KR20110049801A - Methods and Systems for the Evaluation of Memory Substances - Google Patents

Abstract

The present invention provides methods and systems for identifying, evaluating and testing various compounds as memory materials and for identifying, evaluating and testing regimens with training protocols associated with memory enhancement and / or damage. Such methods and systems can test any compound or protocol, including the memory materials and training protocols described herein. Methods may include a combination of training protocols and overall administration of memory materials. In certain embodiments, the invention can include identifying, screening, testing, evaluating or estimating a compound as a drug candidate for a memory material that is effective to enhance or impair memory.

Description

METHODS AND SYSTEMS FOR EVALUATING MEMORY AGENTS

Cross-References to Related Documents

This application claims 35 U.S. C. Under U.S. Provisional patent application, U.S.S.N. 61 / 081,845 (filed July 18, 2008) claims priority, the entirety of which is incorporated herein by reference.

Technical field

The present invention relates to methods and systems for evaluating, identifying or evaluating a memory agent and / or for training a protocol administered to an individual.

Memory is a collection of past experiences. You can register new information, store it, and in some cases recall the stored information later. The collection of facts and events represents a form of self-conscious or obvious memory. You can also remember some less conscious techniques, for example, riding a bike, playing a musical instrument, or associating a simple cue with rewards or corporal punishment, which become more implicit forms of memory. In both cases, the memory formation process seems to go through three general steps. Acquisition relates to the initial awareness of a new experience. Then the short-term memory of these newly acquired experiences seems to be temporary and unstable. Under appropriate conditions, often requiring repetition and rest, short-term experiences can be “consolidated” into more persistent long-term memory.

A routine example of such a storage process involves the use of telephone numbers. When you first find a new number, recite a few times until you learn the number. Then, if the mind is not distracted, the number is successfully dialed in seconds or minutes. Meanwhile, almost everyone forgets that number. Within a few minutes to a few hours after dialing a phone number, you will no longer be able to remember it (short-term memory). However, if the phone number is a friend's new phone number, you will dial the number repeatedly every day. After sufficient repetition, the telephone number can be recalled through long-term memory.

The human brain is evolutionarily designed to change the brain's response to feel the internal and external environment of the brain, to arouse causal relations through stimuli, and to adapt. To accomplish this remarkable work, a self-regulating network of 40,000 genes is involved in the development of a cellular network of 100 trillion connections between 100 billion neurons. This complexity is overwhelming. Nevertheless, neurologists began to understand how the human brain works in part by studying the brain's function in other animals.

The new experience will obviously be quite complex, but if the scale is reduced to a simple, controllable study of “association” between two identified stimuli (eg Pavlovian learning) in the laboratory, the memory formation process can range from insects to mammals. Up to several different animals can be studied. These animal studies contributed to our current understanding of how memory formation progresses and to the neuroanatomical and biochemical aspects of the brain. Notably, these studies have established that most of the basic biochemistry and behavioral phenomenology of memory formation has been evolutionarily preserved throughout the animal world, although it is neurologically detectable between species.

Animal studies have shown that memory formation generally progresses through several functionally distinct transient phases. Acquisition (LRN) usually requires repetition, and is considered to recruit the maximum participation of the underlying neural circuits that normally register this experience. Once learning occurs, early memory is rapidly degenerated, and sensitive to disruption by distraction and anesthesia (eg, anesthesia-sensitive memory, ASM). In genetic dissection, it has been shown that ASM can be further broken down into short-term memory (STM) and medium-term memory (ITM).

In contrast, long-lasting memory is resistant to disruption (eg, anesthesia-resistant memory (ARM)), and lasts for a long time. Importantly, proteins in the appearance of long-term memory (LTM) in animal studies Overall, it was found that synthesis is required: Again, in fruit fly studies, (i) LTM requires protein synthesis (ARM is not required), and (ii) ARM and LTM are genetically independent of memory. It has been shown that it can be dissected into stages.

Animal studies have also shown that during the development a basic wiring diagram of the brain emerges. However, most connections (synapses) in neurons do not function as they are. In order to refine synaptic connections, fine tuning the functions of the various circuits requires continuous experience. Activity-dependent synaptic reorganization persists in the adult brain. Learning of new experiences initiates neuronal activity and ultimately synaptic reorganization occurs among neurons in related circuits. This synaptic change (synaptic plascitiy), distributed throughout the neuronal circuit, appears to be the physical basis of long-term memory.

Experience-dependent changes in neuronal activity initiate complex biochemical responses in neurons. The ultimate effect of this biochemistry is to create short-term (transient) and long-term (long-lasting) synaptic plasticity. Basic research continues to identify more signaling pathways and other cell biology that underlie this synaptic plasticity. The first of the findings is the cAMP pathway, a cascade of enzymatic reactions that transform intracellular changes into the functional state of many proteins by extracellular neurotransmitter release. Neurotransmitters, their corresponding receptors, and cytoplasmic PKA appear to regulate short-term plasticity, and activation of the transcription factor CREB (cAMP response element binding protein) is required to make long-term plasticity.

In all species studied so far, the CREB gene is expressed as a downstream activator or downstream inhibitor of two opposite type-CRE-mediated gene transcription. An important role of CREB in the formation of long-term memory was originally established in major behavioral experiments in Drosophila . Normal fruit flies can learn the relationship between specific odors and foot shock abuse. After one training session, most flies will avoid existing shock-related odors if they have to choose between a conventional shock-related odor and a neutral odor in the T-maze. This learning is quite strong at first, but then the memory is routine and disappears in one day. In contrast, receiving 10 training sessions with a 15-minute rest interval will result in bonafide protein synthesis-dependent LTM that lasts more than a week in normal flies. In flies genetically engineered to produce high levels of CREB inhibitors, LTM is clearly blocked without effect on learning or early memory. In flies genetically engineered to produce high levels of CREB activators, LTM was not affected after spaced training, but surprisingly, LTM was induced after only one training session. In sum, overexpression of CREB activators enhances memory by reducing the number of training sessions required to induce LTM formation—functionally equivalent to image (photo) memory. Molecular studies have begun in several animal models to further elucidate plastic biochemistry, link various aspects of synaptic plasticity and memory formation, and in general, such as brain plasticity: changes in neuronal activity, and new experiences are registered in specific neural circuits. This leads to a perspective: this neuronal activity induces the complex intercellular and intracellular biochemical responses that are still being discovered. However, to date some key aspects of this process appear to be embryologically conserved. Stimulation of NMDA receptors appears to be central to inducing both local and transient biochemical responses at synapses and long-lasting biochemical responses (phosphorylation of CREB) in the cell nucleus. Several intracellular signaling pathways appear to be initially induced by neuronal activity, but nevertheless, CREB appears to be the main "downstream" target that regulates the conversion of early memory to persistent memory. Down-regulation inhibits the gene transcription response that is the source of synaptic growth processes that create persistent functional and structural plasticity at synapses, but up-regulation of CREB enhances it. It appears to act as a central switch that determines when neuronal activity in a circuit is permanent and causes structural changes, the main factor affecting this switch is repetition: when the CREB pathway is up-regulated, it is required for the formation of long-term memory The number of training sessions taken is reduced.

Four basic objectives have led to methods of screening drugs that modulate the "CREB pathway". First, in terms of effectiveness, the screen should be high throughput capable of robot control in 96-well plate format. Second, since any identified drug is ultimately used in humans, the preferred target was the endogenous (neuronal) type of human CREB. Third, when designing the screen, all relevant upstream signaling pathways that may affect CREB function during memory formation have not yet been identified. Thus, the extensive screen for monitoring CREB-dependent gene expression with functional translation of the CREB pathway appears to further include any invisible, but potentially important, biochemical signaling. Finally, the screen needed to identify drugs that did not only affect CREB function, rather than synergized with co-activation of cAMP signaling. This requirement is designed to mimic the inherent behavioral experiments. In Paris, overexpression of CREB alone did not affect fly behavior. However, training itself was required to create enhanced memory because experience-induced increases in neuronal activity activated the cAMP pathway.

These considerations were incorporated into the high throughput screen (HTS) of cultured human neuroblastoma cells stably transfected with the CRE-luciferase reporter gene. In this way, changes in CREB-dependent transcription of luciferase can be monitored via fluorescence. In order to mimic the associative requirement for memory formation (at the biochemical level), there is no change in luciferase activity on its own, but quasi-activation of forskolin to activate the cAMP pathway (as neural activity does in vivo). Drugs were found to be synergistic (> 2 fold) by the optimal dose. Using a robotic system to screen these cells in 96-well plates, a large library of drug compounds was effectively screened, creating a number of “active hits”. To confirm, these active hits were evaluated at four different concentrations with forskolin present. Several identified activities emerged from this screening process.

Interestingly, it was found that the effective molecular target is phosphodiesterase-4 (PDE4). In general and specifically, this finding makes sense. Most biochemical pathways work in well-regulated optimal conditions, with several molecular components (positive modulators) that accelerate signal delivery, and other components (negative modulators) that delay signal delivery. Given the opposite function of CREB activators and inhibitor isoforms, this concept applies to the regulation of CREB switches during long-term memory formation. Similar to genetic mutations, most drugs actually act as inhibitors of target function (“antagonists” for drugs and “hypomorphs” for mutations). Thus, it may be expected to often find drug compounds that antagonize negative regulators of the pathway from screens for enhanced function. Of course, this is the case for PDE inhibitors; Reduction of PDE activity leads to an increase in cAMP normally synthesized by adenylylcyclase (AC), and elevated cAMP induces activation of CREB through PKA. Moreover, PDE inhibitors may be expected to have only minimal effects in the absence of forskolin-induced activity of AC (or neuroactive-induced activation of AC in vivo).

The advanced PDE-4 inhibitor, HT-0712, has proven to be a clinically proven compound with several other CREB pathway enhancers in addition to HT-0712. To date, behavioral assessments in rodent models and biochemical experiments of memory formation have shown that these compounds effectively enhance memory.

It is important to develop, identify and evaluate such compounds and therapies. Such therapy may involve the administration of a compound comprising compounds that act as memory agents, as well as memory enhancers that specifically upregulate specific training protocols and / or CREB pathways.

SUMMARY OF THE INVENTION

The present invention includes methods comprising evaluating the effect of a memory administered to a subject, wherein the assessment comprises providing the subject with a series of stimuli and evaluating the subject's response to the stimulus. The assessment includes a first period of time in which the subject is provided with an associated stimulus and the subject's response is assessed to establish a criterion; And a second period comprising providing the subject with an associated stimulus and evaluating the subject's response to make a long term successful measurement. In some embodiments, the second period of time begins at least about one day after the end of the first period of administration. Evaluating a subject's response can include comparing criteria for long-term success measures. A set of stimuli can include multiple stimulus pairs. In some embodiments, each of the plurality of stimulation pairs may comprise a positive element and / or a negative element. In other embodiments, each of the plurality of stimulus pairs may comprise an element having an identifiable association with other elements of that pair. In some embodiments, such identifiable associations may be face-name associations or word-word associations. The subject can be an animal. In certain embodiments, the animal can be a primate, mammal, mouse or rat. In certain embodiments, the primate can be a human, monkey, lemur, macaque or tailless monkey. In certain embodiments, the first time period can be from about 1 day to about 10 days. In certain embodiments, the second period of time can be about one day. In certain embodiments, there may be a number of individuals that may include a control group and an experimental group. In certain embodiments, the evaluation may comprise comparing the control to the experimental group. In certain embodiments, assessing a subject's response includes calculating and / or measuring one or more aspects selected from the group consisting of: a ratio for long term success measures to criteria, long term memory retention , The amount of time required to obtain a particular criterion, and / or the amount of time between the first and second periods while maintaining a constant, near-constant or improved rate of long-term success measure for the criterion.

In another embodiment, the invention may relate to a method comprising selecting a memory material as a drug candidate, wherein the screening method comprises administering the memory material to an individual; And providing a set of stimuli to the individual, and evaluating the response of the individual's response to the stimulus. The assessment may further include: providing the subject with a set of related stimuli, and evaluating the subject's response to the stimulus to establish criteria, and the second period of time To provide a set of relevant stimuli, and to assess the individual's response to the stimulus to establish long-term success measures. In certain embodiments, the second period begins at least about 1 day after completion of the first period. In certain embodiments, assessing the individual's response includes calculating and / or measuring one or more aspects selected from the group consisting of: a ratio for long term success measures to long term, long term memory retention, specific criteria The amount of time required to obtain and / or the amount of time between the first and second periods while maintaining a constant, near-constant or improved rate of long-term success measure for the criterion. In certain embodiments, a set of stimuli can include multiple stimulus pairs. In certain embodiments, the subject may be an animal, including a mammal, including primates, mice or mice.

In another embodiment, the present invention relates to a method comprising the following steps: testing a memory material with a long term memory enhancer, wherein the test comprises: administering the memory material to the individual; It provides the individual with a set of stimuli and assesses the individual's response to the stimulus. In certain embodiments, the evaluation comprises a first period of time comprising providing the subject with an associated stimulus and evaluating the subject's response to establish a criterion; And a second time period comprising providing the subject with an associated stimulus and evaluating the subject's response to make a long term successful measurement. In certain embodiments, the second period begins at least about 1 day after completion of the first period. In certain embodiments, assessing a subject's response includes calculating and / or measuring one or more aspects selected from the group consisting of: a ratio for long term success measures to long term, long term memory retention, specific criteria The amount of time required to obtain and / or the amount of time between the first and second periods while maintaining a constant, near-constant or improved rate of long-term success measure for the criterion. In certain embodiments, a set of stimuli may comprise multiple stimulus pairs. In certain embodiments, the subject may be an animal comprising an animal selected from the group consisting of primates, mammals, mice or mice.

In a further embodiment, the invention may be a method comprising evaluating the effect of a memory material, wherein the evaluating comprises providing a first subject and a second subject, wherein the first subject and the second subject are the same species ; Administering the memory material to the first individual; Providing the first individual and the second individual with a plurality of pairs of stimuli during the first period of time, and establishing the criteria for each of the first and second individuals, and the response of the first and second individuals to the stimulus. Evaluating; Providing a plurality of pairs of stimuli to a first individual and a second individual during a second period of time, and to establish long-term success measures for each of the first and second individual, Evaluating the response; Comparing the first and second individuals to their respective long term success measures. In certain embodiments, the second period begins at least about 1 day after completion of the first period. In certain embodiments, the second period begins between about 1 day and about 10 days after completion of the first period. In certain embodiments, the comparison of the criteria for long term success measure comprises: establishing a ratio of the long term success measure of the first individual to the first individual criterion; Establishing a ratio of the long term success measure of the second individual to the criteria of the second individual; Then, the first ratio and the second ratio are compared. In certain embodiments, a set of stimuli may comprise multiple stimulus pairs. In certain embodiments, the plurality of stimulus pairs comprise a positive element and / or a negative element. In other embodiments, each of the plurality of stimulus pairs includes an element having an identifiable association with other elements of that pair. In some embodiments, such identifiable associations may be face-name associations or word-word associations. In some embodiments, each of the first and second individuals can be an animal, including an animal selected from the group consisting of primates (humans, monkeys, lemurs, macaques or tailless monkeys), mammals, mice, mice. In certain embodiments, the first period of time can be from about 1 day to about 10 days. In certain embodiments, the second period of time can be about one day. In certain embodiments, each of the first and second individuals is a plurality of individuals.

In some embodiments, the invention is similar to that mentioned above, but includes methods that focus on the evaluation of training protocols rather than memory materials. Some embodiments relate to a method comprising evaluating the effectiveness of a training protocol administered to a subject, wherein the evaluating includes providing the subject with a set of stimuli and assessing the subject's response to the stimulus. Other embodiments relate to a method of testing a training protocol as a long term memory enhancer, wherein the test comprises administering the training protocol to a subject; Providing a set of stimuli to the individual, and evaluating the subject's response to the stimulus. A further embodiment relates to a method comprising evaluating the effectiveness of a training protocol, wherein the evaluating comprises providing a first subject and a second subject, wherein the first subject and the second subject are the same species; Administering a training protocol to the first individual; Providing the first individual and the second individual with a plurality of pairs of stimuli during the first period of time, and evaluating their response to the stimulus to establish criteria for each of the first individual and the second individual; Providing the first individual and the second individual with a plurality of pairs of stimuli during the second time period, and evaluating their response to the stimulus to establish long-term success measures for each of the first individual and the second individual ; Comparing the criteria of the first individual and the second individual for each long term success measure.

The invention also includes a system using the method described herein. For example, in some embodiments, the system may include all the elements needed to achieve an embodiment of the methods of the present invention. The system of the present invention selects memory as a drug candidate, evaluates the effect of the memory, evaluates the effectiveness of the training protocol, and / or tests the memory as a long term memory enhancer and / or long-term memory As an adjuvant, it will be useful for testing memory with training protocols.

In some embodiments, the system of the present invention includes multiple sets of stimuli, wherein the multiple sets of stimuli are set to assess the individual's response to these stimuli, such that the effect of the training protocol administered to the individual Evaluate. In certain embodiments, multiple sets of stimuli comprise a plurality of stimulus pairs. In certain embodiments, each of the plurality of stimulation pairs comprises a positive element and / or a negative element. In some embodiments, each of the plurality of stimulus pairs includes an element having an identifiable association with other elements of that pair. In further embodiments, such identifiable associations may be face-name associations or word-word associations. In other embodiments, the system may further comprise testing the protocol, testing the schedule and / or testing the dosage of the memory substance and the schedule of administration.

In another embodiment, the system of the present invention includes multiple sets of stimuli, wherein the multiple sets of stimuli are set to evaluate the individual's response to these stimuli, thereby determining the effect of the memory material administered to the individual. Evaluate. In certain embodiments, multiple sets of stimuli comprise a plurality of stimulus pairs. In certain embodiments, each of the plurality of stimulation pairs comprises a positive element and / or a negative element. In some embodiments, each of the plurality of stimulus pairs includes an element having an identifiable association with other elements of that pair. In further embodiments, such identifiable associations may be face-name associations or word-word associations. In other embodiments, the system may further comprise testing the protocol, testing the schedule and / or testing the dosage of the memory substance and the schedule of administration.

