Conception et synthèse d'iminoglycolipides comme inhibiteurs d'enzymes lysosomales à effet chaperon pharmacologique, Conception and synthesis of iminoglycolipids as inhibitors of lysosomal enzymes acting as pharmacological chaperones

Farah Oulaïdi

Description

Sous la direction de Olivier Martin, Philippe Compain
Thèse soutenue le 28 janvier 2011: Orléans
La thérapie chaperon représente une approche thérapeutique stratégique et innovante, en particulier dans le traitement des maladies lysosomales. Ces maladies génétiques rares ont une gravité variable, qui peut aller de la létalité avant la naissance jusqu’à la nécessité d‟une prise en charge permanente ; elles apparaissent à tous les stades de la vie. Des mimes du substrat appelé iminosucres, vont agir en allant au coeur du site actif de l’enzyme, stabiliser l’enzyme mutée qui est instable mais non inactive. Paradoxalement, la plupart des chaperons pharmacologiques sont des inhibiteurs de l’enzyme visée mais leur administration à faible concentration leur permet de réaliser leur mission de sauvetage de l’enzyme mutée. Dans cette optique, des recherches effectuées au sein de notre laboratoire ont fait état de la synthèse d’iminosucres, tels que les α-1-C-alkyl iminoxylitols qui sont de très bons inhibiteurs de la β-glucocérébrosidase, l’enzyme défaillante dans la maladie de Gaucher, mais aussi qui doublent l’activité enzymatique résiduelle. Une nouvelle voie de synthèse plus efficace a été réalisée afin d’obtenir plus efficacement ce type d’iminosucres et d’autres dérivés. Ces travaux ont également été l’occasion de développer des iminoxylitols structurellement simplifiés qui agissent comme chaperons pharmacologiques toujours pour le traitement de la maladie de Gaucher. Une partie de ces travaux a aussi été consacrée à la recherche d‟inhibiteurs de la β-galactocérébrosidase, l’enzyme impliquée dans la maladie de Krabbé, et qui pourront agir comme chaperons pharmacologiques. Différentes évaluations pharmacologiques ont été réalisées, notamment des tests d’inhibition et la détermination des effets chaperons.
-Maladie lysosomale
-Chaperon pharmacologique
-Β-galactocérébrosidase
Chaperone Mediated Therapy represents an innovative and strategic approach to treat lysosomal storage disorders which a class of rare genetic diseases. Competitive inhibitors for some of these lysosomal enzymes can, at sub inhibitory concentrations, act as chaperones and rescue the mutant proteins. In fact, enzymes carrying some mutations are still catalytically active. α-1-C-alkyl iminoxylitols represent a class of iminosugars which mimic the “gluco” configuration of the substrate and give powerful inhibitors of β-glucocerebrosidase, the enzyme involved in Gaucher disease. Moreover, this class of iminosugars, synthesized by our group, act as pharmacological chaperones and are able to double the residual activity of the N370S mutant. In order to synthesize more efficiently these iminosugars, the synthetic strategy was improved and optimized. Moreover, we focused our investigations on structural variations on our lead compound (α-1-C9 iminoxylitol) and draw important conclusions on structure-activity relationship. Then, we extended our expertise on iminosugars as pharmacological chaperones to another lysosomal glycosidase. In paricular, we targeted β-galactocerebrosidase, the enzyme responsible for Krabbe disease, and synthesized a series of iminosugars which mimic the “galacto” configuration. Biological assays were performed on our compounds to determine their activity as inhibitors and for some of them, their chaperone effects.
-Lysosomal storage disorder
-Pharmacological chaperone
-Β-galactocerebrosidase
Source: http://www.theses.fr/2011ORLE2001/document

Sujets

• Maladie lysosomale

Informations

• Publié par: Thesee
• Nombre de lectures: 42
• Langue: English
• Poids de l'ouvrage: 1 Mo

