1

CONFIDENTIAL

Integrated synbio tools to

enable biopharma production

systems

Ian Fotheringham

Ingenza Ltd.

www.ingenza.com26th July 2017

2

Ingenza Ltd

Capabilities: 41 staff: (chemists and bioscientists)

Strain construction / protein expression / enzyme evolution

Microbial fermentation / Cell culture

Bioprocess development, DSP, chemical synthesis

All integrated for Industrial Biotechnology / Synthetic Biology application

Customers/Partners where synthetic biology applies:

Chemical companies developing biobased chemical syntheses

Pharma companies outsourcing pathway optimization, enzyme evolution

Academic groups transitioning early stage research to POC and spinout

COGS, new product/functionality, IP protection, sustainability

New synthetic routes, COGS, product diversification…funding (SME)

PoC, feasibility, viability, funding (spinout)

3

• For each target chemical, polymer intermediate or pharmaceutical

• Expression of target enzymes in novel pathways

• Rapid iteration to improve productivity

• Enzyme adaptation

• Confirmed process Feasibility, Economics and Scalability

Engineered pathways to high value chemical building blocks

v vvvB P

vv

Combinatorial assembly

‘Omics interpretation, Revision

PhylogeneticEnzyme Sampling

Screening / Process Testing

ScalableBioprocess

Pilot and Full ScaleManufacturing

4

Delivering predictability through methodologyAdaptable hierarchical screening

Calibrated screen dev

eloped for biotransfo

rmation

Adapted to optimis

e

biologic synthesis

Progression to drug

substance manufactu

re

46,000 novel

codon revisions

ReporterLibraryHighest producers

identified

5

inABLE® – Combinatorial gene assembly

Key advantages:

One-step assembly

No PCR (errors)

Re-usable library of parts

No purification, fully automated

Synergistic with versatile HTP screening (solid/liquid phase)

Part Cv Part DPart A Part B v

vPart Cv Part DBPart A P Part B

Part DvPart CvPart BPart A

Enzyme activity

Toxinresistance

Crossfeeding

Colorimetric

AutomatedLCMS

Design/preparation

Part/linker fusion

Pathway assembly

6

POC for bioprocess patents to key intermediates

Target enzymes expressed

Biotransformations and products characterised

> 40 independent steps in novel configurations

Reactions exemplified … IP established

Enabled multiple development programs to targets

Feasibility

Viability

Scalability

Enzymatic activity determination/measurement

Enabling innovative bioprocess developmentBiobased chemical manufacture

7

Delivering predictability through methodology‘Omics informed design and revision

X X

X X

Phylogenetics, metabolomi

cs

structural biology

Precise genome integr

ation

Reduced processing

Bioethanol efficiency s

avings

Hierarchical screen/t

est

v vvvB P

vvinABLE® assembly

Serine integrase

adapted to yeast

8

Delivering predictability through methodologyPredicting optimal translation

Modeling heterolo

gous mRNA transla

tion

Assembly and testin

g with target biolo

gic

v v

vvB P

vv

kDa16011060

40

3020

1510

20% uplift

> 9 g/L

5 L fed-batch fermentor validation

High volume/high value product –now comme

rcial

Aberdeen University, UK.Prof. Ian StansfieldProf. Carmen Romano

“tRNA abundance/ribosome hopping”

9

Target protein almost completely insoluble in E.coli:

Resistant to all attempts to achieve soluble production

Unaffected by host tRNA adaptation, shake flask or fermentation conditions

No activity in cell extracts

Target

Insolublefraction

Solublefraction

Efficient protein expression systemsHorses for courses

Optimised Pichia pastoris production system:

No 3rd party IP, cGMP compliant

Efficient, high cell density fed-batch

fermentation

Product secreted, soluble and active, now

commercial

10

Ingenza and Plymouth University researchers

• nM potency for Gram + pathogens (MRSA)

• Nasal decolonisation market

• Single dose efficacy vs (15 dose ) market leader

cfu/n

ostri

l

U n tre a te d

V e h ic le

N 1 0 1 , 0.8 % , O

N C E

N 1 0 1 , 0.0 4 % , B

ID

N 1 0 1 , 0.2 % B ID

Mu p iro c in , 2

% B ID1 0 0

1 0 1

1 0 2

1 0 3

1 0 4

1 0 5

1 0 6

1 0 7

L o g R = 2 .1 0 .0 9 0 .0 6 2 .5 9

Untreated Control NI01

Single dose

Antimicrobial discovery / production / engineeringEpidermicin NI01 bacteriocin

Highly promising product class

Low native yield

Adaptable target range:

Potency towards Gram - pathogens

J Antimicrob Chemother (2017) 72 (3): 778-781.

11

> 50x yield

1 2 3 4 5

6 M

Fully active

$1.5 M funding for process scale-up and peptide adaptation

• Spinout (“Kaiju”) progressing lead compound to toxicology

• Machine learning informed adaptation to enhance epidermicin properties: Ingenza-Plymouth-NPL-STFC-IBM

• Solid/liquid phase screens in P. pastoris

Gene configuration, integration, induction

Antimicrobial discovery / production / engineeringEpidermicin NI01 bacteriocin

> 50x productivity, viable, scalable, fermentation protocol

12

Need and opportunity :

500,000 (WHO) untreated haemophiliacs (75%) mainly in poorer nations

Requirement for low cost recombinant Factor VIII

Current cell line yield unable to support low cost market entry

$8 BN market today

Partnership of ProFactor Pharma and Ingenza: synergy of know-how / assets

High producing Factor VIII cell l ine, process

Identify novel construct and manufacturing process

Factor VIII

13

Codon optimised FVIII - CHO

Proprietary FVIII +VWF chaperone

Efficient, scalable SUF perfusion protocol

Proprietary DSP

10x industry standard productivity FVIII

+

B-domain

deletion

High producing Factor VIII cell l ine, process

14

Ingenza:

Worldwide partnerships with leading manufacturers

Enabling new products /markets /functionalities / University spinouts

Adaptable enabling technologies:

Combinatorial inABLE® genetic assembly

Gene design, targeted gene integration, ‘Omics

Scalable fermentation bioreactor protocols including single-use

Our Technology and Business focus is on:

Long-term high-value relationships

Scalable, cost-effective, sustainable bio-manufacturing

Synthetic Biology design plus proven Industrial Biotechnology methods

Bioprocess efficiency Requires “Abilities”(feasibility, adaptability, predictability, scalability)