WO2017046267A1 - Principes actifs pharmaceutiques à utiliser pour le traitement de sclérose latérale amyotrophique (sla) et procédé de diagnostic associé - Google Patents

Principes actifs pharmaceutiques à utiliser pour le traitement de sclérose latérale amyotrophique (sla) et procédé de diagnostic associé Download PDF

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WO2017046267A1
WO2017046267A1 PCT/EP2016/071867 EP2016071867W WO2017046267A1 WO 2017046267 A1 WO2017046267 A1 WO 2017046267A1 EP 2016071867 W EP2016071867 W EP 2016071867W WO 2017046267 A1 WO2017046267 A1 WO 2017046267A1
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als
antibody
ligand
lateral sclerosis
treatment
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PCT/EP2016/071867
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German (de)
English (en)
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Walter Schubert
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ToposNomos Ltd.
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
    • C07K16/2803—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the immunoglobulin superfamily
    • C07K16/283—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the immunoglobulin superfamily against Fc-receptors, e.g. CD16, CD32, CD64
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P21/00—Drugs for disorders of the muscular or neuromuscular system
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
    • C07K16/2803—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the immunoglobulin superfamily
    • C07K16/2815—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the immunoglobulin superfamily against CD8
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
    • G01N33/6896—Neurological disorders, e.g. Alzheimer's disease
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/30—Immunoglobulins specific features characterized by aspects of specificity or valency
    • C07K2317/31—Immunoglobulins specific features characterized by aspects of specificity or valency multispecific
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/30—Immunoglobulins specific features characterized by aspects of specificity or valency
    • C07K2317/33—Crossreactivity, e.g. for species or epitope, or lack of said crossreactivity

Definitions

  • ALS amyotrophic lateral sclerosis
  • the invention relates to drugs for use in the treatment of amyotrophic lateral sclerosis (ALS), the treatment of ALS, methods for the diagnosis and / or therapy control of ALS, and a composition, a biochip and / or a kit for the diagnosis and / or therapy control of ALS ,
  • ALS amyotrophic lateral sclerosis
  • ALS Amyotrophic lateral sclerosis
  • Object of the present invention is to provide active ingredients for the treatment of amyotrophic lateral sclerosis (ALS). Another object of the invention is to provide a treatment method of ALS. Another object of the invention is to provide a method for diagnosis and / or therapy control of ALS. Another object of the invention is to provide a composition, a biochip and / or a kit for the diagnosis and / or therapy control of ALS.
  • ALS amyotrophic lateral sclerosis
  • a first aspect of the invention relates to an antibody and / or ligand which comprises at least one of the group CD16, CD8, STTP1, NeuN, Bax, Bcl2, CD1 1b, CD138, CD16A, CD29, CD2, CD45RA, CD49d, CD54, CD56 , CD57, CD58, CD62L, CD3, HLADR, immunoglobulin G, MHCII, MHCI, SIRT1, RAC1, BMX, GAK, JNK2, MAPKK6, OTUB2, PRKAR2A, SMAD2, SMAD4 and STAP2, for use in the treatment of Amyotrophic Lateral Sclerosis ( WHEN).
  • WHEN Amyotrophic Lateral Sclerosis
  • the antibody and / or ligand two, three, four, five or more from the group CD16, CD8, STTP1, NeuN, Bax, Bcl2, CD1 1b, CD138, CD16A, CD29, CD2, CD45RA, CD49d, CD54, CD56, CD57, CD58, CD62L, CD3, HLADR, immunoglobulin G, MHCII, MHCI, SIRT1, RAC1, BMX, GAK, JNK2, MAPKK6, OTUB2, PRKAR2A, SMAD2, SMAD4 and STAP2.
  • the antibody and / or ligand is bi-, tri-, quad-, pent-, hex-, hept-, oct-, denominated or multispecific.
