SK167396A3 - Improved methods for the determination and adjustment of prolactin daily rhythms - Google Patents

Abstract

Disclosed are methods for detecting and adjusting abnormalities in prolactin daily rhythms. Provided is a method comparing a prolactin profile of a vertebrate subject to the prolactin profile of other vertebrate subjects having a normal prolactin profile; and adjusting the prolactin profile of the subject to cause the profile of the subject to conform to (or approach) the normal prolactin profile. Also provided is a method for evaluating the daily prolactin profile of a human subject wherein the method provides for comparing a set of at least two prolactin levels of the human and comparing them to corresponding prolactin levels for healthy humans (or a preestablished average prolactin curve for healthy humans) at the same time points. The method also provides for determining whether a human subject has an abnormal prolactin profile.

Description

(57) Annotation:

Described are methods for determining and adapting abnormalities in daily prolactin rhythms, a method for comparing the prolactin profile of a vertebrate subject to that of other vertebrate subjects having a normal prolactin profile, adjusting the prolactin profile to achieve that subject's normal profile , or approached the normal prolactin profile. A method for assessing the daily prolactin profile of a human subject, the method comprising comparing at least two levels of human prolactin and comparing them to the corresponding healthy human prolactin levels (or a predetermined average healthy human prolactin curve) at the same time points. The method provides for the determination of an abnormal prolactin profile in a human.

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METHOD OF DETERMINING AND ADJUSTING THE DAILY PROLACTIN Rhythms

Technical field

The present invention relates to improved methods of reducing body fat deposition in animal vertebrates or humans, and reducing at least one of the diseases including insulin resistance, hyperinsulinemia, hyperlipidemia, hyperglycemia and other metabolic disease, particularly such disease associated with type II diabetes. The present invention is particularly directed to

(i) normalizing the daily rhythms of prolactin in humans and vertebrates; (ii) diagnosing aberrant daily rhythms of prolactin in humans and vertebrates; and (iii) determining the appropriate adaptation necessary to achieve normalization of these prolactin rhythms.

Such treatment includes daily administration to the patient of at least one prolactin stimulator and / or prolactin inhibitor at a predetermined time of day (when administered once daily only) or at different predetermined times of day (when administered multiple times daily). This therapy usually results in long-term adaptation of aberrant or abnormal prolactin rhythms, so that normal prolactin cycles are confirmed or stimulated, at which point therapy may be discontinued while adaptation persists.

The present invention is also directed to improved methods for modifying a neural oscillator (or neural oscillators) in which (or) the daily prolactin rhythm is expressed or labeled.

Adapting the daily rhythms of prolactin and the daily rhythms of key neural oscillators causes reduction and control over an extended period of time in various metabolic or other disorders.

BACKGROUND OF THE INVENTION

Data on obesity and lipid metabolism disorders will be presented here.

Body fat storage t

Human obesity can be defined as a body weight that exceeds 20% of the desired body weight for individuals of the same sex, height and skeleton (LB Salans in Endocrinology and Metabolism, 2nd Ed., Pp. 1203-1244, McGraw-HilI, New York ( 1987), see also RH Williams, Textbook of Endocrinology, pp. 904-916 (1974) In other animals and in humans, obesity can be determined based on body weight patterns that are consistent with prolactin profiles indicating that sample members who are young, lean and healthy (that is, they have no disorder except metabolic disorders) have a daily level of plasma prolactin profiles that follows a regular pattern with little or no standard deviation.

Obesity or excessive fat deposition is consistent with and may induce various disorders of lipid metabolism, for example hypertension, type II diabetes, atherosclerosis and the like.

Also, in the absence of clinical obesity (as defined above), reduced body fat deposition (especially visceral fat deposition) in humans on a long-term or permanent basis would be significantly beneficial, both cosmetically and physiologically.

Reducing body fat deposition in domestic animals (as well as pet animals), especially on a long-term or permanent basis, would also seem to be of considerable economic benefit to humans, especially since farmed animals supply the bulk of human food and animal fat can eventually store as fat in humans.

While controlled diet and physical exercise can lead to moderate results in reducing body fat deposition, earlier summary work by the inventors of the present invention (including earlier related patent applications and US patents given below) is not actually an effective or practical treatment found on suppression of obesity and other lipid metabolism disorders.

Hyperlipoproteinemia is a condition in which the concentration of one or more cholesterol-bearing lipoproteins or triglycerides (such as chylomicrons, very low-density lipoproteins or VLDL and low-density lipoproteins or LDL) in plasma exceeds the usual limit. This upper limit is usually defined as 95% of the randomly selected population. Elevated levels of these agents are also in a positive correlation with atherosclerosis and often result in a heart attack or stroke, adding to about half of all deaths in the United States. Strong clinical evidence has been presented that consistent with decreasing plasma lipoprotein concentration, the risk of atherosclerosis is reduced (A. Noma et al., Atherosclerosis, 49, 1 (1983), D. Illingworth and W. Conner in Endocrinology and Metabolism, McGraw- Hil, New York (1987)). Such a significant amount of research is devoted to finding treatments that reduce plasma cholesterol and triglyceride levels.

