Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving oxidoreductase
Reexamination Certificate
2000-03-23
2004-05-25
Eyler, Yvonne (Department: 1646)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving oxidoreductase
C435S006120, C435S007100, C435S007210, C435S069100, C436S501000, C530S350000, C536S023100, C514S002600
Reexamination Certificate
active
06740500
ABSTRACT:
BACKGROUND OF THE INVENTION
The present invention relates to methods of identifying or screening for drugs or agents useful in treatment of psychiatric or neurological disorders, and relates in particular to screening for non-steroidal drugs or agents.
The present invention is based, in part, on experimental work, some of which is reported in the following articles, the disclosures of which are incorporated herein by reference:
(a) Cheney, D. L., Uzunov, D., Costa, E., Guidotti, A. (1995a); Gas chromatographic mass fragmentographic quantitation of 3&agr;-hydroxy-5&agr;-pregnan-20-one (allopregnanolone) and its precursors in blood and brain of adrenalectomized and castrated rats.
J. Neurosei,
15:4641-4650.
(b) Cheney, D. L., Uzunov, D., Guidotti, A. (1995b): Pregnanolone sulfate antagonizes dizocilpine amnesia: Role for allopregnanolone.
NeuroReport
6:1697-1700.
(c) Guidotti, A., Uzunov, D., Auta, J., Costa, E. (1996): Application of gas chromatography mass fragmentography with negative ion chemical ionization technology to measure neurosteroids and their biosynthesis rate in the rat brain In: Genazzani, A. R., Petraglia, F., Purdy, R. H., editors.
The Brain: Source and Target for Sex Steroid Hormones
. New York:Parthenon, pp 24-41.
(d) Uzunov, D. P., Cooper, T. B., Costa, E., Guidotti, A. (1996): Fluoxetine-elicited changes in brain neurosteroid content measured by negative ion mass fragmentography.
Proc. Natl. Acad. Sci. USA
93:12599-12604.
(e) Uzunova, V., Sheline, Y., Davis, J. M., Rasmusson, A., Uzunov, D. P., Costa, E., Guidotti, A. (1998): Increase in cerebrospinal fluid content of neurosteroids in patients with unipolar major depression who are receiving fluoxetine or fluvoxamine.
Proc. Natl. Acad. Sci. USA
95: 3239-3244.
(f) Matsumoto, K., Uzunova, V., Pinna, G., Taki, K., Uzunov, D. P., Watanabe, H., Mienville, J. M., Guidotti, A., Costa, E. (1999): Permissive role of brain allopregnanolone content in the regulation of pentobarbital-induced righting reflex loss.
Neuropharmacol,
38(7):955-964.
It is becoming increasingly clear that naturally-occurring steroidal modulators of neuronal function do exist in the brain acting as very potent endogenous regulators of the function of the brain's most abundant inhibitory (y-aminobutyric acid) or excitatory (glutamatergic) neurotransmitter systems. The central nervous system (CNS) has the capacity to synthesize such steroids de novo from cholesterol independently of peripheral hormonal sources. These steroids are referred to herein as “neurosteroids.” Neurosteroids that are pharmacologically potent modulators of neuronal activity are defined herein as neuroactive steroids. The pharmacological response induced by some neurosteroids is similar to that of the benzodiazepines (Valium-like drugs), and in non-technical language are likely to be the brain's own naturally occurring Valium-like compounds. Specifically, the neurosteroid 5&agr;-pregnan-3&agr;-ol-20-one (allopregnanolone, ALLO) at low nanomolar concentrations positively modulates &ggr;-aminobutyric acid type A (GABA
A
) receptor function, rapidly affecting brain excitability.
An example of biosynthesis of neurosteroids from cholesterol is illustrated in
FIG. 1
, which shows the major metabolic pathway for neurosteroid biosynthesis in the CNS. Pregnenolone (PREG) is formed from cholesterol by the cytochrome P
45O
side chain cleavage (P
450SCC
) enzyme. PREG is transformed into progesterone (PROG) by 3&bgr;-hydroxysteroid dehydrogenase isomerase (3&bgr;-HSD) and PROG is transformed into 5&agr;-dihydroprogesterone (5&agr;-DHP) by 5&agr;-reductase. Finally, ALLO can be synthesized from 5&agr;-dihydroprogesterone. (5&agr;-DHP) by the action of the 3&agr;-hydroxysteroid oxidoreductase (3&agr;-HSOR) enzyme. This enzyme has two reversible functions—oxidative and reductive. The reduction of 5&agr;-DHP to ALLO is dependent upon the availability of the cofactors NADPH or NADH, whereas the opposite step, which catalyzes the oxidation of ALLO to 5&agr;-DHP is NAD/NADP dependent. Currently, the major pathway of ALLO metabolism and deactivation in the brain is thought to be the degradation of ALLO to 5&agr;-DHP by the oxidative action of the 3&agr;-HSOR. We have demonstrated that ALLO is unevenly distributed in brains of adrenalectomized and castrated (ADX/CX) rats, suggesting the existence of local regulatory mechanisms for its production and metabolism.
