NABATIVI –
Novel Approaches to Bacterial Target
Identification, Validation and Inhibition
Alessandra Bragonzi, PhD
Infections and Cystic Fibrosis Unit,
Division of Immunology, Transplantation and Infectious Disease
San Raffaele Scientific Institute
OUTLINE
• The problem
• Anti-microbial drug resistance
• Pharmaceutics' response
• EU’s response
• How we built the NABATIVI project
• International collaboration
• Multidisciplinary approach
• Scientific excellence
• Major achievement
2
OUTLINE
• The problem
• Anti-microbial drug resistance
• Pharmaceutics' response
• EU’s response
• How we built the NABATIVI project
• International collaboration
• Multidisciplinary approach
• Scientific excellence
• Major achievement
3
The world-wide threat of resistant pathogens
4
What is boosting antimicrobial resistance?
superbugs
Insufficient surveillance, prevention and control
Insufficient research and development activities
Insufficient funding
Insufficient coordination of EU efforts
Absence of new drugs
5
Why is antimicrobial resistance a global concern?
• kills
• hampers the control of infectious diseases
• threatens a return to the pre-antibiotic era
• increases the costs of health care
• jeopardizes health-care gains to society
6
Facts on antimicrobial resistance
• 4 million patients every year
• 25 000 deaths
• economic losses € 1.5 billion
7
Who is responsible?
Enterococcus faecium
Staphylococcus aereus
Klebsiella pneumoniae
Acinetobacter baumanni
Pseudomonas aeruginosa
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Enterobacter species
OUTLINE
• The problem
• Anti-microbial drug resistance
• Pharmaceutics' response
• EU’s response
• How we built the NABATIVI project
• International collaboration
• Multidisciplinary approach
• Scientific excellence
• Major achievement
9
Pharmaceutics’ response to antimicrobial-resistance
• The industry is cutting research in antimicrobial discovery and development
• Only few big pharmaceutical companies are still involved in antibiotic discovery
• Antibiotic R&D is a lengthy, costly, and risky process due to:
- length of time (10-15 years) from the discovery phase to market
- huge cost of bringing a new drug to market ($800 million to 1.7 billion)
- low reimbursement due to the small market
- low success rate
10
Payne et al. Nature Reviews Drug Discovery 6, 2007
Antibacterial pipeline, Big Pharma
New systemic antibacterial agents approved by the
US Food and Drug Administration per 5-year period,
through 2012.
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Boucher H W et al. Clin Infect Dis. 2013;56:1685-1694
How do we fill the gap ?
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OUTLINE
• The problem
• Anti-microbial drug resistance
• Pharmaceutics' response
• EU’s response
• How we built the NABATIVI project
• International collaboration
• Multidisciplinary approach
• Scientific excellence
• Major achievement
13
EU’s response –
6th and 7th Framework Programme
• Basic Science
• Prudent use of antibiotics
• New antimicrobials
• Point of care diagnostic tests
• Vaccines
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EU’s response –
Translational research for health (unit F3)
• 2.3.1 – Anti-microbial drug resistance
• Management of Gram negative multi-drug resistant infections.
2.3. TRANSLATIONAL RESEARCH IN MAJOR INFECTIOUS DISEASES:
TO CONFRONT MAJOR THREATS TO PUBLIC HEALTH
2.3.1. Anti-microbial drug resistance including fungal pathogens
2.3.1-1: Novel targets for drugs against Gram negative bacteria.
The objective is to identify and validate novel drug targets in order to select lead compounds, which may be
derived from natural sources or from synthetic compounds, for future development of a new class of antiinfective drugs against Gram-negative bacteria. Significant industrial involvement, particularly by SMEs, is
foreseen in this topic.
Funding scheme: Collaborative projects (Small or medium-scale focused research projects with maximum
EC contribution of € 6,000,000/project).
15
SME ≥30%
OUTLINE
• The problem
• Anti-microbial drug resistance
• Pharmaceutics' response
• EU’s response
• How we built the NABATIVI project
• International collaboration
• Multidisciplinary approach
• Scientific excellence
• Major achievement
16
How we built the NABATIVI project
• International Collaboration
Academic
Alessandra Bragonzi,
San Raffaele Scientific Institute, Milano
SMEs
Natalia Nekohotieva,
KDevExploratory, Stockholm
Miguel Cámara,
University of Nottingham
Gerd Döring,
Eberhard Karls Univerität Tübingen
Do Quoc-Tuan,
Greenpharma S.A., Orléans,
John A. Robinson and Leo Eberl,
University of Zürich
Giovanni Bertoni,
Università degli Studi di Milano
Peter E. Nielsen,
University of Copenhagen
17
Daniel Obrecht,
Polyphor, Allschwil
How we built the NABATIVI project
• Multidisciplinary approach
Molecular
microbiology
Clinical
research
Genomics
Biochemistry
Pre-clinical
research
Drugs
Structural
biology
Highthroughput
technology
Bioinformatics
18
Cell biology
NABATIVI approaches in response to Call FP7-HEALTH2007-B
Discovery Phase Approach A: “from target to lead compounds”
•Target identification/validation is done at the beginning
Discovery Phase Approach B: “from drugs to targets”
• Target identification runs in parallel to the discovery phase
Discovery phase
A
Target
identification
Library
selection/design
Screening
cascade
Hit
identification
Hit-to-lead
Lead
optimization
NDA
Launch
B
Pre-clinical and clinical phases
Pre-clinical
development
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Phase I
Phase II
Phase III
Pseudomonas aeruginosa selected from ESKAPE pathogens
• 5 million cases in Europe, USA and Japan every year
• Responsible of ≈ 30% of world-wide hospital-acquired
infections
• High risk of disease in people with cystic fibrosis, in
immunocompromised, burn patients, patients with cancer,
and with HIV
• Multi-drug resistant strains
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Scientific approach –
Genome - based approach for target identification
PAO1
6.3 Mb
5,570 ORFs
6,7% function experimentally demonstrated
(Class 1)
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Stover et al. Nature. 2000 Aug 31;406(6799):959-64.
NABATIVI scientific approach –
Step 1: genome-wide screening for targets identification
Transposon libraries
Antisense libraries
Screenings
University of Nottingham
Università degli Studi di Milano
Results
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NABATIVI scientific approach –
Step 2: pathogenicity in different model system
Genome-wide screening
57.360
University of Nottingham
San Raffaele Scientific Institute
Università degli Studi di Milano
University of Zürich
404
43
27
5
Eberhard Karls Univerität Tübingen
San Raffaele Scientific Institute
Eberhard Karls Univerität Tübingen
San Raffaele Scientific Institute
23
NABATIVI scientific approach –
Step 3: target inhibition by high-throughput technology
Genomic Target database
(GTD)
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NABATIVI scientific approach –
Step 3: target inhibition by high-throughput technology
Genomic Target database
(GTD)
Purified target-based HTS:
Determination of the druggability of
selected targets
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Greenpharma S.A., Orléans,
•
•
•
•
Plant extracts (800)
Pure natural compounds (480)
Synthetic compounds (45000)
Database AMBINTER 18 M
compounds
NH
B
N
O
KDevExploratory, Stockholm
O
NH
B
PNA
N
O
O
Antisense targeting of selected targets
by peptide nucleic acids
25
University of Copenhagen
Greenpharma S.A., Orléans,
NABATIVI approaches in response to Call FP7-HEALTH2007-B
Discovery Phase Approach B: “from drugs to targets”
• Target identification runs in parallel to the discovery phase
• Most antibiotics have been identified by this approach
Synthesis of a
library of
Protegrin I
analogues by
PEM technology
MIC determination
Target
identification
Library
selection/design
Screening
cascade
Initial hits with broad
spectrum
antimicrobial activity
and lack of
hemolytic activity
Hit
identification
Selective antiPOL7080
Pseudomonas
compounds
POL7001
with novel
SAR
Hit-to-lead
Lead
optimization
LptD, a β-barrel outer membrane transporter is the target of POL7001 and POL7080
Pre-clinical
developmen
t
Phase I
Phase II
Phase III
Efficacy studies in animal
POL7080:
models of sepsis and
preparations for a first Phase IIa clinical
respiratory infections
trial
NDA
Launch
under NABATIVI
NABATIVI scientific approach –
Step 4: target identification running in parallel to the
discovery phase
H3 N
H3N
H3 N
NH
H 2N
NH
H2N
N
H
O
H
N
O
N
H
O
O
H
N
O
N
N
H
Me
NH
O
O
O
H
N
H2 N
H2N
O
N
H
H
N
O
O
N
H
H3N H3N
H
N
O
N
H
HO
O
N
H
N
O
N
H
LptD
Srinivas N, et al. Science. 2010
Bragonzi A. Sci. Transl. Med. 2010
Polyphor Ltd.
University of Zurich
• LptD is an essential gene in P. aeruginosa and the target for novel drug POL7001
and POL7080
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NABATIVI scientific approach –
Step 4: translational research up to clinical trial
Polyphor Ltd.
San Raffaele Scientific Institute
Discovery phase
POL7080
POL7001




