ES2927133B2 - Treatment for Parkinson's disease - Google Patents

Description

DESCRIPTION

Parkinson's disease treatments

TECHNICAL SECTOR

The present invention is framed within the field of biomedicine. It specifically refers to insulin growth factor receptor agonists and their use in the treatment of Parkinson's disease.

BACKGROUND OF THE INVENTION

Parkinson's disease is a disorder of voluntary movement in which muscles become stiff and slow, movement becomes clumsy and difficult, and uncontrollable rhythmic jerks of muscle groups produce a characteristic tremor. The disease is thought to be caused by a degeneration of presynaptic dopaminergic neurons in the brain. The absence of adequate release of the chemical transmitter dopamine during neuronal activity thus leads to parkinsonian symptomatology.

Currently, the most widely used treatment for Parkinson's disease is the administration of L-DOPA, a dopamine precursor that acts indirectly by replacing the missing dopamine. However, the use of L-DOPA has disadvantages, for example, patients often suffer from side effects such as dyskinesia and on-off effects, and it is necessary to administer L-DOPA together with a peripheral dopa decarboxylase inhibitor. , such as carbidopa or benzaseride. These inhibitors prevent the peripheral breakdown of levodopa into dopamine, thereby allowing more of the drug to enter the brain and limiting peripheral side effects. This treatment improves the quality of life of patients, but does not stop the progression of the disease. Furthermore, such treatment is associated with a number of adverse effects, such as nausea, vomiting, abdominal distension, and psychiatric side effects (e.g., toxic confusional states, paranoia, and hallucinations).

An alternative form of therapy is the administration of postsynaptic dopamine agonists, for example ergot alkaloids such as bromocriptine, however this approach is also associated with side effects.

For example, patients receiving bromocriptine often experience problems with dyskinesia and in a small number of cases experience vasospastic phenomena and angina. Additionally, bromocriptine also causes psychiatric side effects, such as hallucinations.

In view of the above, it is clear that there is still a need to provide effective and safe medications for the treatment of Parkinson's disease and disorders associated with parkinsonism.

BRIEF EXPLANATION OF THE INVENTION

The authors of the invention have discovered that IGF-2 has antioxidant and neuroprotective effects on tyrosine hydroxylase (TH)+ positive cells (dopaminergic cells), especially at the mitochondrial level, where IGF-2 protects cells from oxidative damage and It inhibits cell death in specific motor areas, preventing the appearance of motor symptoms of Parkinson's disease.

More specifically, the inventors have shown that, in dopaminergic cells of the substantia nigra incubated with 1-methyl-4-phenylpyridinium (MPP+), treatment with IGF-2 led to a decrease in certain parameters that were increased as a consequence of the damage caused. by the presence of MPP+, such as cell death, lipid peroxidation, protein oxidation, superoxide radical production, levels of the inducible mitochondrial enzyme SOD, as well as mitochondrial damage. Likewise, IGF-2 was able to restore the activity of the diaphorase enzyme, increase the activity of glutathione-S-transferase, as well as restore the expression of the TH enzyme, through the IGF receptor. These results have also been confirmed in mice exposed to the toxic 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine (MPTP), where the inventors have shown that IGF-2 reduces neuronal damage, latency time in the Rotarod motor function test, as well as mortality. The inventors have also demonstrated that the action of IGF-2 occurs through the IGF-2R receptor, thereby supporting the suitability of IGF-2R agonists for treating Parkinson's disease and other disorders associated with parkinsonism.

Thus, in a first aspect the invention relates to a specific agonist of the insulin-like growth factor-2 (IGF-2) receptor or to a pharmaceutically acceptable salt thereof for use in the prevention and/or treatment of disease of Parkinson and disorders associated with parkinsonism.

In a second aspect, the invention relates to a composition comprising a specific agonist of the insulin-like growth factor-2 (IGF-2) receptor and a compound suitable for the treatment and/or prevention of the disease. Parkinson and disorders associated with parkinsonism.

In a third aspect, the invention relates to the composition of the second aspect of the invention for use in medicine.

In a fourth aspect, the invention relates to the composition of the second aspect of the invention for use in the treatment of Parkinson's disease and disorders associated with parkinsonism.

Finally, the invention relates to an in vitro method to identify a compound suitable for the treatment and/or prevention of Parkinson's disease and disorders associated with parkinsonism comprising

(a) contacting a candidate compound with a cell that expresses the IGF-2 receptor under conditions suitable for the binding of the compound and the IGF-2 receptor and

(b) determine the activity of the IGF-2 receptor,

where if the IGF-2 receptor activity after coming into contact with the candidate compound is greater than a reference value, the candidate compound is suitable for the treatment and/or prevention of Parkinson's disease and Parkinson's-associated disorders. parkinsonism.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1: Expression quantitative trait loci (eQTL) between rs680 and IFG-2 in the frontal cortex.

Figure 2: Immunocytochemistry of the IGF-2 receptor in SN4741 dopaminergic neurons.

Figure 3: Experimental design for the study of the neuroprotective effects of IGF-2 on neuronal survival and redox homeostasis.

Figure 4: Experimental design for the study of the neuroprotective effects of IGF2 in a mouse model of Parkinson's.

Figure 5: Pathways of oxidative homeostasis, survival and neuronal function with the parameters studied.

Figure 6: Cytotoxicity of MPP+ in SN4741 neurons after 6 hours of incubation, in the presence of IGF-2, BMS (Bristol-Myers Squibb, CAS No.468740-43-4) (IGF-I receptor antagonist and insulin ), AB (IGF-2r selective antibody). Cell damage was measured by quantifying the release of LDH expressed in % of the control. a: p<0.05 vs. control, b: p<0.05 vs. MPP, c: p<0.05 vs. MPP+IGF-2, d: p<0.05 vs. MPP+IGF- 2+BMS, e: p<0.05 versus IGF-2.

Figure 7: Oxidative damage assessed as lipid peroxidation and oxidized proteins per milligram of protein in SN4741 dopaminergic neurons. a: p<0.05 vs. control, b: p<0.05 vs. MPP, c: p<0.05 vs. MPP+IGF-2, d: p<0.05 vs. MPP+IGF- 2+AB

Figure 8: Superoxide radical production in SN4741 dopaminergic neurons, determined by flow cytometry using the dihydroethidine probe.

Figure 9: Measurement of the activity of the mitochondrial superoxide dismutase enzyme in SN4741 dopaminergic neurons. a: p<0.05 vs. control, b: p<0.05 vs. MPP, c: p<0.05 vs. MPP+IGF-2, d: p<0.05 vs. MPP+IGF- 2+AB.

Figure 10: Measurement of mitochondrial membrane potential by flow cytometry using JC-1 as a marker, and cytochrome c oxidase activity in SN4741 dopaminergic neurons. a: p<0.05 vs control, b: p<0.05 vs MPP, c: p<0.05 vs MPP+IGF-2, d: p<0.05 vs MPP+IGF-2+AB.

Figure 11: Electron microscopy of the mitochondria of SN4741 dopaminergic neurons. CO: untreated neurons, IGF-2: neurons treated with 25 ng/ml of IGF-2, MPP: neurons treated with 400 | ^j M for MPP+, MPP+IGF-2: neurons treated with IGF-2 in the presence of MPP.

Figure 12: Measurement of the enzymatic activity of the enzymes glutathione S-transferase and diaphorase in SN4741 dopaminergic neurons. a: p<0.05 vs. control, b: p<0.05 vs. MPP, c: p<0.05 vs. MPP+IGF-2, d: p<0.05 vs. MPP+IGF-2 +AB.

Figure 13: Confocal microscopy of tyrosine hydroxylase (TH) immunocytochemistry in SN4741 dopaminergic neurons. a: p<0.05 vs control, b: p<0.05 vs MPP.

Figure 14: Study of the effect of Leu[27]IGF, a complete IGF-2r agonist, on the following parameters: neuronal survival (A), ROS production (B), mitochondrial membrane potential (C), proteins oxidized (D), antioxidant enzyme activity (E) and neuronal TH expression (F). a: p<0.05 vs control, b: p<0.05 vs MPP.

Figure 15: Stereological analysis and representative images of TH immunohistochemistry in substantia nigra compact neurons in C57BL/6J mice treated with IGF-2 or vehicle in the case of controls, in the presence or absence of MPTP (plus probenecid) for 44 days. * p<0.05 compared to other groups.

Figure 16: Determination of motor activity using Rotarod in mice treated with IGF-2 in the presence or absence of MPTP. Mean Rotarod latency to fall (A) and correlation between Rotarod latency to fall and TH-positive neurons in the SNc (B). * p<0.05 compared to other groups.

Figure 17: Effect of IGF-2 administration on the survival of mice in the MPTP-induced toxicity model.

PREFERRED EMBODIMENT OF THE INVENTION

Specific IGF-2R agonist and its uses

In a first aspect, the invention relates to a specific agonist of the insulin-like growth factor 2 (IGF-2) receptor or a pharmaceutically acceptable salt thereof for use in the prevention and/or treatment of the disease. Parkinson and disorders associated with parkinsonism.

A "specific insulin-like growth factor-2 receptor (IGF-2R) agonist" or "specific IGF-2R agonist" refers to a ligand that binds to the IGF-2 receptor with higher affinity than to the other insulin receptor subtypes and which, when bound, mimics the effects of endogenous ligand binding. In a particular embodiment, the specific IGF-2R agonist binds to the same IGF-2R binding site as IGF-2. In a particular embodiment, said binding site is in segment 11 of the IGF-2R ectodomain, with residues in domain 13 that increase the affinity of the interaction (Olson LJ et al., Glycobiology 2015, 25, 591-606, Dahms et al., J Biol Chem 1994, 269(5): 3802-9 and Garmroudi and MacDonald, J Biol Chem 1994, 269(43): 26944-52). In a particular embodiment, the specific IGF-2R agonist mimics the effect of binding from IGF-2 to IGF-2R. In a particular embodiment, the specific IGF-2R agonist binds to the same IGF-2R binding site as IGF-2 and mimics the effect of IGF-2 binding to IGF-2R. The term specific IGF-2R agonist includes the endogenous ligand of IGF-2R, that is, IGF-2.

As the expert will know, to determine if a compound is a specific agonist of the IGF-2R it is necessary to use two types of assays: assays capable of determining if said compound specifically binds to the IGF-2R and assays capable of determining if said compound mimics the IGF-2R. effect of the endogenous ligand of the IGF-2R, in this case, the effect triggered by the binding of IGF-2 to the IGF-2R. Assays capable of determining whether a compound specifically binds to IGF-R2 include, without limitation, ligand binding assays and immunohistochemistry, such as, for example, the assays described in Asha Amritraj et al., Endocrinology, 2012, 153, 4784 -4798 and Cheryl Hawkes et al., J Neuroscience 2006; 26: 585-596. Several effects triggered by the binding of IGF-2 to IGF-2R have been described, such as:

- calcium entry in mouse embryo fibroblasts (Nishimoto I, et al., J Biol Chem. 1987; 262(25):12120-6) and in rat skull osteoblasts (Martinez DA, et al., J Cell Biochem, 1995; 59(2):246-57) and

- increased protein phosphorylation (Hammerman MR, Gavin JR 3rd, J Biol Chem. 1984; 259(21):13511-7) and

- increased alkalinization in the cells of the proximal renal tubule (Mellas J, Gavin JR 3rd, Hammerman MR, J Biol Chem. 1986; 261(31):14437-42),

- stimulation of Na+/H+ exchange and inositol trisphosphate production in canine kidney cells (Rogers SA, Hammerman MR, Proc Natl Acad Sci USA.

