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Anti-infective drug discovery at Evotec
Evotec, Anti-infective drug discovery, 2016
Evotec, an ideal partner in
anti-infective drug discovery
The different ways to work with us
On your specific
target or programme
Starting from a phenotypic
assay concept
On an existing
Evotec programme
Access to Evotec drug
discovery expertise and
capabilities to support
your programme
Access to Evotec phenotypic
screening expertise followed
by target deconvolution
leading into a drug discovery
programme
Sponsor an
established theme in
the anti-infective field
Flexible commercial solutions:
multiple business models available to suit our partners
Access to expert discovery platform as stand-alone activities or as part of
integrated drug discovery programmes
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Evotec’s foundations in anti-infective drug
discovery are built upon many years of experience
State-of-the art capabilities & extensive expertise
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Experienced anti-infective drug discovery team with >45 FTEs
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Experience encompasses multiple compound classes: Small molecules, natural products,
biologics, peptides, antibodies, combinations, biocides, vaccines
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Multiple drug discovery approaches from phenotypic screening to target-based discovery:
Folate, non-mevalonate, aromatic biosynthesis, protein synthesis, ribosome, virulence
attributes and resistance pathways
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Extensive portfolio of drug discovery capabilities:
 Medicinal chemistry and structure-based drug design
 Hit finding & library screening
 In vitro microbiology
 Molecular profiling/ Mechanism of Action determination
 Evostrain™: a unique strain collection
 In vivo models of infection, strongly coupled with PK/PD profiling expertise translating
discovery data to clinical trial design
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Contribution to
the discovery
and development
of multiple ‘antiinfective’ agents
including preclinical and
clinical
candidates
through to
marketed drugs
Maximising success in Hit Identification campaigns
HTS & MTS for BSL 2 biological agents
 >15 years of expertise in screening, from assay development to hit finding and
hit profiling with 40 HTS campaigns in the anti-infective space including
phenotypic and target based assays on bacteria or yeasts
 Primary screening for actives identification
 Screening against BSL 2 biological agents: human cells or micro-organisms
 Assay development and miniaturization
 HTS in 384 and 1536 well format
 Screening collection adapted to the targets or approaches (25K to 900K)
 Characterisation of actives/hits: diverse range of secondary assays
 Support for back screening and hit expansion
 State-of-the-art robotic platforms:
 HTS: RTS-2 robotic platform (up to 100 plates/run, 30-100K/run)
 MTS: 1 Beckman robotic system and 1 Agilent workstation
(42 plates/run, 15K/run)
 BSL 2 cabinet and containment
 Access to Evotec and Sanofi compound collections (400K & 700K compounds)
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Strong screening expertise with multiple
microorganisms and targets
Phenotypic and target-based assays
 Phenotypic assays: growth inhibition of microorganisms and cells
 Up to 6 strains screened in a single run
 Gram positive bacteria (Staphylococcus aureus incl. MRSA,
Streptococcus, Enterococcus spp …)
 Gram negative bacteria (Escherichia coli, Pseudomonas
aeruginosa, Klebsiella spp …)
 Mycobacterium species (Mycobacterium bovis …)
 Yeasts & fungi (Candida & Aspergillus spp …)
 Target-based assays
 Biochemical assays
 Cell-based assays with incubation of cells and pathogen
(infection of human cells by bacteria and intracellular killing)
 Cellular assays with reporter gene
(luciferase, beta-lactamase …)
 Readouts: fluorescence, luminescence, optical density, HCS …
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Track record
Assay type (%)
24
Target-based
Phenotypic
76
Micro-organism type (%)
44
23
Gram positive bacteria
Gram negative bacteria
Mycobacterium
Yeast & Fungi
69
Best-in-class microbiological
profiling and characterisation
Breadth and depth of microbiology expertise
 Antimicrobial Susceptibility Testing (AST) and hit follow-up
 Primary screening (0.1-1mg compound)
 Panels include fully susceptible and multidrug resistant strains; e.g. KPC,
NDM-1, ESBL; VRSA
 Large and evolving collection of highly characterised clinical isolates and
strains (EvostrAIn™)
 MICs and MBCs
 Standardised methods (CLSI, EUCAST, BSAC)
 Detailed screening against broad panels of pathogens
 Bespoke methods developed for non-conventional drugs
 Combination studies
 Chequerboard assays
 Assessment for synergy, additive activity, and antagonism
 Mathematical modelling of data
 Strong commitment to quality: Participate in the UK National External Quality
Assessment Service for Microbiology (NEQAS)
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EvostrAIn™ – a unique resource
for anti-infective drug discovery
Evotec’s collection of highly characterised strains and clinical isolates
 Evotec strain collection is a constantly evolving
resource of thousands of primary clinical isolates and
reference strains
 Broad collection of bacteria and fungi
 Clinics and culture collections
 Global, recent sources
 High degree of characterisation (susceptibility profiles,
mechanisms of resistance and in vivo drug response)
 Highly valuable collection used to establish spectrum of
activity and potential clinical use of new antibiotics and
antifungals
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EvostrAIn™
Overview: Gram positives, Gram negatives, Fungi
Bacteria: Gram positive pathogens
 Staphylococcus aureus including MRSA,
VISA and VRSA strains
 β-Haemolytic streptococci groups
A, B, C and G
 Streptococcus pneumoniae (incl. penicillin, macrolide, fluoroquinolone, cephalosporin & MDRSP resistant strains)
 Vancomycin Resistant Enterococci
(VRE)
 Bacillus species
 Listeria species
 Corynebacterium and
Propionibacterium species
 Clostridium difficile (multiple ribo-types
including 012, 027 and 078)
 Other Clostridia (incl. C. perfringens)
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Bacteria: Gram negative pathogens
 E. coli including Extended Beta
lactamase producing strains
 Klebsiella pneumoniae Carbapenemase
producing strains (KPCs & MDR)
 Acinetobacter baumannii incl. MDRAB
 Pseudomonas spp. including multiresistant strains
 Haemophilus influenzae
 Bacteroides spp.