In certain embodiments, the methods and systems of the present invention utilize one or more memory materials selected from the group consisting of a compound having the following structure or a pharmacologically acceptable salt thereof:

Wherein each Y ₁ , Y ₂ , Y ₃ , and Y ₄ are independently H; Straight or branched C ₁ -C ₇ alkyl, monofluoroalkyl or polyfluoroalkyl; Straight or branched C ₂ -C ₇ alkenyl or alkynyl; C ₃ -C ₇ cycloalkyl or C ₅ -C ₇ cycloalkenyl; -F, -Cl, -Br, or -I; -NO ₂ ; -N ₃ ; -CN; -OR ₄ , -SR ₄ , -OCOR ₄ , -COR ₄ , -NCOR ₄ , -N (R ₄ ) ₂ , -CON (R ₄ ) ₂ , or -COOR ₄ ; Aryl or heteroaryl; Or any of Y ₁ , Y ₂ , Y ₃ , and Y ₄ on adjacent carbon atoms Two of which may constitute a methylenedioxy group, where each R ₄ Is independently H; Straight or branched C ₁ -C ₇ alkyl, monofluoroalkyl or polyfluoroalkyl; Straight or branched C ₂ -C ₇ alkenyl or alkynyl; C ₃ -C ₇ cycloalkyl, C ₅ -C ₇ cycloalkenyl, aryl or aryl (C ₁ -C ₆ ) alkyl; Wherein A is A ′, straight or branched C ₁ -C ₇ alkyl, aryl, heteroaryl, aryl (C ₁ -C ₆ ) alkyl or heteroaryl (C ₁ -C ₆ ) alkyl;

Where A 'is

or

Wherein R ₁ and R ₂ are each independently H, straight or branched C ₁ -C ₇ alkyl, —F, —Cl, Br, I, —NO ₂ , or —CN;

Wherein R ₃ is H, straight or branched C ₁ -C ₇ alkyl, F, —Cl, —Br, I, —NO ₂ , —CN, —OR ₆ , aryl or heteroaryl;

Wherein R ₅ is straight or branched C ₁ -C ₇ alkyl, N (R ₄ ) ₂ , —OR ₄ or aryl;

Wherein R ₆ is straight or branched C ₁ -C ₇ alkyl or aryl;

Wherein B is aryl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, indolinyl, indol-4-yl, indol-5-yl, indol-6-yl, indol-7-yl, Isoindolyl, benzo [b] furan-4-yl, benzo [b] furan-5-yl, benzo [b] furan-6-yl, benzo [b] furan-7-yl, benzo [b] thiophene -4-yl, benzo [b] thiophen-5-yl, benzo [b] thiophen-6-yl, benzo [b] thiophen-7-yl, indazolyl, benzimidazolyl, benzo [b] Thiazolyl, purinyl, imidazo [2, lb] thiazolyl, quinolinyl, isoquinolinyl, quinazolinyl, 2,1,3-benzothiazolyl, furanyl, thienyl, pyrroyl, oxazolyl , Thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, oxadiazolyl, triazolyl, thiadiazoleyl, benzoxazolyl, benzisoxazolyl, cynolinyl, quinooxalinyl, 1, 8-naphthyridinyl, putridinyl, or phthalimidyl; In the proviso, the ortho carbon atom or carbon atom for the nitrogen atom of the imine bond is one or more of F, -Cl, --Br, I, --CN, methyl, ethyl or methoxy. May be substituted;

Where n is an integer from 1 to 4;

Wherein aryl is —F, —Cl, —Br, —I, —NO ₂ , —CN, straight or branched C ₁ -C ₇ alkyl, straight or branched C ₁ -C ₇ monofluoroalkyl, straight or branched chain C ₁ -C ₇ polyfluoroalkyl, straight or branched C ₂ -C ₇ alkenyl, straight or branched C ₂ -C ₇ alkynyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₇ monofluorocycloalkyl , C ₃ -C ₇ polyfluorocycloalkyl, C ₅ -C ₇ cycloalkenyl, -OR ₄ , SR ₄ , -OCOR ₄ , -COR ₄ , -NCOR ₄ , -CO ₂ R ₄ , -CON (R ₄ Or phenyl or naphthyl including phenyl or naphthyl substituted with one or more of (CH ₂ ) _n -O- (CH ₂ ) _m CH ₃ .

In certain embodiments, the methods and systems of the present invention utilize one or more memory materials selected from the group consisting of compounds having the structure:

or

Where Me is methyl, cPent is cyclopentyl, and carbons 3 and 5 are in the S configuration.

Degree. 1a-1b illustrate graphs showing results from embodiments of the invention described in Example 1. FIG. Results are expressed as mean days ± sem per treatment, with * p <0.05 vs. Vehicle.
Degree. 2a-2b illustrate graphs showing results from embodiments of the invention described in Example 1. Results are expressed as mean days ± sem per treatment.
Degree. 3a-3b illustrate graphs showing results from embodiments of the invention described in Example 1. Results are expressed as mean days ± sem per treatment.
Degree. 4a-4d describe exemplary visual stimulus pairs used in the embodiment described in Example 1.
Degree. 5 is a graph showing the behavior of healthy middle-aged volunteers in the paired associative (face-name) memory analysis embodiment described in Example 2. FIG. Subjects were asked to remember 40 pairs of faces and names (occupations) for 6 days (d2-d7). During training days, subjects were allowed to see 40 pairs only once. First, they asked questions about their memory and then gave them the correct answers to continue the training. After training, only face-name pair 1-20 (d10) was used to quantify memory retention for 3 days. Face-name pairs 1-20 (d14b) were quantified for 7 days memory retention. The 7-day memory retention for face-name pairs 21-40 was quantified twice, with 1 minute intervals between memory sessions (d14a and d14a2).

In the following paragraphs, the invention will be described by way of example with reference to the accompanying drawings. Throughout the specification, preferred embodiments and examples are to be considered illustrative rather than limiting of the invention. As used herein, “invention” refers to any one of the embodiments of the invention described herein and equivalent thereof. Moreover, reference to various features of "the invention" throughout this document does not imply that all claimed embodiments or methods must include the referenced features.

The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the scope of the invention. Here and in the appended claims, the singular forms “a,” “an,” and “the” include plural referents unless the context clearly dictates otherwise. Thus, for example, a “a compound” refers to one or more compounds and includes equivalents known to those skilled in the art.

Unless otherwise stated, all technical and scientific terms used herein have the same meanings as commonly recognized by one of ordinary skill in the art to which this invention belongs. Although particular methods, devices, and materials are described, any methods and materials similar or equivalent to those described herein can also be used to accomplish and test the present invention.

The present invention provides methods and systems for identifying, evaluating and testing various compounds as memory materials and for identifying, evaluating and testing regimes as training protocols. Such methods and systems can test any compound or protocol, including the memory training protocol described herein. This method may include a combination of training protocols and general administration of memory materials. In certain embodiments, the invention may include identifying, selecting, testing, evaluating or evaluating a compound as a drug candidate for a memory substance, such as, for example, a substance effective for enhancing memory.

Drug candidates may be identified through a drug discovery processor and may be a synthesized, characterized, optimized and / or screened or analyzed compound. Once a compound has been identified as having potential for a given activity or effect using the methods or systems of the present invention, it may be a drug candidate, ie it has the meaning of being finally commercialized and enters a clinical trial. Becomes

How to identify, evaluate and test memory and training protocols

In certain embodiments, the invention relates to the use of a task of assessing the memory capacity of one or more animals. For example, the animal's learning ability, short or long term memory retention can be tested or evaluated. As used herein, the term “memory” refers to long term memory without additional modifiers (eg, short term memory). As used herein, the term "memory agent" is a memory enhancer or memory inhibitor. Memory enhancers and inhibitors are described in U.S. Patents 5,929,223, 6,051,559, 6,689,557, and 6,890,516, and U.S. Pat. Patent application number. As described in 11 / 066,125 (each of which is incorporated by reference in its entirety), includes, but is not limited to, these molecules, compounds and / or substances that function in the CREB pathway.

As used herein, “animal” includes mammals as well as other animals, vertebrates and invertebrates (eg, birds, fish, reptiles, insects (eg, Drosophila species), mollusks (eg, small)). As used herein, “mammals” are used to feed babies, give birth (mammal or placental mammals), or lay eggs (metatharian or non-placental mammals), mononotresmes, Refers to any vertebrate, including marsupials and placental animals. Examples of mammalian species include humans and primates (e.g. monkeys, chimpanzees, humans, tailless monkeys, lemurs, or macaques), rodents (e.g. mice, mice, guinea pigs) and ruminants (e.g. cattle, pigs). , Words), but is not limited to such.

The animal may be an animal with some degree of memory dysfunction or an animal with normal memory performance (eg, an animal without memory loss (any memory abnormality or loss)). One or more animals may be used, and such animals may be divided into one or more groups (eg, one or more controls and / or one or more experimental groups). It can be any age range.

The task involves providing a particular stimulus to the subject. The stimulus can be any suitable sensory stimulus, including but not limited to hearing, sight, touch, smell, taste and / or combinations thereof. In some embodiments, the stimulus may be visual and may be one or more symbols, letters, numbers, lines, shapes, colors, pictures, or a combination thereof. Stimuli are associated in a bundle, with specific associations between the elements in the bundle. Stimuli may be associated in about two, three, four, five, six, seven, eight, nine, or ten groups. In some such embodiments, the stimuli may be associated in pairs. In certain embodiments, each stimulus in a pair is of the same type (eg, auditory, visual or tactile), and in other embodiments, the pair may be of a different type. In certain embodiments, an element in a pair may have a somewhat identifiable association with other elements in the pair. One or more auditory, logical, or other aspects of the pair may be identifiable associations such that one element of the pair and its corresponding element may be associated. In other embodiments, there may be no identifiable association. For example, a pair can be a word-word pair, where one word has no identifiable association with another word (eg, a comb-apple). Examples of visual paired stimuli are shown in FIGS. 4A-D. Another example of a stimulus pair with an identifiable association may be a face-name pair and / or an occupation-face / name pair. In some embodiments, there may be a job listed with the face-name pair.

In certain embodiments, one or more stimulus images can be presented to the subjects. In certain embodiments, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen , 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 50, 100, 120, 150, or any appropriate number of stimuli Pairs can be presented. In certain embodiments, one element of a pair is designated positive, meaning that the individual is rewarded when selecting the element. In another embodiment, one element of the pair is designated negative, meaning that the subject receives a negative feedback when selecting the element. In some embodiments, the individual selects the element by touching the element by hand, but in other embodiments, any other suitable screening method may be used. In some embodiments, the reward may be food, but in other embodiments, the reward may be any suitable for triggering a positive sound, access to a particular toy or item, or triggering a positive response from an individual. In certain embodiments, the negative feedback is any suspension of shock, sound, punishment or benefit or any appropriate action or similar that provides a negative feedback to the individual.

The type of feedback, selection of stimulus pairs, pair of stimulus pairs, type and age of the subject and other aspects of the task can be modified to test and / or evaluate memory of different aspects of the subject. For example, each element may be used with a pair of stimuli that have an identifiable association with other elements in the pair to assess the individual's associated memory.

In other embodiments, other evaluation methods other than reward and / or negative feedback evaluation methods may be used. For example, in some embodiments, when the subject is a human, the stimulus may be corrected by a pair or other association (eg, paired stimuli such that there is an intended identifiable association between one element of the pair and the other). It can have the same association as determining whether it is wrong or wrong. For example, the subject may be taught the face-name pair, and the stimulus pair will be provided to the subject with the exact pair of face-names scattered. The entity will tell if the presented face-name pair is the exact face-name pair that it learned. In such embodiments, no separate compensation is necessary, and the compensation may be a simple feedback on the accuracy of the human response. The stimulus pair may be provided in any suitable manner. For example, pairs may be presented in random, organized or virtual-random order during each test session.

Subjects may be tested at any suitable frequency over any suitable time period. For example, subjects may be used once, twice, three times, four times, five times, six times, seven times, eight times, nine times, ten times, eleven times, twelve times, thirteen times, and fourteen times during the test period. , 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 50, 60, 75, 100, 150, 200 Test may be performed 250 times, 365 times, or any other suitable number of times (eg, if the test is daily, the subject may be tested once, twice, three times). In such a test, the subject may see any suitable number of stimuli or pairs of stimuli. The subject may be tested daily, once every two days, weekly, every two weeks, once a month, or at any appropriate frequency. Moreover, any suitable number of subjects can be tested. In some embodiments, a test is performed to teach or encode information about the correlation between paired stimuli to the individual. For example, a test can be run to teach an individual which elements of a pair are positive and / or negative elements. In other embodiments, the test can be performed to teach the exact pairs of elements (eg, exact face-name, occupation-name or occupation-face / name association) within a set of stimuli. In such embodiments, multiple test phases may be required. Following the first teaching phase is the second phase, where the knowledge of the individual is tested by asking whether the pair presented is correct by appropriate methods, depending on the type of individual (eg human v. S. Rat).

Subjects will be evaluated in any suitable way. In certain embodiments, individuals are provided with one or more pairs of stimuli, where one element of the pair is positive and the other element is negative. In such embodiments, individuals who have learned that either element of a pair is positive and / or negative are evaluated based on the number of times they select a positive and / or negative element and / or do not select a positive and / or negative element . In other embodiments, the stimulus pairs may be related to each other (eg, face-name relationships, job-name relationships, job-face / name relationships, word pairs, letters or numbers sets or groups of names or symbols). In such embodiments, the ability to reestablish the appropriate pair (e.g., place the name in a corresponding pictorial element or similar) can be assessed to the individual who has learned the correct association between the elements of the pair. In some embodiments, the pair may be altered in some way, such that the subject may be asked to confirm how the pair was changed (eg, the response may be “untouched”, “rearranged”, “One can be new” or “both new”).

In certain embodiments, the long term memory of the individual can be tested. In such embodiments, reference results can be established. Such criteria can be established in various ways. For example, in embodiments in which a stimulus pair is provided to the subject in which one element of the pair is positive and / or one element of the pair is negative, the stimulus pair is the subject until the subject identifies positive and / or negative elements at a certain frequency. Will be provided to The frequency here will serve as a criterion so that later results can be evaluated. Similarly, the frequency with which the relationship between the elements of the stimulus pairs is precisely used may be used to establish the criteria. In certain embodiments, an individual is assessed until reaching a certain success rate. For example, about 50%, 55%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, 100% of the baseline, or any suitable ratio or exact result Can be used as a target. In other embodiments, the individual may be tested a certain number of times over a certain period of time, and a final assessment may be used to establish the criteria. For example, about 1-23 hours or about 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 50, 60, 75, 100, 150 , 200, 250, or 365 days or any suitable time or number of days may be used to establish the criteria. In addition, in order to establish the standard, the individual is about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 Times, 15 times, 16 times, 17 times, 18 times, 19 times, 20 times, 21 times, 22 times, 23 times, 24 times, 25 times, 50 times, 60 times, 75 times, 100 times, 150 times, 200, 250, 365, or any other suitable number of times. Criteria can be established using the mean, median or any other statistical means of the individual's actual results.

In certain embodiments, once a criterion is established, the individual is not evaluated or exposed to any stimulus pair used to establish the criterion for a particular time. The amount of time is about 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 15 days, 16 Days, 17 days, 18 days, 19 days, 20 days, 21 days, 22 days, 23 days, 24 days, 25 days, 50 days, 60 days, 75 days, 100 days, 150 days, 200 days, 250 days, 365 days, or any suitable number of days.

After the delay period, the subject is tested again using the same pair of stimuli used to establish the criteria. In certain embodiments, these tests are repeated under the same, similar or different conditions as used to establish the criteria. Moreover, the individual's behavior can be assessed by establishing long term success measures in any suitable manner. In some embodiments, the method of establishing a long term success measure is described to establish a criterion, although a shorter time is generally used, in which the long term success measure is an indicator of the individual's memory rather than learning the relevant criteria. Similar to Individuals can be assessed by comparing long-term success measures with criteria. Statistical or similar analyzes can be used to evaluate such results.

In certain embodiments, the methods of the present invention can be used to assess the effect of a potential memory material and / or a potential training protocol. For example, the method of the present invention can be used to identify a reduction in the amount of training required to reach a selected performance level. In some embodiments, an individual is first evaluated in the absence of a memory material, followed by successive administrations of the memory material. In addition, the subject's control group was tested using a specific pair of stimuli and compared the results with the subjects who received a given amount of memory. Evaluation of memory materials and / or training protocols can take many forms. For example, memory materials may reduce the time or days of training sessions or time required to establish or meet certain criteria or to achieve any suitable effect.

In some embodiments of the invention, the memory material is assessed as changing (eg, increasing or decreasing) the number of days or training sessions to reach a performance criterion. As used herein, “criterion” refers to the measurement of a desired result and / or desired level of performance, for example, the ratio of an accurate response (eg, a correctly paired stimulus) from a test subject or test population. . Criteria may be changed or selected as desired by the facilitator of protocols, tests, tasks and / or other embodiments of the present invention. In these embodiments the complexity of the administered training can be manipulated by changing the criteria, the number of stimuli to be paired and / or the type of stimulus, the number of days of training, the quality of the training, and / or the type of training. In a further embodiment, training in the absence of memory material is sufficient to make CREB-dependent modifications in long term memory, for example, to make CREB dependent reinforcement of long term memory.