1988; 85(11):4037-41),

-increased uptake of amino acids in muscle cells (Shimizu M, et al., Am J Physiol. 1986; 251(5 Pt 1):E611-5),

- increased glycogen synthesis in hepatoma cells (Hari J, et al., EMBO J. 1987; 6(11):3367-71) and

- the synthesis of proteoglycans in human chondrosarcoma cells (Takigawa M, et al., Endocrinology. 1997; 138(10):4390-400),

- calcium mobilization in rabbit articular chondrocytes (Poiraudeau S, et al., J Cell Biochem. 1997; 64(3):414-22), cell motility in rhabdomyosarcoma cells (El-Badry OM, et al., Cell Growth Differ. 1990; 1(7):325-31 and Minniti CP, et al., J Biol Chem. 1992; 267(13):9000-4), aromatase activity in placental cytotrophoblasts (Nestler JE, Endocrinology. 1990; 127(5):2064-70),

- the migration of human extravillous trophoblasts (McKinnon T, et al., J Clin Endocrinol Metab. 2001; 86(8):3665-74), insulin exocytosis by pancreatic p cells Zhang Q, et al., Proc Natl Acad Sci U SA. 1997; 94(12):6232-7)

- regulation of the endogenous release of acetylcholine and GABA in the adult rat brain (Hawkes C, et al., J Neurosci. 2006; 26(2):585-96 and Amritraj A, et al., Neuroscience. 2010; 170(3):722-30).

Non-limiting examples of assays capable of determining whether a compound that binds IGF-2R mimics the effect of IGF-2 binding to IGF-2R include, without limitation, the following:

- Measurement of the proliferation of CHO cells after stimulation with the specific agonist. Typically, stimulation of CHO cells with 75 ng/ml of IGF-2 achieves a maximum stimulation rate equal to that obtained with insulin at 8 ng/ml (Mamounas M et al., 1989, PNAS, vol. 86 , pp 9294-9298). Those skilled in the art could also measure IGF-2R activation and design specific assays capable of analyzing its activation.

- Measurement of intracellular phosphoprotein kinase Ca (PKCa) levels or its dependent substrates MARCKS and GAP-43, as described in Hawkes et al., The Journal of Neuroscience, 2006, 26(2): 585-596.

"Insulin-like growth factor 2 receptor" (IGF2R), also called "cation-independent mannose 6-phosphate receptor" (CD-MPR), is a protein that in humans is encoded by the IGF2R gene. The human gene appears in the database under accession number ENSG00000197081 (Ensembl published 102, November 2020). IGF2R is a multifunctional protein receptor that binds to insulin-like growth factor 2 (IGF2) on the cell surface and to proteins containing mannose-6-phosphate (M6P) residues in the trans-Golgi network.

The IGF system includes four receptors: the insulin receptor (IR), IGF-1R, IGF-2R, and hybrid receptors consisting of half IR and half IGF-1R. These Receptors interact with three main ligands: insulin, IGF-1 and IGF-2. IR, IGF-1R, and IGF 2R have the highest binding affinity to their respective ligands, while the binding of insulin to IGF-1R and of IGF-1 to IR is at least 100-fold lower. IGF-1 and IGF-2 signaling is mediated by the IGF-1R; but IGF-1 has a binding affinity at least 3 times greater than IGF-2. In another embodiment, the specific IGF-2R agonist binds to the IGF-2 receptor with an affinity approximately 1000 times greater than IGF-I and does not interact with insulin. Specific IGF-2R agonist can specifically increase IGF-2R-mediated signaling activity. By specific it is meant that the IGF-2R agonist preferentially activates the IGF-2R relative to the IGF-1R, that is, at a given concentration, the specific IGF-2R agonist increases IGF-2R-mediated signaling in greater extent than that which increases IGF-1R-mediated signaling. In some embodiments, the specific IGF-2R agonist binds to IGF-2R with an affinity that is at least 1.2-fold, at least 1.5-fold, at least 2-fold, at least 5-fold, at least 10-fold. , at least 15 times, at least 20 times, at least 25 times, at least 30 times, at least 35 times, at least 40 times, at least 45 times, at least 50 times, at least 75 times, at least 100 times stronger than its binding affinity to IGF-1R. In a particular embodiment, the specific IGF-2R agonist binds to IGF-2R with an affinity that is at least 20-fold to 50-fold stronger than its affinity for binding to IGF-1R, more particularly between 24-fold and 45-fold. times. The affinity of an agonist to bind IGF1R or IGF-2R can be determined using a binding assay such as, for example, that described by Beukers et al., Endocrinology 1991; 1201-1203; Rongshi Li et al., J Med Chem. 2009; 52: 4981-5004; Hawkes C et al., J Neuroscience 2006;26:585-596. In some embodiments, the specific IGF-2R agonist does not detectably bind to the IGF-1R. In some embodiments, the specific IGF-2R agonist activates IGF-2R signaling to a level that is at least 1.2-fold, at least 1.5-fold, at least 2-fold, at least 5-fold, or at least 10-fold. times greater than that which activates IGF-1R signaling. In some embodiments, the specific IGF-2R agonist does not activate a detectable level of the IGF-1R. Non-limiting examples of specific IGF-2R agonists include, without limitation, IGF-2, Leu[27]IGF-2, M6PRBP1, (Arg6) IGF-2 and Des[1-6] Insulin-like Growth Factor- 2 (IGF-2).

"Pharmaceutically acceptable salt" refers to those salts that retain the biological efficacy and optionally other properties of free bases and that are obtained by reaction with inorganic or organic acids such as hydrochloric acid, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, malic acid, maleic acid, succinic acid, tartaric acid, acid citrus and similar.

In the event that embodiments of the disclosed compounds form salts, these salts are within the scope of the present invention. Reference to a compound or to a compound used in the composition or in the pharmaceutical composition of the invention described herein is understood to include reference to salts thereof, unless otherwise indicated. The term "salt(s)", as used herein, denotes acidic and/or basic salts formed with inorganic and/or organic acids and bases. Additionally, when a compound contains both a basic moiety and an acidic moiety, zwitterions ("internal salts") may form and are included in the term "salt(s)" as used herein. Pharmaceutically acceptable (eg, non-toxic, physiologically acceptable) salts are preferred, although other salts are also useful, for example, in isolation or purification steps that may be employed during preparation. Salts of the compounds may be formed, for example, by reacting the compound with an amount of acid or base, such as an equivalent amount, in a medium such as one in which the salt precipitates or in an aqueous medium followed by lyophilization.

Embodiments of the compounds containing a basic moiety can form salts with a variety of organic and inorganic acids. Examples of acid addition salts include acetates (such as those formed with acetic acid or trihaloacetic acid, e.g. trifluoroacetic acid), adipates, alginates, ascorbates, aspartates, benzoates, benzenesulfonates, bisulfates, borates, butyrates, citrates, camphorates, camphoresulfonates, cyclopentanepropionates, digluconates, dodecyl sulfates, ethanesulfonates, fumarates, glucoheptanoates, glycerophosphates, hemisulfates, heptanoates, hexanoates, hydrochlorides (formed with hydrochloric acid), hydrobromides (formed with hydrogen bromide), hydroiodides, 2-hydroxyethanesulfonates, lactates, maleates (formed with acid maleic), methanesulfonates (formed with methanesulfonic acid), 2-naphthalenesulfonates, nicotinates, nitrates, oxalates, pectinates, persulfates, 3-phenylpropionates, phosphates, picrates, pivalates, propionates, salicylates, succinates, sulfates (such as those formed with acid sulfuric acid), sulfonates (such as those mentioned herein), tartrates, thiocyanates, toluenesulfonates such as tosylates, undecanoates, and the like.

Embodiments of compounds containing an acid moiety can form salts with a variety of organic and inorganic bases. Examples of basic salts include ammonium salts, alkali metal salts such as sodium, lithium and potassium salts, alkaline earth metal salts such as calcium and magnesium salts, salts with organic bases (e.g. organic amines) such as benzathines dicyclohexylamines. , hydrabamines (formed with N,N-bis(dehydroabietyl)ethylenediamine), N-methyl-D-glucamines, N-methyl-D-glucamides, t-butylamines and salts with amino acids such as arginine, lysine and the like.

The term "prevention", "preventing" or "prevent", as used herein, refers to the administration of the specific IGF-2R receptor agonist to a subject who has not been diagnosed with Parkinson's disease or the disorders associated with parkinsonism, but who would normally be expected to develop the disease or be at increased risk of developing it. Prevention aims to avoid the appearance of the disease. Prevention can be complete (for example, the complete absence of the disease). Prevention may also be partial, so that, for example, the onset of a disease in a subject is less than what would have occurred without administration of the inhibitor of the present invention. Prevention also refers to reducing susceptibility to a clinical condition.

The term "treatment", as used herein, includes any type of therapy that aims to terminate, prevent, improve and/or reduce susceptibility to a clinical condition as described herein, for example, Parkinson's disease or disorders associated with parkinsonism. Therefore, "treatment", "treat" and the like, as used herein, refer to obtaining a desired pharmacological and/or physiological effect, encompassing any treatment of a pathological condition or disorder in a mammal, including a human. The effect may be prophylactic in terms of total or partial prevention of a disorder or symptom thereof and/or may be therapeutic in terms of partial or complete cure of a disorder and/or adverse effect attributable to the disorder. That is, "treatment" includes (1) preventing the disorder from occurring or recurring in a subject, (2) inhibiting the disorder, such as stopping its development, (3) stopping or ending the disorder or at least the symptoms associated with it, so that the person no longer suffers from the disorder or its symptoms, such as causing regression of the disorder or its symptoms, for example, by restoring or repairing a lost, absent or defective function, or stimulating an ineffective process, or (4) alleviating, alleviating or improving the disorder or the symptoms associated with it, where improve is used in a broad sense to refer to at least a reduction in the magnitude of a parameter. "Treatment" can also mean prolonging survival compared to what would be expected if no treatment is received. Among the people who need treatment are those who already suffer from the disease or disorder, as well as those who are prone to suffering from it or those for whom prevention is desired.

In a particular embodiment, the specific insulin-like growth factor-2 (IGF-2) receptor agonist or a pharmaceutically acceptable salt thereof is for use in the prevention and/or treatment of Parkinson's disease.

As used herein, "Parkinson's disease" or "PD" refers to a progressive, neurodegenerative brain disorder that ultimately affects an individual's muscle movement. A hallmark of PD is the degeneration of dopaminergic neurons in the substantia nigra pars compacta and the consequent striatal dopamine deficiency.

Many of the signs and symptoms associated with Parkinson's disease can precede typical Parkinson's disease, in some cases by many years. The involvement of the dopaminergic substantia nigra, which underlies the primary motor characteristics of the disease, occurs at a time when the disease is very advanced at a neuropathological level, an observation that may explain the difficulties in successfully testing new drugs in search for possible disease-modulating properties only after it is evident. The motor features of Parkinson's disease are characterized by muscle rigidity, tremor, gait and posture abnormalities, slowing of physical movement (bradykinesia), and, in extreme cases, loss of physical movement (akinesia). Primary symptoms are the result of decreased stimulation of the motor cortex and other areas of the brain by the basal ganglia, usually caused by insufficient formation and action of dopamine, which is produced in dopaminergic neurons in the brain. The motor features of Parkinson's disease are only one component of a much more widespread disorder that causes an abundance of non-motor signs and symptoms, such as olfactory dysfunction, REM sleep behavior disorder (RBD), constipation, depression and cognitive deficits. Importantly, many of these signs and symptoms can precede motor symptoms by years or even a decade or more.