  • the proteins mentioned form super-individual and individual clusters in ALS-specific cells in various combinations and can therefore be cross-linked therapeutically and thereby blocked, as a result of which the ALS-specific cells lose their functionality. This allows a particularly specific and thus low-side-effect or even side-effect-free treatment of ALS.
  • STTP1 here denotes the optionally patient-specific signal protein (signal transduction protein 1, eg kinase), which mediates the signal cascade extending from the cell surface to the cell nucleus and together with the module "CD16a and CD8" on the surface of the Cell where it is colocalized with CD16a and CD8 STTP1 can be isolated by means of the MELK and / or ICM technique known per se (multi-epitope ligand cartography or Imaging Cycler Microscopy) can be individually determined or monitored for the relevant patient.
  • signal transduction protein 1 signal transduction protein 1, eg kinase
  • STTP1 is at least one protein from the group RAC1, STAP2 and SMAD2.
  • This can block ALS-specific protein clusters which, in addition to one or more cell surface proteins (eg CD8, CD16, CD45RA) are additionally directly associated with one or more signal chain molecules from the group RAC1, STAP2 and / or SMAD2.
  • RAC1 Ras-related C3 botulinum toxin substrate 1 as a member of the RAC subfamily regulates a variety of cellular events, including cell growth control, cytoskeletal reorganization, and activation of protein kinases.
  • Signal transducing adapter protein 2 and mothers against decapen- ticidal homologue 2 (SMP) 2 are also part of the signal transduction chain and regulate signal transduction and transcription of central signaling pathways in ALS-specific cells.
  • the antibody and / or ligand comprises or is at least one variable single-chain fragment (scFv) and / or a multivalent antibody fragment (scFv multimer).
  • Antibody fragments offer the advantage of high binding affinity and specificity for a wide range of targets and haptens.
  • Single-chain fragments can also be cross-linked or expressed as diabodies (60 kDa), triabodies (90 kDa), tetrabodies (120 kDa), etc., whereby different linker lengths between V domains are possible.
  • a particular advantage for the treatment of ALS is that molecules of 60-120 kDa increase the penetration of ALS-specific cells and have faster clearance rates than the corresponding Igs (150 kDa).
  • the antibody and / or ligand is a diabody, triabody, tetrabody, pentabody, hexabody, heptabody or octabody.
  • a second aspect of the invention relates to a pharmaceutical composition for use in the treatment of amyotrophic lateral sclerosis (ALS) comprising at least one antibody and / or ligand according to the first aspect of the invention.
  • the pharmaceutical composition may comprise a salt or solvate thereof and / or one or more of pharmaceutically acceptable carriers, diluents and excipients.
  • the pharmaceutical composition binds at least one antibody and / or ligand, the CD8, CD16 and RAC1, and / or at least one antibody and / or ligand, the CD8, CD16 and STAP2 and / or SMAD2 binds, includes.
  • super-individual ALS-specific targets CD8 / CD16 / RAC1
  • individual ALS-specific targets CD8 / CD16 / STAP2, CD8 / CD16 / SMAD2, CD8 / CD16 / STAP2 / SMAD2
  • a third aspect of the invention relates to an IgG inhibitor for use in the treatment of Amyotrophic Lateral Sclerosis (ALS).
  • An example of this are antibodies against IgG and (optionally recombinant) streptococcal protein G.
  • a fourth aspect of the invention relates to a CD16 ectodomain shedding molecule for use in the treatment of amyotrophic lateral sclerosis (ALS).
  • a fifth aspect of the invention relates to a lamellipodia extension and / or migration inhibitor for use in the treatment of amyotrophic lateral sclerosis (ALS).
  • Lamellipodia are leaf-shaped thin pseudopodia on the anterior hemisphere of creeping and similarly motile cells, including ALS-specific cells.
  • a lamellipodium consists of a thin membrane duplicature that encloses a dense grid of parallel aligned bundles of actin filaments. With their plus ends (growth side), the actin bundles are anchored to the anterior chamber of the lamellipodium in the cell membrane and push them forward by continuously growing further actin bricks, while they are continuously broken down at the rear end, their minus side.