Extrait

THÈSE PRESENTÉE A L’UNIVERSITÉ D’ORLÉANS
POUR OBTENIR LE GRADE DE

DOCTEUR DE L’UNIVERSITÉ D’ORLÉANS

par
Farah OULAÏDI

ÉCOLE DOCTORALE SCIENCES ET TECHNOLOGIES
Discipline: Chimie Organique




CONCEPTION ET SYNTHÈSE D’IMINOGLYCOLIPIDES
COMME INHIBITEURS D’ENZYMES LYSOSOMALES
À EFFET CHAPERON PHARMACOLOGIQUE

Soutenue le 28 janvier 2011


Thèse dirigée par :
M. Olivier R. MARTIN Professeur, Université d‟Orléans - Directeur de thèse
M. Philippe COMPAIN , ECPM, Strasbourg 1 - Co-Directeur de thèse
Rapporteurs :
M. Mikael BOLS Professeur, Université de Copenhague, Danemark
Mme Marielle LEMAIRE Professeur, UBP, Clermont-Ferrand
Jury :
M. Olivier R. MARTIN Professeur, Université d‟Orléans
M. Philippe COMPAIN , ECPM , Strasbourg 1
M. Mikael BOLS Professeur, Université de Copenhague, Danemark
Mme Marielle LEMAIRE Professeur, UBP, Clermont-Ferrand
M. Yves LE MERRER Professeur, Université Paris V
M. Jean-Claude JACQUINET Directeur de recherche, INSERM, Université d‟Orléans

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tel-00623109, version 1 - 13 Sep 2011
A THESIS SUBMITTED FOR THE DEGREE OF
DOCTOR OF ORGANIC CHEMISTRY

UNIVERSITY OF ORLÉANS, FRANCE

BY
Farah OULAÏDI

DOCTORAL SCHOOL OF SCIENCES AND TECHNICS






CONCEPTION AND SYNTHESIS OF IMINOGLYCOLIPIDS
AS INHIBITORS OF LYSOSOMAL ENZYMES
ACTING AS PHARMACOLOGICAL CHAPERONES

thDefended on Friday, January 28 2011


Directed by :
Mr Olivier R. MARTIN Professor, University of Orléans
Mr Philippe COMPAIN , ECPM, Strasbourg 1
Referees :
Mr Mikael BOLS Professor, University of Copenhagen, Denmark
Mrs Marielle LEMAIRE Professor, UBP, Clermont-Ferrand
Thesis comitee :
Mr Olivier R. MARTIN Professor, University of Orléans
Mr Philippe COMPAIN , ECPM, Strasbourg 1
Mr Mikael BOLS Professor, University of Copenhagen, Denmark
Mrs Marielle LEMAIRE Professor, UBP, Clermont-Ferrand
Mr Yves LE MERRER , University of Paris V
Mr Jean-Claude JACQUINET Research director, INSERM, University of Orléans

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tel-00623109, version 1 - 13 Sep 2011
Acknowledgements

I would like to thank my supervisors, Prof. Olivier R. Martin and Prof. Philippe Compain, for
their advice and guidance during the course of this work. Both supervisors took a personal
involvement in the outcome of this thesis and their efforts have been greatly appreciated. I am
particularly thankful to Prof. Olivier R. Martin for allowing me some extra time to finish my
experimental work and for his cheerful enthusiasm and ever-friendly nature that I was able to
finish my research work in excellent conditions.
I would like to express gratitude to CNRS and Region Centre for a fellowship.
Besides my supervisors, I would like to thank the members of my thesis committee: Prof.
Marielle Lemaire, Prof. Mikael Bols, Prof. Yves Le Merrer, and Dr Jean-Claude Jacquinet for
the precious time they spent reading the manuscript and for providing me their valuable
suggestions.
My sincere thanks also go to Prof. Aldo Orlacchio (University of Perugia, Italy) for offering
me an internship opportunity in his group and leading me to work on biological
experimentation.
I thank my fellow labmates: Gary, Ana, Cyril, and especially Estelle and Sophie for the
stimulating discussions and for all the fun we have had in the last three years. Also, I thank
my friends: Serour, Sheima, Aurelien, Karim, Samir, Henriette and Saida. I would also thank
my love Olivier who always managed to understand how things were going and find the time
to help me along.
My deepest gratitude goes to my family, for their unflagging love and support throughout my
life; this dissertation was simply impossible without them. I am indebted to my father, Ben
Achir Oulaidi, for his care and love. Although he is no longer with us, he is forever
remembered. I am sure he shares our joy and happiness in the heaven.

