ANTIBACTERIAL POLYMYXIN B ANALOGS

Programa Cooperación Farma-Biotech Jornada 1-2012: Áreas Terapéuticas de Inflamación, Infección y Respiratorio

Barcelona, 14 de marzo de 2012

Antibacterial Polymyxin B analogs

Francesc Rabanal Anglada

A project of:

Managed by:

Content

1. The Research Group 2. The Product

a) Target indications b) Innovative mechanisms of action c) Differential features facing the market d) Current status of development e) IPR protection f) Pitfalls & Risks to be considered

3. Partnering Opportunities


Research Group - Team and collaborators

Dr. Francesc Rabanal (PI) (Dept. Organic Chemistry, UB) Design and Synthesis

Dr. Yolanda Cajal (Dept. Physical Chemistry, UB)

Biophysical Studies MoA


Antimicrobial activity evaluation (Dept. Microbiology, UB) Dr. Ángeles Manresa Antimicrobial activity in MDR bacteria (UB, CRESIB, CEK, Hospital Clínic) Dr. Jordi Vila In vivo studies (CERETOX/ UTOX-UB, Parc Científic de Barcelona) Dr. Miquel Borràs

The Product. Target Indication

Need of new antibiotics ▫ Infectious diseases Second cause of death in the world (third in the

developed world) ▫ Inadequate use of antibiotics Increase in resistant bacteria

Antibiotics loose effectiveness ▫ Reduced pipeline of new antibiotics


AntiMicrobial Peptides (AMP) - Innovative mechanism of action • New antibiotics with novel MoA • AMPs: act on bacterial membrane (no enzymes) lower risk of resistance (or reversible resistance)

Zasloff, Nature, 2002

Jornada 1-2012: Áreas Terapéuticas de Inflamación, Infección y Respiratorio

...facing de market

• Need of new antibiotics • Remergence of Polymyxin



Objectives and differential features • Design and synthesis new antibiotics: lipopeptides based on the

structure of polymyxin/colistin

• Looking for... Activity in resistant and multi-resistant bacteria Lower toxicity Broader spectrum of activity Chemical synthesis designed for optimal scale up

Target IDTarget

ValidationLead

Optimization Pre-Clinical

Drug Discovery Animal Test

Clinical Trials Commercialization


Objectives and differential features

Design & Synthesis Of New Compounds

In vitro Efficacy

(MIC)

MoA SAR

In vitro TOX

IN VIVO EFFICACY

IN VIVO TOX. (LD50)