Another set of plasma lipoproteins found in vertebrates are high density lipoproteins or HDL. HDL is used to remove free cholesterol from plasma. High concentrations of HDL, as a percentage of total plasma cholesterol, are associated with a reduction in the risk of atherosclerosis and heart disease. Thus HDLs are known in the professional press as good cholesterol. Therefore, the therapeutic strategy involves attempts to reduce both LDL and VLDL in plasma (i.e., reduce total plasma cholesterol) and increase the HDL fraction of total plasma cholesterol. Several lines of research have shown that a simple increase in HDL is beneficial even if there is no reduction in LDL or VLDL (G. P. Bell et al., Atherosclerosis, 36, 47-54 (1980), R. Fears,

Biochem. PharmacoL, 33, 219-228 (1984); G. Thompson, Br. Heart J., 51, 585-588 (1989); H. Blackburn, N. E. J. M., 309, 426-428 (1983)).

Current therapies for hyperlipoproteinemia include a low fat diet and the exclusion of aggravating circumstances such as established lifestyle. If hyperlipoproteinemia is secondary (i.e. due to, for example, insufficiency of lipoprotein lipase or LDL receptor, various endocrine pathologies, alcoholism, kidney disease and liver disease), the control of such secondary diseases is also the center of treatment. Hyperlipoproteinemia is also treated with drugs that usually alter the levels of specific components of total plasma cholesterol as well as lowering the total plasma lipid component. Recently introduced hyperlipoproteinemia drugs include lovastatin (MEVACOR®), which causes selective inhibition of the enzyme involved in the production of cholesterol, 3-hydroxy-3-methylglutaryl coenzyme and (HMGCoA) reductase. This drug specifically lowers total cholesterol and may cause a slight (5 to 10%) increase in HDL concentrations. However, the benefit of this therapy varies from patient to patient.

In addition, the use of the HMG-OoA enzyme inhibitor is associated to some extent with side effects such as liver toxicity, renal myoglobinuria, renal collapse and lenticular inflammation. The risk of such side effects necessitates close monitoring of patients (for example, renal function is tested every month).

Another medicine prescribed for hyperlipoproteinemia is clofibrate. The efficacy of clofibrate also varies from patient to patient, and the use of this medicine is often accompanied by nephrotic syndromes, myalgia, nausea and abdominal pain.

diabetes

Diabetes, one of the most insidious serious diseases, can sustain suddenly or remain undiagnosed for years, attacking blood vessels and nerves.

Diabetics as a group suffer more often from blindness, heart disease, stroke, kidney disease, hearing loss, gangrene and impotence. This disease and its complications account for one third of all doctor visits and diabetes, and its complications are the leading cause of premature death in the United States and the Western world.

Diabetes adversely affects the way the body uses sugars and starches that are converted to glucose during digestion. Insulin, a hormone produced by the pancreas, makes glucose available as a source of energy for body cells. In muscle, adipose and connective tissue, insulin allows glucose to enter cells by acting on cell membranes. Injected glucose is normally metabolized in the liver to carbon dioxide and water (50%), to glycogen (5%), and to fat (30 to 40%), which is stored in fat stores. Fatty acids circulate from adipose tissue, return to the liver for re-synthesis of triacylglycerol and metabolize to ketones, which are used in tissues. Fatty acids are also metabolized by other organs, with the main route of utilization of carbohydrates.

The net effect of insulin is to promote the storage and use of carbohydrates, proteins and fat. Insulin deficiency is a common and serious pathological condition in humans. In insulin dependent diabetes (IDDM or type I), the pancreas produces little or no insulin, and insulin must be injected daily to survive the diabetic. In non-insulin-dependent diabetes (NIDDM or type II), the pancreas retains the ability to produce insulin and may actually produce more than normal amounts of insulin, but the amount of insulin is relatively insufficient or not quite effective due to the resistance of the cells to insulin.

There are widespread abnormalities in both forms of diabetes. In most NIDDM patients in whom abnormalities can be observed, the underlying drawback is that it occurs

1. reduced glucose entry into various peripheral tissues; and

2. increased glucose excretion into the liver.

As a result, extracellular glucose excess and intracellular glucose deficiency occur. There is also a decrease in amino acid entry into the muscle and an increase in lipolysis. Hyperlipoproteinemia is also a complication of diabetes. The cumulative effect of these abnormalities associated with diabetes causes severe damage to blood vessels and nerves.

The ineffectiveness of the treatment was found to control either hyperinsulinemia or insulin resistance before the work of the inventors. Hyperinsulinemia is higher than the normal level of insulin in the blood. Insulin resistance can be defined as a condition in which a normal amount of insulin is produced and achieves a subnormal biological response. In diabetic patients treated with insulin, insulin resistance is considered to be present whenever the therapeutic dose of insulin exceeds the excreted insulin in normal subjects. Insulin resistance also occurs in a situation defined above the normal level of insulin, i.e. hyperinsulinemia, where the blood glucose level present is normal or elevated.

The basic unit of measurement of biological time, an aene rhythm, is present at all levels of the organization. Daily rhythm has been reported in many hormones including adrenal steroids, such as glucocorticosteroids, especially cortisol and prolactin, a pituitary secreted hormone. An earlier article discussing the prior art states that although a correlation has been established between hormonal rhythms and other rhythms, there is little direct evidence that the time of daily presence or peak hormone levels has significant physiological relevance, see Temporal Synergism of Prolactin and Adrenal Steroids, Albert H. Meier, General and Comparative Endocrinology, Supplement 3, 1972, Copyright 1972, Academic Press, Inc. The article states that peak prolactin concentration occurs at different times of the day in poor and fat animals. The article then describes an avian physiological response to prolactin injections given daily over several days. These responses include increasing and decreasing body fat stores depending on the daily time of injection. In addition, the time of day when prolactin injection helps to lose body fat is consistent with the time of day when prolactin is highest in poor birds. In addition, the time when prolactin injections aid in body fat deposition is consistent with the time when proactin is highest in fatty birds. It has been found that prolactin stimulates gain only by injection at certain times of the day and that the time response differs between poor and fat animals.

