Adenium Biotech Management: - Peter Nordkild, MD, CEO, ex Novo Nordisk, Ferring, Egalet - Søren Neve, PhD, project director, ex Lundbeck, Novozymes Board of Directors: - Stephan Christgau, PhD, chairman, Novo A/S - Anker Lundemose, MD, ex Novo Nordisk, ex OSI Pharmaceuticals - Khalid Islam, PhD, ex Arpida - Ejner Bech Jensen, MSc, VP R&D Novozymes A/S Current Investor: - Novo A/S
Bad Bugs need new drugs The total US market of hospital acquired infections was in 2006 estimated at USD 7.9 billion. 65% of hospital acquired infections are caused by Gram-negative bacteria (Clin Infect Dis 2005;41:848 854) Resistance
Resistance is inevitable and requires novel antibiotics Antibiotic Year deployed Resistance observed Sulfonamides 1930s 1940s Penicillin 1943 1946 Streptomycin 1943 1959 Chloramphenicol 1947 1959 Tetracycline 1948 1953 Erythromycin 1952 1988 Vancomycin 1956 1988 Methicillin 1960 1961 Ampicillin 1961 1973 Cephalosporins 1960s Late 1960s
Key antibacterial needs 2011 Source: Datamonitor 02/2011.
Arenicin A new Gram-negative antibiotic
Arenicin program highlights Spin out from Novozymes AMP group Novel mode of action, no cross resistance to existing antibiotics Bactericidal on broad range of multidrug resistant Gram-negative bacteria Development plan includes orphan drug opportunity Few competing bactericidal products in development Strong lead/back up product candidates Stable IV formulations Strong IP (2025-2030) Addresses significant unmet Gram-negative clinical need Large, growing and non-generic hospital market of USD 8 billion Hospital and primarily ICU based specialist target group requiring small sales force
Healthcare associated infections in the US 2010 Source: Datamonitor 02/2011
Nosocomial indications - volume/value Source : Datamonitor 03/2007
ESKAPE Pathogens Enterococcus faecium Staphylococcus aureus Klebsiella species Acinetobacter baumannii Pseudomonas aeruginosa Enterobacter E.coli E. aerogenes E. cloacae
Generation of new variants
Arenicin-3 NZ17000 Isolated from lugworm (Arenicola marina) 21 amino acids Very stable beta-hairpin structure MW 2.613 kda pi ~ 11.27
Variant generation through consecutive screening of yeast libraries 250.000 variants Evaluation of variants Output 1. screen S. cerevisiae (250.000) - plate screen vs. E. coli in the presence of 5% blood - retest vs. other Gram- bacteria 2. screen purified peptide (250) - MICs vs. 11 bacteria +/- serum - killing kinetic - protein binding 3. screen chemical synthesis (10) - a range of in vitro & in vivo parameters - MTD, peritonitis, cystitis etc NZ17143 and NZ17211 with distinct features Improved protein binding properties Improved toxicological properties 2 new Arenicin variants
Antimicrobial spectrum and protein binding of NZ17000, NZ17143 and NZ17211 Arenicin variant E. coli (μg/ml) K. pneumoniae (μg/ml) E. cloacae (μg/ml) P. aeruginosa (μg/ml) S. maltophilia (μg/ml) A. baumanii (μg/ml) M. catarrhalis (μg/ml) NZ17000 0.25 1 1 0.5 0.5 0.5 0.5 99 NZ17143 0.25 4 4 2 8 4 2 85 NZ17211 0.5 8 8 4 8 4 2 80 Protein Binding
Antimicrobial activity against selected resistant Gram-negative strains Bacteria No NZ17000 NZ17143 NZ17211 Carbapenem resistant Klebsiella pneumoniae (KPC2) Carbapenem resistant Pseudomonas aeruginosa Carbapenem resistant Klebsiella pneumoniae (NDM-1) MIC (µg/ml) MIC (µg/ml) MIC (µg/ml) 2 1 2 4 1 1 4 4 1 2 2 2 Tigecycline resistant Salmonella Hadar 1 1 1 1
MoA - localization of Arenicin 15 min 30 min 45 min Control Fosfomycin Polymycin B Arenicin-3 A. E. coli exposed for 30 min to NZ17000 and stained with TRITC. Treatment with NZ17000 results in influx of TRITC into the E. coli B. E. coli exposed for 30 min with TRITC labelled NZ17000. Clusters of NZ17000 were localized in the bacterial membrane 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.