Design

Synthesis

In vitro Efficacy

MoA

NH2

HNO

NH

NH2

O

HN

ONH

OH2N

NHO

NH

O

SS

NH

ONH2

HN

O

OH

NH

O

NH2

HN

O

ONH2


Polymyxin B

Design Rational…


Polymyxin B

Design Rational…


Polymyxin B

Design Rational…

Analogs Design

Designed lipopeptide

NH2

HNO

NH

NH2

O

HN

ONH

OH2N

NHO

NH

O

HN

O NH

ONH2

HN

O

OH

NH

O

NH2

HN

O

HO


Polymyxin

NH2

HNO

NH

NH2

O

HN

ONH

OH2N

NHO

NH

O

SS

NH

ONH2

HN

O

OH

NH

O

NH2

HN

O

ONH2

Nº Peptide sequenceGram+ Gram-

S. aureus P. aeruginosa E. coli

pxb CH3-octanoyl-Dab-Thr-Dab-Dab-Dab-Phe-Leu-Dab-Dab-Thr >32 2 1

0 nonanoyl-Dab-Thr-Dab-Cys-Dab-Phe-Leu-Dab-Dab-Cys >32 8 4

1 nonanoyl-Arg-Thr-Dab-Cys-Dab-Phe-Leu-Arg-Dab-Cys 32 16 8

2 nonanoyl-Arg-Thr-Arg-Cys-Dab-Phe-Leu-Arg-Dab-Cys 8 8 4

3 nonanoyl-Dab-Thr-Dab-Cys-Dab-Phe-Met-Dab-Dab-Cys 32 16 4

34 nonanoyl-Arg-Thr-Dab-Cys-Dab-Trp-Leu-Arg-Dab-Cys 8 8 4

79 decanoyl-Arg-Thr-Dab-Cys-Dab-Phe-Leu-Arg-Dab-Cys 16 8 8

85 dodecanoyl-Arg-Thr-Dab-Cys-Dab-Phe-Leu-Arg-Dab-Cys 32 8 8

16 nonanoyl-Arg-Thr-Dab-Cys-Dab-Phe-Leu-Arg-Dab-Cys 16 8 4

93 decanoyl-Lys-Thr-Arg-Cys-Lys-Trp-Leu-Arg-Lys-Cys 16 >64 64

100 decanoyl-Arg-Thr-Arg-Cys-Dab-Trp-Nle-Arg-Dab-Cys 4 64 8

101 nonanoyl-Dab-Thr-Arg-Cys-Dab-Phe-Leu-Arg-Dab-Cys 8 16 2

103 ------------------------------------------------------------------ 4 2 1104 ------------------------------------------------------------------ 4 1 4105 ------------------------------------------------------------------ 4 1 2


Antibacterial Activity – in vitro (measured as Minimum Inhibitory Concentration, MIC, in μg/mL)

Up to 90 analogs synthesized and tested


Antibacterial Activity – in vitro (measured as Minimum Inhibitory concentration, MIC, in μg/mL)

Comparison with reference marketed products. Species Analog

103Analog

104Analog

105

PxBG(-)

control

VancomycinG(+) control

DaptomycinG(+) control

GRAM -Pseudomonas aeruginosa 2 1 1 2 - >32

Escherichia coli 1 4 4 1 - >32

GRAM+

Mycobacterium phlei 4 4 2 16-32 - >32

Staphylococcus aureus 4 4 4 >32 1 2

• Resistance panel: ▫ 40b (clinical isolate): resistance to IMP ▫ 38a (clinical isolate): resistance to Ceftazidime; Ciprofloxacin; Imipenem ; Piperacillin-tazobactam ▫ NMD, New Delhi metallo-beta-lactamase: Amoxicillin 256 mg/L; Amoxicillin clavulanate 32 mg/L ;

Piperacillin/tazobactam 256 mg/L; Cefoxitin 256 mg/L; Cefotaxime 256 mg/L; Ceftazidime 256 mg/L; Cefepime 256 mg/L; Imipenem 8 mg/L; Meropenem 16 mg/L; Doripenem 6 mg/L; Ertapenem 24 mg/L; Aztreonam 256 mg/L; Gentamicin 8 mg/L; Amikacin 32 mg/L, Tobramycin 8 mg/L; Ciprofloxacin 32 mg/L



In multi-drug resistant bacteria. Resistant and multi-resistant Gram negative bacteria Analog

103Analog

104Analog

105

P. aeruginosa40b Carbapenem-resistant strain 4 2 4

38a Highly-resistant strain 0,5 1 16

E. coli

MAC 21a Intermediate resistance to quinolones 0,5 1 2

VAL 10 Intermediate resistance to quinolones 0,5 0,5 1

VAL 5 Intermediate resistance to quinolones 0,5 0,5 1

NDM Highly-resistant strain 0,5 0,5 1

Moderate antibacterial activity in colistin-resistant Acinetobacter baumannii suggests an alternative mechanism of action



In Colistin resistant a. baumannii strains. Colistin-Resistant Acinetobacter baumannii Colistin Analog

103Analog

104Analog

105

A. Baumannii

ATCC-wt Low resistance strain 1 8 2 4

ATCC In vitro mutant of ATCC with resistance to colistin

256 64 16 32

77778 In vitro mutant of a clinical strain with resistance to colistin

256 256 128 128

Ab 10 Clinical isolate resistant to colistin 512 32 16 8

Ab 19 Clinical isolate resistant to colistin 512 32 16 16

Mechanism of action

• By Flow cytometry

Propidium iodide Bis-(1,3-Dibutylbarbituric Acid)Trimethine Oxonol DIBAC4(3)


E. coli

S. aureus

Flow-cytometric tests show a different behaviour of lipopeptides in front Gram+ and Gram- bacteria


Mechanism of action results

CONTROL, 120 min PEPT. 103, 120 min PXB, 120 min PEPT. 100, 120 min

In vivo Toxicity

• Acute toxicity test in mice for peptide 103

• LD50 = 283 mg/Kg

(subcutaneous) Polymyxin LD50 = 59,5 mg/Kg


Protocol OECD 425. 5 reversals in 6 consecutive animals. Animals (3) administered at 200 mg/kg survived after 14 days; Necropsy indicated no visual damage in vital organs. Mice (3) administered at 400 mg/Kg died


