WO2012055851A1 - Chondrones utilisés en thérapie des cartilages - Google Patents

Chondrones utilisés en thérapie des cartilages Download PDF

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Publication number
WO2012055851A1
WO2012055851A1 PCT/EP2011/068618 EP2011068618W WO2012055851A1 WO 2012055851 A1 WO2012055851 A1 WO 2012055851A1 EP 2011068618 W EP2011068618 W EP 2011068618W WO 2012055851 A1 WO2012055851 A1 WO 2012055851A1
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WO
WIPO (PCT)
Prior art keywords
chondrons
cartilage
use according
chondrocytes
osteoarthritis
Prior art date
Application number
PCT/EP2011/068618
Other languages
German (de)
English (en)
Inventor
Bernd Rolauffs
Wilhelm Aicher
Original Assignee
Eberhard-Karls-Universitaet Tuebingen Universitaetsklinikum
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Application filed by Eberhard-Karls-Universitaet Tuebingen Universitaetsklinikum filed Critical Eberhard-Karls-Universitaet Tuebingen Universitaetsklinikum
Publication of WO2012055851A1 publication Critical patent/WO2012055851A1/fr

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
    • A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
    • A61L27/38—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells
    • A61L27/3804—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells characterised by specific cells or progenitors thereof, e.g. fibroblasts, connective tissue cells, kidney cells
    • A61L27/3817—Cartilage-forming cells, e.g. pre-chondrocytes