Referring to
FIG. 2
, in addition to the conversion to 5&agr;-DHP by the 3&agr;-HSOR, an alternative route for the deactivation of allopregnanolone is through the metabolism to 20&agr;-hydroxyallopregnanolone (5&agr;-pregnan-3&agr;,20&agr;-diol)(20&agr;-hydroxy ALLO) by the action of the 20&agr;-hydroxysteroid dehydrogenase (20&agr;-HSD). The 20&agr;-HSD also metabolizes progesterone to 20&agr;-hydroxyprogesterone (4-pregnen-20&agr;-hydroxy-3-one)(20&agr;-hydroxy PROG), thus decreasing the endogenous pool of progesterone available to be metabolized to 5&agr;-DHP and ultimately to allopregnanolone. Since both actions of the 20&agr;-HSD would result in a significant decrease in the CNS levels of allopregnanolone, non-steroidal compounds that selectively inhibit the 20&agr;-HSD would prevent the 20&agr;-HSD-mediated depletion of the neuroactive steroid allopregnanolone in the brain.
Measurements of allopregnanolone in brain microdialysates obtained from freely moving rats have revealed that allopregnanolone is present in the extracellular compartment. These findings strongly suggest that allopregnanolone is released by glial cells and neurons and it can accumulate in the synaptic cleft in concentrations sufficient to activate the GABA
A
receptor. It remains to be understood whether an active reuptake mechanism involved in maintaining a physiological synaptic concentration of allopregnanolone exists. If such a mechanism does exist, then potential inhibitors of the reuptake of allopregnanolone would increase its accumulation in the vicinity of its target receptor.
Several lines of evidence indicate that a potentiation of serotonergic neurotransmission underlies the therapeutic response to various types of antidepressants. Fluoxetine and other selective serotonin reuptake inhibitors (SSRIs) have a spectrum of clinical efficacy that is different, greater, and quite often superior to that of other antidepressants. Hence, it is becoming increasingly clear that the ability of SSRIs to enhance serotonin activity is not the only mechanism responsible for the large spectrum of favorable clinical actions of the SSRIs.
Furthermore, the etiologies of premenstrual dysphoric disorder (PMDD) and premenstrual syndrome (PMS) are of considerable relevance to neurosteroids as their symptoms (anxiety, mood fluctuations, susceptibility to seizures, etc.) are associated with a precipitous decline in circulating levels of progesterone and its GABA
A
receptor active metabolite ALLO. In a recent study, it has been concluded that subjects with PMS manifest significantly lower levels of serum ALLO in the luteal phase when compared to controls. Interestingly, fluoxetine has been shown to be effective in the treatment of premenstrual dysphoria, displaying a much faster therapeutic response than the usual lag of three to six weeks required for SSRIs to become effective in alleviating symptoms of depression, suggestive that an enhancement of serotonergic neurotransmission is not the only mechanism underlying, the therapeutic action of fluoxetine in premenstrual dysphoria.
However, substantial difficulties prevent a direct therapeutic intervention with neurosteroids. For example, the results of the systemic administration of the 3&agr;,5&agr;-reduced derivatives of PROG, acting as positive allosteric modulators of GABA action at GABA
A
receptors, indicate that the doses required to elicit a clear anxiolytic, antidysphoric, and antiepileptic activity may also produce profound sedation, motor impairment, or ataxia. Furthermore, the endogenous neurosteroids have a very short metabolic half-life and thus a limited bioavailability. All these properti
Davis John M.
Uzunov Doncho P.
Brannock Michael
Eyler Yvonne
Seyfarth Shaw LLP
LandOfFree
Method of screening for non-steroidal neuropsychiatric agents does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Method of screening for non-steroidal neuropsychiatric agents, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Method of screening for non-steroidal neuropsychiatric agents will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-3214843