Laboratory of Enteric and Sexually Laboratory of Enteric and Sexually Transmitted DiseasesTransmitted Diseases Historical PerspectiveHistorical Perspective

OriginOrigin: Began in 1994 - Established to review the : Began in 1994 - Established to review the rapidly increasing number of enteric disease products rapidly increasing number of enteric disease products and expected STD products.and expected STD products.

Mission:Mission:a. Conduct basic/applied research a. Conduct basic/applied research

molecular bases of pathogenesismolecular bases of pathogenesis host’s immune responses to infectionhost’s immune responses to infection develop models to measure vaccine safety, immunogenicity develop models to measure vaccine safety, immunogenicity

and efficacyand efficacy

b. Utilize knowledge baseb. Utilize knowledge base enhance product manufacturing analysesenhance product manufacturing analyses better assess product safety and efficacybetter assess product safety and efficacy

Program Review:Program Review:Last scientific program site-visit review - Aug. Last scientific program site-visit review - Aug.

20022002

Laboratory of Enteric and Sexually Laboratory of Enteric and Sexually Transmitted DiseasesTransmitted Diseases Regulatory ResponsibilitiesRegulatory Responsibilities

Responsible for review of IND’s and BLA’s for bacterial enteric, Responsible for review of IND’s and BLA’s for bacterial enteric, UTI and STD products and certain other assigned products.UTI and STD products and certain other assigned products.

LESTD is responsible for covering products related to intestinal LESTD is responsible for covering products related to intestinal disease agents (e.g. disease agents (e.g. ShigellaShigella, , SalmonellaSalmonella spp., all pathogenic spp., all pathogenic Escherichia coli’s, Campylobacter, Vibrio cholerae, Helicobacter Escherichia coli’s, Campylobacter, Vibrio cholerae, Helicobacter pylori, hookwormpylori, hookworm), the use of live attenuated enteric bacteria as ), the use of live attenuated enteric bacteria as vaccine vectors or as anti-cancer therapy, UTI pathogens, vaccine vectors or as anti-cancer therapy, UTI pathogens, probiotic/LBP therapy (e.g. Tx cancers, IBD, CF), L-aspariginase, probiotic/LBP therapy (e.g. Tx cancers, IBD, CF), L-aspariginase, bacteriophages or bovine/chicken immunoglobulin concentrates for bacteriophages or bovine/chicken immunoglobulin concentrates for therapeutic use, and genetic hybrid plant vaccines. [ Involves therapeutic use, and genetic hybrid plant vaccines. [ Involves oral, parenteral, intrarectal, intravaginal, intranasal, oral, parenteral, intrarectal, intravaginal, intranasal, transcutaneous routes + new adjuvants].transcutaneous routes + new adjuvants].

Annual workloadAnnual workload - -New candidate products = >1/monthNew candidate products = >1/monthReview time commitment:Review time commitment:

Stibitz and Kopecko - ~50% review effortStibitz and Kopecko - ~50% review effortCarr and Osorio - 25-30% review effortCarr and Osorio - 25-30% review effortBhattacharyya - ~ 80% review effortBhattacharyya - ~ 80% review effort

Laboratory of Enteric and Sexually Transmitted Diseases

Personnel and Budget - (2003-2006)Personnel and Budget - (2003-2006)

PersonnelPersonnel -Molec. Pathogenesis Gp. - increased from 2 to 7Gene Regulation Gp. - declined from 5 to 3STD Gp. - inactivated due to lack of STD vaccine

activity and departure of C. Deal.Immune Mechanisms Gp. - created in 2002, new research

initiatives being developed

BudgetBudget - FDA intramural research budget continued its decrease.

Fortunately, outside grant support replaced the intramural budget decline.

Problem: lack of internal funding consistency, outside grantors limited in number (e.g. special NIH funding).

Public Health IssuePublic Health Issue:: Enteric bacterial diseases cause ~350 million US Enteric bacterial diseases cause ~350 million US

episodes of diarrhea/yrepisodes of diarrhea/yr kill more than 2 million children worldwide annuallykill more than 2 million children worldwide annually Limited data on pathogenesis and immune responses has Limited data on pathogenesis and immune responses has

slowed the development of enteric vaccine productsslowed the development of enteric vaccine products

Research Relevance to DHHS and FDA Priorities:Research Relevance to DHHS and FDA Priorities:LESTD research programs are entitled: LESTD research programs are entitled: “ “ Enteric Bacterial Pathogens – Improving Safety and Enteric Bacterial Pathogens – Improving Safety and

Efficacy of Combination Vaccines for Diarrheal Efficacy of Combination Vaccines for Diarrheal Diseases and Select Agents”Diseases and Select Agents”

““Bacterial Vaccine Safety: Biomarkers of Virulence Bacterial Vaccine Safety: Biomarkers of Virulence and Attenuation in and Attenuation in Bordetella pertussisBordetella pertussis (Whooping (Whooping Cough) and Anthrax Bacteria.”Cough) and Anthrax Bacteria.”