In an article entitled Circadian and Seasonal Variation of Plasma Insulin and Cortisol Concentrations in the Syrian Hamster, Mesocricetus Auratus by Christopher J. de Souza and Albert H. Meier, Chronobiology International, Vol. 4, no. 2, p. 141-151 (1987) describes a study of daily changes in plasma insulin and cortisol in scotosensitive and scotorefractive Syrian hamsters reared under short and long daylight conditions to determine possible seasonal changes in their daily rhythms. The baseline insulin concentration was found to be greater in females than in male photosensitive hamsters in short daylight. It is reported that the observed differences may be at the expense of the observed strong fat stores in female hamsters maintained under short daylight conditions. The plasma concentrations of cortisol and insulin varied throughout the day in the groups of test animals, but were not equivalent. Daily changes in insulin and cortisol differed significantly with respect to gender, seasonal status and day length. Neither the daily dosage pattern nor the glucose concentration appears to change with seasonal or daytime. It is reported that the time of day or season does not appear to adversely affect glucose levels. It is concluded that the daily rhythms of cortisol and insulin are regulated by various neural pacemaker systems, and that changes in the phase relationships of the daily systems are partly due to seasonal changes in body fat stores. The daily rhythms of prolactin and glucocorticosteroid hormones, such as cortisol, have been found to play an important but far from fully understood role in regulating daily and seasonal changes in body fat stores and in organizing and integrating overall animal metabolism, see Circadian Hormone Rhythms in Lipid Regulation, Albert. H. Meier and John T. Bums, Amer. Zool., 16, 649-659 (1976).

In the related earlier US patent application no. 192,332 co-pending agents, filed May 10, 1988, describe and claim methods for controlling disorders caused by lipid metabolism by administering prolactin (or both prolactin and glucocorticosteroid (GC) to the blood stream of animals or humans on a daily basis in amounts and for a period of time a period sufficient to correct and restore the neural shock oscillation of the prolactin daily rhythm, which then increases insulin sensitivity The injections of prolactin (or prolactin and glucocorticosteroid) are timed to produce a peak in the daily prolactin (or both prolactin and glucocorticosteroid) profiles that with a prolactin peak (or a peak of both prolactin and glucocorticosteroid) in a lean human insulin-sensitive person, to increase insulin sensitivity and reduce body fat deposition. Ola Prolactin O for obese patients to achieve fat if necessary.

In the related earlier U.S. patent application Ser. 463,327 identical authors, filed on January 10, 1990 (and Continuing Part No. 07 / 813,125, filed on December 23, 1991, published as WO 93/12793 on July 8, 1993), a method of restoring and renewing is described and claimed neural phase oscillations of the brain that control both prolactin and glucocorticosteroid in fat animals (or obese man) by administering a dopamine agonist at a predetermined time of the day such that peaks of prolactin (and / or glucocorticosteroid) in fat animals or (obese man) will be phase-shifted to achieve at least one of the following options in a poor animal (or slender) that will increase in body fat, body weight, hyperinsulinemia or hyperglycemia, and / or insulin sensitivity .

In the related earlier U.S. patent application Ser. 719,745 identical authors, filed on June 24, 1991 (published as WO 93/00092 on January 7, 1993), disclose and claim methods for modifying and restoring neural phase brain oscillations that control prolactin levels, including

(a) as the administration of a dopamine agonist to a patient for as long as the peaks of the normal prolactin profile result in a lowering of the prolactin level to a low daily level;

b) thus administering a prolactin stimulator to a patient just prior to peak prolactin levels in normal patients, in order to induce the patient's prolactin profile to mimic the shape and timing of the profile in a lean person suffering from at least one of the aforementioned metabolic disorders.

U.S. Pat. 719 745 also discloses and claims the further administration of a thyroid hormone in patients who are treated with a dopamine agonist and a prolactin stimulator, particularly those patients who are chronically or seasonally hypotroid.

In co-pending U.S. patent application Ser. No. 995,292, filed December 22, 1992 (International Patent Application PCT / US93 / 12701), discloses and claims certain improved methods for diagnosing aberrant prolactin rhythms, for determining alignment to abnormal daily rhythms, and for normalizing abnormal prolactin rhythms.

In co-pending U.S. patent application Ser. No. 171,897, filed Dec. 22, 1993 (International Patent Application PCT / US94 / 14994), discloses and claims an intense composition comprising a prolactin inhibitor that is rapidly released from a patient's blood stream.

In co-pending U.S. patent application Ser. No. 178,569, filed Jan. 7, 1994 (International Patent Application PCT / US95 / 00663), discloses and claims a method of treating various disorders caused by lipid metabolism, including synergistic combinations of diet and administration of a prolactin inhibitor.

Various aspects of the present invention have not been claimed in the earlier applications. In addition, various improvements and advantageous refinements have been made to the application of the working instructions and their detection now increases the effectiveness of the treatment.