  • lamellipodia extension and / or migration inhibitors are, for example, compounds which block RAC1 and / or SIRT1 or prevent their expression.
  • the SIRT1 inhibitor nicotinamide (NAM) or SIRT1 "small interfering RNAs" suppress the lamellipodia extension via platelet-derived growth factor (PDGF).
  • PDGF platelet-derived growth factor
  • a sixth aspect of the invention relates to an inhibitor of the CD16 gene (FCGR3A) and / or the NeuN gene (RBFOX3) for use in the treatment of Amyotrophic Lateral Sclerosis (ALS).
  • a seventh aspect of the invention relates to a method for treating amyotrophic lateral sclerosis (ALS), in which a patient a pharmacologically effective dose of an antibody and / or ligands according to the first aspect of the invention and / or a pharmaceutical composition according to the second aspect of the invention and / or an IgG inhibitor according to the third aspect of the invention and / or a lamellipodia extension and / or migration inhibitor according to the fourth aspect of the invention and / or a CD16 ectodomain shedding molecule according to the fifth aspect of the invention and / or an inhibitor of the CD16 gene ( FCGR3A) and / or the NeuN gene (RBFOX3) according to the sixth aspect of the invention.
  • Preferred is a systemic or intravenous administration. Also included is
  • An eighth aspect of the invention relates to a method of identifying cells indicative of Amyotrophic Lateral Sclerosis (ALS) comprising the steps of providing a sample of a patient's cell and checking whether the cell sample contains at least one cell comprising NeuN and / or CD16, wherein the patient suffers from ALS when the cell sample contains at least one cell containing NeuN and / or CD16.
  • ALS Amyotrophic Lateral Sclerosis
  • a blood sample and / or a tissue sample of the patient is provided as the cell sample.
  • the blood sample is taken between 8 and 10 o'clock in the morning, the patient should be fasting.
  • Further advantages arise when it is checked whether the cell sample contains at least one cell in addition to NeuN and / or CD16 additionally one or more of CD8, STTP1, Bax, Bcl2, CD1 1b, CD138, CD16A, CD29, CD2, CD45RA , CD49d, CD54, CD56, CD57, CD58, CD62L, CD3, HLADR, immunoglobulin G, MHCII, MHCI, SIRT1, RAC1, BMX, GAK, JNK2, MAPKK6, OTUB2, PRKAR2A, SMAD2, SMAD4 and STAP2.
  • At least one composition which contains at least one antibody and / or ligand and / or a MELK or ICM method multi-epitope ligand cartography / imaging cycler microscopy
  • a biochip and / or a kit and / or device is used.
  • the MELK or ICM process can be carried out, for example, by means of a device known per se, as described in patent EP 0810428 B1 and in US Pat. No. 6,150,173.
  • the cell sample with the composition containing the at least one antibody and / or ligand acted by at least one part of the composition by at least one hollow body passed to the cell sample, at least once at least partially into the Retracted hollow body and at least partially passed back to the cell sample.
  • the hollow body can be formed, for example, as a pipe, hose, pipette or pipette tip and the like.
  • the process of retraction and return can be repeated basically once, twice, three times, four times, five times, six times or more frequently (N times, where N is an integer> 0), the time intervals and volume flows kept constant or varied in each round can be.
  • the process of retraction and return is preferably carried out with one and the same hollow body, that is, for example, without pipette tip change.
  • the process of retraction and return is preferably carried out with one and the same hollow body, that is, for example, without pipette tip change.
  • not only a part but the entire volume of the composition applied to the cell sample is withdrawn and redirected to the cell sample. This results in 1 ... N cycles of incubation with the same antibody and / or ligand type.
  • composition is passed onto the cell sample at an angle between 1 ° and 90 ° and / or withdrawn from the cell sample.