The diagnosis of Parkinson's disease can be made following the Movement Disorder Society (MDS) Clinical Diagnostic Criteria for Parkinson's disease (PD) (Ronald B Postuma et al., Mov Disord 2015, Oct 30(12) , 1591-601). The Movement Disorders Society Criteria for PD are intended for use in clinical research, but can also be used to guide clinical diagnosis. The reference point for these criteria is expert clinical diagnosis; The criteria aim to systematize the diagnostic process, so that it is reproducible between centers and applicable by clinicians with less experience in the diagnosis of PD. Although motor abnormalities remain fundamental, there has been increasing recognition of non-motor manifestations; these are incorporated into both the current criteria and, in particular, the separate criteria for prodromal PE. Like the previous criteria, the Movement Disorders Society PD criteria maintain motor parkinsonism as the main feature of the disease, defined as bradykinesia plus resting tremor or rigidity. Once parkinsonism is documented, the determination of PD as a cause of parkinsonism is based on three categories of diagnostic features: absolute exclusion criteria (which rule out PD), red flags (which must be supplemented with additional supporting criteria to allow diagnosis of PD) and supporting criteria (positive characteristics that increase confidence in the diagnosis of PD). As the skilled person will know, as knowledge of PD expands, the Movement Disorder Society criteria will need ongoing revision to adapt to new developments.

The term bradykinesia is defined as slowing of movements and decreased amplitude or speed (or progressive hesitations/stops) as movements are continued. Bradykinesia can be assessed by finger tapping (MDS-UPDRS (Unified Parkinson's Disease Rating Scale) 3.4), hand movements (3.5), pronation/supination movements (3.6), the toes (3.7) and the tapping of the feet (3.8). Although bradykinesia also occurs in the domains of voice, face, and axial/throat movement, bradykinesia of the extremities must be documented to establish the diagnosis of PD. Bradykinesia, as defined here, combines into a single term the definitions of bradykinesia (slowness) and akinesia/hypokinesia (decreased range of movement); both are usually present in the examination, although not always simultaneously (that is, the Patients cannot move at a normal speed with a normal amplitude). In parkinsonism caused by PD, a decrease in speed or amplitude is seen as movements are continued, a feature that is sometimes not seen in parkinsonism caused by other diseases.

As stated in the MDS-UPDRS, stiffness is judged based on "slow passive movement of the major joints with the patient in a relaxed position and the examiner manipulating the extremities and neck." Stiffness refers to "lead pipe" resistance; that is, to velocity-independent resistance to passive movement that does not solely reflect lack of relaxation (i.e., distinct from spasticity or paratonia).

Although the cogwheel phenomenon is often present (and may reflect tremor that is incidentally perceived when evaluating pitch), isolated "cogwheel tremor" without "lead pipe" stiffness does not meet the minimum stiffness requirements.

Rest tremor refers to a 4 to 6 Hz tremor in the limb at rest, which is suppressed during movement initiation. Resting tremor can be assessed throughout the interview and examination (MDS-UPDRS 3.17, 3.18). Kinetic and postural tremors alone (MDS-UPDRS 3.15 and 3.16) do not meet the criteria for parkinsonism. In PD, a parkinsonian resting tremor of the hand with prolonged stance (i.e., a “reemerging” tremor) may also be seen; however, to meet the criteria, tremor during rest must also be observed. In patients with associated postural or kinetic tremor, care must be taken that the extremity is completely relaxed during the examination. The frequency of a true rest tremor is usually slower than the associated action tremor.

Disease models for PD include, but are not limited to, toxin-based models such as MPTP mice, MPTP monkeys, 6-OHDA rats, Rotenone, Paraquat/maneb, and MET/MDMA; genetic models such as animals with altered genes for a-synuclein, LRKK ₂ , PINK1, PARKIN, DJ-1, ATP13A2, SHH, Nurrl, Engrailed 1, Pitx3, C-Rel-NFκΒ, MitoPark, Atg7 and/or VMAT2.

In a particular embodiment, the specific insulin-like growth factor-2 (IGF-2) receptor agonist or a pharmaceutically acceptable salt thereof is for use in the prevention and/or treatment of parkinsonism or associated disorders. to parkinsonism. The term "disorders associated with parkinsonism" includes several other conditions that have the characteristics of Parkinson's disease and are variously called "parkinsonism,""Parkinson-likedisease,""secondaryparkinsonism,""Parkinson'ssyndrome," or "atypical Parkinson's."

Parkinsonism is a general term that refers to a group of neurological disorders that cause movement problems similar to those seen in Parkinson's disease, such as tremors, slowness of movement, and rigidity. Under the category of parkinsonism there are a number of disorders. Early in the disease process, it is often difficult to tell whether a person has idiopathic (meaning "of unknown origin") Parkinson's disease or a syndrome that mimics it. These syndromes tend to progress more rapidly than Parkinson's disease, present with additional symptoms such as early falls, dementia, or hallucinations, and do not respond or respond only briefly to levodopa therapy. The underlying causes of Parkinson-like disease or parkinsonism are numerous, and diagnosis can be complex. A wide range of etiologies can give rise to a similar set of symptoms, including some toxins, a few metabolic diseases, and a handful of neurological conditions unrelated to Parkinson's. A common cause is the side effect of medications, mainly neuroleptic antipsychotics, especially phenothiazines (such as perphenazine and chlorpromazine), thioxanthenes (such as flupenthixol and zuclopenthixol), and butyrophenones (such as haloperidol (Haldol)), piperazines (such as ziprasidone) and, rarely, antidepressants. Other causes are, among others, olivopontocerebellar degeneration, progressive supranuclear palsy, corticobasal degeneration, temporofrontal dementia; induced by drugs such as antipsychotics, prochlorperazine, metoclopromide; carbon monoxide poisoning; head trauma and parkinsonism in Huntington's disease. In some cases, alpha-synucleinopathies can lead to Parkinson's-like disease, secondary parkinsonism, Parkinson's syndrome, or atypical Parkinson's.

Examples of disorders associated with parkinsonism include, but are not limited to, drug-induced parkinsonism, early-onset Parkinson's disease, supranuclear palsy, multiple system atrophy, Lewy body disease or dementia with Lewy bodies, primary lateral sclerosis, multiple sclerosis, corticobasal degeneration and vascular parkinsonism.

Drug-induced parkinsonism (DIP), as used herein, refers to a drug-induced movement disorder caused by any medication that interferes with dopamine transmission. DIP is one of the most common causes of non-degenerative parkinsonism. Drug-induced parkinsonism usually affects both sides of the body equally, while PD is virtually always asymmetric, affecting one side of the body more than the other. The prototypical drugs are dopamine receptor blocking agents, specifically those that block D2.

"Young-onset Parkinson's disease" refers to a rare, genetic parkinsonian disorder characterized by an age of onset between 21 and 45 years, rigidity, painful cramps followed by tremor, bradykinesia, dystonia, gait discomfort and falls, and other non-motor symptoms. Slow disease progression and a more pronounced response to dopaminergic therapy are also observed in most forms of this disease.

The term "supranuclear palsy (PSP)" or "progressive supranuclear palsy" refers to a rare brain disorder that affects movement, control of walking and balance, speech, swallowing, vision, of mood and behavior, and thought. The disease is the result of damage to nerve cells in the brain. The disorder's long name indicates that the disease worsens (progressive) and causes weakness (paralysis) by damaging certain parts of the brain above groups of nerve cells called nuclei (supranuclear). These nuclei especially control eye movements. One of the classic signs of the disease is the inability to aim and move the eyes correctly, which individuals may experience as blurred vision. Both PSP and Parkinson's disease cause stiffness, movement difficulties, and clumsiness, but PSP is more rapidly progressive than Parkinson's disease. People with PSP often stand exceptionally straight or occasionally even tilt their heads back (and tend to fall backwards). This is called "axial stiffness." Those with Parkinson's disease often lean forward. Speech and swallowing problems are much more common and severe in PSP than in Parkinson's disease, and they tend to appear earlier in the course of the disease. Eye movements are abnormal in PSP, but almost normal in Parkinson's disease. Both diseases share other characteristics: onset in older adults, bradykinesia (slowness of movement) and muscle rigidity. The tremor, very common in individuals with Parkinson's disease, is rare in PSP. Although individuals with Parkinson's disease benefit markedly from the drug levodopa, people with PSP respond minimally and only briefly to this drug. Additionally, people with PSP show a buildup of the protein tau in affected brain cells, while people with Parkinson's disease show a buildup of a different protein, called alpha-synuclein. PSP symptoms are caused by a gradual deterioration of brain cells in a few specific areas of the brain, primarily in the region called the brainstem. One of these areas, the substantia nigra, is also affected in Parkinson's disease, and damage to this region of the brain partly explains the motor symptoms that PSP and Parkinson's have in common. The hallmark of the disease is the buildup of abnormal deposits of the protein tau in nerve cells in the brain, so that they do not function properly and die.

As used herein, "multiple system atrophy (MSA)" refers to a progressive neurodegenerative disorder characterized by a combination of symptoms that affect both the autonomic nervous system (the part of the nervous system that controls involuntary actions, such as blood pressure or digestion) and movement. Symptoms reflect the progressive loss of function and death of different types of nerve cells in the brain and spinal cord. Symptoms of autonomic failure that may be seen in MSA include fainting and heart rate problems, erectile dysfunction, and bladder control. Motor disturbances (loss or limitation of muscle control or movement, or limitation of mobility) may include tremors, stiffness, and/or loss of muscle coordination, as well as difficulties with speech and walking (the way a person walks). Some of these features are similar to those seen in Parkinson's disease, and early in the course of the disease it can often be difficult to distinguish these disorders. A hallmark of MSA is the accumulation of the protein alpha-synuclein in glia, the cells that support nerve cells in the brain. These alpha-synuclein deposits occur primarily in oligodendroglia, a type of cell that makes myelin (a covering of nerve cells that allows them to rapidly conduct electrical signals). This protein also accumulates in Parkinson's disease, but in nerve cells. Because both present a buildup of alpha-synuclein in cells, MSA and Parkinson's disease are sometimes called synucleinopathies.

"Lewy body disease" or "dementia with Lewy bodies (DLB)" is one of the most common types of progressive dementia. The main features of MCI are progressive cognitive decline, "fluctuations" in alertness and attention, visual hallucinations, and parkinsonian motor symptoms such as slowness of movement, difficulty walking, or rigidity. People can also suffer from depression. The symptoms of DLB are caused by the accumulation of Lewy bodies (clumps of alpha-synuclein protein) inside the nuclei of neurons in areas of the brain that control certain aspects of memory and motor control. Researchers don't know exactly why alpha-synuclein builds up in Lewy bodies or how Lewy bodies cause DLB symptoms, but they do know that alpha-synuclein buildup is also linked to Parkinson's disease. multiple system atrophy and several other disorders, which are called "synucleinopathies." The similarity of symptoms between DLB and Parkinson's disease, and between DLB and Alzheimer's disease, can often make a definitive diagnosis difficult for the doctor. In addition, Lewy bodies are also often found in the brains of Parkinson's and Alzheimer's patients. These findings suggest that either DLB is related to these other causes of dementia, or a person can have both diseases at the same time. DLB usually appears sporadically, in people with no known family history of the disease. However, rare familial cases have occasionally been described.

