Screening for natural products from marine microorganisms from research to commercialisation

Transcription

1 Screening for natural products from marine microorganisms from research to commercialisation Ulrike Lindequist Institut für Pharmazie und Institut für Marine Biotechnologie e.v. Rendez Vous de Concarneau, ctober 2013

2 Number of marine natural compounds by groups of organisms (Blunt et al., Nat. Prod. Rep., 2009, 26, ) Presently 16-18% of all published novel marine compounds are of microbiological origin (Blunt et al., Nat. Prod. Rep. 2010, 27, )

3 Biologically guided Screening strategies Search for selected biological activities (antibacterial, cytostatic, enzyme inhibiting etc.) of an extract mostly by in vitro methods Fractionation of extracts according to the observed activities Structure elucidation of the isolated bioactive compounds Risk: compounds are already known Chemically guided Search for unknown chemical structures in an extract by NMR, MS etc. Isolation and structure elucidation of the novel compounds Tests for biological activities Risk: compounds without interesting biological activities Genetically guided ( Genome mining ) Search for genes and gene clusters coding for interesting biosynthetic proteins, e.g. polyketide synthases Risk: long way, high risk

4 Biological Screening Antibacterial/antifungal Antiviral Antitumor Bone cells (steoporosis) Keratinocytes (Wound healing, UV protection, cosmetics)

5 Groups of marine microorganisms Marine bacteria, mainly actinomycetes Marine fungi, mainly ascomycetes Marine cyanobacteria H N ClH 2 C H H Salinosporamid A

6 Cyanobacteria Gram-negative photoautotrophic prokaryotes Free-living or living in invertebrates Cause of toxic water blooms Important producents: Lyngbya sp. Focus on cytostatic, antimicrobial and enzyme-inhibiting activities Many cyclic or linear peptides often containing lipophilic side chains or macrolides Mixed biosynthetic pathways (polyketide synthase/nonribosomal peptide synthetase) Till now of some importance for food/feed industry, cosmetic and technical purposes

7 Cyanobacteria: examples Cl CH 3 NH 2 Cl H H H H Cl H 2 N Cl CH 3 Carbamidocyclophanes from Nostoc sp. CAVN 10 Carbamidocyclophan A: MIC St. aureus: IC 50 MCF 7 cells : IC 50 FL cells: 0.1 mmol 0.8 µmol 3.1 µmol Ha T. N. Bui et al.: J Nat Prod 70, 499 (2007), Dissertation Michael Preisitsch

8 Cyanobacteria: examples H PAA H Tyr H NH N H 2 Ichthyopeptin A NH H 3 C Thr Gln NH N H Ahp CH 3 N H 3 C NH CH 3 Ile H N H IC 50 Influenza Virus A: 12.5 µg/ml CH 3 CH 3 Val N-Me-Phe PAA Ichthyopeptin B H Microcystis ichthyoblabe Ahp H Ile Leu CH 3 CH 3 N H C N CH 3 3 H NH H 3 C N NH NH H 3 C NH N-Me-Phe Thr Val NH Asn 2 H 3 C CH 3 Zainuddin E et al.: J. Nat. Prod. 70, 1084 (2007)

9 Problem: Multiresistant bacteria (MRSA, VRE, ESBL etc.) *European Antimicrobial Resistance Surveillance Network (EARS-Net), 2010

10 What can we do? Better hygiene New antibiotics Effective antiseptics Prevention of colonisation: an example for commercialisation

11 Prevention of colonisation - Screening of about 120 different marine microbial strains (biological screening) - Selection of microalgal strain Bio 33, isolated from the Baltic Sea - Microencapsulation of the cultivated biomass into Maresome (patent protected) - Preparation of Maresome ointments - In vitro assays - In vivo assays Bio 33

12 Bio33-Maresome * Microparticles consisting of the whole biomass of cultivated microalgae strain No additives Prepared by microencapsulation using the lipids of the microalgae Multifactorial mode of action: Stabile system in aqueous dispersion with negative zeta potential repulsion of bacteria Increased surface: better release kinetics and bioavailability of bioactive compounds (antibacterial fatty acids etc.) Multiresistant strains more sensitive because of higher extracellular lipase activity of MRSA and possibly other factors Positive effects on skin Prevention of contamination of the skin by multiresistant pathogenic microorganisms (MRSA etc.) *in the following declared as Maresome Trade name Maresome refers to principle of preparation. Lukowski et al.: PCT-03/00747

