Active agent having the potential for use in the treatment of alzheimer's and certain neurodegenerative disorders

Abstract

The invention relates to the development of an active agent increasing the activity of α-carbonic anhydrase (hCA) enzymes, having the potential to be used in the treatment of Alzheimer's and certain neurodegenerative disorders.

Description

[0001] ACTIVE AGENT HAVING THE POTENTIAL FOR USE IN THE TREATMENT OF ALZHEIMER'S AND CERTAIN NEURODEGENERATIVE DISORDERS

[0002] Technical Field

[0003] The invention relates to the development of an active agent increasing the activity of a-carbonic anhydrase (hCA) enzymes, having the potential to be used in the treatment of Alzheimer's and certain neurodegenerative disorders.

[0004] Prior Art

[0005] Alzheimer's disease (AD) constitutes 75% of dementia cases and is a fundamental cause of dementia. With the aging of the population, it has started to become a disease of concern for all health authorities. Currently, the prevalence of AD is estimated to be 40.2 per thousand. According to the World Health Organization (WHO), it is predicted that more than 55 million people experience memory problems and that this figure will rise to 78 million by the end of the 2030s, and the number will rise to 139 million in the year 2050. AD is fundamentally characterized by neurodegenerative disorder and progresses with clinically cognitive and memory loss, progressive deterioration in daily activities, various neuropsychiatric symptoms, and behavioral disorders. It has been proven that the levels of different cerebral isoforms (mainly hCA I and II) of a-carbonic anhydrase (hCA) are significantly reduced in Alzheimer's patients.

[0006] The method used primarily in the treatment of Alzheimer's is drug therapy. With this treatment, memory loss, cognitive decline, and behavioral disorders caused by the disease can be alleviated. In particular, cholinergic drugs improve the communication between nerve cells by increasing acetylcholine levels in the brain. These drugs can help to support memory and thinking abilities in Alzheimer'spatients. Drugs such as Donepezil, Rivastigmine, and Galantamine are widely used in mild and moderate Alzheimer's disease. In advanced stages, the drug named memantine is used. Memantine protects brain cells by reducing the glutamate excess seen in Alzheimer's patients. These drugs contribute to the preservation of cognitive functions while alleviating the symptoms of the disease.

[0007] The cause of most Alzheimer's cases is largely unknown, except for 1-2% of cases in which deterministic genetic differences are detected. Several competing hypotheses attempt to explain the underlying cause; the most dominant hypothesis is the amyloid beta (AP) hypothesis.

[0008] The oldest hypothesis on which most drug therapies are based is the cholinergic hypothesis, which suggests that Alzheimer's disease is caused by the reduced synthesis of the neurotransmitter acetylcholine. The loss of cholinergic neurons seen in the limbic system and cerebral cortex is an important feature in the progression of Alzheimer's disease. In 1991, the amyloid hypothesis proposed that extracellular amyloid beta (AP) accumulations were the fundamental cause of the disease. Support for this assumption stems from the location of the amyloid precursor protein (APP) gene on chromosome 21 and from the fact that trisomy 21 (Down syndrome) patients, who have an extra gene copy, almost universally exhibit the earliest symptoms of Alzheimer's disease by the time they reach at least 40 years of age. APOE4, which is a specific isoform of apolipoprotein, is a major genetic risk factor for Alzheimer's disease. While apolipoproteins increase the breakdown of beta amyloid, some isoforms are not very effective in this task (such as APOE4), leading to excessive amyloid accumulation in the brain.

[0009] Intensive studies have been conducted for many years regarding the investigation of the underlying causes of Alzheimer's disease and the synthesis of drug molecules for the treatment of the disease. However, most of the drugs developed to date are molecules inhibiting the acetylcholinesterase (AChE) enzyme, and these molecules have various side effects. Therefore, there is a need for the synthesis of newmolecules targeting different enzymes and acting through different mechanisms in the treatment of various neurodegenerative disorders, particularly Alzheimer's.

[0010] When the studies included in the prior art are examined, a need has been felt for the development of an active agent increasing the activity of hCA enzymes, having the potential for use in the treatment of Alzheimer's and certain neurodegenerative disorders.

[0011] Object of the Invention

[0012] The object of this invention is to develop an active agent increasing the activity of hCA enzymes, having the potential for use in the treatment of Alzheimer's and certain neurodegenerative disorders.

[0013] Detailed Description of the Invention

[0014] A compound of Formula A having the potential for use in the treatment of Alzheimer's and certain neurodegenerative disorders, wherein

[0015]

[0016] (Formula A)

[0017] N: Nitrogen,Fe: Iron,

[0018] CH2: Methylene,

[0019] CH3or H3C: Methyl radical,

[0020] Cl: Chlorine,

[0021] P: Phosphorus,

[0022] O: Oxygen.

[0023] The IUPAC name of the molecule subject to the invention is stated below.

[0024] Mono-ferrocenyl-spiro(N / O)-trispiro(N / N)-cyclotetraphosphazene

[0025]

[0026] The first starting compound (1), mono-ferrocenyldiaminoalcohol, Fe(C5H5)2CH2N(CH2)3OH (1), was obtained from the reduction of this Schiff base with NaBH4in methanol medium, as a result of the interaction offerrocenecarboxaldehyde with 3 -amino- 1 -propanol in methanol medium. The sodium salt (li) was obtained in the interaction of the N / O donor monoferrocenyldiaminoalcohol compound with NaH in tetrahydrofuran (THF) medium.