Pre-IND

Clinical phase

• Novel drug POL7001 is active against MDR P. aeruginosa strains
• In pre-clinical studies including models of airway infections and septicemia, POL7001
showed higher efficacy when compared to clinically approved antibiotics
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POL7080 indications for P. aeruginosa infections
Indications for Pseudomonas infections
Ventilator-associated
pneumonia (VAP)
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Urinary tract infections
Pseudomonas infection
in cystic fibrosis
(Mukoviscidosis)
NABATIVI - Major achievements
A genomic target database of P. aeruginosa is assembled
 Novel hits and PNAs are identified and are currently under validation
Discovery phase
A
Target
identification
Library
selection/design
Screening
cascade
Hit
identification
Hit-to-lead
Lead
optimization
 LptD as novel target for drug with novel mechanism of action is identified
 POL7080 successfully completed Phase I and will start Phase II at the end of 2013
Pre-clinical and clinical phases
B
Pre-clinical
development
30
Phase I
Phase II
Phase III
NDA
Launch
NABATIVI – novel drug
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NABATIVI – hit the target !
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Critical success factors for NABATIVI project
• Integrated multidisciplinary research at European level was
successful in novel drug discovery;
Molecular
microbiology
• Academia and a small innovative company worked
together productively to fill the gap left by the migration of
big pharmaceuticals away from antibacterial development;
• Combined scientific approaches, including exploitation of
post-genomic information, were successful in identifying
novel drugs;
• Optimal translation of the results into the clinic through
appropriate pre-clinical models
33
Clinical
research
Pre-clinical
research
Genomics
Drugs
Biochemistry
Structural
biology
Highthroughput
technology
Bioinformatics
Cell biology