synthetic biology technology to boost green chemicals ... · in the synthetic genomics lab to,...
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Philippe Gabant • PhD in Molecular Biology ULB + Master in Innovation Management
HEC Brussels• Inventor of patent portfolio about selection by « poison-antidote »
(owned by ULB)• Founder and CEO of Delphi Genetics (DG) 2001 - 2010• Head of Business Development DG from 2010 until June - 2012
Guy Hélin
• Master in Business Engineering (Solvay Business School, ULB)
• Consultant for technology focused SME (including MBO participation)
• Consulting / Senior Management for large corporations in Belgium and Europe
• CFO (part-time) from 2001 to 2012 of DG and CBO (Chief Business Officer) 2012 until May - 2014
Scientific management
Business management
The Founders:Working together > 15 years
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Scientific background
Genetic selection (definition)• A selection is an approach allowing only a sub-population of interest to survive• The result is an enrichment of only individuals presenting the behavior of
interest (i.e. production of a certain compound)
Development of selections1. In the 1990 selection for bacteria (clones) having recombinant plasmids
(Inventor: Dr. Philippe Gabant – commercialized through Life Technologies)2. In the 2000 selection for biopharmaceutical expression system without
antibiotics (Inventor: Dr. Philippe Gabant- commercialized through Delphi Genetics)
3. From 2013, selection technology by Syngulon adapted to industrial biotechnology (Dr. Gabant, inventor and CSO)
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Toxin-Antitoxin Technology 1
Genetic engineering selection standard
Key issues• Selection of recombinants /
expressing clones• Alternative to antibiotic selection
(to follow regulatory guidance)
Minor issues• Genetic containment (via physical
containment, sterile environment)
Non-issues: control of contaminants, partners control, security/competitors
(ULB patents, Licensing by Delphi Genetics)
Plasmid
Antidote (CcdA)
Target
Gene of interest
Poison (CcdB)
Need: replacement of antibiotic selection
Approach: use of the bacterial poison gene (ccdB) and its natural antidote (ccdA). During production bacteria loosing the plasmid are killed (suicide mechanism)
Applications: antigen production, recombinant biocompounds production, pDNA
Closed and controlled batch production – mono-cellular
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Toxin-Antitoxin Technology 2
Technology licensed by Delphi Genetics to:
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Applications of microorganisms in industries
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“ it may be possible to create high-value substances such as biofuels bymodifying simple organisms such as E. coli bacteria.” May 2013, “Next-generation genomics” in Disruptive technologies: Advances that willtransform life, business, and the global economy (McKinsey GlobalInstitute)
A Biobased Chemistry
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• Improvements in energy storage
• An intensification of biotechnological processes
Synthetic Biology: An approach with many industrial applications
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Synthetic Biology: industrialization concept “IT versus genes”
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Synthetic Biology: Industrialization concept “IT versus Genes”
“Blank” chassisConstructed by modules (parts)
Behavior code basedNon self replicative
Possible contamination by external code
“Evolutionary” based chassisConstructed by modules (parts)
Behavior code basedSelf replicative
Possible contamination by external code
Similarities with IT exists but fundamental differences
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The facts
1. Tools developed initially for sequencing human genome economically become accessible
2. Need for new technologies to meet the specific constraints in industrial biotechnology
Vision of Syngulon
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To Provide:
Genetic technologies in order to make microbial strains involved in industrial process more efficient and safe.
Mission of Syngulon
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Industrial biotech environment
GenosphereInside
Outside
Genetic code of interest
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The Problem:
How to control industrial Microorganisms?