SUMMARY OF THE INVENTION

One feature of the present invention relates to a method of normalizing the daily prolactin profile of a first vertebrate subject in need of a step treatment;

- comparing the prolactin profile of the first vertebrate subject to the prolactin profile of other vertebrate subjects having a normal prolactin profile, and

adjusting the prolactin profile of the first vertebrate subject to cause the profile of that first subject as a general adjustment (or stimulation) of the prolactin profile in the second subjects.

Variations of this feature of the invention include;

a) comparing only a set of at least 2 (preferably at least 4, usually 4 to 6) first subject prolactin levels measured at times over at least two (preferably at least three) key day intervals with corresponding prolactin levels in healthy subjects at the same time points during the day and

b) adjusting the first subject's prolactin levels to adjust (or stimulate) the corresponding prolactin levels to that of healthy subjects.

By way of example, the first subject's prolactin levels may be measured at least twice during each early morning key interval and early evening key interval, and at least one or more preferably at least two additional prolactin levels may be measured during the night interval.

A second aspect of the invention relates to a method of assessing the daily prolactin profile of a human subject, comprising the steps of:

(a) a comparison of the subject's prolactin profile, wherein the profile is determined over a continuous period of 24 hours, with the prior determination of a normalized prolactin profile for healthy human subjects; and

(b) detecting an abnormal daily profile of human prolactin, by determining the following indicators:

(i) at any point in the waking hours of the subject (human), the prolactin level is greater than 1 standard mean error (SEM) above the mean prolactin level of healthy subjects (or, in the case of standard waking hours above mean prolactin or normalized prolactin profile) and (ii) at any time during sleep, the subject's prolactin level is less than 1 SEM below the mean prolactin level of normal (healthy) subjects at the same time point.

Preferably, the comparison time point will be the actual prolactin measurement points (i.e., the prolactin levels of the subject to be tested will correspond to blood prolactin levels and no extrapolations are performed) and most preferably the determination will be based on two prolactin measurement points at a single site. In another embodiment, the determination will preferably be based on whether one measured prolactin level is greater than 2 SEM during waking hours or less than 2 SEM during sleep time.

A change to this second aspect of the invention includes:

(a) comparing only a set of at least two levels of human prolactin (measured at predetermined time points during each of at least two key time intervals) with corresponding prolactin levels (e.g. in the prolactin profile) for healthy humans at the same time points throughout the day; and

(b) determining the abnormal prolactin profile in humans by determining the following characteristics:

(i) at any time point during waking hours, the subject's prolactin level is greater than 1 SEM above the corresponding mean prolactin level of healthy subjects (or above the corresponding normalized prolactin profile) at the same time point; and (ii) at any given predetermined time points; that fall into sleep, the subject's prolactin level is less than 1 SEM below the corresponding prolactin level of healthy subjects at night time (or below the corresponding level of normalized prolactin profile at night time) at the same time point.

A third aspect of the invention relates to a method of determining whether an adjustment will be required to normalize or generally to achieve compliance or approach an abnormal prolactin profile or level to a normalized (or healthy) profile or prolactin level, the method comprising:

(a) collection of a plurality of blood samples taken from a subject for a continuous period of time, taking these samples at predetermined time intervals (or at predetermined time points) within that period;

(b) testing the prolactin content of each of the said samples;

(c) plotting the prolactin content of each of said samples versus the time at which said sample was captured over a given period of time to obtain a greater number of experimental points (i.e., a prolactin profile versus time);

(d) formation of a prolactin profile by joining (or otherwise translating a curve) through these experimental points; and

e) comparing the prolactin profile with a predetermined normal prolactin profile.

Steps c) and d) together constitute the degree of expression of the prolactin content as a function of time to form the subject's prolactin profile.

\

Changing this third aspect of the invention involves measuring only the key prolactin levels to create experimental points, omitting step d), and in step e) comparing each experimental point to the corresponding prolactin level at the same time point at a predetermined normalized prolactin profile or prolactin level healthy subjects at the same time.

A fourth aspect of the invention relates to a method of determining an adjustment that causes an abnormal daily prolactin profile of a patient to adapt to or approach a normal daily prolactin profile of an individual, the method comprising:

a) comparison of the patient's prolactin profile with a predetermined normalized prolactin profile for healthy (normal) subjects;

(b) determining whether at least one of the following indicators is achieved:

(i) the patient's prolactin level at any point in time (preferably at any point in the prolactin measurement, more preferably at any two times of prolactin measurement) during waking hours exceeds the corresponding prolactin level of normal (healthy) subjects by at least 1 SEM at the same time point such a patient at any point in time (preferably at any point in the prolactin measurement, more preferably at any two times the prolactin measurement) during sleep is below

-r a corresponding prolactin level of normal healthy individuals of at least 1 SEM at the same time,

(c) determining the time at which at least one agent selected from the group consisting of a prolactin inhibitor and a prolactin stimulator is to be administered to said subject; and

d) selecting an amount of such a stimulator and / or a prolactin inhibitor to adjust the aberrant aberrant prolactin level of the subject so that the prolactin profile of the subject adapts or approaches the prolactin profile of a normal (healthy) subject.

Again, it is preferred that the assay is based on a single time measurement of aberrant prolactin if the change from the normal profile is at least 2 SEM.