Current status of development

Design & Synthesis Of New Compounds

In vitro Efficacy

(MIC)

MoA SAR

In vitro TOX

IN VIVO EFFICACY

IN VIVO TOX. (LD50)

Design

Synthesis

In vitro Efficacy

MoA

NH2

HNO

NH

NH2

O

HN

ONH

OH2N

NHO

NH

O

SS

NH

ONH2

HN

O

OH

NH

O

NH2

HN

O

ONH2

IPR Protection

• 2 different patents • ES 200802626 Granted

• ES 2.374.779 A1 Filed - Priority March 2010

PCT/ES2011/070153 Published as WO2011110716 - ISR Positive

• Ownership

– University of Barcelona 100%



Weaknesses • No Oral Administration (not confirmed) • Early stage project. Few in vivo tests performed. • Need of partner to accelerate development

Threats • Efficacy in in vivo model. • PK/PD & ADME development. • Additional Toxicology development • Patent going to National phases in Sep. 2012

Pitfalls & Risks to be considered


Strengths • Good antibacterial activity front both pathogen and multi-drug

resistant bacteria • Better in vivo and in vitro tox. profile than PxB • MoA independent of membrane receptors • Easy Synthesis • Intl. patent file WO2011110716. Priority march 2010- Positive ISR. • Ownership 100% UB

Opportunities • Market Needs of new antibacterial compounds with wide spectrum /

low toxicity profile. • In vivo efficacy tests in 2012

Pitfalls & Risks to be considered


Market competitors PolyMedix, for S. aureus Several products in preclinical studies for indications such as tuberculosis, malaria, bacterial infections. PMX-30006 is a potent broad spectrum antibiotic, which is active against a number of bacterial strains. PMX-30006 is being developed for the treatment of bacterial infections. Cubist ; CB-182804, macrolactam PxB CB182804 is a lipopeptide which interacts with cell membranes and rapidly shuts down bacterial DNA, RNA and protein synthesis. CB182804 was under development as intravenous injection for the treatment of serious infections caused by multi-drug resistant (MDR) gram-negative bacteria. – Discontinued in phase I in 2009. Northern Antibiotics (complex synthesis: macrolactam) NAB 7061 is a polymyxin derivative that sensitizes target bacteria to other antibiotics such as rifampin and clarithromycin. NAB 7061 is being developed for the treatment of serious gram-negative hospital infections.


Partnering Opportunities

The project is available to licensing out through a collaboration and license agreement.

Contact details

Salvador Mena Project Manager

Tel: +34 93 403 97 95 [email protected]

mailto:[email protected]�

Acknowledgments

Funding has been provided by: ▫ Ministerio de Ciencia e Innovación (CTQ2008-06200) ▫ Generalitat de Catalunya (VALTEC 08-1-0016, ACC10) ▫ Fundació Bosch i Gimpera (UB)


THANK YOU FOR YOUR ATTENTION!


Product Name Companies 2010 2011

Zyvox Pfizer Inc 1,176.00 1,267.00

Cubicin Cubist Pharmaceuticals Inc, Novartis AG 624.90 664.00

Bactroban GlaxoSmithKline plc 194.90

Teflaro AstraZeneca Plc, Forest Laboratories Inc 2.70 56.00

Distaclor Eli Lilly & Co, Saudi Pharmaceutical Industries & Medical Appliances Corporation (SPIMACO) 53.40 54.00

Dificid Optimer Pharmaceuticals Inc 15.00


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Summary: -Proof of concept in vitro for EC, PA and SA -Activity in resistant and MDR bacteria

-In vivo toxicity (acute, sc) better than PxB

-Optimized chemical synthesis and scale up (i. e, octreotride, lanreotide) -Next step: proof of concept in vivo (efficacy)


Optimized chemical synthesis and scale up (i. e, octreotide, lanreotide)

NH2

HNO

NH

NH2

O

HN

ONH

OH2N

NHO

NH

O

SS

NH

ONH2

HN

O

OH

NH

O

NH2

HN

O

ONH2

HN O

NHO

HN

O

NHO

NH

O

SS

NH

O

H2N

ONH

OH

HN

NH2

OH2N

Lanreotide:H-D-2-Nal-Cys-Tyr-D-Trp-Lys-Val-Cys-Thr-NH2

Lanreotide is a synthetic analog of somatostatin with properties similar to octreotide

Health & Medicine

ANTIBACTERIAL POLYMYXIN B ANALOGS