[0027] + NH NHMe MeOH

[0028] MeOH + NH NHMe NaBH4

[0029]

[0030] The bidentate diamine compound (2) used as a side group was obtained from the reduction of the Schiff base, formed as a result of the interaction of piperonal with 7V-methyl-l,3-propanediamine, with NaBH4in methanol medium.*yTHF

[0031]

[0032] Subsequently, 2,4-ansa- and spiro- mono-ferrocenyl-cyclotetraphosphazene compounds (3 and 4) were synthesized from the reaction of the sodium salt (li) of 3 -(A-ferrocenylmethylamino)-l -propanol (1) with tetramer in a 1:1 stoichiometric ratio in the presence of Et3N in THF solvent and under argon atmosphere at -10°C. The solution was allowed to stir for 96 hours, filtered, and its solvent (THF) was removed in a rotary evaporator. Afterwards, the reaction solution was filtered, the solvent was removed from the medium by evaporating to dryness in a rotary evaporator, and the obtained mixture of mono-ferrocenyl derivatives in solid state (ansa and spiro mixture) was dissolved with toluene and separated into its components with the aid of a silica gel filled column prepared again with toluene. Subsequently, the product mixture obtained with the column was dissolved in n-hexane(heptane): ethyl acetate (5:1) solvent mixture and applied to a silica gel filled column prepared with the solvent mixture in the same ratio, and mono-ferrocenyl-spiro- (4) and 2,4-ansa- (3) cyclotetraphosphazene products were isolated in pure state, respectively. Crystallization with hexane was performed for the yellow colored solid compounds.

[0033] The target piperonalyl mono-ferrocenyl-tetraspiro cyclotetraphosphazene (6) compound was synthesized as a result of the reactions of the synthesized partially substituted spiro- (4) compound with an excess amount of the N / N donor mono-piperonalyldiamine compound (2) in the presence of triethylamine in dry THF medium under reflux under argon atmosphere. The reaction mixture was stirred under reflux for four days. The reaction solution was filtered, and its solvent (THF) was removed in a rotary evaporator. Product (6) was separated in pure state as a result of column chromatography performed with toluene: THF (1:5). The compound separated first from the column is the fully substituted tetraspiro (6) cyclotetraphosphazene compound. The yellow colored solid was crystallized with acetonitrile.

[0034] The CA activator properties of the compounds were investigated under in vitro conditions based on the principle that CA enzymes exhibit esterase activity.

[0035] Esterase activity was determined by measuring the absorbance change occurring during the hydrolysis of the / ?-nitrophenyl acetate compound used as substrate to the / ?-nitrophenol compound under the catalysis of CA isoenzymes (hCA I and hCA II) at 348 nm (SHIMADZU, Japan). Measurements were performed at 25°C temperature for a period of 3 minutes. The reaction medium was prepared from a mixture of 1300 pL of 50 mM Tris (pH 7.4), 1000 pL of 3 mM / ?-nitrophenyl acetate, 600 pL of pure water, and 100 pL of hCA enzyme so as to have a total volume of 3 mL. The amount of / ?-nitrophenol formed under the catalysis of hCA enzymes was determined by taking the difference between the absorbance value at the end of the three-minute period and the initial absorbance value. The samemeasurement was also repeated in an enzyme-free medium, the amount of p-nitrophenol formed as a result of the autohydrolysis of / ?-nitrophenyl acetate was determined, and correction was made for the enzyme-catalyzed reaction.

[0036] To determine the effects of the phosphazene derivative on hCA I and hCA II, 0.1% stock solution of the compound were prepared in DMSO. (DMSO does not exhibit inhibitory or activator effect on CA enzymes.) hCA I and hCA II esterase activities were determined in the presence of the phosphazene derivative. Similarly, the amount of / ?-nitrophenol formed was also determined.

[0037] For the determination of KA constants, three different activator concentrations increasing hCA activity by 30%, 50%, and 70% were determined. For each activator concentration, esterase activity was measured at five different p-nitrophenyl acetate concentrations (0.6 mM, 0.8 mM, 1.0 mM, 1.2 mM, and 1.4 mM). The same measurements were also performed in CA catalyzed medium containing no activator. The data were linearized using the Microsoft Excel package program, and K constants were determined using the following equation:

[0038] V = Vmax / (1 +(Km / [S]) (1 + [A] / KA))

[0039] Furthermore, the degradation products of the compound in the body are phosphate, ammonia, and organic acids, which are products that are non-harmful and can be easily excreted. It has been determined that these effects are an important feature that a drug must possess in order to be used for therapeutic purposes.

Claims

CLAIMS1. A compound of Formula A increasing the activity of a-carbonic anhydrase (hCA) enzymes, wherein(Formula A)N: Nitrogen,Fe: Iron,CH2: Methylene,CH3or H3C: Methyl radical,Cl: Chlorine,P: Phosphorus,O: Oxygen.

2. The compound according to claim 1 for use in the treatment of Alzheimer's.

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