  • the composition is applied either at a right angle to the surface of the cell sample ("hanging drop") or
  • a ninth aspect of the invention relates to a composition and / or biochip and / or kit and / or apparatus for use in a method according to the eighth aspect of the invention.
  • the device can be, for example, a suitably adapted Toponome Imaging System (TIS), as described for example in the patent EP 810 428 B1.
  • TIS Toponome Imaging System
  • the device and / or the kit may be structurally simple and thus inexpensive tube systems with which antibody and / or ligand-containing compositions can be applied to and aspirated onto a patient's cell sample to test for the presence of ALS-specific binding patterns.
  • Figures 1a to 1f show different topographical images as more specific, abnormal
  • Figures 2a to 2f show the step process of these abnormal cells leading to the ALS specific disease
  • 3 shows different strategies for the treatment of ALS; and 4 shows a comparison between a classical incubation of a cell sample in the hanging drop and an incubation with improved diffusivity.
  • ALS Toponome Analyzes of cell surfaces of immune cells in the blood have shown that ALS has a specific abnormal topon. An essential feature is the existence of aberrant T-lymphocytes, which in addition to the CD8 receptor also express the CD16 receptor complex. Exactly the same cell forms are found in the morphologically well-preserved ALS postmortem tissue within postcapillary venules of the spinal pyramidal tract ( Figure 1 a, b; Box).
  • CD8 / CD16 complexes are required by "forward transport" for the transmigration of these cells through the endothelial cell layer via vesicle budding in the cell surface to promote transmigration as part of an abnormal "homing code” (CD16)
  • This complex is degraded after transmigration, since CD8 positive T cells in the pyramidal tract parenchyma do not have the CD16 complex, but instead can be found as single cells between the myelinated neu- tral cells.
  • Fig. 2 a, CD8 + CD16 - blue cell) Increased 3D magnification of this cell has a cell process that has penetrated deep into the surface of a morphologically intact axon (Fig.
  • This cell process expresses CD3 and CD8 ( Figure 2, arrows 1, 2.)
  • a geometrically accurate 3D calculation yields the cell extension in FIG. 2 f (arrow 3) and the axon in Fig. 2 f (marked with asterisks).
  • a similar cell process can be found starting from the same cell in Fig. 2 d (lower half): Here has the Cellular process (red) apparently permeated the intact myelin sheath (white). For comparison, Fig. 2 e shows an intact myelin sheath.
  • CD16 is necessary to control the abnormal homing aberrant CD8 cells into the pyramidal tract, and when this homing process is complete, the cells degrade (lose) the CD16 complex, then migrate as CD8 + CD16 cells between the cells
  • This process is obviously primarily autonomous, as the cells are never observed in the context of ZeH "debris," which is a sign of primary neuronal degradation, so the cells play a primary pathogenetic role supported by the fact that downregulation of CD16 leads to a halt of disease progression: when CD16 is no longer available as in Figure 1f, the endothelial homing process can no longer be performed are therefore described as a disease of the Immunzelltoponoms that by means of a highly selective "homing codes" the ALS speci fish pyramidal tract lesions (axotomy) generated. Many genetic findings and protein aggregations described in ALS have also been described in experimental axotomy, so that immunocelltoponomically-mediated axotomy explains these findings.
  • the invasive cells according to FIG. 1 c furthermore coexpress the following molecules: Bax, Bcl2, CD1b, CD138, CD16A, CD29, CD2, CD45RA, CD49d, CD54, CD56, CD57, CD58, CD62L, CD8, HLADR, immunoglobulin G, MHCII , MHCI, NeuN, SIRT1, RAC1, BMX, GAK, JNK2, MAPKK6, OTUB2, PRKAR2A, SMAD2, SMAD4 and STAP2.