Fold change MoA - ATP efflux after treatment with Arenicin Extracellular ATP after 10 min 25 Arenicin: 20 15 10 5 Ar col pip Perturbs the membrane potential increasing the permeability of the bacterial membrane 0 0 16 64 256 1024 4096 x MIC Inhibits the protein synthesis 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.
CFU/ml CFU/ ml of Pseudomonas Time kill of NZ17000 vs E.coli and Pseudomonas Time kill kinetics for Escherichia coli ATCC25922 NZ17000 in ca-müller Hinton broth 10 9 10 8 10 7 10 6 10 5 10 4 10 3 10 2 10 1 0 30 60 90 120 150 180 Time (minutes) Growth control 10 x MIC Arenicin-3 (5µg/ml) 10 x MIC Gentamicin (5µg/ml) NZ17000 Time killing kinetics for Pseudomonas aeruginosa ATCC27853 10 8 10 7 10 6 10 5 10 4 10 3 10 2 10 1 0 30 60 90 120 150 180 Time (minutes) Growth Control 0.25 x MIC NZ17000 ( 0.125µg/ml) 2 x MIC NZ17000 (1µg/ml) 10 x MIC NZ17000 (5µg/ml)
Dose response of NZ17000 in the UTI mouse model ED 50 ~1.5 mg/kg in urine and ~ 1.8 mg/kg in the bladder
Dose response of NZ17143 in the UTI mouse model ED 50 < 0.8 mg/kg in urine, bladder and in the kidneys
Dose response of NZ17211 in the UTI mouse model ED 50 < 0.8 mg/kg in urine, bladder and in the kidneys
NZ17143 NZ17211 NZ17000 Dose response in neutropenic mouse peritonitis model against a multi-resistant E. coli at 5 hours after treatment Blood Peritoneal fluid
Log 10 CFU/ml Log 10 CFU/ml Log 10 CFU/ml Efficacy of NZ17143/NZ17211 against multi-resistant E. coli in the murine Urinary Tract Infection model (UTI) 8 7 Urine 2 day post infection Urine day 2 post infection 8 7 Bladder 3 days post infection Bladder day 3 post infection 4 Kidney 3 days post infection Kidneys day 3 post infection 6 6 3 5 4 5 4 2 3 2 3 2 1 vehicle NZ17143 12.5 mg/kg NZ17211 20 mg/kg Meropenem 40 mg/kg vehicle NZ17143 12.5 mg/kg NZ17211 20 mg/kg Meropenem 40 mg/kg vehicle NZ17143 12.5 mg/kg NZ17211 20 mg/kg Meropenem 40 mg/kg
Pharmacokinetic properties after IV administration Arenicin-3 variant Protein binding (%) T ½ (min) AUC (min*ug/ml) C max (ug/ml) Bioavailability (%, SC vs IV) NZ17000 >99 130 532 7.7 12 NZ17143 85 69 324 7.5 70 NZ17211 80 60 432 9.9 60
Toxicological overview of NZ17000, NZ17143 and NZ17211 Variant NZ17000 (mg/kg) NZ17143 (mg/kg) NZ17211 (mg/kg) MTD (IV) 15 25 30 NOAEL (IV) 7,5 15 20
Arenicin summary New mode of action Spontaneous mutational frequency for E. coli is 3X10-9 and P. aeruginosa >10-8 Potent in vitro activity against a wide spectrum of Gramnegative bacteria Rapidly bactericidal MBCs ~ MICs No cross resistance to known antibiotics No or little inoculum effect Favorable efficacy in experimental animal models of infection Septicemia against E. coli and P. aeruginosa UTI against E. coli Thigh infections against E. coli.