1. Corporate presentation,October 2013

2. Ideal Target Product Profile for multidrug resistant broad spectrum Gram-negative antimicrobial
Novel mode of action
Bactericidal
Selective and specific
Low frequency of resistance
Active against GAIN pathogens
Drugable 2

3. Several G+ but only one G- identified !
Arenicin selection process
Variant library generation
(~ variants)
~40 AMP’s identified
Several G+ but only one G- identified !
> 500 organisms screened for antimicrobial activity
NZ17074
Second variant library (~ variants)
1500 hits but only 10 variants selected
First Hit
Clin Cand – AA139 3

4. Mode of action summary
Arenicin acts at least partly due to non-lipid A-mediated penetration and disruption of both Gram negative membranes
Inhibition of cytosolic processes in protein synthesis suggested in TraDIS studies
Arenicins mode of action is different from Colistins 4

5. Arenicin interacts with the lipid transporter protein MLAC
MlaC is a periplasmic binding protein maintaining phospholipid homeostasis of the dual cell membrane
Gene analysis of E.coli shows that an MlaC L11R mutation is required to prevent the interaction between Arenicin and MlaC
Resistant strains regain sensitiviy to Arenicin as mutant is not stable 5

6. Arenicin causes loss of cell surface structure and partial cytoplasm clearing in E. coli
Transmission Electron Microscopy (TEM) of the Arenicin effect on E.coli (ATCC 25922)
E.coli. No treatment. Black arrow, cytoplasmic membrane; Red arrow, outer membrane; Green arrow, pili. Bar, 200 nm
E. coli incubated with 32 μg/mL NZ17000 for 40 min induced loss of cell surface structures and partial clearing of cytoplasm Bar, 200 nm
University of Queensland 2013 6

7. Arenicin causes ATP release without dramatic changes in cell morphology
At OD600 =0.4 E.coli cells were exposed to 32ug/ml Arenicin, 64ug/ml Fosfomycin and 16ug/ml Polymycin B. Even at very high concentration of Arenicin-3, no dramatic morphological changes of the cells were observed.
Extracellular ATP after 10 min
x MIC
Fold change
Arenicin-3 (Ar), colistin (col), and piperacillin (pip) induced release of ATP from E. coli. Exponential cells were incubated with drug for 10 minutes and ATP measured. y-axis is fold change relative to untreated (0xmic) and x-axis is fold MIC applied.
Novozymes A/S, 2010 7

8. Arenicin and Colistin have different effects on osmotic shock genes
Symbol
Gene ID
Colistin µg/ml
Arenicin µg/ml
Description 2½ 5 10
Osmotic shock genes
osmB
B1283 63 29 8 3
lipoprotein, osmotically inducible,
osmC
B1482 -
Peroxiredoxin
osmY
B4376
Osmotically-inducible protein Y precursor
osmE
B1739
Osmotically-inducible lipoprotein E precursor
bdm
B1481 11 6
Biofilm-dependent modulation proteinosmotically inducible
sra
B1480 4
rpsV, osmotically inducible
rcsA
B1951 2
positive regulator for ctr capsule biosynthesis,
Whole Genome TraDIS preliminary data suggest inhibition
of key cytosolic processes in gene translation
Novozymes A/S, 2010 8

9. In vitro efficacy summary
Potent in vitro activity against GAIN pathogens
Rapidly bactericidal – MBCs ~ MICs
Extremely low spontaneous mutational frequency
Small and mostly reversible increase of MIC in serial passage studies comparable with Colistin
Little inoculum effect
Moderate effect of serum on MIC
Limited effect of Survanta on MIC
No synergistic or antagonistic effect with other antibiotics
No cross resistance with strains with acquired resistance to Colistin 9

10. Potent in vitro activity against GAIN pathogens
MIC90 determinations (MDR clinical isolates)
# strains
AA139
Colistin
Meropenem
Ceftazidime
Ciprofloxacin
Gentamicin
Tigecycline
N=325
MIC (µg/ml)
E.coli
N=55 1
0.25 4
>32
>4
0.5
K.pneumonia
N=75 8
>16
P.aeruginosa 2 ND
A.baumanii
N=120
Eurofins medinet 2012 10