Definitions

  • the present invention relates to the use of endogenous structures for cartilage, bone, meniscus and disc therapy and a method for obtaining these endogenous structures.
  • Cartilage damage such as diseases or degenerations can have many causes. Common to all damage, however, is the loss and alteration of the matrix located between and above the chondrocytes. As a result, the cartilaginous surface initially becomes rough, then cartilage substance is lost, it develops an arthrosis, which in turn limits the function of the affected joint.
  • Chondrosis is generally referred to as the degenerative changes of the cartilage.
  • intervertebral discs there is a narrowing of the intervertebral space between one or more vertebral segments compared to the remaining vertebral bodies, whereby the height of the intervertebral spaces decreases.
  • the cause may be either congenital or acquired changes, the latter being degenerative, inflammatory, or operative.
  • osteochondrosis if the underlying bone involved, it is called osteochondrosis. Changes in the cartilaginous and osseous tissues in the large and small joints of the musculoskeletal system lead to osteoarthritis, which can also be caused by congenital or acquired changes.
  • the cause may be degenerative, inflammatory, post-traumatic factors and diseases of the central nervous system in question.
  • osteochondritis dissecans The resulting in an arthritis / osteoarthritis from the cartilaginous cartilage pieces, or the resulting state is referred to as osteochondritis dissecans, which also occurs in the context of an accident or today medically identifiable causes and also causes the development of osteoarthritis / osteoarthritis.
  • the ACT also leads to the formation of only a hyaline-like cartilage, the quality of which neither clinically nor histologically corresponds to the hyaline cartilage. This is reflected in a reduced biomechanical quality of the hyaline-like (fibrous) cartilage, whereby further degeneration and thus also the development of osteoarthritis can only be deferred, but can not be reversed. It is therefore an object of the present invention to provide a comparison with the prior art improved therapy option with which even larger defects can be treated.
  • this object is achieved by the use of chondrons for the treatment of cartilage, bone, meniscus or disc damage or diseases and the resulting osteoarthritis / osteoarthritis.
  • chondrons can be isolated with the body's own or allogenic and xenogeneic chondrons.
  • chondrons provides a means by which on the one hand larger cartilage defects can be treated and on the other hand the long-term survival of the chondrocytes contained in the chondrons is improved.
  • chondron is understood as meaning the unit comprising chondrocyte and the surrounding pericellular environment, i. the pericellular glycocalyx and the pericellular capsule, understood. Chondrocytes and their pericellular environment form a structural and functional unit.
  • chondrocytes are isolated, expanded in the laboratory, applied to a carrier material and implanted in the cartilage defect.
  • the isolation of the chondrocytes destroys the pericellular matrix, and thus the structure of the chondron.
  • the isolated chondrocytes produce insufficient amounts of glycosaminogly- and collagen type II, whereby the biomechanical properties of this cartilage formed from the ACT are inferior to those of the original cartilaginous tissue.
  • the present inventors have recognized that the isolation of physiologically complete chondrons from the cartilage tissue and their use to form cartilage replacement tissue offers an improved approach to the therapy of cartilage damage compared to the method known in the art.
  • chondrons for the treatment of cartilage damage or diseases of the large joints such. the knee, hip, jump, shoulder or elbow joint or the spine, in particular for the treatment of disc damage, are used.
  • cartilage damage or diseases of joints in these areas include, for example, osteochondritis dissecans, osteoarthritis, arthritis and osteoarthritis, ie inflammatory and degenerative joint diseases.
  • chondrons With the use of chondrons according to the invention, such diseases, ie cartilage degeneration and the resulting osteoarthritis / arthrosis can be treated in a targeted manner, as well as a progression of osteoarthritis can be prevented or delayed.
  • Chondrons as explained above, form a functional unit of chondrocytes with their immediate surrounding pericellular matrix, and can be obtained therefrom mechanically and enzymatically from cartilaginous tissue. Both metabolically and biomechanically, chondrons are of great functional importance because their structure plays a major role in biomechanical, biophysical and biochemical interactions between chondrocyte and ECM.
  • Chondron pellets form significantly more glycosaminogly- kane and collagen type I as chondrocyte pellets, which is why they are particularly suitable for the claimed use in comparison to the chondrocytes hitherto used isolated in the ACT.
  • the inventors have in fact recognized that due to the preserved, original pericellular matrix of the chondrons, the formation of a hyaline-like matrix in cartilage damage can be promoted much more strongly than a pericellular matrix, which must be newly synthesized by expanded chondrocytes.
  • chondrons in the diseases / injuries listed offers the advantage that they provide as an important aggrecan reservoir the essential markomolecule for the hydrodynamic cartilage function and for cartilage proteoglycans.
  • the chondrons also provide mechanical protection for chondrocytes from biomechanical overload.
  • chondrons are non-immunogenic, and therefore much more compatible in their use than these.
  • the use of the invention thus offers the advantage that not only young patients or those with isolated, smaller cartilage defects can be treated, but now older patients and / or patients with major cartilage damage, currently still by an implantation of artificial joints or only be treated by conservative measures such as painkillers etc.
  • the chondrons are used for the treatment of osteoarthritis / osteoarthitis or osteochondrosis dissecans.
  • osteoarthritis or arthrosis is understood to mean a joint disease with destruction of the cartilage surfaces.
  • Ostochondrosis dissecans is understood to mean the detachment of a cartilaginous piece (flake) by, for example, an accident or also by other unexplained causes. which can lead to the development of arthritis / osteoarthritis.
  • These diseases occur especially in the knee joint, in the upper ankle joint, in the hip joint, in the shoulder joint and in the elbow joint, not only in humans, but also in animals, such as in dogs, (racing) horses, cats, cows or pigs.
  • chondrons which were obtained from autologous, so the body's own, cartilage material.
  • This embodiment has the advantage that it can be used on the body's own and thus patient's own material, which possibly minimizes unwanted reactions of the body to the chondrons.
  • chondrons obtained from heterologous, allogenic or xenogenic cartilage material are used.
  • allogeneic material is meant cartilage material from a donor of the same species as the patient to be treated, and "xenogeneic” material from cartilage material from a donor of a species other than the patient into whom the chondrons are to be reincorporated.
  • This embodiment has the advantage that it is also possible to treat patients in whom cartilage material which is insufficiently good or in quantity can be provided for the isolation of chondrons.