 Neisseria gonorrhoeae and
N. meningitidis
 Intestinal pathogens: Vibrio spp,
Campylobacter spp, Salmonella spp,
Shigella spp, Yersinia spp.
 Legionella spp.
 Mycobacterium
Fungi
 Aspergillus spp. (including strains
resistant to azoles, polyenes and
echinocandins with known mechanisms
of resistance)
 Candida spp. (including strains resistant
to azoles, polyenes and echinocandins
with known mechanisms of resistance)
 Mucorales
 Cryptococcus
 Dermatophytes including Malassezia spp
and Trichophyton spp
 Fusarium
Protozoa
 Acanthamoeba spp
Highly validated and bespoke studies for target
identification and molecular profiling
Mechanism of action determination
 Resistant mutant generation to assist Mechanism-of-Action studies
 Serial passage in presence of drug or isolation from gradient agar plates
 Whole-cell random mutagenesis of bacteria
 Determination of mutation frequencies/stability or mutation
 Sequencing of mutants
 Verification that mutation correlates with resistance. Mutations can be
recreated in wild-type backgrounds where tools are available.
 Generation of targeted gene knockout mutants in a number of bacterial species
 Screening of mutant libraries for increased susceptibility or resistance
 Other molecular microbiology techniques available at Evotec include:
 Gene cloning & vector construction, expression studies (e.g. novel drug targets)
 Site-directed mutagenesis or random mutagenesis of target genes
 RT-PCR analysis of gene expression, use of reporter constructs
 State-of-the-art proteomics and biomarker platform for cellular target profiling
and mode-of-action studies
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Capabilities and expertise to reduce cycle time and
efficiently progress anti-infective programmes
An integrated synthetic and medicinal chemistry platform
Medicinal
chemistry
know-how
 Experience encompasses multiple compound classes
 Heterocyclic molecules, natural products, linear or cyclized peptides, carbohydrate
antigens) and targets (including Ribosome, DNA gyrase, Topo IV, Kinases)
 Optimising anti-bacterial activities for complex peptides & carbohydrates
 Transforming an anti-Gram (+) compound into an anti-Gram (-) compound
Synthetic
excellence




Purification
platform
 Guaranteed high chemical and chiral purity of molecules; multiple purification technologies
including SFC
 Separation of chiral compounds (intermediates or final compounds)
 Higher resolution techniques for difficult purifications (peptides, macrolides …)
Analytical
Chemistry
 Analytical support for >20 antibacterial & antifungal programmes (LI and LO)
 Expertise in analysis of multiple compound classes: small molecules, natural products,
peptides, unusual peptides, oligosaccharides, steroids, macrolides …
 Deconvolution of mixtures and structural elucidation of unknown natural products
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SBDD and computational design of antimicrobials
Compound library enhancement strategies
Rapid synthetic execution; ability to address difficult chemistry
Strong expertise in peptide synthesis (from 200mg to 10-15g scale)
A fully comprehensive in vivo
pharmacology offering
In vivo pharmacology/DMPK
 State-of-the-art animal facilities (BSL 2) equipped to house rodents,
immunocompromised animals, genetic models and other models
 In vivo services include: target validation, tolerability studies, DMPK
studies, PK 1)/PD studies and MoA studies, efficacy screening, in vivo
pharmacology profiling, pre-clinical imaging
 Custom design protocols and model development
 Full range of endpoints: Burden, (culture, qPCR), histology,
immunohistochemistry, morphometric analysis, cell and cytokines,
biomarkers
 Real time imaging of microbes during infection: IVIS
 Ongoing programme of new model development
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Pharmacokinetic profiling in
anti-infective drug discovery
From in vivo characterisation to bioanalysis and biomarker assessment
 Standard and specialised PK studies in multiple rodent species:
 Administration routes; intravenous (infusion), per oral, intraperitoneal, subcutaneous, intra
cerebrospinal, intramuscular, pulmonary (nebulized, aerosolized), iPrecio & Alzet pumps
Compound XXX plasma concentrations following intravenous and oral administration to male Han Wistar rats at nominal doses of 2 and 4 mg/kg Plasma Concentration (nM)
1000
100
intravenous
oral