The term "primary lateral sclerosis (PLS)" refers to a rare neuromuscular disease with slowly progressive weakness in voluntary muscle movement. PLD belongs to a group of disorders known as motor neuron diseases. PLD affects the upper motor neurons (also called corticospinal neurons) in the arms, legs, and face. It occurs when nerve cells in the motor regions of the cerebral cortex (the thin layer of cells that cover the brain and are responsible for higher-level mental functions) gradually degenerate, causing movements to be slow and laborious. The disorder usually affects the legs first, followed by the body, trunk, arms and hands, and finally the bulbar muscles (muscles that control speaking, swallowing, and chewing). Symptoms include muscle weakness, stiffness and spasticity, clumsiness, slowness of movement, and problems with balance and speech. ELP is more common in men than in women, with a gradual and varied onset that usually occurs between 40 and 60 years of age. The PLA progresses gradually over several years, or even decades. Scientists do not believe that PLA has a simple hereditary cause. Diagnosis of PLD requires extensive testing to exclude other diseases. When symptoms begin, PLD can be confused with amyotrophic lateral sclerosis (ALS) or spastic paraplegia. Most neurologists follow the clinical course of an affected individual for at least 3 to 4 years before making a diagnosis of PLD.

The term "multiple sclerosis (MS)" refers to the most common disabling neurological disease among young adults, with symptoms usually appearing between the ages of 20 and 40. In MS, immune system cells that normally protect us from viruses, bacteria, and diseased cells mistakenly attack myelin in the central nervous system (brain, optic nerves, and spinal cord), a substance that makes up the protective sheath ( called myelin sheath) that covers nerve fibers (axons). MS is a chronic disease that affects people differently. MS attacks the myelin-protected axons of the central nervous system, often called white matter. MS also damages nerve cell bodies, which are found in the brain's gray matter, as well as the brain's own axons, spinal cord, and optic nerves that transmit visual information from the eye to the brain. As the disease progresses, the outermost layer of the brain, called the cerebral cortex, shrinks (known as cortical atrophy). The term multiple sclerosis refers to the distinctive areas of scar tissue (sclerosis, also called plaques or lesions) that result from the attack on myelin by the immune system. These plaques are visible using MRI in the white and/or gray matter of people with MS. The plates can be as small as the head of a pin or as large as a golf ball. During an MS exacerbation, most of the myelin, and to a lesser extent the axons within the affected area, is damaged or destroyed by different types of immune cells (also known as inflammation). The symptoms of MS depend on the severity of the inflammatory reaction, as well as the location and extent of the plaques, which appear mainly in the brain stem, the cerebellum (which is involved in balance and coordination of movement, among others). functions), the spinal cord, the optic nerves, and the white matter surrounding the brain ventricles (fluid-filled spaces).

As used here, "corticobasal degeneration" refers to a progressive neurological disorder characterized by nerve cell loss and atrophy. (shrinkage) of multiple areas of the brain, including the cerebral cortex and basal ganglia. Corticobasal degeneration progresses gradually. Initial symptoms, which usually begin at or around age 60, may first appear on one side of the body (unilateral), but eventually affect both sides as the disease progresses. Symptoms are similar to those of Parkinson's disease, such as poor coordination, akinesia (absence of movement), rigidity (resistance to imposed movement), imbalance (impaired balance), and limb dystonia (muscular posturing). abnormal). Other symptoms may also appear, such as cognitive and visual-spatial disturbances, apraxia (loss of the ability to perform familiar and purposeful movements), halting and halting speech, myoclonus (muscle jerks), and dysphagia (difficulty swallowing). A person with corticobasal degeneration ends up being unable to walk.

The term “vascular parkinsonism (PV),” also known as “multiinfarct,” refers to a hypokinetic movement disorder characterized by akinesia/bradykinesia, rest tremors, and extrapyramidal rigidity. "Vascular parkinsonism (PV)" is a form of atypical parkinsonism, in which parkinsonian features are of vascular origin, in contrast to typical Parkinson's disease (PD), of neurodegenerative etiology. It represents between 4.4% and 12% of all cases of parkinsonism. Typically, parkinsonian symptoms (slow movements, tremors, difficulty walking and maintaining balance, stiffness and stiffness) are caused by one or more small strokes, rather than by the gradual loss of nerve cells, as seen in the most typical neurodegenerative Parkinson's disease.

Alternatively, the invention relates to a method for treating and/or preventing Parkinson's disease and disorders associated with parkinsonism, which method comprises administering to a patient in need of said treatment a therapeutically effective amount of a specific agonist of the factor receptor. insulin-like growth-2 (IGF-2) or a pharmaceutically acceptable salt thereof.

The term "therapeutically effective amount" will be defined below when referring to pharmaceutical compositions.

Alternatively, the invention relates to the use of a specific insulin-like growth factor-2 (IGF-2) receptor agonist or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for the treatment of Parkinson's disease. and disorders associated with parkinsonism.

In a preferred embodiment, the specific IFG-2 receptor agonist is selected from the group consisting of IGF-2, Leu[27]IGF-2, M6PRBP1, (Arg6) IGF-2 and Des[1-6] Growth Factor insulin-like-2 (IGF-2).

As used herein, "insulin-like growth factor 2", also known as "IGF-2", "IGF-II" or "somatomedin", refers to insulin-like growth factor -2 from any species, including bovine, ovine, porcine, equine and human, preferably human, and, if it refers to exogenous administration, from any source, whether natural, synthetic or recombinant. It can be prepared by the method described in, for example, EP 128,733. In a preferred embodiment, the IGF-2 is human IGF-2. Mature human IGF-2, a 7.5 kDa peptide containing 67 amino acids, is originally synthesized as a biologically inactive pro-IGF-2 peptide (156 amino acids), which subsequently undergoes regulated endoproteolytic cleavage to the form mature. In addition to mature IGF-2, two "large" variants, IGF-2(1-87) and IGF-2(1-104), have been identified in human and bovine serum. They are 11-17 kDa in size, contain 87 or 104 amino acids, respectively, and differ in glycosylation.

In a particular embodiment, IGF-2 is the protein encoded by the human gene shown in the Ensembl database under accession number ENSG00000167244 (Ensembl version 102, November 2020). In another particular embodiment, IGF-2 is the protein of SEQ ID NO: 1, shown in the UniProt database under accession number P01344 (entry version 246, February 10, 2021, sequence version 1 ,July 21, 1986). In another particular embodiment, IGF-2 is the protein of SEQ ID NO: 2, which corresponds to the protein shown in the UniProt database with accession number P01344 (entry version 246, February 10, 2021, version sequence 1, July 21, 1986) without the first 24 amino acids of the signal peptide. In a preferred embodiment, IGF-2 is the protein of SEQ ID NO: 3, which corresponds to the mature 67 amino acid IGF-2 protein. In another embodiment, IGF-2 is the 87 amino acid protein of SEQ ID NO: 4 or the 104 amino acid protein of SEQ ID NO: 5.

Herein, the term "Leu[27]IGF-2" or "Leu27IGF-2" refers to an analog of human IGF-2 that has a Leu (L) substitution in place of Tyr (Y) in the position 27 (SEQ ID NO: 7) of the IGF-2 amino acid sequence and selectively interacts with the IGF-2R.

As used herein, the term "receptor binding protein Mannose-6-phosphate 1" or "M6PRBP1" refers to a protein that interacts with the cytoplasmic domains of cation-independent and cation-dependent MPRs, and is required for transport from the endosome to the Golgi. This protein also binds directly to GTPase RAB9 (RAB9A), a member of the RAS oncogene family. In a particular embodiment, M6PRBP1 is the protein encoded by the human gene that appears in the Ensembl database under accession number ENSG00000105355. Multiple transcript variants that encode different isoforms for this gene.

The term "(Arg6)IGF-2 or "[Arg6]Insulin-like Growth Factor-2 [R6]IGF-2" refers to an analogue of human IGF-2, which has an Arg substitution instead of Glu at position 6 of the IGF-2 amino acid sequence (SEQ ID NO: 8).

The term Des[1-6] Insulin-like Growth Factor-2 (IGF-2) refers to a 61-amino acid analog of human IGF-2 that lacks the N-terminal hexapeptide Ala-Tyr-Arg-Pro- Ser-Glu (SEQ ID NO: 6)

In a preferred embodiment, the specific IGF-2 receptor agonist for use according to the invention is IGF2 or a functionally equivalent variant thereof.

"Functionally equivalent variant" is understood as all those IGF-2 proteins derived from IGF-2, by modification, substitution, insertion and/or deletion of one or more amino acids, provided that the function is substantially maintained.

The activity of a functionally equivalent variant can be assessed by measuring the activation of the IGF-2 receptor caused by such variants. Useful assays for determining variant activity include, but are not limited to, those based on biological activity triggered by IGF-2 receptor activation, such as measuring Chinese hamster ovary (CHO) cell proliferation ( EC50 ≤ 150 ng/ml) or the stimulation of protein synthesis in rat myoblasts, (Mamounas M et al., 1989, PNAS, vol. 86, pp 9294-9298 and Bach LA et al., Endocrinology.

1995 Nov;136(11):5061-9), in combination with assays suitable for screening specific IGF-2 agonists, such as binding assays and immunohistochemistry, as mentioned above.

Preferably, "variants" of specific IGF-2R agonists are (i) those in which one or more amino acid residues are replaced by a conserved or non-conserved amino acid residue (preferably a conserved amino acid residue) and said amino acid substituted may or may not be encoded by the code genetic, (ii) those in which there are one or more modified amino acid residues, for example, residues modified by attachment of substituents, (iii) those resulting from the alternative processing of a similar mRNA, (iv) the fragments of specific agonists of the IGF-2R and/or (v) those resulting from the fusion of the specific IGF-2R agonist or the variants defined above in (i) to (iii) with another polypeptide, such as a secretory leader sequence, a sequence that is used for purification (e.g. His tag), for detection (e.g. Sv5 epitope tag) or for reaching a specific target cell. Fragments include polypeptides generated by proteolytic cleavage (including multisite proteolysis) of an original sequence. The functionally equivalent variant may be the result of a post-translational or chemical modification. Such variants will be evident to those skilled in the art.

Sequence variants can be both natural and artificial. The term "natural variant" refers to all those variants of the endogenous human specific IGF-2 receptor that appear naturally in other species, that is, the orthologs of IGF-2.

A functionally equivalent variant may be derived from specific IGF-2R agonists comprising the addition, substitution or modification of one or more amino acid residues. By way of illustration, functionally equivalent variants may include the amino acid sequence of the specific IGF-2R agonist comprising the addition of 1 amino acid, 2 amino acids, 3 amino acids, 4 amino acids, 5 amino acids, 10 amino acids, 15 amino acids, 20 amino acids , 25 amino acids, 30 amino acids, 35 amino acids, 40 amino acids, 45 amino acids, 50 amino acids, 60 amino acids, 70 amino acids, 80 amino acids, 90 amino acids, 100 amino acids, 150 amino acids, 200 amino acids, at least 500 amino acids, at least 1000 amino acids or more in the amino terminus of the amino acid sequence of specific IGF-2R agonists; and/or comprising the addition of 1 amino acid, 2 amino acids, 3 amino acids, 4 amino acids, 5 amino acids, 10 amino acids, 11 amino acids, 12 amino acids, 13 amino acids, 14 amino acids, 15 amino acids amino acids, 20 amino acids, 25 amino acids, 30 amino acids , 35 amino acids, 40 amino acids, 45 amino acids, 50 amino acids, 60 amino acids, 70 amino acids, 80 amino acids, 90 amino acids, 100 amino acids, 150 amino acids, 200 amino acids, at least 500 amino acids, at least 1000 amino acids or more at the carboxy end of the specific agonist of IGF-2R. In a particular embodiment, the equivalent functional variant of IGF-2 has at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity with IGF-2. In a more particular embodiment, the functionally equivalent variant of IGF-2 has at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91% , at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity sequence with the protein identified by UniProt accession number P01344.