In certain embodiments, the methods of the present invention can be used to assess the effect of memory impairment. Such embodiments may include similar or identical steps as the method of evaluating the memory material. However, the indicators evaluated in such embodiments will be different. For example, a memory impairment may increase the time required to establish a particular criterion, increase the number of training sessions required to reach the criterion, reduce the ratio of long-term success measures to the criterion, or any other Can be evaluated in an appropriate manner.

Any suitable memory material or memory impairment may be provided by any suitable method via any suitable delivery system, using any suitable delivery schedule and dosage. In addition, any suitable training protocol may be used. Exemplary training protocols, memory materials and memory impairments are discussed in particular in the following paragraphs.

The invention also includes a system, including a system used with the method described herein. For example, in some embodiments, the system may include all the elements necessary to practice an embodiment of the method of the present invention. The system of the present invention selects a memory as a drug candidate, evaluates the effect of the memory, evaluates the effectiveness of the training protocol, tests the memory as a long-term memory enhancer, and stores it as a specific impairment or general memory. It will be useful for testing materials and / or testing memory materials with training protocols as long-term memory enhancers. Such a system may include the use of multiple sets of stimuli (including paired stimuli) described above, and the use of test schedules and protocols described above.

Training protocol

As described herein, evaluation using the methods and systems of the present invention may include administration of two parts (1) specific training protocols and (2) memory materials. In certain embodiments, only a training protocol is used. In other embodiments, both training and administration of memory material occur. Such a combination can enhance the training protocol by reducing the number of training sessions required to make performance gains compared to those obtained by training alone or by eliminating shorter or rest periods between training sessions to gain performance gains. . Such a combination can be achieved by reducing the number or duration of training sessions required for induction in specific neuronal circuits of a training protocol neuronal activity pattern or during synaptic connections of neuronal circuits (CREB) -dependent long-term structural / function Training protocols can be enhanced by reducing the number and / or duration of underlying patterns of neuronal activity required to induce (eg, long-lasting) changes. In this way, the effectiveness of existing training protocols can be improved to create significant and economic advantages.

In learning new languages or learning to play new musical instruments, training protocols (eg, massed training, spaced training) are used. In addition to training, administering memory material reduces the time and / or number of training sessions required to obtain a harvest together in performance. As a result, less practice (training sessions) is required to learn a new language or learn to play a new instrument.

Training protocols are used for repeated stimulation of underlying neuronal activity or neuronal activity patterns in specific neuronal circuits in an individual. Administration of memory with training results in a baseline pattern of neuronal activity or the duration of the training session required to achieve a given result, such as induction of CREB-dependent organ structure / function changes (eg, long-lasting) among synaptic connections of neuronal circuits. And / or reduce the number of times.

Such methods include (a) administering to the animal a substance that enhances CREB pathway function; And (b) training the animal under sufficient conditions to improve the memory performance of interest by the animal, thereby enhancing certain memory aspects of the animal in need thereof (especially humans or other mammals or vertebrates). In certain embodiments, the memory material is a CREB pathway-enhancing drug.

The methods described include (a) administering to the animal a substance that enhances CREB pathway function; And (b) training the animal under conditions sufficient to allow for improvement in memory performance of interest by the animal, thereby treating memory defects associated with central nervous system (CNS) abnormalities or diseases in animals that require such treatment. . CNS disorders and abnormalities include memory impairment associated with aging, neurodegenerative diseases (eg, Alzheimer's disease, Parkinson's disease, Huntington's disease (chorea), other senile dementia), mental illnesses (eg, depression, schizophrenia, autism, attention deficit disorder) Loss of function due to trauma (eg, cerebrovascular disease (eg, seizures, ischemia), brain tumors, head or brain damage), genetic defects (eg, Rubinstein-Taybi syndrome, Down syndrome), learning disabilities, and / or Includes complex thereof.

The described method can be used for repetitive stimulation of neuronal activity or neuronal activity patterns underlying certain neuronal circuits in an animal, which method comprises: (a) administering to the animal a substance that enhances CREB pathway function; And (b) training the animal under conditions sufficient for improvement in memory performance of interest by the animal.

For many tasks in many species, including humans, the spaced training protocol (multiple training sessions with rest periods between each training session) is a massed training protocol (without rest periods between each training session). Stronger, longer lasting memory than multiple training sessions). The behavior of Pavlov olfactory learning in Drosophila-massed training in genetic studies creates long-lasting memory that collapses for at least four days, is not destroyed by over-expression of the CREB-inhibitor transgene, and collapse in radish mutations Has been established ( Tully , T. et. al ., Cell , 79 (l): 35-47 (1994); and Yin , JC et al ., Cell , 79 (l): 49-58 (1994) ). In contrast, spaced training produces long-lasting memory that lasts at least seven days, is protein synthesis-dependent, destroyed by overexpression of CREB-inhibitor transgenes, and is normal in radish mutations ( Tully , T. et al , Cell , 79 (l): 35-47 (1994); and Yin, JC et al , Cell , 79 (l): 49-58 (1994) ). At 1 day after interval training, memory retention consists of both protein synthesis-independent and CREB-independent early memory (ARM) and protein synthesis-dependent and CREB-dependent long-term memory (LTM). Further massed training to induce LTM is insufficient ( Tully , T. et. al ., Cell , 79 (l): 35-47 (1994); and Yin , JC et al ., Cell , 79 (l): 49-58 (1994) ).

A series of increasing increases results from invertebrates to mammals. For example, in mice, molecular manipulation of CREB expression similar to that performed in flies can (i) inhibit or enhance LTM of facilitatory electrophysical responses at sensorimotor monosynapses in cell culture, and (ii) Suppresses or enhances synaptic connections between sensory and motor neurons that normally make up after spaced application of palpation stimuli ( Bartsch , D. et. al , Cell , 83 (6): 979-992 (1995) ). In mice, injection of antisense RNA oligonucleotides into the hippocampus or amygdala blocks LTM formation of two different tasks, each dependent on the activity of these anatomical sites. ( Guzowski , JF et al ., Proc . Natl . Acad . Sci . USA , 94 (6): 2693-2698 (1997); and Lamprecht , R. et al ., J. Neurosci ., 17 (21): 8443-8450 (1997) ). In mice, LTM formation for implicit and explicit tasks is defective in CREB mutant mice ( Bourtchuladze , R. et. al ., Cell , 79 (l): 59-68 (1994) ).

Transgenic mice carrying the CRE-dependent reporter gene (betagalactosidase) are trained in hippocampal-dependent associated fear control or passive avoidance operations to induce CRE-dependent reporter gene expression at the CA1 and CA3 sites of the hippocampus. Training these mice in the amygdala-dependent fear regulation task induces CRE-dependent reporter gene expression in the amygdala, not the hippocampus. Thus, training protocols that induce LTM formation also induce CRE-dependent gene transcription in specific anatomical regions of the mammalian brain ( Impey , S. et al ., Nat . Neurosci ., L (7): 595-601 ( 1998) ).

In these animal models, three prominent cases of LTM enhancement are described. First, overexpression of the CREB-activator transgene eliminates the need for multiple, spaced training sessions and instead induces LTM formation after only one training session (normally with little memory retention after 24 hours). Or none, ( Yin, JC et al , Cell , 81 (1): 107-115 (1995) ). Second, injection of virus-expressed CREB-active mass transfer gene into rat amygdala is sufficient to enhance memory after massed training for fear-enhanced startling response, thus providing a rest interval in spaced training. Eliminates the need for ( Josselyn , SA et al ., Society for Neuroscience , Vol . 24, Abstract 365.10 (1998) ). Third, if mutant mice receive differently spaced training protocols, LTM formation can be normally formed in CREB-deficient mice (Bourtchuladze, R. et al., Cell, 79 (l): 59-68 (1994)). Kogan , JH et al ., Curr . Biol, 7 (1): 1-11 (1997) ).

CREB also appears to be involved in various forms of developmental and cellular plasticity in the vertebrate brain. For example, neuronal activity increases CREB activity in the cerebral cortex ( Moore , AN et. al ., J. Biol. Chem ., 271 (24): 14214-14220 (1996) ). CREB is also used for hippocampus ( Murphy , DD et al ., Proc . Natl. Acad . Sci . USA , 94 (4): 1482-1487 (1997 ), Glazewski , S. et. al ., Cereb . Cortex, 9 (3): 249-256 (1999) ), striatum ( Liu , FC et al ., Neuron , 17 (6): 1133-1144 (1996) ), and visual age ( Pham , TA et al. al ., Neuron , 22 (l): 63-72 (1999) ) mediate developmental plasticity.

CREB appears to be affected by human neurodegenerative diseases and brain damage. For example, CREB activation and / or expression is disrupted in Alzheimer's disease ( Ikezu , T. et. al ., EMBO J., 15 (10): 2468-2475 (1996); Sato , N. et al ., Biochem . Biophys . Res . Commun , 232 (3): 637-642 (1997); and Yamamoto- Sasaki, M. et al ., brain. Res ., 824 (2): 300-303 (1999) . CREB activation and / or expression is also elevated after seizures or ischemia ( Blendy , JA et al , Brain Res ., 681 (l-2): 8-14 (1995); and Tanaka , K. et al , Neuroreport , 10 (11): 2245-2250 (1999)). " Environmental enrichment " is neuroprotective , preventing cell death by acting through CREB ( Young , D. et al ., Nat . Med ., 5 (4): 448-453 (1999) ).

CREB functions during drug sensitivity and dosing off. For example, CREB is affected by ethanol ( Pandey , SC et al ., Alcohol Clin . Exp . Res ., 23 (9): 1425-1434 (1999); Constantinescu , A. et al ., J. Biol . Chem ., 274 (38): 26985-26991 (1999); Yang , X. et al ., Alcohol Clin . Exp . Res., 22 (2): 382-390 (1998); Yang , X. et al ., J. Neurochem ., 70 (l): 224-232 (1998); and Moore , MS et al ., Cell , 93 (6): 997-1007 (1998)), by cocaine (Carlezon, WA, Jr. et al ., Science, 282 (5397): 2272-2275 (1998)), by morphine ( Widnell , KL et al ., J. Pharmacol . Exp . Ther., 276 (l): 306-315 (1996)), by methamphetamine ( Muratake , T. et al ., Ann NY Acad . ScL , 844: 21-26 (1998)) and by cannabinoid ( Calandra , B. et al ., Eur . J. Pharmacol , 374 (3): 445-455 (1999); and Herring , AC et al ., Biochem . Pharmacol , 55 (7): 1013-1023 (1998) ).

The signal induction pathway that can stimulate the CREB / CRE transcription pathway is the cAMP regulatory system. Correspondingly, mice lacking adenylate cyclase 1 (AC1) and AC8 enzymes do not learn ( Wong ST et al ., Neuron , 23 (4): 787-798 (1999) ). In these mice, administration of forskolin to the CA1 site of the hippocampus restores learning and memory of hippocampal-dependent tasks. Furthermore, treatment of older mice with drugs that raise cAMP levels (rollifram and D1 receptor agonist) improves the loss of hippocampal-dependent memory and aging of cellular organs ( Barad , M. et. al ., Proc . Natl . Acad . Sci . USA , 95 (25): 15020-15025 (1998) ). These latter data suggest that older mice with learning disabilities have defects in cAMP signaling ( Bach , ME et. al ., Proc . Natl . Acad . Sci . USA , 96 (9): 5280-5285 (1999) ).

An increasing series of evidence indicates that neurons continue to proliferate in the adult brain ( Arsenijevic , Y. et al., Exp . Neurol ., 170: 48-62 (2001); Vescovi , AL et al ., Biomed . Pharmacother ., 55: 201-205 (2001); Cameron , HA and McKay , RD, J. Comp . Neurol , 435: 406-417 (2001); and Geuna , S. et al ., Anat . Rec , 265: 132-141 (2001) ) and this proliferation is suggested to respond to a variety of experiences ( Nilsson , M. et. al ., J. NeurobioL , 39: 569-578 (1999); Gould , E. et al ., Trends Cogn . ScL, 3: 186-192 (1999); Fuchs , E. and Gould , E., Eur . J. Neurosci ., 12: 2211-2214 (2000); Gould, E. et al ., Biol . Psychiatry , 48: 715-720 (2000); and Gould , E. et al ., Nat . Neurosci ., 2: 260-265 (1999) ). The experimental strategy now is to transplant neuronal stems into the adult brain for various therapeutic indications ( Kurimoto , Y. et. al ., Neurosci . Lett ., 306: 57-60 (2001); Singh , G., Neuropathology, 21: 110-114 (2001); and Cameron , HA and McKay , RD, Nat . Neurosci ., 2: 894-897 (1999) ). Much is known about neurogenesis in the embryonic stage ( Saitoe , M. and Tully , T., " Making connections between synaptic and behavioral plasticity in Drosophila ", In Toward a theory of Neuroplasticity , J. McEachern and C. Shaw , Eds . ( New York : Psychology Press .), Pp. 193-220 (2000) ). Neuronal differentiation, neurogenesis prolongation, and early synaptic target recognition all appear to occur in an active-independent manner. However, synapse production and synaptic growth then require ongoing neuronal activity to fine-tune synaptic connections in a functionally relevant manner. This finding suggests that functional (final) integration of transplanted neural stem cells requires neuronal activity. Thus, ACT can be used to act on appropriate neuronal circuits to fine-tune synaptic connections of newly acquired, transplanted stem cells (differentiated into neurons). “The proper action of neuronal circuits” means induction in the appropriate neuronal circuits with patterns of neuronal activity, corresponding to those made by specific cognitive training protocols. Cognitive training protocols can be used to induce such neuronal activity. Alternatively, neuronal activity may be induced by direct electrical stimulation of the neuronal circuit. "Nurnal activity" and "nerve activity" are used interchangeably herein.

Memory substances, such as CREB pathway-enhancing drugs, can enhance CREB pathway function, which is required to incorporate newly acquired information into the LTM. "Enhanced CREB pathway function" means the ability to enhance or improve CREB-dependent gene expression. CREB-dependent gene expression can be enhanced or improved by increasing endogenous CREB production, e.g., by direct or indirect stimulation of endogenous genes to produce increased amounts of CREB or by functioning (biologically) May be enhanced or improved by an increase in CREB). For example, U.S. Pat. No. 5,929,223; U.S. Pat. No. 6,051,559; And International Publication No. See WO9611270 (Apr. 18, 1996) —all incorporated by reference. Administration of memory materials may reduce the training required to obtain improved performance compared to that obtained with training alone. "Performance gain" means improvement in terms of memory performance.

The present invention provides a method of identifying, testing and / or enhancing enrichment of a particular memory aspect in a mammal (particularly a human or other mammal or vertebrate) (in some embodiments, in an individual in need thereof), where such Enrichment comprises (a) administering a memory substance to the animal, eg, a memory substance that enhances CREB pathway function; And (b) train animals under sufficient conditions to improve on their specific memory performance.

Training can consist of one or multiple training sessions and is appropriate to make improvements in the performance of the memory task of interest. For example, if you want to improve on language acquisition, training will focus on language acquisition. If you want to improve your ability to learn to play a musical instrument, training will focus on learning to play a musical instrument. If one wants to improve on a particular athletic skill, training will be focused on the acquisition of that particular athletic skill. Certain memory tasks of interest are paired with appropriate training.

The invention also provides a method of assessing, testing, and / or confirming the enhancement of memory performance in an animal by repeated stimulation of neuronal activity or neuronal activity underlying certain neuronal circuits in the animal, the method comprising: (a ) Administering to the animal a memory that enhances CREB pathway function; And (b) training the animal under conditions sufficient to stimulate or induce neuronal activity or neuronal activity pattern in the animal. In this case, training refers to training appropriate for stimulating or inducing neuronal activity or patterns of neuronal activity in mammals.

"Multiple training sessions" means two or more training sessions. The memory may be administered before, during or after one or more training sessions. In certain embodiments, the memory material is administered before and during each training session.