11. Extremely low spontaneous mutational frequency to Arenicin
Organism
Isolate ID
Resistance Frequency (4XMIC)
Resistance Frequency (8XMIC)
AA139
E. coli
ATCC 25922
≤2,50E-12
≤8,90E-11
K. pneumoniae
≤4,16E-10
≤1,38E-11
P. aeruginosa
ATCC 27853
≤2,61E-12
≤2,68E-12
A. baumannii
≤2,65E-12
≤4,80E-10
Eurofins medinet 2012 11

12. Limited effect of mucin (Survanta) on in vitro efficacy
NBS plates- MHB only
Compound ID
GN_01
E.coli
ATCC 25922
GN_03
K. pneumoniae
ATCC
GN_04
ATCC 13883
GN_34
A.baumannii
ATCC 19606
GN_42
P.aeruginosa
ATCC 27853
GN_43
Polymixin R
GN_44
ATCC 10536
GN_45
BAA_2146
MIC [μg/mL]
Colistin MCC_000094B
≤0.03
0.03/0.06
1/2
32/64
0.03/0.125
0.06
Ciprofloxacin  MCC_000166
0.25
0.5/1 2
0.25/0.5
>64
AA139
0.5
0.125/0.25
0.125
NBS plates-  MHB + 5% Survanta
Compound ID
MIC [μg/mL]
Colistin MCC_000094B
0.06
1/0.5
0.125
0.5/0.25
0.5 64
Ciprofloxacin  MCC_000166
≤0.03 2
>64 1
2/1
AA139
0.125/0.06
16/8
8x MIC increase
Decrease in MIC
University of Queensland 2013 12

13. Pharmacokinetic/pharmacodynamic summary
Arenicin efficacy is driven by Cmax
Clinical therapy should thus be based on slow bolus injection
Hepatic clearance does not seem to play a role
AA139 has a good volume of distribution corresponding to the extracellular volume
AA139 has a half life of 4.3 hours
AA139 has a low penetration into ELF (<5%) 13

14. PK/PD dose fractionation study shows that Cmax drives the efficacy of Arenicin (NZ17230)
(5mg/kg over 3 days seems to exert maximal effect)
Euprotec 2013 14

15. In vivo efficacy summary
Excellent efficacy against K.pneumoniae and E.coli in UTI with ED50 of 0.5-1mg/kg (BID I.V. administration)
Modest efficacy against K.pneumoniae, P.aeruginosa and A. baumannii in pneumonia based on QID I.V. administration
Very good efficacy against K.pneumoniae in pneumonia based on aerosol administration 15

16. Excellent efficacy in UTI
ED50 for AA139 in the bladder
E. coli
K. pneumonia
Euprotec 2013 16

17. Klebsiella Pneumonia NCTC13442
Very good activity of Arenicin against K. pneumonia in a neutropenic pneumonia model following aerosol admin
Klebsiella Pneumonia NCTC13442
Variant
log reduction
MIC
AA139
-3.89 1
Colistin
-1.75
Euprotec 2013 17

18. Toxicological summary
Selective and specific for bacteria
Wide therapeutic window – a factor of 25
MTD level of 25 mg/kg versus ED 50 of 1 mg/kg
Adverse effects related to histamin release
Changes in proximal tubuli the only, dose dependent and reversible pathological finding
Changes in NGAL correspond with pathological kidney findings
No cardiotoxic effect 18

19. Product profiles of Meropenem, Colistin and Arenicin
Indications
Meropenem
Colistin
Arenicin
Pneumonia
+++
Complicated urinary tract infections
Coverage
XDR E.coli ++
XDR P.aeruginosa
XDR A.baumannii +
KPC K.pneumonia -
Colistin G- Bacteria
Administration
Oral no IV
yes IT
Adverse events
Renal/Hepatic
(yes)
(no)
Neurological
Hypersensitivity
Miscellaneous
Bactericidal 19