A variation of this fourth aspect of the invention includes the assay required in step b) and the selection required in step d) based on comparing the patient's key prolactin levels at predetermined times of day (or night) with corresponding healthy or normal prolactin levels at the same times.

BRIEF DESCRIPTION OF THE DRAWINGS

The present invention is further described with reference to the accompanying drawings, in which:

Fig. 1 shows a graph of mean plasma prolactin levels (ng / ml) for healthy subjects (male: curve M, female: curve F) versus time of day (hours).

In FIG. Figure 2 shows an abnormal basis prolactin profile in a male patient (pre-therapy) (plotted as a thin dashed line), an improved post-treatment prolactin profile (thin line), and a normalized normal prolactin profile (thick line), all graphed . (Units of prolactin are given in nanograms per ml and time of day is expressed in hours).

Fig. 3 is the same type of graphical representation as in FIG. 2, but for another male patient.

Fig. 4 is an illustrative illustration of various types of common aberration profiles a) to d) of prolactin in humans.

The invention is described in detail below.

All patents, patent applications, and references cited herein are hereby incorporated by reference in their entirety, insofar as their description would be physically present in the disclosure of this invention. However, in the event of a conflict, the present description has a decisive influence.

Recently, metabolic disorders, as mentioned above, have been found to be associated with aberrant patterns in daily levels (and fluctuations) of prolactin and neural oscillators, such as those associated with dopamine and serotonin. Healthy (normal) subjects, i.e., lean humans or poor animals of a selected species who do not suffer from such metabolic abnormalities, have highly indicative daily prolactin profiles, which are characterized by low and relatively constant prolactin levels during waking hours (during the day) with followed by a sharp rise to peak during sleep (night) and a subsequent gradual decrease, to a level corresponding to the waking hours, in the morning. As a result, healthy subjects have prolactin levels that are in the 1 SEM of the normalized prolactin M or F level profile of FIG. 1 (depending on whether it is male or female), preferably for at least two levels of prolactin, measured at different times or at 2 SEM of the normalized prolactin profile at at least one measured prolactin level.

In this context, lean or poor means an obese subject that is not abnormally light in weight. In contrast, an obese person is defined as a person whose body weight is more than 20% above the ideal body weight for a given population (R. H. Williams, Textbook of Endocrinology, pp. 904-916 (1974)). A person with abnormal underweight is an individual whose body weight is at least 10% below the ideal body weight. Ideal body weight (IBW) can be determined using standard tabulated age, height, and weight data from the Metropolitan Life Insurance Company.

The subject's prolactin profile sa. from the collected blood samples of the subject at intervals over a continuous period of time (preferably at 1 to 3 hours intervals for a period of about 12 to 24 hours), testing each blood sample for prolactin content, plotting blood sampling time against the amount of prolactin present in each sample; generating experimental points for each sample, and joining these experimental points (or otherwise fitting these experimental points with a curve) to form a prolactin profile.

According to another embodiment (for the vast majority of subjects), a set of only a few required levels of prolactin should be obtained by capturing blood samples from the subject at independently spaced times at least once and preferably at least twice during each of the at least two key intervals during a 24 hour period. One such key interval is early morning (6.00 to 9.00), another key interval is early evening (16.00 to 20.00). Independently spaced samples may be collected, for example, at 7.00 and 8.00 hours during the first interval and at 16.00 and 2.00 hours during the second interval. Desirably, at least one additional sample is taken at night, during a third time interval of 23.00 to 2.00, for example at 23.00 and 24.00. At least one additional optional prolactin measurement may be performed at yet another interval, for example at a late evening interval, from 8:00 pm to 11:00 pm. Prolactin levels achieved over key intervals will be referred to as key prolactin levels.

This alternative method is advantageous in clinical practice to achieve 24-hour prolactin profiles for several reasons. Firstly, such a procedure that avoids overnight hospitalization or staying in the blood center is significantly less expensive than taking blood for 24 hours. Second, a reduced number of blood samples is beneficial for the patient, since it significantly disrupts his daily program. Key prolactin levels are used as indicators of the prolactin profile in patients and as a replacement for them.

Although females generally have higher levels of prolactin and sharper peaks than males, the shape of the normal prolactin profile in both sexes is qualitatively similar and noticeably unchanged from normal to normal (in the same sex) in the same species.

In contrast, individuals suffering from one or more metabolic disorders have (often highly aberrant) daily prolactin profiles and levels. These profiles and prolactin levels not only differ significantly from normal, but may also differ from each other.

The vast majority (at least about 80%) of patients with at least one of the aforementioned metabolic disorders have abnormal daily prolactin profiles, which generally fall within one of the four 24-hour patterns schematically shown in FIG. 4:

a) flat high level (no peak) (in this context, high means that it is higher than the level of prolactin during the day or hours of waking),

b) flat low level (no peak) (in this context, low means so low that it is lower than the level of prolactin during the day or hours of waking),

(c) a peak during the day and a lower level at night (in this context, low means that it is low compared to the normal average prolactin value during sleep, between 1.00 and 4.00 hours); and

d) a peak during the day and a second peak during the night.

According to an alternative method of measuring key prolactin levels, a patient with type a), c) and d) abnormalities should be diagnosed early in the morning and early evening. If the subject has an abnormal prolactin type b) profile as described above, the subject should have one or two overnight blood donations that should not omit an overnight stay at the hospital or blood center.