Specific training protocols to be evaluated in accordance with the present invention are known and are readily available in the art. For example, Kami , A. and Sagi , D., " Where practice makes perfect in text discrimination : evidence for primary visual cortex plasticity ", Proc . Natl . Acad . Sci . USA , 88: 4966-4970 (1991); Kami , A. and Sagi , D., " The time course of learning a visual skill , " Nature , 365: 250-252 (1993); Kramer , AF et al ., " Task coordination and aging : explorations of executive control processes in the task switching paradigm , " Acta Psychol . (. Amst), 101: 339-378 (1999); Kramer , AF et al , " training for executive control : Task coordination strategies and aging , " In Aging and Skilled Performance : Advances In Theory and applications, W. Rogers et al ., eds . (Hillsdale, NJ: Erlbaum) ( 1999); Rider , RA and Abdulahad, DT, " Effects of massed versus distributed practice on gross and fine motor proficiency of educable mentally handicapped adolescents , " Percept . Mot . Skills , 73: 219-224 (1991); Willis , SL and Schaie , KW, " training the elderly on the ability factors of spatial orientation and inductive reasoning , " Psychol . Aging , 1: 239-247 (1986); Willis , SL and Nesselroade , CS, " long - term effects of fluid ability training in old - old age , "Develop. Psychol , 26: 905-910 (1990); Wek , SR and Husak , WS, " Distributed and massed practice effects on motor performance and learning of autistic children , " Percept . Mot . Skills, 68: 107-113 (1989); Verhaehen , P. et. al ., " Improving memory performance in the aged through mnemonic training : a meta - analytic study , " Psychol . Aging , 7: 242-251 (1992); Verhaeghen, P. and Salthouse , TA, " Meta - analyses of age - cognition relations in adulthood : estimates of linear and nonlinear age effects and structural models , " Psychol . Bull , 122: 231-249 (1997); Dean , CM et al ., " Task - related circuit training improves performance of locomotor tasks in chronic stroke : a randomized , controlled pilot trial , " Arch . Phys . Med. Rehabil , 81: 409-417 (2000); Greener , J. et. al ., " Speech and language therapy for aphasia following stroke , " Cochrane Database Syst . Rev. , CD000425 (2000); Hummelsheim , H. and Eickhof , C, " Repetitive sensorimotor training for arm and hand in a patient with locked -in syndrome , " Scand . J. Rehabil . Med ., 31: 250-256 (1999); Johansson , BB," brain plasticity and stroke rehabilitation . The Willis lecture , " Stroke , 31: 223-230 (2000); Ko Ko, C," Effectiveness of rehabilitation for multiple sclerosis , " Clin . Rehabil ., 13 ( Suppl . 1): 33-41 (1999); Lange , G. et al ., " Organizational strategy influence on visual memory performance after stroke : cortical / subcortical and left / right hemisphere contrasts , " Arch . Phys. Med . Rehabil ., 81: 89-94 (2000); Liepert , J. et. al ., " Treatment - induced cortical reorganization after stroke in humans , " Stroke , 31: 1210-1216 (2000); Lotery , AJ et al ., "Correctable visual impairment in stroke rehabilitation patients , " Age Ageing , 29: 221-222 (2000); Majid , MJ et al ., " recognition rehabilitation for memory deficits following stroke "( Cochrane review ), Cochrane Database Syst . Rev. , CD002293 (2000); Merzenich , M. et al., " Cortical plasticity underlying perceptual , motor , and recognition skill development : implications for neurorehabilitation , " Cold Spring Harb . Symp . Quant . Biol ., 61: 1-8 (1996); Merzenich, MM et al ., " Temporal processing deficits of language - learning impaired children ameliorated by training , " Science , 271: 77-81 (1996); Murphy , E.," Stroke rehabilitation, "JR Coll . Physicians Lond ., 33: 466-468 (1999); Nagarajan , SS et al ., "Speech modifications algorithms used for training language learning - impaired children , "IEEE Trans . Rehabil . Eng ., 6: 257-268. (1998); Oddone , E. et. al ., " Quality Enhancement Research Initiative in stroke : prevention , treatment , and rehabilitation , " Med . Care 38: 192-1104 (2000); Rice - Oxley , M. and Turner - Stokes , L., " Effectiveness of brain injury rehabilitation, " Clin . Rehabil ., 13 ( Suppl l): 7-24 (1999); Tallal , P. et. al ., " Language learning impairments : integrating basic science , technology , and remediation , " Exp . Brain Res., 123: 210-219 (1998); Tallal , P. et al ., " Language comprehension in language - learning impaired children improved with acoustically modified speech , " Science , 271: 81-84 (1996) , each of which is incorporated by reference in its entirety.

Accordingly, the present invention relates to a method of enhancing long-term memory in an animal (especially a human or other mammal or vertebrate) in need of such treatment, which method (a) assesses, estimates and / or Or administering the identified, tested or evaluated memory to the animal by a method of testing, screening the memory as a drug compound or testing the memory as a long term memory enhancer, and (b) long-term memory in the animal. Training the animal under conditions sufficient to enhance the activity.

In one embodiment, the invention relates to a method of treating aging-associated memory damage in an animal in need of such treatment, the method comprising: (a) assessing, estimating and / or testing the effect of a memory material and Sufficient conditions for administering the identified, tested or evaluated memory material to the animal by selecting a memory material as a compound or testing the memory as a long term memory enhancer, and (b) enhancing the long term memory in the animal. Training animals under the following conditions.

In another embodiment, the present invention relates to a method of treating memory defects associated with neurodegenerative diseases (eg, Alzheimer's disease, Parkinson's disease, Huntington's disease, other senile dementia) in an animal in need of such treatment, (A) identified, tested or evaluated by a method of evaluating, estimating and / or testing the effectiveness of a memory material, selecting the memory material as a drug compound or testing the memory as a long-term memory enhancer To the animal, and (b) training the animal under conditions sufficient to enhance long-term memory in the animal.

In another embodiment, the present invention relates to a method of treating mental illness (eg, depression, schizophrenia, autism, attention deficit disorder) in an animal in need of such treatment, the method comprising: (a) Administering the identified, tested or evaluated memory to the animal by assessing, estimating and / or testing the effectiveness, selecting the memory as a drug compound or testing the memory as a long term memory enhancer, and (b) training the animal under conditions sufficient to enhance long-term memory in the animal.

In another embodiment, the invention provides for memory defects associated with traumatic dependent loss of memory function in an animal in need of such treatment (eg, cerebrovascular disease (eg, seizures, ischemia), brain tumors, head or brain damage). A method of treatment, the method comprising: (a) evaluating, estimating and / or testing the effectiveness of a memory, screening the memory with drug compounds or testing the memory with long-term memory enhancers Administering the tested, tested or evaluated memory material to the animal, and (b) training the animal under conditions sufficient to enhance long-term memory in the animal.

In another embodiment, the present invention relates to a method of treating memory deficiency (eg, Rubinstein-Taybi syndrome, Down syndrome) associated with a genetic defect in an animal in need of such treatment, the method comprising: (a) memory Administering the identified, tested or evaluated memory material to the animal by assessing, estimating and / or testing the effect of the material, selecting the memory material as a drug compound or testing the memory as a long term memory enhancer And (b) training the animal under conditions sufficient to enhance long-term memory in the animal.

The present invention relates to a treatment for memory deficiency associated with a CNS disorder or condition in an animal undergoing neuronal stem cell manipulation, the method comprising: (a) evaluating, evaluating and / or testing the effects of a memory material and using the drug substance as a drug compound. Administering the identified, tested or evaluated memory to the animal by screening or testing the memory as a long-term memory enhancer, and (b) training the animal under conditions sufficient to enhance long-term memory in the animal. It involves doing.

"Nurnal stem cell manipulation" means that (1) exogenous neuronal stem cells are transplanted into the animal's brain or spinal cord, or (2) endogenous neuronal stem cells are stimulated or induced to proliferate in the animal. Transplanting neuronal stem cells into the animal brain or spinal cord is known and is readily available in the art (eg, Cameron , HA and McKay , RD, Nat . Neurosci ., 2: 894-897 (1999); Kurimoto , Y. et al, Neurosci . Lett ., 306: 57-60 (2001); and Singh , G., Neuropathology , 21: 110-114 (2001 ). Methods for stimulating or inducing proliferation of endogenous neuronal stem cells in animals are known and are readily available in the art (eg, Gould , E. et. al ., Trends Cogn . Sci , 3: 186-192 (1999); Gould, E. et al ., Biol . Psychiatry , 48: 715-20 (2000); Nilsson , M. et al , J. NeurobioL , 39: 569-578 (1999); Fuchs , E. and Gould , E., Eur . J. Neurosci ., 12: 2211-2214 (2000); and Gould, E. et al ., Nat . Neurosci ., 2: 260-265 (1999) ). Certain methods of transplanting neuronal stem cells into the brain or spinal cord of an animal and specific methods of stimulating or inducing proliferation of endogenous neuronal stem cells in an animal are not critical to the practice of the present invention.

The present invention also relates to a method of improving or enhancing memory and / or performance in animals with learning disabilities, language dysfunction, or dyslexia, which method comprises: (a) evaluating, estimating and / or evaluating the effects of memory substances; Or administering the identified, tested or evaluated memory to the animal by a method of testing, screening the memory as a drug compound or testing the memory as a long term memory enhancer, and (b) long-term memory in the animal. Training the animal under conditions sufficient to enhance the activity.

Memory material ( Memory Agents )

As used herein, memory materials are compounds that have pharmacological activity associated with memory disorders and / or memory dysfunctions, and include drugs, chemical compounds, ionic compounds, organic compounds, organic ligands including licensors, saccharides, recombination and Synthetic peptides, proteins, peptoids, nucleic acid sequences including genes, nucleic acid products, and other molecules and compositions. Memory materials can, for example, enhance or impair short-term or long-term memory. For example, the memory material is described in US Patent Application No. 11 / 679,782 "Therapeutic Piperazones" (February 27, 2007); US patent application number. 11 / 608,746 "Indolone Compounds Useful To Treat Cognitive Impairment" (December 8, 2006); And US patent application number. 11 / 679,775 "Therapeutic Compounds" (February 27, 2006), US Provisional Patent Application No. 60 / 942,992, "Therapeutic Pyrazoloquinoline Urea Derivatives" (June 8, 2007), each of the above-mentioned patents The application is incorporated in its entirety by reference.

As used herein, memory impairments are compounds that have pharmacological activity associated with sustained memory (eg, post-traumatic stress disorder) and are organic ligands, including drugs, chemical compounds, ionic compounds, organic compounds, and officials. Saccharides, recombinant and synthetic peptides, proteins, peptoids, nucleic acid sequences including genes, nucleic acid products, and other molecules and compositions. The memory impairments and memory reinforcements of the present invention can be used with any of the methods and systems of the present invention.

For example, memory materials include cell penetrating cAMP analogs (eg, 8-bromo cAMP); Active substances of adenylate cyclase 1 (AC1) (eg, forskolin); Substances that affect G-protein linked receptors such as adrenergic receptors, opioid receptors and their ligands (eg, phenethylamine); Intracellular calcium concentration regulating agents (eg, topsiazine, N-methyl-D-aspartate (NMDA) receptor agonist); inhibitors of phosphodiesterase responsible for cAMP disruption (e.g., rolipram (which inhibits phosphodiesterase 4), iso-buto-metho-santhine (IBMX) (phosphodiesterases 1 and 2) Inhibits)); Regulators of protein kinases and / or protein phosphatase—regulating CREB protein activation and CREB-dependent gene expression. The memory material may also be exogenous CREB, CREB analogues, CREB-like molecules, biologically active CREB fragments, CREB fusion proteins, and / or nucleic acid sequences encoding exogenous CREBs, CREB analogs, CREB-like molecules, biologically active CREB fragments. And / or CREB fusion proteins.

The memory material may also be a CREB function modulator, and / or a nucleic acid sequence encoding a CREB function modulator. As used herein, CREB function modulators have the ability to modulate CREB pathway function. "Modulate" means the ability to change (increase or decrease) or alter CREB pathway function.

The memory material may be a compound capable of enhancing CREB function in the CNS. Such compounds include, but are not limited to, compounds that affect membrane stability, flowability, and certain immune stimuli. In certain embodiments, the memory material may temporarily enhance CREB pathway function in the CNS.

CREB analogs, or derivatives, are defined herein as proteins with amino acid sequences similar to endogenous CREB. An analogous amino acid sequence is defined as meaning an amino acid sequence that is sufficiently homologous to the amino acid sequence of an endogenous CREB, which retains the biological activity of the endogenous CREB but retains one or more "silent" changes in the amino acid sequence. CREB analogs include mammalian CREM, mammalian ATF-1, and other CREB / CREM / ATF-1 subfamily members.

As used herein, CREB-like molecules refer to proteins that functionally mimic CREB. CREB-like molecules need not have amino acid sequences similar to endogenous CREB.

Examples of memory materials may include:

Examples of memory materials may include:

Where "Me" is methyl and "cPent" refers to "cyclopentyl". The formula includes both enantiomers and mixtures thereof. In certain embodiments, (HT-0712), carbons 3 and 5 of the above formula are in the S configuration.

Examples may also include compounds having the structure:

Wherein each Y ₁ , Y ₂ , Y ₃ , and Y ₄ are independently H; Straight or branched C ₁ -C ₇ alkyl, monofluoroalkyl or polyfluoroalkyl; Straight or branched C ₂ -C ₇ alkenyl or alkynyl; C ₃ -C ₇ cycloalkyl or C ₅ -C ₇ cycloalkenyl; -F, -Cl, -Br, or -I; -NO ₂ ; -N ₃ ; -CN; -OR ₄ , -SR ₄ , -OCOR ₄ , -COR ₄ , -NCOR ₄ , -N (R ₄ ) ₂ , -CON (R ₄ ) ₂ , or -COOR ₄ ; Aryl or heteroaryl; Or any of Y ₁ , Y ₂ , Y ₃ , and Y ₄ on adjacent carbon atoms Two of which may constitute a methylenedioxy group, where each R ₄ Is independently H; Straight or branched C ₁ -C ₇ alkyl, monofluoroalkyl or polyfluoroalkyl; Straight or branched C ₂ -C ₇ alkenyl or alkynyl; C ₃ -C ₇ cycloalkyl, C ₅ -C ₇ cycloalkenyl, aryl or aryl (C ₁ -C ₆ ) alkyl; Wherein A is A ′, straight or branched C ₁ -C ₇ alkyl, aryl, heteroaryl, aryl (C ₁ -C ₆ ) alkyl or heteroaryl (C ₁ -C ₆ ) alkyl;

Where A 'is

or

Wherein R ₁ and R ₂ are each independently H, straight or branched C ₁ -C ₇ alkyl, —F, —Cl, Br, I, —NO ₂ , or —CN;

Wherein R ₃ is H, straight or branched C ₁ -C ₇ alkyl, F, —Cl, —Br, I, —NO ₂ , —CN, —OR ₆ , aryl or heteroaryl;

Wherein R ₅ is straight or branched C ₁ -C ₇ alkyl, N (R ₄ ) ₂ , —OR ₄ or aryl;

Wherein R ₆ is straight or branched C ₁ -C ₇ alkyl or aryl;

Wherein B is C ₃ -C ₇ cycloalkyl, C ₅ -C ₇ cycloalkenyl, adamantyl, aryl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, indolinyl, indole-4 -Yl, indol-5-yl, indol-6-yl, indol-7-yl, isoindolyl, benzo [b] furan-4-yl, benzo [b] furan-5-yl, benzo [b] furan -6-yl, benzo [b] furan-7-yl, benzo [b] thiophen-4-yl, benzo [b] thiophen-5-yl, benzo [b] thiophen-6-yl, benzo [ b] thiophen-7-yl, indazolyl, benzimidazolyl, benzo [b] thiazolyl, purinyl, imidazo (2,1-b) thiazolyl, quinolinyl, isoquinolinyl, quinazoli Neyl, 2,1,3-benzothiazolyl, furanyl, thienyl, pyrrolyyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, oxdiazolyl, triazolyl, thia Diazolyl, benzoxazolyl, benzisoxazolyl, cynolinyl, quinooxalinyl, 1,8-naphthyridinyl, putridinyl, or phthalimidyl;

In the proviso, B is aryl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, indolinyl, indol-4-yl, indol-5-yl, indole-6-yl, indole-7 -Yl, isoindolyl, benzo [b] furan-4-yl, benzo [b] furan-5-yl, benzo [b] furan-6-yl, benzo [b] furan-7-yl, benzo [b ] Thiophen-4-yl, benzo [b] thiophen-5-yl, benzo [b] thiophen-6-yl, benzo [b] thiophen-7-yl, indazolyl, benzimidazolyl, benzo [b] thiazolyl, purinyl, imidazo (2,1-b) thiazolyl, quinolinyl, isoquinolinyl, quinazolinyl, 2,1,3-benzothiazolyl, furanyl, thienyl, Pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, oxadiazoleyl, triazolyl, thidiazolyl, benzoxazolyl, benzisoxazolyl, cynolinyl, quinoox If salinyl, 1,8-naphthyridinyl, putridinyl, or phthalimidyl, the ortho carbon atom for the nitrogen atom of the imine bond Carbon atoms may be substituted by F, -Cl, --Br, I, --CN, one or more of methyl, ethyl or methoxy, and;

Where n is an integer from 1 to 4;

Wherein aryl is —F, —Cl, —Br, —I, —NO ₂ , —CN, straight or branched C ₁ -C ₇ alkyl, straight or branched C ₁ -C ₇ monofluoroalkyl, straight or branched chain C ₁ -C ₇ polyfluoroalkyl, straight or branched C ₂ -C ₇ alkenyl, straight or branched C ₂ -C ₇ alkynyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₇ monofluorocycloalkyl , C ₃ -C ₇ polyfluorocycloalkyl, C ₅ -C ₇ cycloalkenyl, -OR ₄ , SR ₄ , -OCOR ₄ , -COR ₄ , -NCOR ₄ , -CO ₂ R ₄ , -CON (R ₄ Or phenyl or naphthyl including phenyl or naphthyl substituted with one or more of (CH ₂ ) _n -O- (CH ₂ ) _m CH ₃ .

In certain embodiments, A is aryl, heteroaryl, or heteroaryl (C ₁ -C ₆ ) alkyl;

이다. Wherein aryl is —F, —Cl, —Br, —I, —NO ₂ , —CN, straight or branched C ₁ -C ₇ alkyl, straight or branched C ₁ -C ₇ monofluoroalkyl, straight or branched chain C ₁ -C ₇ polyfluoroalkyl, straight or branched C ₂ -C ₇ alkenyl, straight or branched C ₂ -C ₇ alkynyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₇ monofluorocycloalkyl , C ₃ -C ₇ polyfluorocycloalkyl, C ₅ -C ₇ cycloalkenyl, -N (R ₄ ) ₂ , -OR ₄ , SR ₄ , -OCOR ₄ , -COR ₄ , -NCOR ₄ , -CO ₂ R ₄ , -CON (R ₄ ) or (CH ₂ ) _n -O- (CH ₂ ) _m CH ₃ . A may be aryl, heteroaryl, heteroaryl (C ₁ -C ₆ ) alkyl or-(CH ₂ ) _n -CC-R ₄ ; Wherein aryl is substituted with OH. In another embodiment, when A is A ', A' is

to be.

In other embodiments, each Y ₁ , Y ₂ , Y ₃ , and Y ₄ is independently H; Straight or branched C ₁ -C ₇ alkyl, CF _3- , —F, —Cl, —Br, or —I, —OR ₄ , —N (R ₄ ) ₂ , or —CON (R ₄ ) ₂ , Where each R ₄ Is independently H; Straight or branched C ₁ -C ₇ alkyl, CF _3- , phenyl; Wherein A is A ′, straight or branched C ₁ -C ₇ alkyl, aryl, heteroaryl, aryl (C ₁ -C ₆ ) alkyl or heteroaryl (C ₁ -C ₆ ) alkyl;

이다.
Where A 'is

to be.