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tel-00623109, version 1 - 13 Sep 2011Contents
Acknowledgement
Contents……………………………………………………………………………………………………….…..1
Biological and medicinal glossary……………………………………………………………………………..5
Abbreviations…………………………………………………………………………………………………....8
Avant propos……………………………………………………………………………………………………...9
Chapter 1 ........................................................................................................................................... 14
From health to disease: the vital role of Glycosphingolipids ............................................................ 14
Introduction ....... 14
I. Lysosomal storage disorders .......................................................................................................... 15
I.1. Lysosome ................................ 15
I.1.a. Synthesis and trafficking of lysosomal enzymes .. 15
I.1.b. Structure and function .......................................................................................................... 17
I.2. Lysosomal storage disorders ... 17
II. Glycosphingolipids ....................................................................................................................... 21
II.1. Structure and function ............ 21
II.2. Sphingolipid Biosynthesis ..................................................................................................... 22
II.3. Glycosphinglipid Biosynthesis .............................. 25
II.4. Glycosphingolipid catabolism ............................... 27
III. Therapies for GSLs Storage ........................................................................................................ 32
III.1. Therapy by substitution ........ 32
III.1.a. Enzyme Replacement Therapy (ERT) ............... 32
III.1.b. Haematopoietic Cell Transplantation (HCT) .................................................................... 34
III.1.c. ERT in combination with HCT .......................................................... 35
III.2. Substrate Reduction Therapy (SRT) .................................................... 35
III.3. Chaperone Mediated Therapy (CMT or ASSC) ................................................................... 37
III.4. Conclusion ............................................................................................ 44
IV. Gaucher disease ........................... 46
IV.1. Causes of the disease ............................................................................................................ 46
IV.2. Clinical aspect ...................... 48
IV.3. Physiopathology ................... 49
IV.4. Related allelic disorder: Parkinson disease .......................................................................... 50
IV.5. Therapy for Gaucher disease ................................................................ 51
IV.6. Conclusion............................................................................................ 52
V. Krabbe disease (Globoid cell leukodystrophy, GLD) .. 53
V.1. Causes of the disease ............................................................................................................. 53
V.1.a. pathogenic mechanism ........ 53
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tel-00623109, version 1 - 13 Sep 2011V.1.b. GALC mutations ................................................................................................................ 54
V.2. Clinical aspect ....................... 56
V.3. Therapy for Krabbe disease ................................................................................................... 57
Objectives .......................................... 58
Chapter 2 ........... 60
Improvement of the synthesis of α-1-C-alkyl-iminoxylitols as inhibitors and pharmacological
chaperones of GCase ......................................................................................................................... 60
I. Synthetic target .............................. 60
II. Previous synthetic pathway .......................................................................................................... 61
III. Addition to imines: improved strategies ...................... 63
III.1. Protection of the anomeric center ......................................................................................... 64
III.1.a. Direct tritylation of the primary alcohol function .............................. 64
III.1.b. Benzyl glycoside ............................................................................................................... 65
III.1.c. Methyl glycoside 66
III.2. Preparation of partially protected derivative 16 ................................................................... 67
III.3. Pn of the imine ....................................... 67
III.3.a. N-Benzylimine ................................................................................... 67
III.3.b. N-Alkylsulfinylimine ......... 68
III.4. Addition of Grignard reagents .............................................................................................. 69
III.4.a. Addition to the N-Benzylimine .......................... 69
III.4.b. Addit