Hours of waking or day are understood to be the time period in which normal (healthy) people (who do not have shifted work overnight or do not shift alternately) have prolactin levels relatively constant and low (from 7am to 10pm).

As used herein, the term "sleep-time" or "night" refers to the time period in which prolactin levels rise to peak (from 1.00 to 4.00 hours) in normal humans and then decrease.

Normal average prolactin levels between 1.00 and 4.00 hours are from 8.0 to 14.0 nanograms per milliliter in males and from 14.0 to 26.0 nanograms per milliliter in females; 1 to FIG. 4th

A prolactin inhibitor should include substances that directly or indirectly inhibit prolactin in a subject (animal vertebrate or human). Examples of prolactin inhibitors that do not in any way limit their calculation include dopamine agonists such as dopamine and certain prolactin inhibiting compounds that are ergot-related.

Examples of prolactin-inhibiting dopamine agonists include 2-bromo-α-ergocriptine, 6-methyl-8B-carbobenzyloxyamino-ethyl-10α-ergoline, 8acylaminoergolines such as 6-methyl-8a- (N-acyl) amino-9-ergoline and 6-methyl-8a- (N-phenylacetyl) amino-9-ergoline, ergocorine, 9,10-dihydroergocorn, D-2-halo-6-alkyl-substituted ergonines such as D-2-bromo -6-methyl-8-cyanomethylergoline and lizuride. In addition, non-toxic salts of the prolactin-inhibiting compounds which are ergot-related, formed from pharmaceutically acceptable acids, may also be used in the practice of this invention. Bromocriptine or 2-bromo-α-ergocriptine is believed to be particularly suitable for practicing the present invention.

Prolactin stimulators should include substances that directly or indirectly stimulate prolactin. Examples of prolactin stimulators that do not in any way limit their calculation include dopamine antagonists such as metoclopramide, haloperidol, pimozide, phenothiazine, domperidone, sulpiridine, chlorpromazine, and serotonin agonists, such as MAO inhibitors such as pargyline, synthetic, methadone, antiemetics such as metoclopramide, antipsychotics such as estrogens and other agents such as tryptophan and 5-hydroxytryptophan, melatonin, fluxitan and dexfenfluoramine. In addition, non-toxic salts of the aforementioned prolactin stimulating compounds formed from pharmaceutically acceptable salts are suitable in the practice of this invention. Metoclopramide is believed to be particularly useful in practicing the present invention.

A prolactin modulator refers to both prolactin stimulators and prolactin inhibitors as well as both.

Furthermore, the development of a normal prolactin profile and key prolactin levels is discussed.

A statistically significant number of healthy and young people (aged 25-35) are selected as follows:

All subjects are healthy and have a normal daily work and rest schedule (no night services or other shift of night work). Subjects are divided into two different groups according to sex. All subjects must normally sleep (between approximately 23.00 and approximately 7.00). Individuals are healthy in the sense that they do not suffer from physiological disorders or pathological changes. In particular, they are not obese (based on normalized tables of age, height and weight), they are known to have normal plasma insulin levels and are eutyroidic, i.e. they have normal thyroxine and triiodothyronine plasma levels, are thyroxine-free (freethyroxine) and thyroid stimulating hormone and are not affected by cancer, autoimmune diseases, or sexually transmitted diseases.

A statistically significant number is meant to mean at least 3 as the smallest number of samples for which statistical formulas may in general have any meaning. Normally, however, a number of samples of at least 6 (at least 10 is more preferred) is preferred because this number of samples generally reduces the standard error (SE) of the prolactin assay.

Blood is collected from each subject at 1 to 3 hour intervals over 24 hours and daily levels of prolactin, total triiodothyronine (T3), total thyroxine (T4), free-T4 (depleted of T4) and thyroid stimulating hormone (TSH) in plasma are collected. measured in each subject. For example, blood is collected over a period of 24 hours at appropriate time intervals (e.g., every 1 to 3 hours), usually starting at 10.00 and ending at 8.00 the following morning.

Appropriate sampling techniques and assay procedures are well known to one of skill in the art and may be selected from published procedures, such as reported by P. Linkowski et al., In J. Clin. Endocrinol. Metab. 61, 429-438 (1985) and van Cauter and Copinschi, Circadian and Episodic Variations, Martinus Nyhoff, The Hague, The Netherlands, p. 1-25 (1981).

During sampling, all subjects must receive the same diet and maintain the same sleep schedule. Prolactin values are plotted against time of day and a normal prolactin curve including SEM is generated. A mathematical expression can be created to describe the curves and surfaces below these curves.

The result of a normal 24 - hour profile for men of approximately 30 years of age is shown as curve M in FIG. The outcome for women of about 30 years of age is reported as the F curve. The prolactin SEM during waking hours for men is 1.0 to 2.0 ng / ml and about 3.0 ng / ml during sleep. For women, the SEM of prolactin during waking hours is from 1.0 to 3.0 ng / ml and from 3.0 to 6.0 ng / ml during sleep.

According to another embodiment, the curves of FIG. 1 can be used as a normalized prolactin profile (as well as normalized prolactin levels at times corresponding to the key prolactin measurement) or only a set of prolactin levels can be measured in healthy patients at key intervals during the day, for example at least twice early in the morning and at least twice early in the evening; falls once or twice at night.

Further, the determination of the daily profile of the hormone and / or the key levels of prolactin of the affected subjects is described.