In certain embodiments, B is C ₃ -C ₇ cycloalkyl or adamantyl. In another embodiment, B is pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, indolinyl, indol-4-yl, indol-5-yl, indole-6-yl, indole-7- 1, isoindolyl, benzo [b] furan-4-yl, benzo [b] furan-5-yl, benzo [b] furan-6-yl, benzo [b] furan-7-yl, benzo [b] Thiophen-4-yl, benzo [b] thiophen-5-yl, benzo [b] thiophen-6-yl, benzo [b] thiophen-7-yl, indazolyl, benzimidazolyl, benzo [ b] thiazolyl, purinyl, imidazo [2, l-b] thiazolyl, quinolinyl, isoquinolinyl, quinazolinyl, 2,1,3-benzothiazolyl, furanyl, thienyl, pi Rollyl, oxazolyl, thiazolyl, imidazoyl, pyrazolyl, isoxazolyl, isothiazolyl, oxdiazolyl, triazolyl, thidiazolyl, benzoxazolyl, benzisoxazolyl, cynolinyl, quinooxali Nil, 1,8-naphthyridinyl, putridinyl, or phthalimidyl. In another embodiment, B is aryl or phenyl, and phenyl is F, -Cl, --Br, -CF ₃ , straight or branched C ₁ -C ₇ alkyl, --OR ₄ , -COR ₄ , -NCOR Optionally substituted with one or more of ₄ , —CO ₂ R ₄ , or —CON (R ₄ ) ₂ .

Further examples include compounds having the following structures:

Further examples include compounds having the following structure or pharmacologically acceptable salts thereof:

Wherein each Y ₁ , Y ₂ , Y ₃ , and Y ₄ are independently H; Straight or branched C ₁ -C ₇ alkyl, monofluoroalkyl or polyfluoroalkyl; Straight or branched C ₂ -C ₇ alkenyl or alkynyl; C ₃ -C ₇ cycloalkyl or C ₅ -C ₇ cycloalkenyl; -F, -Cl, -Br, or -I; -NO ₂ ; -N ₃ ; -CN; -OR ₄ , -SR ₄ , -OCOR ₄ , -COR ₄ , -NCOR ₄ , -N (R ₄ ) ₂ , -CON (R ₄ ) ₂ , or -COOR ₄ ; Aryl or heteroaryl; Or any of Y ₁ , Y ₂ , Y ₃ , and Y ₄ on adjacent carbon atoms Two of which may constitute a methylenedioxy group, where each R ₄ Is independently H; Straight or branched C ₁ -C ₇ alkyl, monofluoroalkyl or polyfluoroalkyl; Straight or branched C ₂ -C ₇ alkenyl or alkynyl; C ₃ -C ₇ cycloalkyl, C ₅ -C ₇ cycloalkenyl, aryl or aryl (C ₁ -C ₆ ) alkyl; Wherein A is A ′, straight or branched C ₁ -C ₇ alkyl, aryl, heteroaryl, aryl (C ₁ -C ₆ ) alkyl or heteroaryl (C ₁ -C ₆ ) alkyl;

Where A 'is

or

이며;

Is;

Wherein R ₁ and R ₂ are each independently H, straight or branched C ₁ -C ₇ alkyl, —F, —Cl, Br, I, —NO ₂ , or —CN;

Wherein R ₃ is H, straight or branched C ₁ -C ₇ alkyl, F, —Cl, —Br, I, —NO ₂ , —CN, —OR ₆ , aryl or heteroaryl;

Wherein R ₅ is straight or branched C ₁ -C ₇ alkyl, N (R ₄ ) ₂ , —OR ₄ or aryl;

Wherein R ₆ is straight or branched C ₁ -C ₇ alkyl or aryl;

Wherein B is aryl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, indolinyl, indol-4-yl, indol-5-yl, indol-6-yl, indol-7-yl, Isoindolyl, benzo [b] furan-4-yl, benzo [b] furan-5-yl, benzo [b] furan-6-yl, benzo [b] furan-7-yl, benzo [b] thiophene -4-yl, benzo [b] thiophen-5-yl, benzo [b] thiophen-6-yl, benzo [b] thiophen-7-yl, indazolyl, benzimidazolyl, benzo [b] Thiazolyl, purinyl, imidazo [2, lb] thiazolyl, quinolinyl, isoquinolinyl, quinazolinyl, 2,1,3-benzothiazolyl, furanyl, thienyl, pyrroyl, oxazolyl , Thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, oxadiazolyl, triazolyl, thiadiazoleyl, benzoxazolyl, benzisoxazolyl, cynolinyl, quinooxalinyl, 1, 8-naphthyridinyl, putridinyl, or phthalimidyl; In the proviso, the ortho carbon atom or carbon atom for the nitrogen atom of the imine bond is one or more of F, -Cl, --Br, I, --CN, methyl, ethyl or methoxy. May be substituted;

Where n is an integer from 1 to 4;

Wherein aryl is —F, —Cl, —Br, —I, —NO ₂ , —CN, straight or branched C ₁ -C ₇ alkyl, straight or branched C ₁ -C ₇ monofluoroalkyl, straight or branched chain C ₁ -C ₇ polyfluoroalkyl, straight or branched C ₂ -C ₇ alkenyl, straight or branched C ₂ -C ₇ alkynyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₇ monofluorocycloalkyl , C ₃ -C ₇ polyfluorocycloalkyl, C ₅ -C ₇ cycloalkenyl, -OR ₄ , SR ₄ , -OCOR ₄ , -COR ₄ , -NCOR ₄ , -CO ₂ R ₄ , -CON (R ₄ Or phenyl or naphthyl including phenyl or naphthyl substituted with one or more of (CH ₂ ) _n -O- (CH ₂ ) _m CH ₃ . In certain embodiments, each Y ₁ , Y ₂ , Y ₃ , and Y ₄ is independently -H; Straight or branched C ₁ -C ₇ alkyl, -CF ₃ , 1F, --Cl, --Br, I, -OR ₄ , -N (R ₄ ) ₂ , or -CON (R ₄ ) ₂

이다. In certain embodiments, A is aryl, heteroaryl, heteroaryl (C ₁ -C ₆ ) alkyl or-(CH ₂ ) _n -CC-R ₄ ; Wherein aryl is substituted with -OH. In another embodiment, A is aryl, heteroaryl, or heteroaryl (C ₁ -C ₆ ) alkyl; And wherein aryl is F, -Cl, -Br, I, -NO ₂ , -CN, straight or branched C ₁ -C ₇ alkyl, straight or branched C ₁ -C ₇ monofluoroalkyl, straight or branched C _1- C ₇ polyfluoroalkyl, straight or branched C ₂ -C ₇ alkenyl, straight or branched C ₂ -C ₇ alkynyl, C ₃ -C ₇ cycloalkyl, C ₃ -C ₇ monofluorocycloalkyl, C ₃ -C ₇ polyfluorocycloalkyl, C ₅ -C ₇ cycloalkenyl, -N (R ₄ ) ₂ , -OR ₄ , -SR ₄ , -OCOR ₄ , -COR ₄ , -CO ₂ R ₄ ,- Is substituted with CON (R ₄ ) ₂ or (CH ₂ ) _n -O- (CH ₂ ) _m CH ₃ . A may also be aryl or aryl (C ₁ -C ₆ ) alkyl. In another embodiment, A is A ', straight or branched C ₁ -C ₇ alkyl, aryl, heteroaryl, arylC ₁ -C ₆ ) alkyl or heteroarylC ₁ -C ₆ ) alkyl; A '

to be.

The compounds described herein may be purely isomeric and / or diastereomeric. The compounds may also be pure Z isomers or pure Z alkene isomers or pure E isomers or pure E alkene isomers.

Additional examples include:

Further examples include compounds having the formula: or pharmacologically acceptable salts thereof:

Wherein R ₁ , R ₂ , R _d , and R ₄ are each independently hydrogen, hydroxy, halo, cyano, -CONR _a R _b , -NR _a R _b , hydroxy (C ₁ -C ₆ ) alkyl, aryl, heteroaryl, heterocycle, amino (C ₁ -C ₆₎ alkyl, optionally substituted with up to five flow rise (C ₁ -C ₆₎ alkyl, and optionally substituted with up to five flow rise (C ₁ -C ₆ ) alkoxy;

Each R _a and R _b is independently hydrogen, (C ₁ -C ₆ ) alkyl, aryl, (C ₁ -C ₆ ) alkylOC (O) —, or arylOC (O) —, or R _a and R _b , together with the nitrogen attached thereto, forms a heterocycle group optionally substituted with one or more R _d ; Wherein the heterocycle group optionally comprises one or more groups selected from the group consisting of O (oxygen), S (O) _z , and NR _c ;

Each z is an integer selected from 0, 1, and 2;

Each R _c is independently hydrogen, (C ₂ -C ₆ ) alkenyl, (C ₂ -C ₆ ) alkynyl, -C (O) O (C ₁ -C ₆ ) alkyl, -C (O) Oaryl , (C ₁ -C ₆ ) alkoxy (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkyl0 (CH ₂ ) _m , hydroxy (C ₁ -C ₆ ) alkyl, aryl, heteroaryl, hetero Cycle, aryl O (C ₁ -C ₆ ) alkyl, -C (O) NR _g (C ₁ -C ₆ ) alkyl, -C (O) NR _g aryl, -S (O), (C ₁ -C ₆ ) Alkyl, -S (O), aryl, -C (O) (C ₁ -C ₆ ) alkyl, aryl C (O)-, (C ₁ -C ₆ ) alkyl optionally substituted with up to 5 fluoro Or (C ₁ -C ₆ ) alkoxy optionally substituted with up to 5 fluoro;

Each m is an integer selected from 2, 3, 4, 5, and 6;

Each R _d is independently of hydrogen, halo, oxo, hydroxy, -C (0) NR _a R _b , -NR _a R _b , hydroxy (C ₁ -C ₆ ) alkyl, aryl, aryl (C _1- C ₆ ) alkyl, (C ₁ -C ₆ ) alkyl optionally substituted with up to 5 fluoro, and (C ₁ -C ₆ ) alkoxy optionally substituted with up to 5 fluoro;

R _c and R _r are each independently hydrogen, (C ₁ -C ₆ ) alkyl, aryl, -S (O), (C ₁ -C ₆ ) alkyl, -S (O), aryl, -CONR _g (C _1- C ₆ alkyl), (C ₁ -C ₆ ) alkyl C (O)-, aryl C (O)-, (C ₁ -C ₆ ) alkylOC (O)-, and arylOC (O)- Selected from the group consisting of;

R _g is hydrogen or (C ₁ -C ₆ ) alkyl;

R ₅ and R ₆ are independently selected from the group consisting of hydrogen, (C ₁ -C ₆ ) alkyl, (C ₂ -C ₆ ) alkenyl, (C ₂ -C ₆ ) alkynyl, and aryl, or R ₅ and R ₆ together with the nitrogen attached thereto form a heterocycle group optionally substituted with one or more R _d ; Wherein the heterocycle group optionally comprises one or more groups selected from the group consisting of O (oxygen), S (O) _z , and NR _c ;

R ₇ is hydrogen, hydroxy, halo, hydroxy (C ₁ -C ₆ ) alkyl, optionally substituted with up to 5 fluoro (C ₁ -C ₆ ) alkyl, and up to 5 fluoro Selected from the group consisting of (C ₁ -C ₆ ) alkoxy;

Ar is aryl, or heteroaryl, each optionally substituted with one or more R ₈ ; And each R ₈ is independently hydrogen, halo, CF ₃ , CF ₂ H, hydroxy, cyano, nitro, (C ₁ -C ₆ ) alkyl, hydroxy (C ₁ -C ₆ ) alkyl, (C _1- C ₆ ) alkoxy, —NR _a R _b , aryl, heteroaryl or heterocycle.

Other examples include compounds having the formula and pharmacologically acceptable salts thereof:

R ₅ and R ₆ may, for example, together with the nitrogen attached thereto, form a piperidinyl, pyrrolidinyl, morpholinyl, or thiomorpholinyl ring in the compound of the above formula.

Another example includes a compound having the formula and a pharmacologically acceptable salt thereof:

Wherein: X is N (R _C ), O (oxygen), C (R _d ) ₂ , or S (O) _z ;

z is an integer selected from 0, 1, and 2;

Each R _d is independently hydrogen, halo, oxo, hydroxy, -C (O) NR _a R _b , -NR _a R _b , hydroxy (C ₁ -C ₆ ) alkyl, aryl, aryl (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkyl optionally substituted with up to 5 fluoro, and (C ₁ -C ₆ ) alkoxy optionally substituted with up to 5 fluoro; And

n is an integer selected from the group consisting of 0, 1, and 2;

If n = 0, X is C (R _d ) ₂ .

Another example includes a compound having the formula and a pharmacologically acceptable salt thereof:

Another example includes a compound having the formula and a pharmacologically acceptable salt thereof:

Where n is 0, 1, or 2.

In certain embodiments, R ₂ is methyl, fluorine, or OMe. In certain embodiments, R ₃ is methyl, fluorine, or OMe. In certain embodiments, R ₂ and R ₃ are fluorine. In other embodiments, R ₂ and R ₃ are methyl.

Another example includes a compound having the formula and a pharmacologically acceptable salt thereof:

Another example includes a compound having the formula and a pharmacologically acceptable salt thereof:

Wherein each Y is independently N or C (R ₈ ). In certain embodiments, R ₅ and R ₆ together with the nitrogen attached thereto, for example, will form a piperidinyl, pyrrolidinyl, morpholinyl, or thiomorpholinyl ring in a compound of the formula Each of which is optionally substituted with one or more R _d .

Another example includes a compound having the formula and a pharmacologically acceptable salt thereof:

In certain embodiments, n can be 0, 1 or 2. In certain embodiments, R ₂ can be methyl, fluorine or OMe. In certain embodiments, R ₃ can be methyl, fluorine or OMe. In certain embodiments, R ₂ and R ₃ can be fluorine. In still other embodiments, R ₂ and R ₃ can be methyl.

Another example includes a compound having the formula and a pharmacologically acceptable salt thereof:

Examples also include compounds having the formula and pharmacologically acceptable salts thereof:

Wherein: R ₁ is H, (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkoxy, (C ₁ -C ₆ ) alkanoyl, (C ₁ -C ₆ ) alkoxycarbonyl, (C _1- C ₆ ) alkanoyloxy, (C ₃ -C ₈ ) cycloalkyl, (C ₃ -C ₈ ) cycloalkyl ((C ₁ -C ₆ ) alkyl, halo (C ₁ -C ₆ ) alkyl, aryl, aryl ( C ₁ -C ₆ ) alkyl, aryl (C ₁ -C ₆ ) alkoxy, aryl (C ₁ -C ₆ ) alkanoyl, het, het (C ₁ -C ₆ ) alkyl, het (C ₁ -C ₆ ) alkoxy Or het (C ₁ -C ₆ ) alkanoyl;

n is 1 or 2;

m is 1 or 2;

W is O, S, or two hydrogens;

X is 0 or NYR ₄ ;

Y is a direct bond, -CH _2- , -C (= 0)-, -C (= S)-, -0-, -C (= 0) O-, -OC (= 0)-, -C ( = 0) NRa-, -S-, -S (= 0) -, -S (= 0) 2 -, or _{-S (= 0) 2 NR a} - , and the;

R ₄ is H, (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkoxy, (C ₁ -C ₆ ) alkanoyl, hydroxy, (C ₃ -C ₈ ) cycloalkyl, (C _3- C ₈ ) cycloalkyl (C ₁ -C ₆ ) alkyl, halo (C ₁ -C ₆ ) alkyl, hydroxy (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkoxycarbonyl, carboxy, aryl, Aryl (C ₁ -C ₆ ) alkyl, het, NR _d R _e , —C (═O) NR _d R _e , NRR _d R _e (C ₁ -C ₆ ) alkyl, or het (C ₁ -C ₆ ) Alkyl;

R _a is H, (C ₁ -C ₆ ) alkyl, (C ₃ -C ₈ ) cycloalkyl, (C ₁ -C ₆ ) alkoxy (C ₂ -C ₆ ) alkyl, or (C ₃ -C ₈ ) cyclo Alkyl (C ₁ -C ₆ ) alkyl;

Z is (C ₁ -C ₆ ) alkyl, halo (C ₁ -C ₆ ) alkyl, (C ₃ -C ₈ ) cycloalkyl, (C ₃ -C ₈ ) cycloalkyl (C ₁ -C ₆ ) alkyl, ( From the group consisting of C ₁ -C ₆ ) alkoxy, halo (C ₁ -C ₆ ) alkoxy, (C ₃ -C ₈ ) cycloalkyloxy, and (C ₃ -C ₈ cycloalkyl (C ₁ -C ₆ ) alkoxy Phenyl ring independently substituted with one or more substituents selected; or

Z is a phenyl ring fused to a ring system comprising 3 to 8 elements selected from saturated, partially unsaturated, or aromatic, monocycle or bicycle carbon, oxygen and NR _b , wherein mono- or The bicycling system is optionally substituted with one or more R _b , wherein the phenyl ring fused to the mono- or bicycling system is selected from (C ₁ -C ₆ ) alkyl, halo (C ₁ -C ₆ ) alkyl, (C ₃ -C ₈ ) cycloalkyl, (C ₃ -C ₈ ) cycloalkyl (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkoxy, halo (C ₂ -C ₆ ) alkoxy, (C ₃ -C ₈₎ cycloalkyloxy, and (C ₃ -C ₈₎ cycloalkyl (C ₁ -C ₆₎ alkoxy is optionally substituted with with one or more substituents independently selected;

R _b is absent or H, (C ₁ -C ₆ ) alkyl, (C ₃ -C ₈ ) cycloalkyl, (C ₁ -C ₆ ) alkoxy (C ₂ -C ₆ ) alkyl, or (C ₃ -C ₈ ) Cycloalkyl (C ₁ -C ₆ ) alkyl;