  • NeuN Fluor-3, Rbfox3, or Hexaribonucleotide Binding Protein-3 is a neuronal, nuclear antigen that is normally used as a biomarker for neurons
  • Nuclear coexpression of NeuN and CD49d, as well as cytoplasmic expression of immunoglobulin G (IgG) along with the CD16 bearing vesicles of Figure 1df) and cell surface expression of CD8 and CD3, indicate that this cell has abnormal differentiation status:
  • T cell CD3, CD8
  • Neuronal cell Neuronal cell in the nucleus
  • CD16 monocyte
  • B lymphocyte IgG
  • Proteins partially colocalized with specific CD8 and / or CD16 in ALS-indicative cells, in some cases with specific individuality or frequency, are given in Table 1. All of these proteins thus represent therapeutic target structures (targets) individually or in combination with CD8 and / or CD16, since switching off these proteins-for example by means of appropriate antibodies / ligands and / or genetic engineering methods-leads to a collapse of the intracellular information flow and thus to a loss of function of the aberrant cells.
  • GAK [GAK] cyclin G associated kinase
  • MAPKK6 mitogen-activated protein kinase kinase 6
  • OTUB2 OTU deubiquitinase, ubiquitin aldehyde binding 2
  • PRKAR2 [PRKAR2A] protein kinase cAMP-dependent type II regulatory sub- A unit alpha
  • RAC1 ras-related C3 botulinum toxin substrates 1 (rho family, small GTP binding protein Rad)
  • SMAD2 [SMAD2] SMAD family member 2
  • SMAD4 [SMAD4] SMAD family member 4
  • STAP2 [STAP2] signal transducing adapter family member 2
  • SIRT1 nicotinamide adenine dinucleotide-dependent protein deacetylase
  • the cell of Figure 2c, f which, as explained above, expresses a CD16 negative phenotype upon invasion of the cell of Figure 1, remains positive for NeuN.
  • the invasive cell described above can be detected as an ALS-specific cell with the abovementioned features in the blood or in tissue samples by means of spatial tomography analysis. Since their invasion behavior into the pyramidal tract is known as set forth above and since it is further known that the cell is transformed by phenotype switching (CD16-) to a NeuN positive T cell which axotomizes the neuron, the detection of cells of the CD16 / CD8 phenotype from Fig. 1 c on the process of axotomy close. It follows that cell invasion of the cell of Figure 1c must be therapeutically prevented by molecular blocking or lysis in order to prevent axotomy and stop the progression of ALS.
  • the described two-step process is a violation of basic rules of cooperation of cells of different cotyledons: These rules are based in intact systems, cells from different cotyledons follow the rule of non-injury of the mutual cell surfaces.
  • the transformed T cell-like cells (cotyledon mesenchyme) injure the surfaces of neurons (cotylus ectoderm). This leads to a physical transcellular cross-linking of the two cotyledonous functions with the consequence of the interruption of the nerve tracts to the voluntary musculature.
  • NeuN in the nucleus of the acting cells (immune cells in the circulation and after invasion into the nervous system of the pyramidal tract) means that these cells A program of aberrant neuronal stem cells follows because NeuN is expressed under physiological conditions only in neurons. Thus, NeuN can be used as a marker for the presence of ALS-causing cells, that is for the diagnosis of ALS.
  • IgG IgG 1 .
  • Provide molecules antibodies / ligands that bind and / or block / inhibit IgG (1/3) or block / inhibit the physiological mechanism of IgG-mediated activation of CD16.
  • An example of this is antibodies to IgG and (optionally recombinant) streptococcal protein G;
  • STTP1 denotes the optionally patient-specific signal protein (signal transduction protein 1, eg kinase), which mediates the signal cascade emanating from the cell nucleus and coupled together with the module "CD16a and CD8" on the surface of the cell or with
  • STTP1 can be individually determined or controlled with the aid of the known MELK and / or ICM technique (multi-epitope ligand cartography or imaging cycler microscopy) for the respective patient.
  • STTP1 may be, for example, one or more proteins from the group RAC1, STAP2 and / or SMAD2 and / or another protein from the signal transduction chain of ALS cells.