The procedure described above can be used to generate a prolactin profile over 24 hours (or a set of key prolactin levels at certain key intervals) for subjects below a clinical evaluation for the therapy of the invention. Subjects are believed to have abnormal daily prolactin rhythms, including those diagnosed as having at least one of the following conditions. Obesity (i.e., more than 20% excess weight, based on age, frame size and sexual characteristics, using tables provided by Metropolitan Ufe Insurance Company of normalized weights by age and age), insulin resistance, hyperglycemia, hyperinsulinaemia, hyperlipidemia or type II diabetes. However, this assessment is not limited to such entities.

Subjects envisaged for therapy should be euthyroid, i.e., have a total T3 level of from about 90 to about 180 ng / dl, a total T4 of from about 4.5 to about 12 g / dl, and free- T4 from about 0.7 to about 1.9 g / dl. If this is not the case, T4 or T3 should be administered (preferably in the morning) before or concurrently with the administration of the prolactin modulator. In other words, it is preferred that the prolactin profile be adjusted in an individual if its T4 and / or T3 levels are normal. Thus, before adjusting the profile or levels of prolactin to a subject by administering a prolactin stimulator or inhibitor, or both, it is preferable to determine that the subject is euthyroid. If the thyroid hormone levels in the subject are below normal (as determined with respect to the above-mentioned normal values), T3 and / or T4 are preferably first administered to the subject until the plasma levels of these hormones have normalized and then treated with a stimulator or a prolactin inhibitor. The amount of thyroid hormone administered corresponds to the dose required to bring the patient into a eutyroid state, usually from about 25 to about 150 pg per patient per day. If it is necessary to achieve appropriate levels of T3 / T4 during the period of administration of the stimulator and / or prolactin inhibitor, administration of T3 / T4 may be continued together with the prolactin modulator therapy means. The hypertyroid subject should also be treated (via the use of antityroid preparations or thyroid ablation) prior to treatment with a prolactin modulator.

Once a daily prolactin level profile has been established for an individual, the profile is compared to a normal profile (e.g., already formed as described in the earlier section or Figure 1). The determination can then be made on the basis of these general criteria described below.

1. From about 7.00 to about 22.00, i.e., during a flat and low portion of the normal prolactin profile at a given point in the subject's prolactin profile, the value of the prolactin (in blood) must not be greater than 1 SEM above the mean at the same time (or greater than 2 SEM above the mean at two different times). This rule is applied to exclude from the final judgment all normal individual prolactin profiles that do not meet the above criteria, either because their prolactin is too high during the day or because their prolactin is too low during sleep or both.

2. From about 22.00 hours to about 7.00 hours, i.e. during a sudden change (and peak at sleep time) of the normal daily prolactin profile, the individual prolactin profile must first peak at approximately the same time or within two to six hours after sleep has commenced, as a normal prolactin peak for subjects in the same category (usually about 1.00 hours) and must also be in the 1 SEM of a normal prolactin profile of a healthy individual (preferably for 2 prolactin readings or another 2 SEM for at least one prolactin subtraction).

It should be noted that, based on the clinical experience of the present inventors, adjusting high levels of prolactin at day time to normal or near normal values will cause a significant improvement in the pathological conditions of the treated patients, although the treatment results in a decrease in the prolactin peak at night. Therefore, in accordance with this observation, observation of criterion 1 above takes precedence over observation of criterion 2.

If only a set of key prolactin measurements are performed for a subject to be tested, the above criteria are applied as described below. If any of the times of day on which key prolactin measurements are taken to determine the subject's prolactin level is greater than 1 SEM (and preferably 2 SEM) above the mean normal prolactin level during the day time at the same time points, then the subject has abnormal daily prolactin profile or rhythm, and if at any time of the night at which key prolactin levels are measured, the subject's prolactin level (calculated over a key period from 23.00 to 2.00 hours) is less than 1 SEM (and preferably less than 2 SEM) below mean normal prolactin levels during sleep at the same time points, then the subject has an abnormal prolactin night profile or rhythm.

To determine whether a subject has an aberrant prolactin profile, the time spent in bed by a subject to determine the subject's prolactin profile should ideally be the same as the bed time for subjects with a normal profile. If. if this is not the case, the subject's profile and the profile of normal individuals may be depicted and one or the other may be shifted such that the onset time of the subject to be tested is identical to that of normal healthy subjects.

Further, the determination of the treatment of the affected subject is described.

The information (prolactin profile and set of key prolactin levels) obtained as described above will be used to

(a) identifying patients who need to adjust their prolactin profile; and

(b) determination of the type and extent of adjustment required.

In general, those individuals suffering from obesity, hyperinsusemia, hyperlipidemia, hyperglycidemia and / or diabetes exhibit abnormal prolactin profiles (or key prolactin levels) as compared to healthy individuals. Simply stated, by comparing a subject's prolactin profile (or key prolactin level) with a normalized prolactin profile or a corresponding healthy (normal) set of prolactin levels, one can identify individuals affected by the abnormal conditions discussed above. By adjusting the abnormal prolactin profile of such individuals (either by administering a prolactin inhibitor or a prolactin stimulator, or both) at a suitable time during the day and at an appropriate (amount) dose, the prolactin profile of the individual can be adjusted to accommodate (or at least approximate) the normal profile. The amount and timing of administration of such doses can be determined based on the information contained in the prolactin profiles (or key prolactin levels) as discussed above.