The functionally equivalent variants of the invention are those that maintain at least 50%, at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% , at least 99%, or at least 100% of the activity of the IGF-2 protein, as defined above.

The terms "identity", "identical" or "percent identity" in the context of two or more amino acid sequences, refer to two or more sequences or subsequences that are the same or have a specified percentage of amino acid residues that are the same. , when compared and aligned (introducing gaps, if necessary) to obtain maximum correspondence, without considering any conservative amino acid substitution as part of the sequence identity. Percent identity can be measured using sequence comparison software or algorithms or by visual inspection. Various algorithms and computer programs are known that can be used to obtain alignments of amino acid or nucleotide sequences. A non-limiting example of a sequence alignment algorithm is that described in Karlin et al., 1990, Proc. Natl. Academic Sci., 87:2264-8, modified in Karlin et al., 1993, Proc. Academic Sci., 90:5873-7, and incorporated into the NBLAST and XBLAST programs (Altschul et al., 1991, Nucleic Acids Res., 25:3389-402). In certain embodiments, Gapped BLAST can be used as described in Altschul et al., 1997, Nucleic Acids Res. 25:3389-402. BLAST-2, WU-BLAST-2 (Altschul et al., 1996, Methods in Enzymology, 266:460-80), ALIGN, ALIGN-2 (Genentech, South San Francisco, California) or Megalign (DNASTAR) are other programs publicly available computer tools that can be used to align sequences. In certain embodiments, the percentage of identity between two nucleotide sequences is determined using the GAP program of the GCG software (for example, using a CMP NWSgapdna. matrix and a separation weight of 40, 50, 60, 70 or 90 and a length weight of 1.2, 3, 4, 5 or 6 ). In certain alternative embodiments, the GAP program of the GCG software package, which incorporates the algorithm of Needleman and Wunsch (J. Mol. Biol. 48:444-53 (1970)) can be used to determine the percentage of identity between two sequences of amino acids (for example, using a Blossum 62 matrix or a PAM250 matrix, and a separation weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2, 3, 4, 5 ). Alternatively, in certain embodiments, the percent identity between nucleotide or amino acid sequences is determined using the Myers and Miller algorithm (CABIOS, 4:11-7 (1989)). For example, percent identity can be determined using the ALIGN program (version 2.0) and using a PAM120 table with residuals, a gap length penalty of 12, and a gap penalty of 4. The appropriate parameters for maximum alignment using a Particular alignment software can be determined by a person skilled in the art. In certain embodiments, the default parameters of the alignment software are used. In certain embodiments, the percent identity "X" of a first amino acid sequence with respect to a second amino acid sequence is calculated as 100 x (Y/Z), where Y is the number of amino acid residues rated as identical match in the alignment of the first and second sequences (as aligned by visual inspection or a particular sequence alignment program) and Z is the total number of residues in the second sequence. If the second sequence is longer than the first, the percent identity can be determined only in the region of overlap between said first and second sequences. In this case, the same previous formula can be used, but using as the Z value the length of the region in which the first and second sequences overlap, said region having a length substantially equal to that of the first sequence.

In a particular embodiment, sequence identity is determined along the entire length of the IGF-2, or along the entire length of the variant, or both.

The term "conservative substitution" or "conserved amino acid residue", as used herein, refers to the substitution of an amino acid present in the native sequence by a naturally occurring or non-naturally occurring amino acid or by a peptidomimetic having properties similar sterics. When the side chain of the native amino acid to be substituted is polar or hydrophobic, the conservative substitution should be made with a natural amino acid, a non-natural amino acid or with a peptidomimetic moiety that is also polar or hydrophobic (in addition to having the same steric properties as the side chain of the substituted amino acid).

As natural amino acids are typically grouped according to their properties, conservative substitutions for natural amino acids can be easily determined taking into account the fact that, according to the invention, the substitution of charged amino acids for sterically similar uncharged amino acids are considered conservative substitutions.

To produce conservative substitutions for unnatural amino acids it is also possible to use amino acid analogues (synthetic amino acids) well known in the art.

When dealing with conservative substitutions, the substituent amino acid must have the same or a similar functional group on the side chain as the original amino acid.

The phrase "non-conservative substitutions" or "non-conserved amino acid residue", as used herein, refers to the substitution of the amino acid as present in the original sequence by another amino acid of natural or non-natural origin. natural, with different electrochemical and/or steric properties. Thus, the side chain of the substituted amino acid may be significantly larger (or smaller) than the side chain of the substituted native amino acid and/or may have functional groups with electronic properties significantly different from those of the substituted amino acid. Examples of non-conservative substitutions of this type include the substitution of phenylalanine or cycohexylmethylglycine for alanine, isoleucine for glycine, or -NH-CH[(-CH ₂ )5-COOH]-CO- for aspartic acid. Non-conservative substitutions that fall within the scope of the present invention are those that still constitute a peptide with anti-proliferative properties.

The specific IGF-2R agonists of the present invention may also comprise non-amino acid groups, such as, for example, hydrophobic groups (various linear, branched, cyclic, polycyclic or heterocyclic hydrocarbons and hydrocarbon derivatives) attached thereto; several protecting groups, which are attached to the terminals of the compound to reduce degradation. Suitable protecting functional groups are described in Green and Wuts, "Protecting Groups in Organic Synthesis", John Wiley and Sons, chapters 5 and 7, 1991.

Chemical groups (non-amino acids) can be included to improve various physiological properties, such as decreased degradation or clearance; the decrease in repulsion by various cellular pumps, the improvement of immunogenic activities, the improvement of various modes of administration (such as the binding of various sequences that allow penetration through various barriers, through the intestine, etc.); increasing specificity, increasing affinity, increasing stability, bioavailability, solubility, decreasing toxicity and the like.

Variants of the invention may encompass native peptides (whether degradation products, synthetically synthesized peptides or recombinant peptides), analogues, derivatives, salts, retro-inverse isomers, mimics, mimetics or peptidomimetics (typically synthetically synthesized peptides), as well as peptoids. and semipeptoids.

The terms "analogue", "derivative" and "mimetic" include molecules that mimic the chemical structure of a peptide structure and retain the functional properties of said structure. Approaches for designing peptide analogues, derivatives and mimetics are known in the art. For example, see Farmer, P. S. in Dmg Design (E. J. Ariens, ed.) Academic Press, New York, 1980, vol. 10, pp. 119-143; Ball, J. B. and Alewood, P. F. (1990) J. Mol. Recognition 3:55. Morgan, B. A. and Gainor, J. A. (1989) Ann. Rep Med Chem 24:243; and Freidinger, R. M. (1989) Trends Pharmacol. Sci.

10:270. See also Sawyer, T. K. (1995) Peptidomimetic Design and Chemical Approaches to Peptide Metabolism in Taylor, M. D. and Amidon, G. L. (eds.) Peptide-Based Drug Design: Controlling Transport and Metabolism, Chapter 17; Smith, A.B.

3rd, et al. (1995)J. Am Chem Soc 117: 11113-11123; Smith, A.B. 3rd, et al. (1994) J Am Chem Soc 116:9947-9962; and Hirschman, R., et al. (1993) J Am Chem Soc 115: 12550-12568.

Variants of the invention may also include derivatives. A "derivative" (e.g., a peptide or an amino acid) includes forms in which one or more reaction groups of the compound have been derived with a substituent group. Examples of derivatives include peptides in which an amino acid side chain, the peptide backbone, or the amino or carboxy terminus has been derived (e.g., peptide compounds with methylated amino bonds). An "analogue" of a compound X includes compounds that retain chemical structures necessary for functional activity, but that also contain certain chemical structures that differ.

A compound "mimetic" includes compounds in which the chemical structures of the compound necessary for functional activity have been replaced by other chemical structures that mimic the conformation of the compound. Examples of peptidomimetics include peptide compounds in which the peptide backbone is replaced by one or more benzodiazepine molecules (see, for example, James, GL et al. (1993) Science 260: 1937-1942) or oligomers that mimic the secondary structure of the peptide by using isoteric amide bonds and/or modifying the native peptide backbone, including chain extension or incorporation of heteroatoms; examples of which include azapeptides, oligocarbamates, oligoureas, beta-peptides, gamma-peptides, oligo(phenylene-ethynylene)s, vinyl sulfonopeptides, poly-N-substituted glycines (peptoids) and the like. Methods for preparing peptidomimetic compounds are well known in the art and are specified, for example, in Quantitative Drug Design, CA Ramsden Gd., Chapter 17.2, F. Choplin Pergamon Press (1992).

In another embodiment, the specific IGF-2 receptor agonist for use according to the invention is comprised in a pharmaceutical composition with a pharmaceutically acceptable carrier.

The term "pharmaceutical composition", as used herein, refers to a composition comprising a therapeutically effective amount of the agents according to the present invention and at least one pharmaceutically acceptable excipient. Pharmaceutical compositions that can be used in the medical uses of the invention can be prepared, for example, as injectables, as liquid solutions, suspensions and emulsions.

The term "therapeutically effective amount", as used herein in relation to the specific IGF-2R agonist for the medical uses of the invention, refers to the sufficient amount of the specific IGF-2R agonist to provide the desired effect. , that is, to achieve an appreciable prevention, cure, delay, reduction in severity or improvement of one or more symptoms derived from a disease, and will generally be determined, among other causes, by the characteristics of the agent itself and the therapeutic effect that it you want to achieve. It will also depend on the subject to be treated, the severity of the disease suffered by said subject, the pharmaceutical form chosen, etc. For this reason, the doses mentioned in this invention should be considered only as guides for the person skilled in the art, who must adjust the doses based on the mentioned variables. In one embodiment, the effective amount produces improvement of one or more symptoms of the disease being treated. treating.

Although individual needs vary, determination of optimal ranges for effective amounts of the compound of the invention is within the common experience of those skilled in the art. In general, the dose necessary to provide an effective amount of said compound, which can be adjusted by one skilled in the art, will vary depending on age, health, physical condition, sex, diet, weight, degree of receptor alteration, frequency of treatment and the nature and extent of impairment or disease, medical condition of the patient, route of administration, pharmacological considerations such as activity, efficacy, pharmacokinetic and toxicological profile of the compound particular drug used, whether a drug delivery system is used, and whether the compound is administered as part of a drug combination.