 Sampling types: jugular vein cannulation, cardiac puncture, tail vein microsampling
10
1
0
4
8
12
16
20
24
 Matrices: blood, plasma, CSF, BALF, whole tissues, bile, urine and faeces
Time (h)
 PK in infected animals assessing impact of disease state on drug exposure
 Data directly translated into efficacy studies to optimise outcomes
 PK experiments designed to accompany PKPD profiling programmes
 State-of-the-art bioanalytics
 Highly sensitive biophysical methods including LC-MSMS
 Bioassay techniques: tracking biological activity
 Biomarker quantification: pathogen/infection specific and host response
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Delivering Rapid proof of concept
and detailed in vivo characterisation
Validated and bespoke pre-clinical efficacy models
 Study of multiple pathogens across large portfolio of infectious disease models
 Infecting pathogens: fully susceptible through to multidrug resistant (MDR)
 Multiple hosts and fully validated models of infection:
 Rat, mouse, guinea pig, hamster, cotton rat, rabbit
 Immunosuppressed and immunocompetent backgrounds
 Multiple sites/routes of infection including lung, thigh, blood (sepsis), skin, urinary tract, GI
tract, vagina, bone
 Routine and specialised routes and methods of infection and drug administration
 Constantly expanding portfolio of ‘specialised models’: e.g. Chronic models, C. difficile
infection (mouse and hamster)
 Endpoints: pathogen burden (culture / qPCR) and host response
 IVIS imaging of microbes during infection
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Translating compounds into the clinic
through PKPD profiling and discovery data
From discovery to clinical trial design
PKPD Profiling platform include
 Correlation of exposure with efficacy & drug response
 Magnitude of effect
 PD drivers and what parameters are important to achieve efficacy
 Understanding of post-antibiotic effect (PAE)
 Mathematical modelling of data
 In vitro PK/PD
 Humanising dosing in efficacy models:
 Significant expertise in the translation of efficacy in pre-clinical models and microbiological
assays to predict efficacy in humans
 Use and analysis of pharmacokinetic measurements determined in the alternative host
system to predict exposure in humans
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Case study: SMT19969: A novel agent for the
treatment of C. difficile infection
In vitro and in vivo pharmacodynamic profiling
Partners
Programme
SMT19969: A new drug for the
treatment of C. difficile infection
SMT19969
10 9
Control
1xMIC
5xMIC
20xMIC
LOD
10 8
CFU/mL
10 7
10 6
10 5
10 4
10 3
BI1 Ribotype 027
24
20
16
12
8
4
0
10 2
Time (hours)
100
100
Vehicle
VAN 10mg/Kg
FDX 2.5mg/Kg
FDX 1mg/Kg
SMT 25mg/Kg
SMT 12.5mg/Kg
60
40
C. difficile Ribotype 027
20
0
0
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Survival (%)
Survival (%)
80
5
14
10
15
20
Days Post Infection
25
30
Vehicle
VAN 10mg/Kg
FDX 25 mg/Kg
FDX 12.5 mg/Kg
FDX 2.5mg/Kg
FDX 1mg/Kg
SMT 25mg/Kg
SMT 12.5mg/Kg
60
40
20
0
C. difficile Ribotype 012
0
5
10
15
20
Days Post Infection
25
30
NOTE: Presented at ICAAC and ECCMID
Target
Outcome
Clostridium difficile
Detailed in vitro and in vivo
characterisation of SMT19969
to support progression into
clinical trials
 Full characterisation demonstrating that SMT19969
 Is bactericidal in vitro at ≥2xMIC
 Demonstrates prolonged PAE at 5xMIC that results in bactericidal
activity after 3 hours exposure
 Maximum clinical efficacy of SMT19969 is likely to occur following
exposure of >5xMIC for 3 hours
 Bridged data from in vivo pharmacokinetic studies demonstrates
such exposure is readily achieved in the GI tract following oral dosing
 Studies support the potential use of SMT19969 in the treatment of C.
difficile infection
 Comprehensive Tier 1 microbiology work up
 In vivo characterisation of SMT19969 in Golden Syrian hamster
model of C. difficile infection (CDI) against multiple strains of
Clostridium difficile
Case study: Characterisation of novel polymyxin B
analogues – in vivo models of Gram negative
bacterial infection
Pseudomonas aeruginosa, Acinetobacter baumanii
Partners
Programme
Target
Outcome
Development of novel
polymyxin B analogues with
improved safety profiles
MDR Gram negative bacteria
In vivo proof of concept
established in a novel
class of antibacterial agent
Efficacy in
neutropenic mouse
thigh model of
infection caused by
A. baumannii NCTC
13301 (OXA-23,
OXA-51-like)
10
9
8
7
6
5
4
3
2
LOD
CA824
20
10
3
1
20
0.