R _c is (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkoxy, (C ₁ -C ₆ ) alkanoyl, (C ₁ -C ₆ ) alkoxycarbonyl, (C ₁ -C ₆ ) alka Noyloxy, (C ₃ -C ₈ ) cycloalkyl, (C ₃ -C ₈ ) cycloalkyl (C ₁ -C ₆ ) alkyl, halo (C ₁ -C ₆ ) alkyl, aryl, aryl (C ₁ -C ₆ ) Alkyl, aryl (C ₁ -C ₆ ) alkoxy, aryl (C ₁ -C ₆ ) alkanoyl, het, het (C ₁ -C ₆ ) alkyl, het (C ₁ -C ₆ ) alkoxy, or het (C ₁ -C ₆ ) alkanoyl;

Each R _d and R _e is independently H, hydroxy, (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkenyl, (C ₂ -C ₆ ) alkynyl, (C ₁ -C ₆ alkoxy , (C ₂ -C ₆ ) alkenyloxy, (C ₂ -C ₆ ) alkynyloxy, (C ₁ -C ₆ ) alkanoyl, (C ₁ -C ₆ ) alkoxycarbonyl, (C ₃ -C ₈ ) Cycloalkyl, (C ₃ -C ₈ ) cycloalkyl (C ₁ -C ₆ ) alkyl, aryl, aryl (C ₁ -C ₆ ) alkyl, NR _f R _g , or aryl (C ₁ -C ₆ ) alkoxy ; And

Each R _f and R _g is independently H, (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkoxy, (C ₁ -C ₆ ) alkanoyl, (C ₁ -C ₆ ) alkoxycarbonyl, (C ₃ -C ₈ ) cycloalkyl, (C ₃ -C ₈ ) cycloalkyl (C ₁ -C ₆ ) alkyl, aryl, aryl (C ₁ -C ₆ ) alkyl, or aryl (C ₁ -C ₆ ) alkoxy Becomes; or

R _f and R _g together with the nitrogen attached thereto form a pyrrolidino, piperidino, piperazino, morpholino, or thiomorpholino ring;

Wherein any aryl or het of R ₁ and R ₄ is (C ₁ -C ₆ ) alkyl, phenyl, (C ₁ -C ₆ ) alkoxy, (C ₁ -C ₆ ) alkanoyl, (C ₁ -C ₆ ) Alkoxycarbonyl, (C ₁ -C ₆ ) alkanoyloxy, (C ₃ -C ₈ ) cycloalkyl, (C ₃ -C ₈ ) cycloalkyl (C ₁ -C ₆ ) alkyl, halo (C ₁ -C ₆ ) Alkyl, (C ₃ -C ₈ ) cycloalkyloxy, (C ₃ -C ₈ ) cycloalkyl (C ₁ -C ₆ ) alkoxy, halo (C ₂ -C ₆ ) alkoxy, cyan, nitro, halo, carboxy or Optionally substituted with one or more substituents independently selected from NR _d R _e , and

Wherein the ring comprising X is optionally substituted at carbon with one or more halo, (C ₁ -C ₆ ) alkyl, or (C ₁ -C ₆ ) alkoxy.

Other examples include compounds having the formula:

Wherein: R ¹ is (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) haloalkyl, or aryl unsubstituted or substituted with one or more R _e ; One of R ² and R ³ is absent, the other is hydrogen, (C ₁ -C ₆ ) alkyl, halo (C ₁ -C ₆ ) alkyl, hydroxy (C ₁ -C ₆ ) alkyl, (C ₃ -C ₈ ) cycloalkyl, amino (C ₂ -C ₆ ) alkyl, or aryl, each of which is unsubstituted or alkyl, halo, haloalkyl or nitro, Het, (C ₃ -C ₈ ) cycloalkyl (C _1- C ₆ ) alkyl, aryl (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) or Het (C ₁ -C ₆ ) alkyl, substituted with one or more groups;

B is aryl, thiophene, heteroaryl, furan or pyrrole;

X is -C (= 0), -C (-S), -C (R ₄ ) ₂ , -C (OH)-, or -S (O) _z ;

Each z is independently 0, 1, or 2;

Y is R ⁴ , -N (R ⁴ ) ₂ , -OR ⁴ , -SR ⁴ , or -C (R ⁴ ) ₃ ;

Each R ⁴ is independently hydrogen, (C ₁ -C ₆ ) alkyl, (C ₂ -C ₆ ) alkenyl, (C ₂ -C ₆ ) alkynyl, (C ₁ -C ₆ ) alkoxy, (C _1- C ₆ ) alkanoyl, (C ₁ -C ₆ ) alkoxycarbonyl, (C ₃ -C ₈ ) cycloalkyl, (C ₃ -C ₈ ) cycloalkyl (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkoxy (C ₂ -C ₆ ) alkyl, hydroxy (C ₂ -C ₆ ) alkyl, cyano (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkylthio (C ₂ -C ₆ ) Alkyl, aryl, aryl (C ₁ -C ₆ ) alkyl, aryloxy (C ₂ -C ₆ ) alkyl, halo (C ₂ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkoxycarbonyl (C ₁ -C ₆ ) alkyl, NR _a R _b , Het, or Het (C ₁ -C ₆ ) alkyl, wherein unsubstituted or one or more or two R ⁴ groups, together with the elements attached thereto, are aryl, Het, or Saturated or unsaturated form a monocycle of 3-8 members or a bicycle ring system of 8-12 members, wherein the ring is one or more selected from carbon atoms and O, S (O) _z , and NR _c Include additional heterozygous elements, where Ring system is optionally optionally substituted by one or more R _d;

Each R _a and R _b is independently hydrogen or (C ₁ -C ₆ ) alkyl;

Each R _c is independently hydrogen, aryl, S (O) ₂ , (C ₁ -C ₆ ) alkanoyl, hydroxy (C ₁ -C ₆ ) alkyl, alkoxy (C ₁ -C ₆ ) alkyl, Het, ( C ₁ -C ₆ ) alkoxycarbonyl or (C ₁ -C ₆ ) alkyl, unsubstituted or substituted with one or more R _e ;

Each R _d is independently halo, hydroxy, cyano, nitro, azido, amino, (C ₁ -C ₆ ) alkylamino, amino (C ₁ -C ₆ ) alkyl, amido, (C ₁ -C ₆ ) Alkyl amido, aryl amido, carboxylic acid, (C ₁ -C ₆ ) alkyl, hydroxy (C ₁ -C ₆ ) alkyl, halo (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) Alkoxy, halo (C ₁ -C ₆ ) alkoxy, (C ₁ -C ₆ ) alkanoyl, (C ₁ -C ₆ ) alkoxycarbonyl, carboxy, (C ₁ -C ₆ ) alkanoyloxy, Het, aryl, Het (C ₁ -C ₆ ) alkyl, or aryl (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkylaryl, sulfonyl, sulfonamido, urea, carbamate; Unsubstituted or substituted with one or more substituents R _e , or two R _d together with an element attached thereto form a ketone or spirocycle carbocycle or heterocycling, or two R _d to Together with the attached element, it forms a bicyclic carbocycle or heterocycle ring, wherein each spirocycle or bicycle ring is unsubstituted or one or more halo, hydroxy, cyano, nitro, azido, (C ₁ -C ₆ ) alkyl, hydroxy (C ₁ -C ₆ ) alkyl, halo (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkoxy, halo (C ₁ -C ₆ ) alkoxy, (C _1- C ₆ ) alkanoyl, (C ₁ -C ₆ ) alkoxycarbonyl, carboxy, (C ₁ -C ₆ ) alkanoyloxy, NR _f R _g , R _f R _g NC (═O)-, phenyl, or phenyl (C ₁ -C ₆ ) alkyl, sulfonyl, sulfonamido, urea, carbamate, wherein R _f And R _g , together with the nitrogen attached thereto, forms a piperidino, pyrrolidino, morpholino, or thiomorpholino ring and is unsubstituted or substituted with one or more substituents R _e ;

Each R _e is halo, hydroxy, cyano, nitro, azido, (C ₁ -C ₆ ) alkyl, Het, aryl, (C ₁ -C ₆ ) alkylHet, (C ₁ -C ₆ ) alkylaryl, (C ₁ -C ₆ ) alkylHet (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkylaryl (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) haloalkyl, (C _1- From C ₆ ) alkoxy, (C ₁ -C ₆ ) haloalkoxy, (C ₁ -C ₆ ) alkanoyl, (C ₁ -C ₆ ) alkoxycarbonyl, carboxy, and (C ₁ -C ₆ ) alkanoyloxy Independently selected;

R ⁵ is H, (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkenyl, (C ₁ -C ₆ ) alkynyl, aryl (C ₁ -C ₆ ) alkyl; And

Each R ⁶ is H, (C ₁ -C ₆ ) alkyl, amino, amido, keto, or aryl (C ₁ -C ₆ ) alkyl, where B is thiophene and R ¹ is trifluoro Methyl, R ² is methyl, R ³ and R ⁶ are absent, R ⁵ is H, X is C (= 0), and Y is N (R ₄ ) ₂ , the two R ⁴ are methyl no.

Further examples include compounds having the formula:

Wherein R ¹ is (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) haloalkyl, or aryl, unsubstituted or substituted with one or more R _e ;

One of R ² and R ³ is absent, the other is hydrogen, (C ₁ -C ₆ ) alkyl, halo (C ₁ -C ₆ ) alkyl, hydroxy (C ₁ -C ₆ ) alkyl, (C ₃ -C ₈ ) cycloalkyl, amino (C ₂ -C ₆ ) alkyl, or aryl, each unsubstituted or substituted with alkyl, halo, haloalkyl or nitro, Het, (C ₃ -C ₈ ) cycloalkyl (C _1- Substituted with one or more groups selected from C ₆ ) alkyl, aryl (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) or Het (C ₁ -C ₆ ) alkyl;

Each Z ¹ , Z ² , and Z ³ is C (R ⁶ ) _P , N (R ⁶ ) _q , O, or S, where Z ¹ is N (R ⁶ ) _q , O, or S, Z ¹ Or at least one of Z ² must be N (R ⁶ ) _q , O, or S;

Each p is independently 0, 1, or 2;

Each q is independently 0 or 1;

X is -C (= 0), -C (= S), -C (R ⁴ ) ₂ , or -S (O) L;

Each z is independently 0, 1, or 2;

Y is R ⁴ , -N (R ⁴ ) _z , -OR ⁴ , -SR ⁴ , or -C (R ⁴ ) ₃ ;

Each R ⁴ is hydrogen, (C ₁ -C ₆ ) alkyl, (C ₂ -C ₆ ) alkenyl, (C ₂ -C ₆ ) alkynyl, (C ₁ -C ₆ ) alkoxy, (C ₁ -C ₆ ) Alkanoyl, (C ₁ -C ₆ ) alkoxycarbonyl, (C ₃ -C ₈ ) cycloalkyl, (C ₃ -C ₈ ) cycloalkyl (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) Alkoxy (C ₂ -C ₆ ) alkyl, hydroxy (C ₂ -C ₆ ) alkyl, cyano (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkylthio (C ₂ -C ₆ ) alkyl, Aryl, aryl (C ₁ -C ₆ ) alkyl, aryloxy (C ₂ -C ₆ ) alkyl, halo (C ₂ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkoxycarbonyl (C ₁ -C ₆ ) Alkyl, NR _a R _b , Het, or Het (C ₁ -C ₆ ) alkyl, wherein each alkyl, aryl, or Het is unsubstituted or substituted with one or more R _d , or two R ⁴ groups are Together with the atoms attached to it form an aryl, Het, or saturated or unsaturated 3-8 membered monocycle or 8-12 membered bicyclic ring system, wherein the ring is optionally substituted with carbon atoms and O, S (O) ₂ , and NR _c selected from One or more additional heteroatoms, wherein each ring system is optionally substituted with one or more R _d ;

Each R _a and R _b is independently hydrogen or (C ₁ -C ₆ ) alkyl;

Each R _c is independently hydrogen, aryl, S (O) ₂ , (C ₁ -C ₆ ) alkanoyl, hydroxy (C ₁ -C ₆ ) alkyl, alkoxy (C ₁ -C ₆ ) alkyl, Het, ( C ₁ -C ₆ ) alkoxycarbonyl or (C ₁ -C ₆ ) alkyl, unsubstituted or substituted with one or more substituents R _e ;

Each R _d is independently halo, hydroxy, cyano, nitro, azido, amino, (C ₁ -C ₆ ) alkylamino, amino (C ₁ -C ₆ ) alkyl, amido, (C ₁ -C ₆ ) Alkyl amido, aryl amido, carboxylic acid, (C ₁ -C ₆ ) alkyl, hydroxy (C ₁ -C ₆ ) alkyl, halo (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) Alkoxy, halo (C ₁ -C ₆ ) alkoxy, (C ₁ -C ₆ ) alkanoyl, (C ₁ -C ₆ ) alkoxycarbonyl, carboxy, (C ₁ -C ₆ ) alkanoyloxy, Het, aryl, Het (C ₁ -C ₆ ) alkyl, or aryl (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkylaryl, sulfonyl, sulfonamide, urea, carbamate, unsubstituted or one Is substituted with one or more substituents R _e , or two R _d together with an atom attached thereto form a ketone or spirocycle carbocycle or heterocycling, or two R _d together with an atom attached thereto To form a bicyclic carbocycle or heterocycle ring Wherein each spirocycle or bicycling is unsubstituted or one or more halo, hydroxy, cyano, nitro, azido, (C ₁ -C ₆ ) alkyl, hydroxy (C ₁ -C ₆ ) alkyl, Halo (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkoxy, halo (C ₁ -C ₆ ) alkoxy, (C ₁ -C ₆ ) alkanoyl, (C ₁ -C ₆ ) alkoxycarbonyl, Carboxy, (C ₁ -C ₆ ) alkanoyloxy, NR _f R _g , R _f R _g NC (═O)-, phenyl, or phenyl (C ₁ -C ₆ ) alkyl, sulfonyl, sulfonamido, urea , Substituted with carbamate, wherein R _f and R _g together with the nitrogen attached thereto form a piperidino, pyrrolidino, morpholino, or thiomorpholino ring and are unsubstituted or one or more substituents Substituted with R _e ;

Each R _e is halo, hydroxy, cyano, nitro, azido, (C ₁ -C ₆ ) alkyl, Het, aryl, (C ₁ -C ₆ ) alkylHet, (C ₁ -C ₆ ) alkylaryl, (C ₁ -C ₆ ) alkylHet (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) alkylaryl (C ₁ -C ₆ ) alkyl, (C ₁ -C ₆ ) haloalkyl, (C _1- From C ₆ ) alkoxy, (C ₁ -C ₆ ) haloalkoxy, (C ₁ -C ₆ ) alkanoyl, (C ₁ -C ₆ ) alkoxycarbonyl, carboxy, and (C ₁ -C ₆ ) alkanoyloxy Independently selected;

R ⁵ is H, (C ₁ -C ₆ ) alkyl, aryl (C ₁ -C ₆ ) alkyl; And

Each R ⁶ is H, (C ₁ -C ₆ ) alkyl, amino, amido, keto, or aryl (C ₁ -C ₆ ) alkyl; In the proviso that X is -C (= 0), Y is N (R ⁴ ) _z , Z ₁ is O, Z ₂ becomes N, and Z ₃ becomes CH, then R ⁴ of Y Not H

Biologically active polypeptide fragments of CREB retain biological activity, but may comprise only a portion of the full length amino acid sequence of CREB. Such fragments can be made by internal deletions as well as carboxy or amino terminal deletions.

Fusion proteins include the CREB protein described herein, which refers to a first moiety linked to a second moiety that does not occur in the CREB protein. The second moiety can be a single amino acid, peptide or polypeptide or other organic moiety, for example an organic moiety or an inorganic moiety, including but not limited to carbohydrates, lipids.

Nucleic acid sequences are defined herein as heteropolymers of nucleic acid molecules. Nucleic acid molecules can be double stranded or single stranded, and can be deoxyribonucleotide (DNA) molecules, such as cDNA or genomic DNA, or ribonucleotide (RNA) molecules. As such, the nucleic acid sequence may, for example, comprise one or more exons in the presence of a suitable intron, as well as one or more appropriate regulatory sequences. In one example, the nucleic acid molecule comprises a single reading frame that encodes the desired nucleic acid product. Nucleic acid sequences can be linked to be operable to appropriate promoters.

Nucleic acid sequences encoding desired CREB proteins, CREB analogs (including CREM, ATF-1), CREB-like molecules, biologically active CREB fragments, CREB fusion proteins, or CREB function modulators may be isolated from nature or derived from native sequences. Deformed or Ausubel et al ., Current Protocol in Molecular Biology, John Wiley & Sons , New York (1998); and Sambrook et al ., Molecular Cloning : A Laboratory Manual , 2 nd edition , Cold Spring Harbor University Press , New York . It can be created from scratch as described in (1989) . Nucleic acids are isolated and fused by methods known in the art, such as, for example, the use and preparation of compatible cloning or restriction sites.

In general, the nucleic acid sequence may be a gene encoding a desired CREB protein, CREB analog, CREB-like molecule, CREB fusion protein, or CREB function modulator. Such genes are generally linked to act on appropriate regulatory sequences that may preferably affect the expression of a CREB protein or CREB function modulator in the CNS. As used herein, “operably linked” is defined as a gene or nucleic acid sequence linked to a regulatory sequence in such a way that expression of the gene (or nucleic acid sequence) is allowed. In general, to be operatively linked does not mean that it is always in contact.

Regulatory sequences include transcriptional promoters, selective action sequences that regulate transcription, sequences that encode appropriate messenger RNA (mRNA) ribosomal binding sites, and sequences that control transcription and translation. In certain embodiments, a recombinant gene (or nucleic acid sequence) encoding a CREB protein, a CREB analog, a CREB-like molecule, a biologically active CREB fragment, a CREB fusion protein, or a CREB function modulator, can be induced or inhibited. Under the control of, a higher degree of control over the product level can be provided.