Those skilled in the art are familiar with the principles and procedures discussed in widely known and available sources such as Remington's Pharmaceutical Science (17th Ed., Mack Publishing Co., Easton, Pa., 1985) and Goodman and Gilman's The terms "pharmaceutically acceptable excipient " or "pharmaceutically acceptable carrier", refer to any compound or combination of compounds that is essentially non-toxic to the subject at the dose and concentration used, and is compatible with the other components of a pharmaceutical composition. Therefore, an excipient is an inactive substance formulated together with the active ingredient of a pharmaceutical composition, with the purpose of giving size and shape to the compositions containing said active ingredients. Shaping and sizing allows for convenient and precise dispensing of a pharmaceutical substance when producing a dosage form. Excipients may also serve various therapeutic enhancement purposes, such as facilitating the absorption or solubility of the compound (drug), or other pharmacokinetic considerations. Excipients may also be useful in the manufacturing process, to aid the handling of the active ingredient in question, for example by facilitating powder flowability or non-stick properties, as well as aiding in vitro stability, such as preventing denaturation during the expected useful life. The selection of suitable excipients depends on the route of administration and the pharmaceutical form, as well as the active ingredient and other factors. An excipient may be a non-toxic solid, semi-solid or liquid filler, a diluent, an encapsulation material or a formulation aid of any conventional type. Among illustrative examples, non-limiting excipients or carriers include water, saline solutions, alcohol, dextrose, vegetable oils, polyethylene glycols, gelatin, lactose, amylose, magnesium stearate, talc, surfactants, silicic acid, viscous paraffin, perfume oil, monoglycerides and diglycerides of fatty acids, esters of petroethral fatty acids, hydroxymethylcellulose, polyvinylpyrrolidone and the like.

In a particular embodiment, the specific IGF-2R agonist for use according to the invention or the pharmaceutical composition containing the specific IGF-2R agonist for use in the present invention is for parenteral administration. Thus, said specific IGF-2R agonist for use according to the invention or the pharmaceutical composition containing the specific IGF-2R agonist suitable for parenteral injection, include physiologically acceptable aqueous or non-aqueous solutions, dispersions, suspensions or emulsions, or may comprise Sterile powders for reconstitution into sterile injectable solutions or dispersions. Examples of suitable aqueous or non-aqueous excipients or carriers, diluents, solvents or vehicles include water, ethanol, polyols (propylene glycol, polyethylene glycol, glycerol and the like), suitable mixtures thereof, triglycerides, including vegetable oils such as olive oil, or injectable organic esters such as ethyl oleate. In a particular embodiment, the specific IGF-2R agonist for use according to the invention or the pharmaceutical composition containing the specific IGF-2R agonist is for use in the present invention for intravenous, intramuscular or subcutaneous administration. Typically, pharmaceutical compositions for intravenous, intramuscular or subcutaneous administration are solutions in sterile isotonic aqueous buffer. If necessary, the specific IGF-2R agonist for use according to the invention or the pharmaceutical composition containing the specific IGF-2R agonist for use in the present invention also includes a local anesthetic to improve any pain at the site of the injection. Typically, the ingredients are supplied separately or mixed in unit dosage form, for example as a freeze-dried dry powder or water-free concentrate in an airtight container such as a vial or sachet indicating the amount of active ingredient. Where the pharmaceutical composition is administered by infusion, it may be dispensed with an infusion bottle containing water or sterile pharmaceutical grade saline. Where the pharmaceutical composition is administered by injection, an ampoule of sterile water for injection or saline may be provided so that the ingredients can be mixed before administration.

Solid dosage forms for oral administration include conventional capsules, sustained release capsules, conventional tablets, sustained release tablets, chewable tablets, sublingual tablets, effervescent tablets, pills, suspensions, powders, granules and gels. In solid dosage forms, the active ingredient (the specific IGF-2R agonist) is mixed with at least one suitable excipient or carrier, such as sodium citrate or dicalcium phosphate or (a) fillers or extenders, such as starches. , lactose, sucrose, mannitol or silicic acid; (b) binders, such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose or acacia; (c) humectants, such as glycerol; (d) disintegrating agents, such as, for example, agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain complex silicates or sodium carbonate; (e) dissolution retarding agents, such as paraffin; (f) absorption accelerators, such as quaternary ammonium compounds; (g) wetting agents, such as, for example, cetyl alcohol or glycerol monostearate; (h) adsorbents, such as, for example, kaolin or bentonite; and/or (i) lubricants, such as, for example, talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, or mixtures thereof. In the case of capsules and tablets, dosage forms may also include buffering agents.

Solid formulations of similar type can also be used as fillers in soft or hard filled gelatin capsules using excipients such as lactose or milk sugar, as well as high molecular weight polyethylene glycols, and the like. Solid dosage forms, such as tablets, capsules and coated granules, can be prepared with coatings or envelopes, such as enteric coatings and others known in the art. They may also contain opacifying agents, and may be formulated to release the active ingredient(s) in a delayed manner. Examples of inlay formulations that can be used are polymeric substances and waxes. The active ingredients may also be in microencapsulated form, if appropriate, with one or more of the mentioned excipients.

Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups and elixirs containing suitable excipients or carriers used in the art. In addition to the active substance (i.e., the specific IGF-2 agonist or a functionally equivalent) the liquid dosage form may contain one or more excipients or carriers commonly used in the art, such as water or other solvents, solubilizing and emulsifying agents, such as, for example, ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, alcohol benzyl benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils, in particular cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil, sesame seed oil, Miglyol ®, glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols, sorbitan fatty acid esters, or mixtures of these substances, and the like. In addition to said inert diluents, the formulation may also include adjuvants, such as wetting agents, emulsifying and suspending agents, sweetening agents, flavoring agents and perfuming agents. The suspensions, in addition to the active ingredient(s), may contain suspending agents, such as, for example, ethoxylated isostearyl alcohols, polyoxylated sorbitol or sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar or tragacanth, or mixtures thereof. substances, and the like.

In another particular embodiment, the IGF-2R agonist specific for use according to the invention or the pharmaceutical composition containing the IGF-2R agonist specific for use in the present invention is for topical administration. For topical administration, said pharmaceutical composition can be formulated as a cream, gel, lotion, liquid, ointment, aerosol solution, dispersion, solid stick, emulsion, microemulsion and the like which can be formulated according to conventional methods using suitable excipients. , such as, for example, emulsifiers, surfactants, thickening agents, coloring agents and combinations of two or more thereof.

The specific IGF-2R agonist for use according to the invention or the pharmaceutical composition containing the specific IGF-2R agonist for use in the present invention may also be administered in the form of transdermal patches or iontophoresis devices. Thus, in a specific embodiment, a compound for use according to the invention is administered as a transdermal patch, for example, in the form of a sustained release transdermal patch. Suitable transdermal patches are described in more detail, for example, in US5262165, US5948433, US6010715 and US6071531.

Various drug delivery systems are known that can be used to administer the specific IGF-2R agonist for use according to the invention or the invention. pharmaceutical composition containing the specific IGF-2R agonist for use in the present invention, including, for example, encapsulation in liposomes, microbubbles, emulsions, microparticles, microcapsules or through other nanotechnological systems such as, for example, polymeric therapeutics and Similar. The required dose may be administered as a single unit or in a sustained release form.

The sustained release forms and the appropriate materials and methods for their preparation are described in the state of the art.

In a particular embodiment, the specific IGF-2R agonist for use according to the invention or the pharmaceutical composition containing the IGF-2R agonist for use in the present invention is administered orally.

The orally administrable form of the specific IGF-2R agonist specific for use according to the invention or the pharmaceutical composition containing the specific IGF-2R agonist for use according to the invention may be in a sustained release form which may further be comprise a coating or matrix. The sustained release coating or matrix includes, without limitation, natural, semi-synthetic or synthetic water-insoluble, modified polymers, waxes, fats, fatty alcohols, fatty acids, semi-synthetic or synthetic natural plasticizers, or a combination of two or more thereof. Enteric coatings can be applied by conventional processes known to those skilled in the art.

The specific IGF-2R agonist or the pharmaceutical compositions comprising it may be in any dosage form that is suitable for administration to the animal body, including the human body, especially in any form that is conventional for oral administration, for example in solid form as (additives/supplements for) food or feed, premixes for food or feed, fortified food or feed, tablets, pills, granules, dragees, capsules and effervescent formulations as powders and tablets, or in liquid form as solutions, emulsions or suspensions such as beverages, pastes and oily suspensions. Pastes can be filled into hard or soft shell capsules. Examples of other forms of application are transdermal, parenteral or injectable forms of administration. Dietary and pharmaceutical compositions may be presented in the form of controlled (delayed) release formulations.

Some examples of foods are dairy products, such as yogurt.

Beverages include both non-alcoholic and alcoholic beverages, as well as liquid preparations added to drinking water and liquid foods. Non-alcoholic beverages include, for example, soft drinks, sports drinks, fruit juices, lemonade, quasi-aquatic beverages (i.e. water-based drinks with a low calorie content), teas and milk-based drinks. Liquid foods are, for example, soups and dairy products (e.g. muesli drinks).

In a preferred embodiment, the specific IGF-2R agonist for use according to the invention is administered through fortified foods. Enriched foods are products to which vitamins, minerals or other substances with a nutritional or physiological effect are added.

The fortified food product of the present invention is suitable for human or animal consumption, and may also be a special purpose food, such as baby food, medical food, sports food, performance food or nutritional bars. In embodiments of the present invention, the fortified food product includes, but is not limited to, gelatin-type confections, preserved fruits and vegetables, dairy products such as milks, infant formulas, yogurts, cheeses, butter, margarine, shakes or ice creams. , by-products such as soy milk, soy yogurt or soy cheese, vegetable milks such as almond or rice milk, beverages, syrups, desserts such as flans, custards or other pasty or creamy foods, and confectionery formulations, such as chewing gum, chocolates, chewy candies, hard candies, boiled candies, caramel, dulce de leche, jellies, gummies, gelatin candies or hard and soft baking products. The specific IGF-2R agonist for use according to the invention or the pharmaceutical composition containing the specific IGF-2R agonist for use in the present invention may be added directly to the fortified food product, or may be supplied in separate sachets. The specific IGF-2R agonist for use according to the invention or the pharmaceutical composition containing the specific IGF-2R agonist for use in the present invention can be added directly to a gelatin-type formulation in ready-to-eat desserts or children's products. .

The fortified food product of the invention contains 0.001% to 10% by weight of the specific IGF-2R agonist, preferably 0.01 to 1%.

In a preferred embodiment, the fortified food is selected from the group consisting of natural milk, artificial milk and colostrum.

Compositions

In a second aspect, the invention relates to a composition comprising a specific agonist of the insulin-like growth factor-2 (IGF-2) receptor and a compound suitable for the treatment and/or prevention of the disease. Parkinson and disorders associated with parkinsonism.

The terms "specific IGF-2R agonist", "Parkinson's disease" and "parkinsonism-associated" have been defined in the context of the first aspect of the invention and apply equally to the present aspect of the invention.

The amount of the specific IGF-2R agonist present in the composition of the invention can vary within a wide range, however, in a particular embodiment, the percentage by weight of the specific IGF-2R agonist with respect to the total composition of the invention is at least 0.10% 0.50%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15% 20%, 25%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or, at least, 95%.

The proportion by weight of the specific IGF-2R agonist and the at least the second component of the composition in the composition of the invention can vary within a wide range, however, in general, the proportion is chosen, taking into account factors such as the condition being treated, the age, sex, weight, etc., of the subject receiving the inventive composition. Preferably, said weight ratio of the specific IGF-2R agonist and the second component of the invention is one that results in a beneficial effect, in particular, an increase in the therapeutic effect of the composition of the invention in relation to each of its components, so that the same result can be achieved with lower doses of each of the components, thus reducing the side effects in the subject receiving the composition of the invention. In one embodiment, the compound of the composition of the invention suitable for the treatment of a disease associated with the formation of amyloid deposits is selected from the group consisting of:

(i) a dopamine precursor and a DOPA decarboxylase inhibitor (DDCI),

(ii) a dopamine agonist,

(iii) a MAO-B inhibitor,

(iv) a catechol O-methyltransferase (COMT) inhibitor,

(v) an anticholinergic, and

(vi) a nicotinic antagonist and/or a non-competitive NMDA antagonist.