  • the chondrons can be used after their provision either directly into the area to be treated with cartilage damage or cultivated in vitro, which can be used after cultivation, the chondrons with or without a corresponding carrier material in the joint.
  • the present invention also relates to the use of chondrons derived from cartilage material for the in vitro production of cartilage. replacement tissue. These tissues can then be implanted in the affected area of the patient from whom the original cartilage material has been removed.
  • the chondrons are obtained as follows: a) in vitro isolation of chondrons from a biological cartilage tissue sample of a patient by means of comminution and / or enzymatic digestion, in particular Dispase II and collagenase P, wherein the Digestion is preferably carried out by incubation with the enzymes for a period of 5 hours and preferably at 37 ° C with stirring, followed by sieving, preferably with a 100 ⁇ m sieve, to obtain chondrons; b) in vitro expansion of the chondrons by culturing the chondrons obtained in step a) in monolayer or three-dimensional culture in standard medium;
  • Step b) can be carried out, for example, the following standard media: F12 + GlutaMAX, Gibco 21885 (250 ml), DMEM + GlutaMAX, Gibco 31765 (250 ml), FBS Superior, Biochrom S0615 (50 ml), PenStrep, Gibco 15140 (10 ml), partricin (fungicides), biochrome A2812 (5 ml), L-ascorbic acid phosphate magnesium salt, Sigma Aldrich A8960 (0.5 ml).
  • standard media F12 + GlutaMAX, Gibco 21885 (250 ml), DMEM + GlutaMAX, Gibco 31765 (250 ml), FBS Superior, Biochrom S0615 (50 ml), PenStrep, Gibco 15140 (10 ml), partricin (fungicides), biochrome A2812 (5 ml), L-ascorbic acid phosphate magnesium salt, Sigma Aldrich A8960 (0.5 ml).
  • cartilage replacement tissue is obtained after adaptation under GMP conditions.
  • the thus isolated chondrons can then be used directly and / with carrier material for the treatment and thus for the generation of cartilage replacement tissue in vivo, or for the in vitro production of cartilage replacement tissue.
  • a "biological sample” is understood to mean any material already taken from a patient's body, preferably tissue, more preferably cartilage tissue, the viable cells, in particular chondrons, chondrocytes or their precursors, cell aggregates, and / or other cell-free material contains.
  • the biological sample may be an endogenous sample, ie a sample of the patient to be treated with the chondrons obtained therefrom, or an allogeneic / xenogeneic sample.
  • the patient is a mammal, preferably a human.
  • a mammal preferably a human.
  • analogous applications in other mammals such as horses, camels, dogs, cats, etc., are also being considered since there is an increasing interest in effective treatment of osteoarthritis / osteochondrosis dissecans / cartilage damage consists of these animals.
  • the chondrons thus obtained for the treatment of cartilage, meniscal, bone or disc damage, - diseases, or -degenerationen, or for the treatment of osteoarthritis / osteochondrosis dissecans, in particular a human, used become.
  • chondrons used according to the present invention may be further administered together with pharmaceutically acceptable carriers and / or excipients, and optionally therapeutically active substances.
  • pharmaceutically acceptable excipients or excipients as used herein means any substance / composition to be administered to a patient in pharmacy which does not or not adversely affect cells, and / or which can support or facilitate the pharmacological use of the chondrons
  • therapeutically active substance is further understood any substance used for the purpose of treating or ameliorating a clinical picture of a patient. It is understood that the above-mentioned features and those yet to be explained not only in the combination specified, but also in other combinations or alone, without departing from the scope of the present invention.
  • Fig. 1 (A) is a schematic representation of the chondron organization and the surrounding collagen fibers with chondrocyte (C), pericellular glycocalyx (PG), pericellular capsule (PC), territorial matrix (M) and interteritorial (IM) matrix ; and (B) a three-dimensional representation of a solitary chondroma; Reference: C. Anthony Poole; 1997;
  • FIG. 2 Fluorescence microscopy image with collagen type VI staining after isolation from human cartilage tissue A) from chondrocytes and B) from chondrons; in the present case two chondrons: B) on the left a chondron, which contains a cell; B) on the right a chondron, which contains two cells (own data);
  • Fig. 3 fluorescence microscopy images with A) a collagen type II stain, B) a collagen type VI stain, C) an aggrecan molecule stain;
  • Fig. 4 A) fluorescence microscopy images of a chondroma consisting of several cells, B) transmitted light microscopy image of the same chondron, C) and D) collagen type II fibers thereof Chondrons as a link to the extracellular cartilage matrix (own data);
  • OA chondrons shows a diagram of the gene expression profiles of chondrons from osteoarthritic joints (OA chondrons) and in comparison of chondrocytes from the same arthritic joints (OA chondrocytes) and chondrocytes from a joint of a young patient, who has an autologous chondrocyte transplantation (ACT) of the above shown chondrocytes (ACT chondrocytes); and
  • Fig. 6 is a diagram for the relative cell number of chondrons
  • Chrondocytes after a certain culture period Chrondocytes after a certain culture period.
  • a chondrocyte in its pericellular environment, the chondron.
  • the chondrocyte and its pericellular environment form a functional unit and have a multifaceted importance for the function of the hyaline cartilage.
  • a chondron consists of a single or multiple chondrocytes (C) enclosed by the pericellular glycocalyx (PG) and a fibrillar pericellular capsule (PC).
  • the capsule defines the pericellular matrix around the chondrocyte.
  • OA osteoarthritic
  • the fluorescence microscopy images in Figure 2 show that living chondrons could be recovered from human tissue with their intact structure; Furthermore, the fluorescence microscopy images in Fig. 3 show that the obtained chondrons produce the fractions, collagen type II, type VI and aggrecan, which are fundamental for cartilage function, in quantities which can be detected by conventional techniques.
  • Fig. 4 shows that chondrons can be obtained with their fiber network. This may be. relevant for improved / superior function of the chondrone.
  • the isolation of the fiber network and the use of chondrons with and without this fiber network is included in the invention.
  • chondrons are superior to chondrocytes.
  • gene expression of chondrons even from osteoarthrotic diseased joints, is shown to be superior to the chondrocytes of the same joint as well as a young and relatively healthy subject to ACT.
  • chondrons show a significantly faster proliferation rate than chondrocytes, so that in a laboratory phase of a given length More living chondrons are available for transplantation than chondrocytes.
  • the introduction of chondrons into the human or animal joint as a therapy has not yet been performed, but can be based on the data presented herein with the previously clinically proven, approved and used in clinical routine daily methods.
  • the isolated chondrons can be introduced instead of the usually used chondrocytes in a carrier, which is sewn or glued or inserted after punching a cartilage defect in this defect.
  • z. B. also be used a gel which is injected into the defect. All methods approved for ACT are also eligible.