As used herein, “promoter” refers to the DNA sequence, usually upstream (5 ′) of the coding portion of a structural gene, to recognize and recognize RNA polymerase and other factors that may be needed early in transcription. Regulating the expression of the coding moiety by providing binding sites. Suitable promoters are well known in the art. Exemplary promoters include SV40 and human elongation factor (EFI). Other suitable promoters are readily available in the art (eg Ausubel). et al ., Current Protocol in Molecular Biology , John Wiley & Sons , Inc. , New York (1998); Sambrook et al ., Molecular Cloning: A Laboratory Manual , 2 nd edition , Cold Spring Harbor University Press , New York (1989); And US Pat . No. 5,681,735 ).

Memory substances can enhance CREB pathway function by a variety of mechanisms. For example, memory agents may affect signal induction pathways that induce CREB-dependent gene expression. Induction of CREB-dependent gene expression can be achieved, for example, through up-regulation of positive effectors of CREB function and / or down-regulation of negative effectors of CREB function. Positive effectors of CREB function include adenylate cyclase and CREB activators. Negative effectors of CREB function include cAMP phosphodiesterase (cAMP PDE) and CREB inhibitors.

The memory material may enhance CREB pathway function by acting biochemically upstream of the active or inhibitory substance form of the CREB protein or directly on them and / or directly on the CREB protein, including the transcriptional complex. . For example, CREB pathway function can be achieved transcriptionally, by post-transcriptional or by increasing transcription and post-transcriptional CREB protein levels; By altering the affinity of the CREB protein for other essential components of the transcriptional complex, such as the CREB-binding protein (CBP protein); By altering the affinity of the CREB protein, including the transcriptional complex for DNA CREB response elements in the promoter portion; Or by inducing passive or active immunity against CREB protein allotropes. The specific mechanisms that enhance CREB pathway function are not critical to the practice of the present invention.

The memory material can be administered directly to the animal in a variety of ways. In certain embodiments, the memory material is administered systemically. Other routes of administration are known in the art and include intravenous, intracerebroventricularly, intrathecal, parenteral, mucosal, implants, including infusions and / or intravenous injections. , Intratraperitoneal, intraoral, intradermal, transdermal (eg, sustained release polymer), intramuscular, subcutaneous, topical, epidural, and the like. Other suitable routes of administration may also be used, such as absorption through epithelial or mucosal linings. Certain memory substances may also be administered by gene therapy, where a DNA molecule encoding a particular therapeutic protein or peptide is administered to the animal via a vector, and the vector expresses the specific protein or peptide in vivo and secretes it at the therapeutic level. Be sure to

As used herein, a vector refers to a nucleic acid vector such as a DNA plasmid, virus or other suitable replicon (eg viral vector). Viral vectors may include retroviruses, adenoviruses, parvoviruses (eg, adeno-associated viruses), coronaviruses, negative strand RNA viruses such as orthomyxoviruses (eg influenza viruses), rhabdoviruses (eg, Rabies and bullous stomatitis virus), paramyxoviruses (such as measles and Sendai), positive strand RNA viruses such as picomavirus and alphaviruses, and adenoviruses and herpesviruses (such as herpes simplex virus types 1 and 2) , Double stranded DNA viruses, including Ibstein-Barr virus, cytomegalovirus), and poxviruses (eg, vaccinia, chicken pox, and canarypox). Other viruses include Norwalk virus, tagavirus, flavivirus, leobivirus, leovirus, parvovavirus, hepadnavirus and hepatitis virus. Examples of retroviruses include avian leukemia-breast, mammalian type C-virus, type B-virus, type D-virus, HTLV-BLV family, lentivirus, spumavirus ( Coffin , JM, Retroviridae : The viruses and their replication , In Fundamental Virology, Third Edition , BN Fields , et al ., Eds ., Lippincott - Raven Publishers , Philadelphia, 1996 ). Other examples include murine leukemia virus, murine sarcoma virus, mouse mammary tumor virus, bovine leukemia virus, feline leukemia virus, feline sarcoma virus, avian leukemia virus, human T-cell leukemia virus, baboon endogenous virus, gibbon tailless leukemia virus , Mason Pfizer monkey virus, monkey immunodeficiency virus, monkey sarcoma virus, Raus sarcoma virus, and lentivirus. Other examples of vectors are described in McVey et al., US Pat. No. 5,801,030, the contents of which are incorporated by reference.

Nucleic acid sequences encoding proteins or peptides (eg, CREB proteins, CREB analogs (including CREM, ATF-1), CREB-like molecules, biologically active CREB fragments, CREB fusion proteins, CREB function modulators) are shown herein. Can be inserted into a nucleic acid vector according to methods generally known in the art (eg, Ausubel et al ., Eds ., Current Protocol in Molecular Biology , John Wiley & Sons , Inc. , New York (1998); Sambrook et al ., Eds ., Molecular Cloning : A Laboratory Manual , 2 nd edition , Cold Spring harbor university Press , New York (1989) ).

The mode of administration is generally the target cell location. In certain embodiments, the mode of administration is administered to neurons.

Memory materials are biologically inert materials such as pharmacologically acceptable surfactants or carriers (eg glycerides), excipients (eg lactose), stabilizers, preservatives, wetting agents, emulsifiers, antioxidants, carriers, diluents and It may be administered with a vehicle. If desired, certain sweetening, flavoring and / or coloring agents may also be added. When various inactive ingredients are combined, the memory material may be referred to as a pharmacological composition.

The memory material may be formulated into a solution, suspension, emulsion or lyophilized powder in association with a pharmacologically acceptable extranasal vehicle. Examples of such vehicles include water, salts, Ringer's solution, isotonic sodium chloride solution, dextrose solution and 5% human serum albumin. Liposomes and water-insoluble vehicles such as fixed oils may also be used. Vehicle or lyophilized powder may include additives to maintain isotonicity (eg sodium chloride, mannitol) and additives to maintain chemical stability (eg buffers and preservatives). Formulations may be stabilized by conventional techniques. Suitable pharmacological carriers are Remington's Pharmaceutical Sciences .

The dose of memory substance administered to the animal is the amount required to affect the change in neurons, especially CREB-dependent gene expression. Dosages to animals, including frequency of administration, may be determined by the pharmacodynamic characteristics, route of administration, and mode of the particular memory material; Prisoner's body size, age, sex, health status, weight, diet; The nature and extent of the condition to be treated or the nature and extent of the memory to be enhanced or controlled, the type of concurrent treatment, the frequency of treatment and the desired effect may vary.

Memory substances can be in single or divided doses (such as a series of doses every few days, weeks, or months) or delayed release, depending on factors such as the nature and severity of symptoms, the type of concurrent treatment, and the desired effect. May be administered. Other treatment protocols or agents can be used with the present invention.

All documents, patents, and patent applications mentioned herein are incorporated by reference in their entirety.

Example  One

A study was conducted to examine the effect of memory material (HT-0712).

The study was conducted using a 6-year-old macaque (about 22 years old, equivalent to about 66 years of age at the start of the study) previously trained to perform specific memory tasks. Memory performance assessment is based on the CANTAB system ( the Cambridge Neuropsychological Test Automated Battery ) and new work.

Differentiation learning-long term reproduction task ( Long Retrieval Task ) A general description of DL LTR )

In this task, animals are provided with two sets of stimuli (eg, one shape and one line) during the daily test session. For each pair of stimuli, one stimulus is optionally designated a positive stimulus (eg, touching the stimulus is compensated for). Each pair of stimuli is given 14 times during a daily session consisting of 28 sessions. Stimulus pairs (shape and line) are provided in a pseudo randomized fashion during each session. Animals must touch the "positive" shape or line to receive sugar pellets accompanied by a pleasant tone. Touching other shapes or lines stops receiving negative tones and sugar pellets. Animals are tested daily until reaching a performance criterion of 85% accurate response during the daily session. Once this criterion is reached, the animal is no longer tested for the next 7 days. At baseline 8 days later, animals are tested again using the same pair of stimuli already learned to assess long-term regeneration of the learned distinctions.

drug injection

All drugs were administered orally with marshmallows while the animals were in specially designed test cages. The vehicle became marshmallow alone.

The treatment schedule is shown in Table 1. Treatment drugs were administered continuously starting with Treatment 1. Each treatment drug (HT-0712 or vehicle) was administered once daily for four days prior to the start of behavioral training, and then during the training session, treatment drug (HT-0712 or vehicle) was administered once daily, 4 before the test. Carried for time. Training and dosing were discontinued for 6 days when each individual animal reached the performance criterion of at least 85% correct response during the daily training session. Then at 8 days after baseline the animals were again tested for the DL LTR task (memory retention test).

A second vehicle study was conducted at the end of the drug trial using the first three animals that completed the 100 mg / kg dose study.

Behavior data analysis

In the total daily trials, the correct response rate and number of days to reach the baseline (at least 85% accurate response in a single test session) were recorded. Accurate response rates were recorded in retention trials for the entire memory retention trial session and sub-analyzed for performance differences in trials 1-14 and trials 15-28.

One-way ANOVA was used to compare the vehicle data with data from drug trials. Post hoc comparisons were made using the Bonferroni t-test.

result

There were no noticeable behavioral or physical side effects after administration of HT-0712 at any dose provided during the study. Five of the six showed improved memory performance as evidenced by the reduction in the number of days to reach the baseline. One (9 years old) did not show improved task performance at any dose of HT-0712 administered. Such non-responder data was removed from statistical analysis.

Vehicle  process

During the first vehicle study, all animals reached baseline for a period of 26.3 ± 4.6 days. The mean baseline reached was an accurate response of 89.3 ± 1.8%. The mean performance in memory retention attempts was an accurate response of 79.7 ± 5.9%. Memory retention trial outcomes were divided into trials 1-14 and trials 15-28, with animals showing an accurate response of 77.5 ± 5.1% and an accurate response of 82.3 ± 7.5%, respectively.

The second vehicle study (after the drug study) was conducted using the first three animals that completed the drug trial. During the second vehicle study, the animals reached baseline over a period of 20.0 ± 4.5 days. These mean baselines were an accurate response of 86.3 ± 1.3%. The performance in memory retention attempts had an accurate response of 64.0 ± 4.0% on average. Memory retention trial outcomes were divided into trials 1-14 and trials 15-28, with animals showing an accurate response of 59.3 ± 8.4% and an accurate response of 69.0 ± 10.4%, respectively.

There was no statistically significant difference between the results of pre and post vehicle trials (t = 1.168, (P = 0.363)).

Memory processing

At the lowest dose of HT-0712 (1 mg / kg) used in this study, animals reached the baseline with a performance level of 91.4 ± 1.8% accurate response over a period of 12.0 ± 3.6 days. The average performance of the animals in memory retention trials was an accurate response of 85.8 ± 4.6%. Memory retention trial outcomes were divided into trials 1-14 and trials 15-28, with animals showing an accurate response of 80.2 ± 6.2% and 91.6 ± 3.4%, respectively.

At the intermediate dose of HT-0712 (10 mg / kg) used in this study, animals reached baseline (90.0 ± 2.6%) over a period of 11.8 ± 2.3 days. The average performance of the animals in memory retention trials was an accurate response of 79.2 ± 3.5%. Memory retention trial outcomes were divided into trials 1-14 and trials 15-28, with animals showing an accurate response of 71.4 ± 3.4% and an accurate response of 87.2 ± 4.8%, respectively.

At the maximum dose of HT-0712 (100 mg / kg) used in this study, animals reached baseline (86.4 ± 0.7%) over a period of 9.0 ± 2.8 days. The average performance of the animals in the memory retention attempt was an accurate response of 65.0 ± 3.5%. Memory retention trial outcomes were divided into trials 1-14 and trials 15-28, with animals showing 58.4 ± 4.1% accurate response and 71.4 ± 6.1% accurate response, respectively.

The effect of treatment days on baseline was significant (repeat measure ANOVA, F (4,3) = 6.327, p = 0.008). There was a significant difference between vehicle and each dose of HT-0712 in post hoc evaluation using the Bonferroni t-test (p <0.050 in each comparison).

There was no significant effect of treatment on performance levels obtained at baseline (F (4,3) = 1.408, p = 0.289).

There was no significant effect of treatment on the performance level obtained in the memory retention test (F (4,3) = 3.199, p = 0.062). The analysis of treatment effects on memory retention trial outcomes at trials 1-14 and 15-28 did not reach statistically significant levels (p = 0.058 and p = 0.167 in turn).

This study was carried out using a new task, the Discriminative Learning-Long Term Regeneration Task (DL LTR), designed to evaluate the effects of experimental memory materials on learning and memory retention in non-human primates.

Vehicle control studies showed that older normal animals required a long time (eg> 20 days) to reach an 85% accurate response in this test. In addition, the increase in performance seen with HT-0712 in post-drug vehicle testing is not due to an increase in the animal's ability or performance in the test because the results of the second vehicle control test are the first vehicle study. Because there is no big difference.

Five of the six animals showed improved performance after HT-0712 administration. Improvements were shown to significantly reduce the number of days required to reach the criterion, which showed that treatment with HT-0712 improved learning ability.

Animal responses can be separated into two general groups. In the first group (animals of age 5 and age 8), the highest dose that improved performance was 1 mg / kg dose of HT-0712. These animals showed a decrease in performance when doses increased from 1 to 10 to 100 mg / kg. In other words, increasing the dose of HT-0712 took longer to reach the baseline (e.g., 4 days after the 1, 10, or 100 mg / kg dose was reached in animals of age 5 , 5 days, 10 days, and 3 days, 12 days, 18 days for animals of age 8). Animals in the second group (age 1, age 2, age 7) showed a more typical dose-response effect, with improved performance as the dose increased (age 1 for doses of 1, 10 and 100 mg / kg). The time to reach the criterion in is 21, 15, and 3 days in turn; the time to reach the criterion at age 2 is 17, 18, and 11 days, and the time to reach the criterion at age 7 is 15 and 9 days. And 3 days).

Currently animals have been tested and evaluated for recognition deficits using both delayed matching to sample (DMTS) and paired associative learning (PAL) tasks. These animals were graded according to their performance level on this task (eg, animals rated from 1 to 6 per task, 1 for maximum performance, 6 for minimum performance) (see Table 10). As such, animals that did not respond to HT-0712 received a grade of 1 for PAL and a grade of 2 for DMTS. The same animal received the best results in the first vehicle study of the current project.

In summary, the results of this study show that HT-0712 significantly improves learning and long-term memory retention of older, non-human primates, using certain systems and methods of the present invention.

Legend

C: standard; RT: memory retention test, RT 1-14 memory retention test attempts 1-14; RT 15-28: Memory retention test attempt 15 to 28

Legend

C: standard; RT: memory retention test, RT 1-14 memory retention test attempts 1-14; RT 15-28: Memory retention test attempt 15 to 28

Per treatment  rough Data

Legend

C: standard; RT: memory retention test, RT 1-14 memory retention test attempts 1-14; RT 15-28: Memory retention test attempt 15 to 28

The ratings for the PAL and DMTS assignments were calculated as follows:

1. PAL: The average number of attempts needed to complete the most challenging level of task (3 stimuli / 3 locations).

2. DMTS: Average number of correct responses to longer delays

Animals are rated 1 to 6 per task, with 1 being the best and 6 being the worst.

Example  2: Single-blind Study in Healthy, Older Humans

Research proposal

Studies using the overall protocol and drug dosing presented in Example 1 can be carried out using humans. As described below, long term retreat tasks may utilize stimuli in the form of face-name and / or working pairs. The human subject can be shown a photograph of the face with its name and / or occupation. Jobs for each name-face pair may be listed. While recognizing, ask the entity if the name matches a job. During the test, human subjects are provided with unique, rearranged, and new face-name pairs (presented without a job) and are asked whether the face and name pairs are unique, rearranged, or new. This test can be used to evaluate a human subject in the same manner as presented in Example 1.

In one embodiment, a single site, double-blind, three crossover study was conducted to test the following three dosage regimens: (1) HT-0712 45 mg po for 7 days; (2) HT-0712 45 mg po + warfarin 2.5 mg po for 7 days; And (3) Warfarin 2.5 mg po. For 7 days. There was a 7 day washout between dosing periods. The study involved three days (72 hours) in the organization during the seven-day feeding period. Seven days of rest The same day, four days were given leave from the clinic. In the subject selection criteria, there were no inclusion and / or exclusion criteria for associative tests.

Study schedule

Group I individuals received 45 mg dose of HT-0712 on Day 1, Day 2, Day 3, Day 4, Day 5, Day 6, and Day 7; 45 mg dose of HT-0712 + 2.5 mg dose of warfarin was given on days 15, 16, 17, 18, 19, 20, and 21; 2.5 mg dose of warfarin was given on 29, 30, 31, 32, 33, 34 and 35 days. Group II subjects received 45 mg dose of HT-0712 + 2.5 mg dose of warfarin on Days 1, 2, 3, 4, 5, 6, and 7; 2.5 mg dose of warfarin was given at 15, 16, 17, 18, 19, 20, and 21 days; And 45 mg doses of HT-0712 were received on 29, 30, 31, 32, 33, 34, and 35 days. Group III subjects received 2.5 mg doses of warfarin at 1, 2, 3, 4, 5, 6, and 7 days; 45 mg doses of HT-0712 were given at 15, 16, 17, 18, 19, 20, and 21 days; And 45 mg dose of HT-0712 + 2.5 mg dose of warfarin was given at 29, 30, 31, 32, 33, 34, and 35 days.