Dopamine precursors include, without limitation, levodopa and its prodrugs, such as melevodopa, the methyl ester of levodopa, which is disclosed in EP 252,290 B.

The term "DOPA decarboxylase inhibitor" refers to a drug that inhibits the peripheral metabolism of levodopa to dopamine by aromatic L-amino acid decarboxylase, such as carbidopa and benserazide, "Dopamine agonist", as used Herein, it refers to a compound that is capable of binding to one or more subgroups of dopamine receptors, resulting in a beneficial therapeutic effect in an individual treated with the agonist. The dopamine agonists described herein are typically agonists of at least the D2 subgroup of dopamine receptors, and may also be agonists of the D1 and/or D3 receptors. Dopamine agonists include, without limitation, apomorphine, lisuride, pergolide, bromocriptine, pramipexole, ropinerole, or rotigotine, or a combination of two or more of these dopamine agonists.

Monoamine oxidase inhibitors (MAOIs) are a class of drugs that inhibit the activity of one or both of the monoamine oxidase enzymes: monoamine oxidase A (MAO-A) and monoamine oxidase B (MAO-B). They are known as highly effective antidepressants, as well as effective therapeutic agents for panic disorder and social phobia. Non-limiting examples of MAOIs include, but are not limited to, non-selective inhibitors of MAO- N MAO-B, such as hydrazine (antidepressant), isocarboxazid (Marplan), hydracarbazine, phenelzine (Nardil), hydrazines, tranylcypromine (Parnate, Jatrosom), selective MAO-A inhibitors, biphemelane (Alnert, Celeport) (available in Japan), Moclobemide (Aurorix, Manerix), Pirlindole (Pyrazidol) (available in Russia), Toloxatone (Humoryl) (available in France) and selective MAO-B inhibitors such as Rasagiline (Azilect), Selegiline (Deprenyl, Eldepryl, Emsam, Zelapar), Safinamide (Xadago) or the reversible MAOIs Linezolid and Methylene Blue.

A catechol-O-methyltransferase (COMT) inhibitor is a drug that inhibits the enzyme catechol-O-methyltransferase. This enzyme methylates catecholamines such as dopamine, norepinephrine and epinephrine. It also methylates levodopa. Non-limiting examples of COMT inhibitors include, without limitation, entacapone (Comtan, Comtess, Stalevo), nebicapone, nitecapone, opicapone (Ongentys), and tolcapone (Tasmar).

Anticholinergics (anticholinergic agent) are a group of substances that block the action of the neurotransmitter called acetylcholine (ACh) at the synapses of the central and peripheral nervous system. Broadly, anticholinergics are divided into two categories according to their specific targets in the central and peripheral nervous system and at the neuromuscular junction: antimuscarinic agents and antinicotinic agents (ganglion blockers, neuromuscular blockers).

The term "anticholinergic" is typically used to refer to antimuscarinics that competitively inhibit the binding of ACh to muscarinic acetylcholine receptors; Such agents do not antagonize binding to nicotinic acetylcholine receptors at the neuromuscular junction, although the term is sometimes used to refer to agents that do so. Some examples of antimuscarinic agents are Antipsychotics (clozapine, quetiapine), Atropine, Benztropine, Biperiden, Chlorpheniramine, Paroxetine, Dicyclomine (dicycloverine), Dimenhydrinate, Diphenhydramine, Doxepin, Doxylamine, Glycopyrrolate, Glycopyrronium, Hyoscyamine, Ipratropium, Orphenadrine, Oxitropium, Oxybutynin, Promethazine, Propantheline bromide, Scopolamine, Solifenacin, Tolterodine, Tiotropium, Tricyclic antidepressants, Trihexyphenidyl, Tropicamide and Umeclidinium. Examples of antinicotinic agents include, without limitation, Bupropion, Dextromethorphan, Doxacurium, Hexamethonium, Mecamylamine and Tubocurarine.

Nicotinic antagonists are a type of anticholinergic drug that inhibits the action of acetylcholine (ACh) at nicotinic acetylcholine receptors. Examples of nicotinic antagonists include, without limitation, Hexamethonium, Mecamylamine, Trimethaphan, Atracurium, Doxacurium, Mivacurium, Pancuronium, Tubocurarine, Mecamylamine, Vecuronium, Succinylcholine, 18-Methoxycoronaridine, Dextromethorphan, Dextrorphan, 3-Methoxymorphinan and Botulinum Toxin.

In a preferred embodiment,

(i) the dopamine precursor is Levodopa and the DOPA decarboxylase inhibitor is Carvidopa;

(ii) the dopamine agonist is selected from pramipexole, ropinirole, rotigotine and apomorphine;

(iii) the MAO inhibitor is selected from selegiline, rasagiline and safinamide;

(iv) the catechol O-methyltransferase (COMT) inhibitor is selected from Entacapone and Tolcapone;

(v) the anticholinergic is selected from benztropine and trihexyphenidyl;

(vi) the nicotinic antagonist and/or a non-competitive NMDA antagonist is Amantadine.

In another aspect, the invention relates to the composition according to the invention for use in medicine.

Alternatively, the invention relates to the use of a composition according to the invention for the manufacture of a medicament.

In another aspect, the invention relates to the composition of the invention for use in the treatment of Parkinson's disease and disorders associated with parkinsonism.

Alternatively, the invention relates to a method of treatment and/or prevention of Parkinson's disease and disorders associated with parkinsonism, which method comprises administering a composition according to the invention to a patient in need of said treatment.

Furthermore, the invention relates to the use of a composition according to the invention for the manufacture of a medicament for the treatment of Parkinson's disease and disorders associated with parkinsonism.

In a particular embodiment, the disorder associated with parkinsonism is selected from young-onset Parkinson's disease, supranuclear palsy, multiple system atrophy, Lewy body disease or dementia with Lewy bodies and corticobasal degeneration.

Invention method

In a final aspect, the invention relates to an in vitro method for identifying a compound suitable for the treatment and/or prevention of Parkinson's disease and disorders associated with parkinsonism comprising

(a) contacting a candidate compound with a cell that expresses the IGF-2 receptor under conditions appropriate for binding of the compound and the IGF-2 receptor and

(b) determine the activity of the IGF-2 receptor,

wherein if the IGF-2 receptor activity after contacting the candidate compound is higher than a reference value, the candidate compound is suitable for the treatment and/or prevention of Parkinson's disease and disorders associated with parkinsonism.

The terms "Parkinson's disease", "parkinsonism-associated" and "IGF-2R" have already been described and apply equally to the present aspect of the invention.

The term "in vitro", as used herein, means that the method is not carried out in the body of a human or animal subject, but rather in cells or fluids isolated from said subject or in a test tube. The term "contact", as used herein, refers to the process by which the candidate compound comes into contact with the cell or cell population that expresses the IGF-2 receptor and includes any possible method that allows the compound candidate interacts with the cell population that expresses IGF-2R. Thus, when the candidate compound comes into contact with the cell or cell population that expresses the IGF-2R, the candidate compound binds to the IGF-2R, leading to its activation. The means of contact of the candidate compound with the cell or cell population that expresses the IGF-2R vary depending on the environment of the cell. For example, the candidate compound can be administered to a cell in culture by adding the candidate compound to the cell culture medium. In a particular embodiment, the candidate compound is administered to a cell culture or extracted tissue.

In a particular embodiment, the candidate compound is introduced into a sample having at least one cell, or population of cells, in a test tube, flask, tissue culture, plate or the like and incubated at a temperature and time sufficient to allow the interaction of the candidate compound with the IGF-2R receptor.

The term "determine" includes the evaluation and measurement of IGF-2R activation, for example, by evaluating any of the parameters tested in the examples of the present application, such as reduction of cell death, lipid peroxidation , protein oxidation, superoxide radical production or levels of the inducible mitochondrial enzyme SOD, mitochondrial damage, as well as restoration of diaphorase enzyme activity, glutathione S-transferase activity or expression of TH enzyme.

The expression "IGF-2 receptor activity after contact with IGF-2 is higher than a reference value" preferably refers to, but does not refer to necessarily, to an IGF-2R activity that is at least 10%, more preferably at least 50%, even more preferably at least 100%, and even more preferably at least 150% higher in the IGF-2R activity. 2R in control. Increased activity, for example, may be the result of increased basal IGF-2R activity, prolonged stimulation, delayed degradation, or overexpression, for example due to increased ligand binding. to promiscuous or inappropriate ligand binding, to constitutive activation of the receptor, to impaired recycling leading to increased signaling or delayed degradation.

In a particular embodiment, the activity of the IGF-2 receptor that is to be determined in the method of the invention is one of the activities that is triggered by the binding of IGF-2 to the IGF-2 receptor. A list of these activities has already been mentioned when describing specific IGF-2 receptor agonists. Non-limiting illustrative examples of assays for determining IGF-2R activity are assays capable of determining whether a compound that binds to IGF-2R mimics the effect of IGF-2 binding to the IGF-2R mentioned above.

The term "control" or "reference value", as used herein, refers to a predetermined criterion used as a reference to evaluate values or data obtained from test samples. The reference value or reference level may be an absolute value; a relative value; a value that has an upper or lower limit; a range of values; an average value; a medium value; an average value; or a value compared to a particular control or reference value. A reference value may be based on an individual sample value, such as a value obtained from a sample of the subject being tested, but at an earlier time. The reference value may be based on a large number of samples, or on a set of samples that include or exclude the sample to be analyzed. In one embodiment, the reference value corresponds to the IGF-2 receptor activity before contacting the cell or cell population with the candidate compound. In another embodiment, the reference value corresponds to the activity of the IGF-2 receptor in a cell or cell population that has not been contacted with the candidate compound.

The invention will be described by means of the following examples, which should be considered merely illustrative and not limiting the scope of the invention.

Examples

Two studies on the effect of IGF-2 treatment are described below. The first of them is on dopaminergic neurons in culture (SN4741 cells) and the second on a Parkinson's model in male mice (C57BL/6J).

1. - Characterization of the cellular model:

An experiment was designed to study the antioxidant and neuroprotective effects of IGF-2 through its specific receptor.

To develop this experiment, the SN4741 cell line of dopaminergic neurons of the substantia nigra was used (provided by Prof. Ernest Arena, Karolinska Institute, Stockholm). These cells express receptors for IGF-2, as demonstrated by the immunocytochemistry of the receptor shown in Figure 2. To inhibit the other receptors for which IGF-2 has less affinity, BS-536924 (TOCRIS) was used. which inhibits the tyrosine kinase effect of IGF-Ir and RIns receptors. Since there are no specific antagonists to inhibit the effects of IGF-2 mediated by its own receptor, an antibody against it (AF-292 R&D System) was used. The protective effects of IGF-2 were studied at the final time (8 h), and the mechanisms involved with REDOX homeostasis, as well as neuronal function at the short time (2.5 h) following the scheme shown in Figure 3. In some experiments, related to essential mechanisms for neuroprotection, the full Leu[27]IGF-2 agonist (Gropep) was included to demonstrate the neuroprotective efficacy of IGF-2r agonists.

2. - Characterization of the animal model:

Male mice of the C57BL/6J strain (10 weeks old, approximately 30 g) were used, distributed into the following 4 groups following the scheme shown in Figure 4:

- Control (n=8): intraperitoneal (ip) administration of vehicle (PBS) for 45 days.