  • a speaking antibody for example a trispecific anti-CD16a-CD8-STTP1 antibody, can then also be prepared by known per se manufacturing method and used for the therapy of the patient in question. Since such a trispecific antibody has an extremely high selectivity for abnormal, ALS-specific cells, such treatment is particularly low in side effects;
  • shedding or ectodomain shedding molecules that cause the proteolytic separation of the extracellular CD16 domains (so-called shedding or ectodomain shedding).
  • An example of this is the administration of Adam17 (metallopeptidase domain 17), which is also called TACE (tumor necrosis factor-a-converting enzyme).
  • Adam17 is a 70 kDa enzyme belonging to the ADAM protein family of disintegrins and metalloproteases.
  • Adam17 or the respective CD16-shedding-capable molecule can optionally be coupled with a mono-, bi-, tri- or multispecific antibody (see point 2) in order to increase its specificity advantageously;
  • Administration of at least one mono-, bi-, tri- or multi-specific anti-CD16 antibody (recombinant, human, de novo etc.);
  • FCGR3A CD16 gene
  • the above-mentioned compounds (drugs) can in principle be administered individually or in any combination.
  • the compounds mentioned can be present as a pharmaceutical composition together with other pharmaceutical substances and / or auxiliaries.
  • the galenics are to be adapted or chosen in a manner known per se.
  • the drug (s) is (are) administered once or several times.
  • a pharmaceutically effective dose is administered all at once or distributed over two or more divided doses. Therapy should be checked through regular monitoring and adjusted if necessary.
  • the therapy control can be carried out, for example, with the aid of the diagnostic method described above.
  • ALS-specific surface proteins CDs
  • surface protein clusters that are cross-linked, inhibited or otherwise blocked.
  • central signaling chain molecules may optionally be cross-linked, inhibited, or otherwise blocked by ALS cells.
  • the CD8 / CD16A CD45RA cell surface proteins may optionally be blocked along with the signal chain molecule RAC1, as these are exclusively associated directly in ALS cells.
  • Such a therapy can be carried out, for example, with the aid of bi-, tri- and / or tetraspecific antibodies or by means of corresponding variable single-chain fragments (scFv) or multivalent antibody fragments (scFv multimers), that is to say by means of so-called diabodies, triabodies or tetrabodies.
  • scFv variable single-chain fragments
  • scFv multimers multivalent antibody fragments
  • Individual targets that can be therapeutically addressed as an alternative or in addition to the ALS-specific cluster described above include individual clusters of CD16 / CD8 with the molecules STAP2 and / or SMAD2 and optionally combinations of the molecules designated in Table 1 together with the CD8 / CD16 cluster or as isolated clusters without CD8 / CD16. These clusters can also be switched off with the help of optionally multispecific antibodies and / or antibody fragments.
  • 4 shows a comparison between a classical incubation of a cell sample in the hanging drop (FIGS. 4a, 4b) and an incubation with improved diffusivity (FIGS. 4c, 4d).
  • a kit or a device with a tube system is used to apply solutions containing antibodies and / or ligands to a tissue or cell sample, wherein the solution in the experiment shown in FIGS. 4c and 4d can be selected at arbitrary intervals within a tube of the tubing system was reciprocated back into the tubing and passed back to the tissue or cell sample to produce an improved interaction of the reagents in the solution with the sample to be incubated.
  • FITC does not disturb the aforementioned hinge region of the antibody because it is a comparatively small molecule. It should be emphasized, however, that instead of or in addition to FITC, other labeling and fluorescent dyes can be used as long as they do not substantially interfere with the functionality of the hinge region.
  • FIG. 4a shows in detail the resulting gray level distribution of the tag of tissue in the "hanging drop" over 30 minutes using a composition with an antibody concentration of 0.5 mg / ml
  • FIG. 4b shows the corresponding histogram of Fig. 4a.
  • FIG. 4 c shows the gray value distribution of the marking of the tissue or cell sample after a total incubation time of 30 minutes.