Paired associations ( Paired Associates ) Test

Materials : Materials consisted of 21 Lenovo notebook computers. Each notebook was set up to remove all self-setting software and reduce the settings that interfere with the internal clock settings and E-Run software. E-runs (v1.2.1.68) were installed on all notebook computers to perform associative tests. 120 black-and-white photographs (male and female) were paired with two word occupations, such as a fire-fighter. Black and white facial signatures were obtained from the facial database of the Lund University Cognitive Science Lab. Only two words or compound word occupations were used. The 120 face-name pairs were divided into three groups of 40. The same 40 face-name pairs were used in each round.

Procedure - Timing of Administration : The memory test began about 6 hours after drug administration. All memory tests were completed before the daily blood collection. As can be seen in Table 12, training was conducted in three rounds (or fragments) of 14 days. The test was conducted for 42 days. At one day in a given round, the subjects became familiar with the computer and were trained how to use the test software. On day 2, an entity was provided with an encoding and retrieval session. On days 3-7, individuals were received for daily memory. At 10 days the subjects were tested for half of the face-name pairs, at 14 days the subjects were tested for the remainder of the face-name pairs, and then for all face-name pairs used in that round. All tested.

Training and testing round process Pair of numbers One training 2 encryption 40 pairs 3 Test 40 pairs 4 Test 40 pairs 5 Test 40 pairs 6 Test 40 pairs 7 Test 40 pairs 8 9 10 Test Pair 1-20 11 12 13 14 Test Part 1: 21-40 pairs
Part 2: Repeat all 40 pairs

Test Course Mastery -At one day, the subjects were accustomed to five computers and trained how to use the test software by receiving mock encryption and playback sessions consisting of nine face-name pairs. These pairs were not repeated again.

Encryption and Replay : At two days, each participant received a single encryption and replay session.

Encryption : During an encryption session, you see 40 faces paired with a job at a time on the computer screen. For example, participants see a job “ice skater” on the face and on the right. Their task for each face and job pair is to create a vivid, remembered image of the person in the picture having the job shown on the right. Thus, for a face paired to a skater, participants can imagine the person making a difficult jump and falling or being very pleased by winning a gold medal. Immediately assessing how easy it is to create a memorized image with a -3 rating, 1 is easy, 2 is difficult, 3 does not produce a memorized image, and after 2 seconds, the next face-occupation pair is presented.

Play (Retrieval): In this part of the experiment, participants saw faces 40 during encryption to-remember test (cued-recall test) was performed by the signal for a job pair. During regeneration, only the photographs were presented to test subjects without associating jobs. Next to the face is a box that allows you to type the job paired to that face during encryption. When the general participants typed their responses, they received feedback on whether they were right or wrong. If the response is correct, a "Correct!" Appears on the screen with the appropriate face and occupation pairs, followed by the next stimulus pair. If wrong, the exact face and occupation pairs are presented on the screen for 3 seconds with feedback, followed by the next stimulus pair. The session ends when all 40 face-occupation pairs are presented. This regeneration session is provided once daily on 3-7 days of study.

Long-term memory assessment : 3 days after each last medication and before release (10, 24 and 38 days), participants were assessed by a signal consisting of 20 face-occupation pairs to measure the long-term stability of face-occupational memory. I received a cued-recall test. Seven days prior to each drug treatment (14, 28 and 42 days), participants were tested on the remaining 20 face-occupation pairs that were not previously tested. Twenty face-occupation pairs were provided after one minute of rest at 14, 28 and 42 days. After one minute of rest, participants were presented with all 40 facial occupation pairs.

Results : Data reflecting the participants' memories was collected and graphically represented in FIG. 5. These data show that long-lasting memory of association tasks can be established by sufficient daily repetitive training.

Blinding process (Blinding Procedure): study double-blind, placebo-controlled test was. Pharmacologists are responsible for maintaining the object handling code.

Thus, it can be seen that various methods are provided. Those skilled in the art will recognize that the present invention may be practiced in addition to the various non-limiting embodiments and preferred embodiments provided for the purposes of the present invention, the invention being limited only by the following claims. do. The invention may be practiced by equivalents to the specific embodiments discussed in this description.

While various embodiments of the invention have been described above, these are presented by way of example and not by way of limitation. Similarly, various drawings may illustrate examples of the invention, which may be helpful in understanding the features and functions of the invention. The invention is not limited to the described embodiments, but may be practiced using various other drawings. Moreover, those skilled in the art will recognize how alternative functional, logical or physical shapes are provided to implement the desirable features of the present invention. In addition, for the method claims, the order in which the steps are presented does not imply that the various embodiments execute the specified functions in the same order unless otherwise indicated.

Although the present invention is described in various embodiments and embodiments, the various features, aspects, and functions described in one or more individual embodiments are not intended to limit the application only to the specific embodiments in which they are described, but instead one or more other embodiments of the invention. Yes, it may be applied to various embodiments, whether such embodiments are described or presented as part of the above described embodiments. Accordingly, the scope and inverse of the present invention is not limited by the exemplary embodiments described above.

The terms and phrases used in this document, and variants thereof, should be construed as limiting, as opposed to limiting, unless expressly stated. In an embodiment, "including" should be interpreted as "including without limitation"; "Examples" are used to present exemplary cases of the subject matter discussed, and do not exclude or limit the listed; The singular "a" or "an" should be interpreted as "at least one" or "one or more" or the like; And the terms "conventional", "traditional", "normal", "reference", "known" and similar terms are construed as limiting the items available at a given time period or at a given time. It should not be construed as including a conventional, traditional, common or reference technique that is used or currently known at any time in the present or future instead. Similarly, when referring to techniques that are obvious or known to those skilled in the art, such techniques include those that are apparent or known to those skilled in the art at any point in time, now or in the future.

Groups linked by “and” are not to be interpreted as having to be in each and every group of items, but rather as and “or” unless stated otherwise. Similarly, items concatenated with “or” should not be construed as exclusive to each other in the group, unless otherwise stated otherwise. Moreover, although items, elements, and components of the present invention are described in the singular, plural concepts are also contemplated within the scope of the present invention unless explicitly stated only in the singular.

Broad words and terms, such as “one or more”, “at least”, “not limited to”, or other similar phrases, should not be construed as intended or requiring a narrow range of cases. As will be apparent to those skilled in the art after reading this document, the described embodiments and various alternatives may be practiced without limitation to the described embodiments.

Claims (24)

Evaluating the effect of the memory substance administered to the subject, wherein the evaluating comprises presenting the subject with a series of stimuli, and evaluating the subject's response to the stimulus. The method of claim 1, wherein the stimulus affects the CREB pathway. The method of claim 2, wherein the stimulus affects long-term memory of the subject. The method of claim 1, wherein the evaluation further comprises selecting a performance criterion and evaluating the number of training days required in the individual to reach the performance criterion. The method of claim 1, wherein the series of stimuli comprises a plurality of pairs of stimuli. 6. The method of claim 5, wherein each stimulus pair comprises an element having an associative association with other elements of the pair. 7. The method of claim 6, wherein the associative association is selected from the group consisting of face-name associations and word-word associations. 8. The subject of claim 1, wherein the subject is a mammal, preferably selected from the group consisting of primates such as humans, monkeys, lemurs, macaques or macaques, mice, mice, Way. The method of claim 1, wherein the subject is a plurality of subjects, preferably comprising a control group and an experimental group. The method of claim 1, wherein the memory material is a compound having the structure: or a pharmacologically acceptable salt thereof.
 

Where each Y_One, Y₂, Y₃, And Y₄Is independently H; Straight or branched chain C_One-C₇ Alkyl, monofluoroalkyl or polyfluoroalkyl; Straight or branched chain C₂-C₇ Alkenyl or alkynyl; C₃-C₇ Cycloalkyl or C₅-C₇ Cycloalkenyl; -F, -Cl, -Br, or -I; -NO₂; -N₃; -CN; -OR₄, -SR₄, -OCOR₄, -COR₄, -NCOR₄, -N (R₄)₂, -CON (R₄)₂, Or -COOR₄; Aryl or heteroaryl; Or any Y on adjacent carbon atoms_One, Y₂, Y₃, And Y₄ Two of which may constitute a methylenedioxy group, where each R₄ Is independently H; Straight or branched chain C_One-C₇ Alkyl, monofluoroalkyl or polyfluoroalkyl; Straight or branched chain C₂-C₇ Alkenyl or alkynyl; C₃-C₇ Cycloalkyl, C₅-C₇ Cycloalkenyl, aryl or aryl (C_One-C₆Alkyl; Where A is A ', straight or branched chain C_One-C₇ Alkyl, aryl, heteroaryl, aryl (C_One-C₆Alkyl or heteroaryl (C)_One-C₆Alkyl;
Where A 'is
 

 or

Where R_One And R₂Are each independently H, straight or branched chain C_One-C₇ Alkyl, -F, -Cl, Br, I, -NO₂Or -CN;
Where R₃Is H, straight or branched chain C_One-C₇ Alkyl, F, -Cl, -Br, I, -NO₂, -CN, -OR₆, Aryl or heteroaryl;
Where R₅Is a straight or branched chain C_One-C₇ Alkyl, N (R₄)₂, -OR₄ Or aryl;
Where R₆Is a straight or branched chain C_One-C₇ Alkyl or aryl;
Wherein B is aryl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, indolinyl, indol-4-yl, indol-5-yl, indol-6-yl, indol-7-yl, Isoindolyl, benzo [b] furan-4-yl, benzo [b] furan-5-yl, benzo [b] furan-6-yl, benzo [b] furan-7-yl, benzo [b] thiophene -4-yl, benzo [b] thiophen-5-yl, benzo [b] thiophen-6-yl, benzo [b] thiophen-7-yl, indazolyl, benzimidazolyl, benzo [b] Thiazolyl, purinyl, imidazo [2, lb] thiazolyl, quinolinyl, isoquinolinyl, quinazolinyl, 2,1,3-benzothiazolyl, furanyl, thienyl, pyrroyl, oxazolyl , Thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, oxadiazolyl, triazolyl, thiadiazoleyl, benzoxazolyl, benzisoxazolyl, cynolinyl, quinooxalinyl, 1, 8-naphthyridinyl, putridinyl, or phthalimidyl; In the proviso, the ortho carbon atom or carbon atom for the nitrogen atom of the imine bond is one or more of F, -Cl, --Br, I, --CN, methyl, ethyl or methoxy. May be substituted;
Where n is an integer from 1 to 4;
Where aryl is -F, -Cl, -Br, -I, -NO₂, -CN, straight or branched chain C_One-C₇ Alkyl, straight or branched C_One-C₇ Monofluoroalkyl, straight or branched chain C_One-C₇ Polyfluoroalkyl, straight or branched chain C₂-C₇ Alkenyl, straight or branched chain C₂-C₇ Alkynyl, C₃-C₇ Cycloalkyl, C_One-C₇ Monofluorocycloalkyl, C₃-C₇ Polyfluorocycloalkyl, C₅-C₇ Cycloalkenyl, -OR₄, SR₄, -OCOR₄, -COR₄, -NCOR₄, -CO₂R₄, -CON (R₄) Or (CH₂)_n-O- (CH₂)_mCH₃Phenyl or naphthyl including phenyl or naphthyl substituted by one or more of; Or one or more pharmacologically acceptable salts selected from the group consisting of compounds having the structures:

or
 

Where Me is methyl, cPent is cyclopentyl, and carbons 3 and 5 are in the S configuration. A method comprising evaluating the effect of a memory substance administered to a subject, wherein the assessment selects performance criteria, presents the subject with a series of stimuli to train the subject, and evaluates the subject's response to the stimuli Determining whether the individual has reached the performance criteria, and determining the number of training days required to reach the performance criteria. The method of claim 11, wherein the performance criterion is a ratio that indicates the quality of the subject's response to the stimulus. 13. The method of claim 12, wherein the performance criteria indicate long term memory acquisition of the individual. A method comprising evaluating the effect of a memory substance administered to a subject, wherein the assessment selects performance criteria, presents the subject with a series of stimuli to train the subject, and evaluates the subject's response to the stimuli Determining whether the individual has reached the performance criteria, repeating the stimulus presentation and evaluation until the individual has reached the performance criteria, and determining the number of days of training required to reach the performance criteria. A method comprising selecting a memory substance as a drug posterior substance, wherein the screening comprises administering the memory substance to a non-human subject, providing a series of stimuli to the subject, and evaluating the subject's response to the stimulus. Including, the method. Tests memory materials as long-term memory enhancers,
Wherein the test comprises administering the memory material to a non-human individual; Presenting a series of stimuli to the individual; And evaluating the subject's response to the stimulus. In a system comprising a series of stimuli,
Wherein the series of stimuli allows for evaluating the subject's response to these stimuli and, therefore, evaluating the effect of the memory material administered to the subject or evaluating the effectiveness of a training protocol administered to the subject. In a method of testing a training protocol as a long term memory enhancer,
The test includes subjecting the subject to a training protocol, and providing the subject with a series of stimuli and evaluating the subject's response to the stimulus. A method comprising assessing the effect of a memory impairment agent administered to a subject, wherein the assessment comprises providing the subject with a series of stimuli and assessing the subject's response to the stimulus. A method comprising screening for a memory impairment as a drug candidate, wherein screening comprises administering the memory impairment to a non-human individual, presenting a series of stimuli to the individual, and assessing the individual's response to the stimulus. Assessing the memory impaired substance. In a system comprising a series of stimuli,
Wherein the series of stimuli allows for evaluating the subject's response to these stimuli and, thus, evaluating the effect of the memory impairment agent administered to the subject. 18. The method of any one of claims 11-17, wherein the memory material is a compound having the structure: or a pharmacologically acceptable salt thereof.

Wherein each Y ₁ , Y ₂ , Y ₃ , and Y ₄ are independently H; Straight or branched C ₁ -C ₇ alkyl, monofluoroalkyl or polyfluoroalkyl; Straight or branched C ₂ -C ₇ alkenyl or alkynyl; C ₃ -C ₇ cycloalkyl or C ₅ -C ₇ cycloalkenyl; -F, -Cl, -Br, or -I; -NO ₂ ; -N ₃ ; -CN; -OR ₄ , -SR ₄ , -OCOR ₄ , -COR ₄ , -NCOR ₄ , -N (R ₄ ) ₂ , -CON (R ₄ ) ₂ , or -COOR ₄ ; Aryl or heteroaryl; Or any of Y ₁ , Y ₂ , Y ₃ , and Y ₄ on adjacent carbon atoms Two of which may constitute a methylenedioxy group, where each R ₄ Is independently H; Straight or branched C ₁ -C ₇ alkyl, monofluoroalkyl or polyfluoroalkyl; Straight or branched C ₂ -C ₇ alkenyl or alkynyl; C ₃ -C ₇ cycloalkyl, C ₅ -C ₇ cycloalkenyl, aryl or aryl (C ₁ -C ₆ ) alkyl; Wherein A is A ′, straight or branched C ₁ -C ₇ alkyl, aryl, heteroaryl, aryl (C ₁ -C ₆ ) alkyl or heteroaryl (C ₁ -C ₆ ) alkyl;
Where A 'is

or

Wherein R ₁ and R ₂ are each independently H, straight or branched C ₁ -C ₇ alkyl, —F, —Cl, Br, I, —NO ₂ , or —CN;
Wherein R ₃ is H, straight or branched C ₁ -C ₇ alkyl, F, —Cl, —Br, I, —NO ₂ , —CN, —OR ₆ , aryl or heteroaryl;
Wherein R ₅ is straight or branched C ₁ -C ₇ alkyl, N (R ₄ ) ₂ , —OR ₄ or aryl;
Wherein R ₆ is straight or branched C ₁ -C ₇ alkyl or aryl;
Wherein B is aryl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, indolinyl, indol-4-yl, indol-5-yl, indol-6-yl, indol-7-yl, Isoindolyl, benzo [b] furan-4-yl, benzo [b] furan-5-yl, benzo [b] furan-6-yl, benzo [b] furan-7-yl, benzo [b] thiophene -4-yl, benzo [b] thiophen-5-yl, benzo [b] thiophen-6-yl, benzo [b] thiophen-7-yl, indazolyl, benzimidazolyl, benzo [b] Thiazolyl, purinyl, imidazo [2, lb] thiazolyl, quinolinyl, isoquinolinyl, quinazolinyl, 2,1,3-benzothiazolyl, furanyl, thienyl, pyrroyl, oxazolyl , Thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, oxadiazolyl, triazolyl, thiadiazoleyl, benzoxazolyl, benzisoxazolyl, cynolinyl, quinooxalinyl, 1, 8-naphthyridinyl, putridinyl, or phthalimidyl; In the proviso, the ortho carbon atom or carbon atom for the nitrogen atom of the imine bond is one or more of F, -Cl, --Br, I, --CN, methyl, ethyl or methoxy. May be substituted;
Where n is an integer from 1 to 4;
Wherein aryl is —F, —Cl, —Br, —I, —NO ₂ , —CN, straight or branched C ₁ -C ₇ alkyl, straight or branched C ₁ -C ₇ monofluoroalkyl, straight or branched chain C ₁ -C ₇ polyfluoroalkyl, straight or branched C ₂ -C ₇ alkenyl, straight or branched C ₂ -C ₇ alkynyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₇ monofluorocycloalkyl , C ₃ -C ₇ polyfluorocycloalkyl, C ₅ -C ₇ cycloalkenyl, -OR ₄ , SR ₄ , -OCOR ₄ , -COR ₄ , -NCOR ₄ , -CO ₂ R ₄ , -CON (R ₄ Or phenyl or naphthyl including phenyl or naphthyl substituted with one or more of (CH ₂ ) _n -O- (CH ₂ ) _m CH ₃ ; Or one or more pharmacologically acceptable salts selected from the group consisting of compounds having the structures:

or

Where Me is methyl, cPent is cyclopentyl, and carbons 3 and 5 are in the S configuration. The method of claim 22, wherein the stimulus affects the CREB pathway. The method of claim 23, wherein the stimulus affects long-term memory of the subject.

Images