- Control+IGF-2 (n=8): subcutaneous (sc) administration of IGF-2, 1.5^g/100g of body weight/day divided into two doses for 44 days.

- MPTP (n=8): ip administration of probenecid 25mg/l00g (ip) plus sc administration of MPTP 2.4mg/l00g, twice a week for 35 days.

- MPTP+IGF-2 (n=11): sc administration of IGF-2, 1.5^g/100g body weight/day divided in two doses plus sc administration of MPTP 2.4mg/l00g twice a week for 44 days.

On day 44, they underwent motor testing and on day 45 they were anesthetized and perfused.

3. - Parameters to study in the cellular model

Figure 5 shows the pathways of REDOX homeostasis with the evaluated parameters in bold. To evaluate the efficacy of the treatment in relation to cell survival, LDH was quantified and the percentage of cell death was analyzed after coincubation with MPP+ in the presence and absence of IGF-2; Oxidative damage was also evaluated by quantifying lipid hydroperoxides and oxidized proteins (Martín-Montanez et al., 2014), as well as the production of superoxide radicals by flow cytometry using the dihydroethidine probe and the activity of the antioxidant enzyme superoxide dismutase. (Martín-Montanez etal., Biochim Biophis Acta, 2014, 1842(7): 1041-51).

The study of mitochondrial function was carried out by measuring the mitochondrial membrane potential by flow cytometry with the JC1 probe and the activity of the cytochrome c oxidase enzyme of the electron transport chain by spectrophotometric methods using the COBAS MIRA autoanalyzer (Martín-Montanez etal., Redox Biol., 2017, 13: 69-81). Analysis of mitochondrial morphology was performed by electron microscopy.

The activities of the antioxidant enzymes GST and diaphorase induced by Nrf2 were also quantified by spectrophotometric methods using the COBAS MIRA autoanalyzer (Martín-Montañez et al., Free Rardic Biol Med.,2019, 137: 116-30).

Finally, neuronal function was evaluated by immunohistochemistry and confocal microscopy analysis of the expression of the enzyme thyroxine hydroxylase (TH) (Andrews et al., Eur J Neurosci., 2006, 24(1): 32-6).

4. - Parameters to study in the animal model

To evaluate the neuroprotective effect of IGF-2 on dopaminergic neurons of the substantia nigra pars compacta, TH expression was examined in mouse brains by immunohistochemistry and subsequent stereological analysis (Andrews et al., 2006, supra). To evaluate motor symptoms, the Rotarod test was used (De Guevara-Miranda et al., DDM Dis Model Mech., 2017, 10(3): 323-36). Studies were performed to quantify survival by analyzing survival curves.

5.- Results

The dopaminergic neurons of the substantia nigra (SN4741) express receptors for IGF-2, as demonstrated in the immunocytochemistry for the receptor shown in Figure 2, which makes them useful to study the parameters determined in the cellular model described above.

Incubation of SN4741 neurons with MPP+ at the doses used induced cellular damage that led to a statistically significant increase in neuronal death of 23% compared to controls (Figure 6). Coincubation of MPP+ in the presence of IGF-2 reduced the percentage of neuronal death to the same level as control neurons. The same results were obtained when these experiments were repeated in the presence of BMS (IGF-2 and insulin receptor inhibitor), indicating that the protective effect of IGF-2 is mediated by the interaction with its specific receptor IGF-2. 2r; This fact was confirmed when an antibody against the IGF-2r receptor was included in the incubation medium, which caused the cell death parameter to reach values very similar to those obtained in the group of neurons incubated with the neurotoxic MPP+ (figure 6). .

Regarding the antioxidant and neuroprotective effect of IGF-2 mediated by its interaction with the specific IGF-2r receptor, the results described below were obtained:

After incubation of neurons with MPP+, there was an increase in lipid peroxidation and protein oxidation compared to controls (Figure 7). In both parameters, coincubation of MPP+ in the presence of IGF-2 led to values similar to those obtained in the control group. Again, using the selective antibody for IGF-2r (AB), it was demonstrated that the effect obtained is due to the interaction of IGF-2 with its specific receptor (Figure 7).

Regarding superoxide radical production, an increase was observed in neurons incubated with MPP+ compared to controls. This increase returned to values equal to the controls when MPP+ was coincubated in the presence of IGF-2. Blocking the IGF-Ir and insulin receptor (BMS) did not modify the antioxidant effect of IGF-2, demonstrating that this behavior was the result of the interaction of IGF-2 with its specific receptor (Figure 8). As a consequence of the increase in superoxide radical, an increase in the inducible mitochondrial SOD enzyme was observed compared to control cells in the group treated with MPP+. Once again, this The increase returned to values equal to those of the controls when MPP+ was coincubated in the presence of IGF-2. This beneficial effect of IGF-2 was reversed by the inclusion of the antibody against IGF-2r in the coincubation medium (Figure 9).

Mitochondrial function was studied by measuring mitochondrial membrane potential, cytochrome c oxidase activity, and morphological studies using electron microscopy. In membrane potential, cytochrome c oxidase, and morphology studies, it was observed that mitochondrial damage induced by MPP+ was reversed by coincubation of MPP+ with IGF-2 (Figures 10-11). Using the same strategies as in the previous experiments (incubation in the presence of BMS or AB), it was demonstrated that the neuroprotective effect of IGF-2 was mediated by the interaction with its specific receptor IGF-2r.

When the activity of antioxidant enzymes dependent on the antioxidant transcription factor Nrf2 was checked, a 70% decrease in diaphorase activity was found in neurons exposed to the toxic MPP+. This decrease was reverted to values similar to the control with coincubation of the toxicant in the presence of IGF-2 (Figure 12). The activity of the glutathione-S-transferase enzyme was not altered by incubation with the toxicant, however, the addition of IGF-2 to the incubation medium produced an increase in the enzyme activity of 74% compared to the control. This effect was mediated by the specific IGF-2r receptor (Figure 12).

In neuroprotection experiments, measured as the expression of the enzyme that limits the synthesis of the neurotransmitter dopamine (tyrosine hydroxylase (TH)), it was found that the incubation of neurons with the toxic MPP+ produced a decrease in the expression of the enzyme dopamine. 63% compared to the control. Coincubation of the toxicant with IGF-2 increased the expression levels of TH above those obtained in the control group (35%). Again, this effect was mediated by the specific IGF-2r receptor, since blockade of IGF-Ir and insulin receptors with BMS did not modify the increase in TH expression (Figure 13).

When the IGF-2r agonist, Leu[27]IGF-2, was used in the cellular experiments, the results were very similar to those obtained with the natural IGF-2 agonist (Figure 14).

These data on the neuroprotective effect achieved in the cellular model were confirmed in the animal model. In mice, quantification from immunohistochemical experiments showed that histological sections from treated animals with the toxic MPTPp they express 70% fewer TH-positive cells in the substantia nigra pars compacta (figure 15). Again, the co-administration of the toxicant together with IGF-2 prevented cell damage, and values of TH-positive cells similar to those obtained in the animals in the control group were obtained. Regarding motor manifestations, animals treated with the toxicant showed much worse performance in the Rotarod test, with a decrease in latency time to fall of 28% compared to controls. Again, the joint administration of the toxicant with IGF-2 recovers the latency time to the fall to the levels of the control group. This improvement in motor symptoms was related to the number of TH positive cells in these animals (r=0.544, p<0.01) (figure 16).

Finally, regarding the mortality of experimental animals, a mortality rate of 33% was observed in the group treated with MPTPp, while there were no deaths in the group that was co-administered with IGF-II (p<0 .05) (figure 17).

Claims (16)

1. A specific agonist of the insulin-like growth factor 2 (IGF-2) receptor or a pharmaceutically acceptable salt thereof for use in the prevention and/or treatment of Parkinson's disease and disorders associated with parkinsonism. 2. The specific IGF-2 receptor agonist for use according to claim 1 characterized in that the agonist is selected from the group consisting of IGF-2, Leu[27]IGF-II and M6PRBP1. 3. The specific agonist of the IGF-2 receptor for use according to claim 2 characterized in that the agonist is IGF-2 or a functionally equivalent variant thereof. 4. The specific agonist of the IGF-2 receptor for use according to claim 2 characterized in that the agonist is Leu[27]IGF-II or a functionally equivalent variant thereof. 5. The specific agonist of the IGF-2 receptor for use according to any of claims 1 to 4 characterized in that the use is in the prevention and/or treatment of Parkinson's disease. 6. The specific IGF-2 receptor agonist for use according to any of claims 1 to 5 characterized in that the disorder associated with parkinsonism is selected from the group consisting of drug-induced parkinsonism, early-onset Parkinson's disease, supranuclear palsy, multiple system atrophy, Lewy body disease and dementia with Lewy bodies, primary lateral sclerosis, multiple sclerosis, corticobasal degeneration and vascular parkinsonism. 7. The specific IGF-2 receptor agonist for use according to any of claims 1 to 6 characterized in that the agonist is administered orally. 8. The specific IGF-2 receptor agonist for use according to claim 7 characterized in that the agonist is administered through fortified foods. 9. The specific IGF-2 receptor agonist for use according to claim 8 characterized in that the fortified food is selected from the group consisting of natural milk, artificial milk and colostrum. 10. The specific IGF-2 receptor agonist for use according to any of claims 1 to 6 characterized in that the agonist is comprised in a pharmaceutical composition with a pharmaceutically acceptable carrier. 11. A composition comprising a specific agonist of the insulin-like growth factor 2 (IGF-2) receptor and a compound suitable for the treatment and/or prevention of Parkinson's disease and disorders associated with parkinsonism, in where . The compound suitable for the treatment of a disease associated with the formation of amyloid deposits is selected from the group consisting of: (i) a dopamine precursor and a DOPA decarboxylase inhibitor (DDCI) (ii) a dopamine agonist, (iii) a MAO-B inhibitor, (iv) a catechol O-methyltransferase (COMT) inhibitor, (v) an anticholinergic, and (vi) a nicotinic antagonist and/or a non-competitive NMDA antagonist. 12. The composition according to claim 11 characterized in that: (i) the dopamine precursor is Levodopa and the DOPA decarboxylase inhibitor is Carvidopa; (ii) the dopamine agonist is selected from pramipexole, ropinirole, rotigotine and apomorphine; (iii) the MAO inhibitor is selected from selegiline, rasagiline and safinamide; (iv) the catechol O-methyltransferase (COMT) inhibitor is selected from Entacapone and Tolcapone; (v) the anticholinergic is selected from benztropine and trihexyphenidyl; (vi) the nicotinic antagonist and/or a non-competitive NMDA antagonist is Amantadine. 13. The composition according to any of claims 11 to 12 for use in medicine. 14. The composition according to any of claims 11 to 13 for use in the treatment of Parkinson's disease and disorders associated with parkinsonism. 15. The composition according to claim 14 characterized in that the disorder associated with parkinsonism is selected from the group consisting of early-onset Parkinson's disease, supranuclear palsy, atrophy multisystem disease, Lewy body disease or dementia with Lewy bodies, and corticobasal degeneration. 16. An in vitro method for identifying a compound suitable for the treatment and/or prevention of Parkinson's disease and disorders associated with parkinsonism comprising (a) contacting a candidate compound with a cell that expresses the IGF-2 receptor under conditions appropriate for binding of the compound and the IGF-2 receptor and (b) determine IGF-2 receptor activity; characterized in that if the IGF-2 receptor activity after contacting the candidate compound is higher than a reference value, the candidate compound is suitable for the treatment and/or prevention of Parkinson's disease and disorders associated with parkinsonism.