Laboratory of Enteric and Sexually Laboratory of Enteric and Sexually Transmitted DiseasesTransmitted Diseases Relevance of Research Program To CBER’s Public Relevance of Research Program To CBER’s Public Health MissionHealth Mission

Laboratory of Enteric and Sexually Laboratory of Enteric and Sexually Transmitted DiseasesTransmitted DiseasesResearch Programs - 1Research Programs - 1

Molecular Pathogenesis SectionMolecular Pathogenesis Section - -

I.I. Studies of Studies of SalmonellaSalmonella Typhi Ty21a as a safe Typhi Ty21a as a safe vector for expression of foreign antigensvector for expression of foreign antigens

II.II. Pathogenesis and immune responses to Pathogenesis and immune responses to Campylobacter jejuniCampylobacter jejuni infection in humans infection in humans

III.III.Minor research projects - aided by key Minor research projects - aided by key collaborations.collaborations.

I. Studies of I. Studies of SalmonellaSalmonella Typhi Typhi Ty21a as a safe vector for Ty21a as a safe vector for

expression of foreign antigensexpression of foreign antigens

Key GoalsKey Goals

Define key attenuating features of Ty21aDefine key attenuating features of Ty21a (Genomic sequence and microarray (Genomic sequence and microarray analyses)analyses)

Optimizing a temperature-stabilized Optimizing a temperature-stabilized formulation and delivery systemformulation and delivery system(NIAID-supported collaboration with Aridis (NIAID-supported collaboration with Aridis Pharmaceuticals LLD)Pharmaceuticals LLD)

Demonstrate stable expression of Demonstrate stable expression of targeted targeted Shigella Shigella or anthrax genes in a or anthrax genes in a live platform vector systemlive platform vector system

2. 2. Pathogenesis and immune Pathogenesis and immune responses to responses to Campylobacter Campylobacter

jejunijejuni infection in humans infection in humans

Overall goals of project:A. Examine C. jejuni attachment,

invasion, and exocytosis events by TEM and SEM.

B. Define host signal transduction pathways involved in C. jejuni internalization into epithelial cells

C. Study C. jejuni interaction with human DCs for cytokine/chemokine responses involved in inflammation

Laboratory of Enteric and Sexually Laboratory of Enteric and Sexually Transmitted DiseasesTransmitted DiseasesResearch Programs - 2Research Programs - 2

Immune Mechanisms SectionImmune Mechanisms Section

New DirectionsNew Directions – Evaluate – Evaluate approaches to achieving mucosal approaches to achieving mucosal immunizationimmunization

Immune Mechanisms SectionImmune Mechanisms SectionMucosal Immunization against Enteric PathogensMucosal Immunization against Enteric Pathogens

1. Study methods useful for inactivation of enteric bacteria

2. Demonstrate immunogenicity of antigen delivery systemsa. Transcutaneous immunizationb. Bacterial ghostsc. Mucosally delivered whole cells

3. Develop Animal Models for Evaluating Vaccine Efficacy Against Enteric Pathogens

Laboratory of Enteric and Sexually Laboratory of Enteric and Sexually Transmitted DiseasesTransmitted DiseasesResearch Programs - 3Research Programs - 3

Gene Regulation SectionGene Regulation Section - -

I. Virulence Gene Regulation - using I. Virulence Gene Regulation - using B. B. pertussispertussis as model system. as model system.

II. Development of Genetic Tools for the II. Development of Genetic Tools for the Analysis and Manipulation of Analysis and Manipulation of Bacillus Bacillus anthracisanthracis..

I. Virulence Gene Regulation - using I. Virulence Gene Regulation - using B. pertussisB. pertussis as model systemas model system

A. Molecular mechanisms of virulence gene activationby the BvgAS two-component system

Progress: Completed a study of eight different BvgA-regulated virulence gene promoters for high-resolution mappingof BvgA-binding sites using BvgA conjugated to Fe-BABE.

Future Directions: More detailed studies of specific promoters and comprehensive genetic studies to elucidate critical interactions of BvgA with RNA polymerase.

I.I. Virulence Gene Regulation - using Virulence Gene Regulation - using B. pertussisB. pertussis as model systemas model system

B. Development of Genetic Tools

Progress: Have constructed a powerful allelic exchange systemfor manipulation of unmarked B. pertussis strains (for usein animal studies)

I.I. Virulence Gene Regulation - using Virulence Gene Regulation - using B. pertussisB. pertussis as model systemas model system

C. Use of bioluminescence to characterize B. pertussis infectionin a mouse aerosol challenge model.

Progress: Have demonstrated utility of in vivo imaging system for examining pertussis infection - can follow individual mice over time.Characteristic growth in upper respiratory (nose) tract followed bygrowth in lungs and trachea.

Future Directions: Implementation to characterize the effectof mutations that affect key/novel virulence genes and regulatory phenotypes.

2. Development of Genetic Tools for the 2. Development of Genetic Tools for the Analysis and Analysis and Manipulation of Manipulation of Bacillus anthracisBacillus anthracis

1. Developed routine allelic-exchange procedures

2. Constructed over 70 directed mutants of B. anthracis.

Future Directions:

1. Development of additional tools: expression vectors, promoter assay vectors, transposon delivery vectors.

2. Application of tools in a genomic search for new virulence genes (funded by MARCE developmental grant)

3. Characterization of underlying causes for sub-optimal stability of rPA anthrax vaccine (funded by CBER/NIAID partnership)