KEY ISSUES
1. Gene containment2. Yield increase by industrial Genetic Bio-control 3. Contamination prevention against microbial invaders 4. Genetic drift
Self replicative
Industrial and large-scale applications need a selection to design a Genetic Firewall
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Outside
Inside
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• Killing the industrial Microbes (microrefineries)
outside of the refineries: Security and Gene
containment
• Control bacterial invaders: genetic shielding
• Increase production stability of the microrefineries
The Solution
New Genetic control designed for Industrial biotechnology
"Give me a fulcrum and I will move the world," from Archimedes, gets updatedin the synthetic genomics lab to, "Give me a selection, and I will change the world." George M. Church in "Regenesis: How Synthetic Biology Will Reinvent Nature and Ourselves” (Oct 2012)
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New Selection context = New Genes need to be used
• Micro-refineries are in interaction with the
environment: Security and Gene containment issues
• Control bacterial invaders: genetic shielding
• Increase production stability of micro-refineries
• Development of a genetic firewall based on
Bacteriocins Selection Loci
• Solution and response to need:
Syngulon’s novel technology designedfor industrial biotechnology
Genosphere
Microcin B17 used in E. coli BL21
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Bacteriocins:
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Microcin B17 in action: microfluidic view
WITHOUT MccB17(pACY)
WITH MccB17 (1h)(pCID)
Growth of E coli (BL21) in the B04 CellASIC ONIX microfluidic plate (Merck Millipore). Cells were perfused with LB medium and imaged on a 100X oil immersion phase contrast objective (Zeiss Observer Z1).
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Production of Pediocin PA-1 (PedA) by Yeast Saccharomyces cerevisiae
Pediococcus Pentosaceus(HELA: CWBI-B29)
Pediococcus Pentosaceus was used as an indicator organism in the MRS soft agar overlays.
Yeast colonies were 2 days old and overlaid plates were incubated 18h at 37°c.
The inhibition zones against P. Pentosaceus were visible around the yeast coloniestransformed with plasmid (pACTII-PedA) containing PedA gene.
pACTII pACTII-PedA pACTII-ColE1
Syngulon Technology for industrial biotech
Key issues• Genetic containment • Genetic security / competitors• Selection of recombinants presenting
the designed behaviour• Alternative to antibiotic used to fight
environmental contaminants)• Control of contaminants (bacterial
invaders, divergent micro-refineries, escaping host)
• Genetic drift• Partners control
Non-issues (because impossible) : physicalcontainment, sterile
(Syngulon US Patent 9,333,227)
Immunity
Target
Genetic code of interest
Bacteriocin (molecule
naturaly secreted to kill
or control other
bacteria)
Need : selection not only focused on the host of interest (suicidal approach) but also allowing the production microorganisms (micro-refineries) to control its growing environment and limit the spreading of micro-refineries outside the production plan
Approach : bacterial bacteriocinsloci are adapted for a “muti-dimensional” selection controlling the hosts and environment during production
Applications : genetic vigilance of the production plant allowing continuous production
Open / semi-open continuous production – could be multicellular
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Plasmid
43/46 are Kan resistant
Strain transformed with “auto-selection” plasmid (pMccV/ColV)
Negative control
After transformation, bacteria are spread on medium without selection agent
Picked on LB + Kanamycin
Verification of plasmid contain
All candidates (5/5) contain the plasmid
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Auto-selection with MccV/ColV operon
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Bacterial Genetics and Physiology Lab (LGPB)Université Libre de Bruxelles (ULB)
Pr Laurence Van MelderenDr Frédéric Goormaghtigh
ACKNOWLEDGEMENTSScientific Advisory Board
Pr Joseph Martial (Chairman), ULg, Liège (BE)Pr Bruno André, ULB, Brussels (BE)Adj-Pr Mike Chandler, University of Georgetown (USA)Pr Pascal Hols, UCL, Louvain-la-Neuve (BE)Pr Didier Mazel, Institut Pasteur, Paris (FR)Dr Régis Sodoyer, ex-Sanofi Pasteur, Lyon (FR)Pr Laurence Van Melderen , ULB, Charleroi (BE)Pr Ruddy Wattiez, UMons, Mons (BE)
SYNGULON Team
Guy Helin, Co-Founder, CEODr Philippe Gabant, Co-Founder, CSO
Dr Mohamed El Bakkoury, CTO YeastBertrand Delahaye, R&D ScientistThomas Gosset, Ir, R&D EngineerDr Johann Mignolet, R&D Project Manager