  • the same volume of the same composition as in FIG. 4a / FIG. 4b a total of ten times every 3 minutes over a total of 30 minutes in short pulses (Jet Flow) applied, sucked off and reapplied.
  • Jet Flow short pulses
  • 4c, 4d is achieved by activating the hinge region of the antibody.
  • a significant superiority of this form of application compared to the "classical" hanging drop method is seen with an efficiency increase of about 67% .
  • hitherto invisible subcellular protein arrangements have been identified, which are marked with arrows in FIG Identify clustering.

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Abstract

L'invention concerne des principes actifs pharmaceutiques à utiliser pour le traitement de sclérose latérale amyotrophique (LSA), et un procédé de diagnostic de la SLA.
PCT/EP2016/071867 2015-09-17 2016-09-15 Principes actifs pharmaceutiques à utiliser pour le traitement de sclérose latérale amyotrophique (sla) et procédé de diagnostic associé WO2017046267A1 (fr)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
DE102015115747 2015-09-17
DE102015115747.0 2015-09-17
DE102016103562.9 2016-02-29
DE102016103562.9A DE102016103562A1 (de) 2015-09-17 2016-02-29 Pharmazeutische Wirkstoffe zur Verwendung in der Behandlung von Amyotropher Lateralsklerose (ALS) und Diagnoseverfahren

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WO2017046267A1 true WO2017046267A1 (fr) 2017-03-23

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2021156832A1 (fr) * 2020-02-07 2021-08-12 Biorchestra Co., Ltd. Utilisation d'inhibiteurs de miarn-485 pour traiter la sclérose latérale amyotrophique (sla)

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WO1999029731A1 (fr) * 1997-06-05 1999-06-17 Walter Schubert Utilisation de substance a effet immunomodulateur pour le traitement de la sclerose lateral amyotrophique
US6150173A (en) 1996-05-29 2000-11-21 Schubert; Walter Automated determining and measuring device and method
EP0810428B1 (fr) 1996-05-29 2004-04-28 Walter Dr. Schubert Appareil automatique et procédé de mesure et d'identification de molécules ou de fragments de molécules
WO2006100033A1 (fr) * 2005-03-21 2006-09-28 Mpb Meltec Patent- Und Beteiligungsgesellschaft Mbh Procede permettant d'identifier des motifs de combinaison de proteines specifiques de cellule
WO2014001324A1 (fr) * 2012-06-27 2014-01-03 Hoffmann-La Roche Ag Méthode de sélection et de production d'entités de ciblage personnalisées fortement sélectives et multi-spécifiques renfermant au moins deux entités de liaison différentes et leurs utilisations

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6150173A (en) 1996-05-29 2000-11-21 Schubert; Walter Automated determining and measuring device and method
EP0810428B1 (fr) 1996-05-29 2004-04-28 Walter Dr. Schubert Appareil automatique et procédé de mesure et d'identification de molécules ou de fragments de molécules
WO1999029731A1 (fr) * 1997-06-05 1999-06-17 Walter Schubert Utilisation de substance a effet immunomodulateur pour le traitement de la sclerose lateral amyotrophique
WO2006100033A1 (fr) * 2005-03-21 2006-09-28 Mpb Meltec Patent- Und Beteiligungsgesellschaft Mbh Procede permettant d'identifier des motifs de combinaison de proteines specifiques de cellule
WO2014001324A1 (fr) * 2012-06-27 2014-01-03 Hoffmann-La Roche Ag Méthode de sélection et de production d'entités de ciblage personnalisées fortement sélectives et multi-spécifiques renfermant au moins deux entités de liaison différentes et leurs utilisations

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2021156832A1 (fr) * 2020-02-07 2021-08-12 Biorchestra Co., Ltd. Utilisation d'inhibiteurs de miarn-485 pour traiter la sclérose latérale amyotrophique (sla)

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