie RA, Loomis RJ, Moliva JI, Stewart-Jones G, Himansu S, Kong WP, Nason MC, Morabito KM, Ruckwardt TJ, Ledgerwood JE, Gaudinski MR, Kwong PD, Mascola JR, Carfi A, Lewis MG, Baric RS, McDermott A, Moore IN, Sullivan NJ, Roederer M, Seder RA, Graham BS. Evaluation of the mRNA-1273 Vaccine against SARS-CoV-2 in Nonhuman Primates. N Engl J Med. 2020 Oct 15;383(16):1544-1555.
Van Rompay KKA, Keesler RI, Ardeshir A, Watanabe J, Usachenko J, Singapuri A, Cruzen C, Bliss-Moreau E, Murphy AM, Yee JL, Webster H, Dennis M, Singh T, Heimsath H, Lemos D, Stuart J, Morabito KM, Foreman BM, Burgomaster KE, Noe AT, Dowd KA, Ball E, Woolard K, Presicce P, Kallapur SG, Permar SR, Foulds KE, Coffey LL, Pierson TC, Graham BS. DNA vaccination before conception protects Zika virus-exposed pregnant macaques against prolonged viremia and improves fetal outcomes. Sci Transl Med. 2019 Dec 18;11(523):eaay2736.
Iwamoto N, Mason RD, Song K, Gorman J, Welles HC, Arthos J, Cicala C, Min S, King HAD, Belli AJ, Reimann KA, Foulds KE, Kwong PD, Lifson JD, Keele BF, Roederer M. Blocking α4β7 integrin binding to SIV does not improve virologic control. Science. 2019 Sep 6;365(6457):1033-1036.
Kong R, Duan H, Sheng Z, Xu K, Acharya P, Chen X, Cheng C, Dingens AS, Gorman J, Sastry M, Shen CH, Zhang B, Zhou T, Chuang GY, Chao CW, Gu Y, Jafari AJ, Louder MK, O'Dell S, Rowshan AP, Viox EG, Wang Y, Choi CW, Corcoran MM, Corrigan AR, Dandey VP, Eng ET, Geng H, Foulds KE, Guo Y, Kwon YD, Lin B, Liu K, Mason RD, Nason MC, Ohr TY, Ou L, Rawi R, Sarfo EK, Schön A, Todd JP, Wang S, Wei H, Wu W; NISC Comparative Sequencing Program, Mullikin JC, Bailer RT, Doria-Rose NA, Karlsson Hedestam GB, Scorpio DG, Overbaugh J, Bloom JD, Carragher B, Potter CS, Shapiro L, Kwong PD, Mascola JR. Antibody Lineages with Vaccine-Induced Antigen-Binding Hotspots Develop Broad HIV Neutralization. Cell. 2019 Jul 25;178(3):567-584.e19.
Donaldson MM, Kao SF, Foulds KE. OMIP-052: An 18-Color Panel for Measuring Th1, Th2, Th17, and Tfh Responses in Rhesus Macaques. Cytometry A. 2019 Mar;95(3):261-263.