Abstract

L'invention concerne l'utilisation de chondrones pour traiter des altérations du cartilage, de l'os, du ménisque ou des disques intervertébraux, en particulier l'ostéoarthrite, l'arthrose, l'ostéochondrite disséquante, en particulier chez l'homme et le cheval.
PCT/EP2011/068618 2010-10-27 2011-10-25 Chondrones utilisés en thérapie des cartilages WO2012055851A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
DE102010050460.2 2010-10-27
DE201010050460 DE102010050460A1 (de) 2010-10-27 2010-10-27 Chondrone zur Knorpeltherapie

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WO2012055851A1 true WO2012055851A1 (fr) 2012-05-03

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Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
DE102009018640A1 (de) * 2009-04-17 2010-10-21 Tetec Tissue Engineering Technologies Ag Implantat und therapeutische Zusammensetzung zur Behandlung von Schäden und/oder Erkrankungen im Bereich des menschlichen und/oder tierischen Stütz- und Bewegungsapparates

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
CHRISTOPHER M. LARSON: "Retention of the native chondrocyte pericelular matrix results in significantly improved matrix production", MATRIX BIOLOGY, vol. 21, 2002, pages 349 - 359, XP002668069 *
K.H. TER STEGE: "Artificial chondrons in cartilage tissue engineering", 16 February 2007, AO RESEARCH INSTITUTE, Davos-Switzerland, pages: 1 - 37, XP002668067 *
LUCIENEN A. VONK: "Preservation of the Chondrocyte's Pericelular Matrix Improves Cell-Induced cartilage Formation", JOURNAL OF CELLULAR BIOCHEMISTRY, vol. 110, 8 March 2010 (2010-03-08), pages 260 - 271, XP002668068 *
Z.ZHANG ET AL.: "Comparison of gene expresion profile between human chondrons and chondrocytes: a cDNA microarray study", OSTEOARTHRITIS AND CARTILAGE, vol. 14, 2006, pages 449 - 459, XP002668079 *

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