13 Prevention of transfer of MRSA from a donator to an acceptor Number of colonies Control Donator Acceptor Influence of Maresome on the colonisation of MRSA North German Epidemic Strain in the model Mouse ear Lukowski et al.: Skin pharmacology and physiology 21, , 2008

14 Comparison to other microalgae strains number of colonies control Chlorella Spirulina Bio 33 Influence of Maresome on the colonisation of MRSA North German Epidemic Strain in the model Cow udder teat Lukowski et al.: Skin pharmacology and physiology 21, , 2008

15 Influence on the colonization of several MRSA strains 1500 number of colonies Control Col Mu 50 N315 Nord. Epe. Strain Influence of Maresome on the colonisation of several MRSA strains in the model Cow udder teat Lukowski et al.: Skin pharmacology and physiology 21, , 2008

16 Influence on the colonization of Pseudomonas aeruginosa 1200 colony forming units Control Algae ointment Influence of Maresome on the colonisation of P. aeruginosa in the model Cow udder teat

17 Influence on the transfer of Candida maltosa colony forming units Donor Acceptor Control Influence of Maresome on the transfer of C. albicans in the model Cow udder teat

18 Clinical observation Patient with phlebolipolymph edema and erysipel Before treatment Up to 3 days after treatment Pathogenic skin flora (Cfu/cm 2 ) S. aureus P. aeruginosa 60 0 S. agalactiae 50 0 Physiologic skin flora (Cfu/cm 2 ) Coagulase neg. Staphylococci 3 x G. Lukowski, Lindequist U, Mundt Jülich WD, Jünger M, Daeschlein,G, CSMDERM XVI ESCAD, 2010

19 Clinical observation Patient: 82y, ulcus cruris venosum left lower limb d1 Control (cfu/ plate) d2 with ointment (cfu/ plate) d3, d4, d5 with ointment (cfu/ plate) S. aureus Pseudomonas aeruginosa Klebsiella oxytoca Application 1 times per day

20 Comparison with positive and negative controls Effects of a 30min and 120min treatment on MSSA on healthy skin 120,0 100,0 % 80,0 60,0 40,0 20,0 0,0 51,3 3,1 4,8 1,2 0,2 30min 31,8 120min Gentamicin Maresome Base Control

21 Effect on dermal barrier function TEWL Median ,2 14,55 12, ,5 11,1 9,7 11,2 Algae-intment Betamethaso... untreated Day 1 Day 3 Day 8 TEWL = transepidermal water loss (g/m 2 *h)

22 Cosmetic trial with atopic patients skin care improvement % of all responds strongly improved improved not improved after 4 Weeks after 8 Weeks Interview with 104 patients, ambulant, 2011

23 Summary Bio33-Maresome Protection against bacterial (re)infections Prevention of (re)colonization with pathogens (MRSA etc.) Preservation of physiological skin flora Prevention of nosocomial infections and in cosmetic procedures Ideal skin care for patients with atopic dermatitis, eczema, superficial folliculitis, diabetic and phlebologic problems

24 Maresome : from research to commercialisation: Research Successful screening interesting biological effect Meeting high medicinal need First clinical results Patent protected formulation Protected brand name (Maresome ) Search for possible industrial collaborators

25 Maresome : from research to commercialisation: Commercialisation: Present situation Licensation for use as cosmetics to a company In the company: Scale up of production Formulation Packing Commercialisation Sale as cosmetic product in Germany and and Mexico

26 Maresome : from research to commercialisation Can we be satisfied with the present situation? No! This product deserves application in hospitals, nursing homes etc. Necessary are: Licensation of the patent for use in hospital hygiene Broader clinical trials Licensation as medical product

27 Challenges on the way from research to commercialisation Sustainable production + Scale up + Patent protection + Money - Clinical trials - Regulatory aspects, licensation -

28 What is necessary in general? Expanding the knowledge on marine life and ecological relationships Sustainable production in accordance with ecology: biotechnology, gentechnology, synthesis Improvement of teaching and education Close interactions between basic and applied research, and between research and industry Development of intelligent management and application strategies Finances

29 Thanks to all colleagues, especially PD Dr. Sabine Mundt and her group, students and collaborators Thank you for your attention! Contact: Prof. Dr. Ulrike Lindequist