25
10
3
1
st
ar
t
en
d
0.
25
1
PMB
Treatment groups
(mg/kg SC, 2, 6, and 10 h, harvest at 16 h)
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 Multiple PMB analogues profiled in models of infection
(thigh. Lung): Pseudomonas aeruginosa, Acinetobacter baumanii
 CA824 is a semi-synthetic PMB analogue generated through
replacement of the N-terminal fatty acyl and diaminobutyate at position
1 with an iso-butyl piperazine.-2-carboxylate
 In vitro antibacterial activity of CA824 is comparable to PMB while in an
in vitro cytotoxicity assay against HK-2 cells it has an improved IC50
compared to PMB (F-734, F-739)
 In mouse lung infection models the antibacterial activity of CA824 was
superior to PMB against A. baumannii NCTC 13301 and P. aeruginosa
ATCC 27853
 In a mouse thigh infection model the antibacterial activity of CA824
against A. baumannii NCTC 13301 was comparable to PMB
Poster presentation F-735 ICAAC 2015, San Diego: Novel Polymyxin Derivative CA824: Efficacy in neutropenic mouse thigh and lung infection models
Case study: In vivo models of
Gram negative bacterial infection
PKPD profiling of new antimicrobial drugs
Partners
Programme
Target
Outcome
Early PKPD profiling
Gram negative infections
PKPD Profiling to support
clinical studies
 Dose fractionation experiments indicated: increased frequency of dosing of BAL30072, aztreonam, meropenem &
ceftazidime, results in greater efficacy as measured by a reduction in bacterial burden in a murine thigh infection model
 Designed and selected the appropriate models, dosing regimens, and showed how they can be used to
investigate the in vivo pharmacodynamic properties of developmental drugs against a selection of multi-resistant
Gram-negative organisms
 Profiled BAL30072 in vitro and in vivo against MDR bacteria including those expressing
ESBL, NDM-1, CTX-M, VIM10/VEB, AmpC and OXA
 Demonstrated that the in vivo efficacy of BAL30072 against most Enterobacteriaceae
and P. aeruginosa isolates was time dependent and rapid in vivo bactericidal activity was
induced when fT>MIC is greater than 50% for these species
 Produced data to show that BAL30072 could be an effective treatment option for MDR
and carbapenemase-producing Gram-negative bacteria and that further clinical
studies were warranted
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NOTE: Presented at ICAAC and ECCMID
Case study: In vivo models of
Gram positive bacterial infection
Staphylococcus aureus
Partners
Programme
Target
Outcome
Institut für Medizinische
Mikrobiologie, Universität
Zürich
Apramycin and discovery of
novel aminoglycosides
Gram positive and Gram
negative infections +
mycobacteria
In vivo proof of concept
established in a novel class of
antibacterial agent
 Development of Staphylococcus aureus septicaemia models with client
specific strains
 In vitro and in vivo characterisation of Apramycin and analogues
 Demonstrated that promising in vitro activity, including strains resistant
to multiple antibiotics, extends to in vivo activity in different models of
infection including experimental S. aureus septicemia
 The low ototoxicity of apramycin, together with its promising in vitro and in vivo
antibacterial activity has led to further preclinical investigation of this
unique aminoglycoside for treatment of infectious diseases
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40-O-substitutions determine selectivity of aminoglycoside antibiotics, Nature Comms, 2014.
In Vivo Efficacy of Apramycin in Murine Infection Models, Antimicrobial Agents and Chemotherapy, 2014, 58, 11
Identification and Evaluation of Improved 4=-O-(Alkyl) 4,5-Disubstituted 2-Deoxystreptamines as Next-Generation Aminoglycoside Antibiotics, mBio, 2014, 5, 5
Why us?
Evotec – The right partner in anti-infective drug discovery
A track record of
success means that
we consistently
deliver on our
clients’ needs
State-of-the-art
capabilities and
scientific excellence
will maximise your
chances of success
Fully integrated drug
discovery platform
and project management expertise
will accelerate your
drug discovery
programme
Flexible commercial solutions:
multiple business models available to suit our partners
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Evotec is a low-risk
outsourcing partner
who is continually
investing in its
platform to the
benefit of the
customer
Your contact:
[email protected]