The adjusted profile approaches a normal or healthy profile if all or part of the abnormal profile is shifted in the correct direction by at least 2 ng / ml. For example, if the abnormal level of human subject prolactin is 18 ng / ml between 7.00 and 10.00 hours and (after adjustment) is reduced to 16 ng / ml for the same time period, the adjusted profile approximates to a healthy profile. This is important to reduce the area under the prolactin curve during the day (usually at least 20%) and avoid the prolactin peak during the day.

Determination of treatment has three aspects:

(a) selecting the desired effect (i.e., selecting a prolactin stimulator, increasing the levels of prolactin or a prolactin inhibitor, decreasing prolactin levels, or both increasing prolactin at night and decreasing prolactin during the day),

(b) the timing of each dose administered; and

(c) the amount of each dose to be administered.

For example, generally, if a patient is initially diagnosed as requiring a prolactin inhibitor, he should initially receive 0.8-1.6 mg of bromocriptine per day from a (preferably) fast-release single dose (or divided doses). The dose may be adjusted to a treatment lasting approximately 4 weeks, for example at a time when a fresh prolactin profile (or a set of key prolactin levels) is determined and for example if the patient still has a high prolactin level, he might receive an increased dose (preferably) of a rapidly released prolactin inhibitor. at the appropriate time of the day. The time of administration may also be adjusted (depending on the patient's current prolactin profile or key prolactin levels) and, for example, would change the time of bromocriptine administration.

The comparison of the individual prolactin profile (or levels) with the normal prolactin profile (or levels) shall be made according to (a) and (b) of the previous paragraph as follows: If at any time of the day the prolactin level is too high (whether or not peak), administration of a prolactin inhibitor is required.

If the nocturnal prolactin profile is absent or if the peak is not significantly expressed, administration of a prolactin stimulator is required.

A preferred prolactin inhibitor (or dopamine agonist) is an accelerated release inhibitor, in particular an accelerated release composition comprising bromocriptine as disclosed in copending U.S. patent application Ser. 08/171 897.

A preferred prolactin stimulator is metoclopramide.

Whether a 24-hour prolactin profile is generated for the subject to be treated or only key prolactin levels are measured, the following detailed guidance for initial determination of the bromocriptine administration schedule over a treatment period of approximately 26 weeks will be followed.

(a) Week 1 to Week 6

First Dose: If at 7.00, 8.00, 16.00, or 19.00 hours in any of the patients the prolactin levels are equal to or greater than 5.0 ng / ml for men or 7.0 ng / ml for women, then 0.8 is administered. mg of rapidly released bromocriptine daily at 6.00 am.

Second dose: Starting at week 3, a second dose containing 0.8 mg of accelerated release bromocriptine daily at 10.30 am is also administered.

(b) Week 7 to Week 12

First dose: If at 7.00, 8.00, 16.00 or 19.00 any of the levels of prolactin are still equal to or greater than 5.0 ng / ml for men or 7.0 ng / ml for women, then 1.6 mg of accelerated bromocriptine is administered daily at 6.00 am. Otherwise, 0.8 mg of accelerated release bromocriptine is administered daily at 6.00 am.

Second dose: In addition, if the prolactin level at 19.00 hours is equal to or less than 1.5 ng / ml for men or women, then a second dose of 0.8 mg of accelerated release bromocriptine is administered at 8.30 hours daily instead of 10.30 hours. If the prolactin level at 7 pm is greater than 1.5 ng / ml for men or women, then the second dose is continued daily at 10.30 am.

If the prolactin level is less than 1.0 ng / ml at 19.00 hrs for men or women, then no second dosing is performed.

c) Week 13 to Week 36

For both the first and second dosages, the same rules are given as for Week 7 through Week 12, with the subject being monitored for:

(i) If at either 16.00 or 19.00 hours the prolactin level is equal to or greater than 5.0 ng / ml for men or 7.0 ng / ml for women, then an additional 0.8 mg of rapidly released bromocriptine is added to the first dose without regardless of whether the patient already receives a total of 2.4 mg bromocriptine. In this case, an additional 0.8 mg of rapidly released bromocriptine is added to the second dose.

(ii) If at 19,00 hours the prolactin level is less than 1,5 ng / ml for males or females, then the second dosing time shall be adjusted to be given 2 hours earlier; and (iii) if at 8.00, 16.00 and At 7pm, prolactin levels are less than 1.0 ng / ml for men or women, then 0.8 mg of accelerated release bromocriptine is deducted from the second dose or, if no second dose is administered, 0.8 mg of accelerated release bromocriptine is deducted from first dose. In the vast majority of patients, the first dose must contain at least 0.8 mg of accelerated release bromocriptine.

The dosage amount time schedule given above is meant as a guide for bromocriptine administration, and one skilled in the art can further adjust the exact time schedule and amount of bromocriptine administered based on the patient's actual prolactin profile or key prolactin levels to be treated. For example, if the patient is not responding (or not responding adequately) to the administered dose (or doses) (e.g. 0.8 mg), the dose may be (or may be) increased (e.g. to 1.6 mg).

If necessary, metoclopramide (generally daily dosed in the range of 0.5 to 5.0 mg per person, preferably daily dosed in the range of 0.5 to 2.0 mg per person) may be administered about one hour before bedtime. the bed. A prolactin stimulator is generally not administered, with the sole exception that the patient's average prolactin level between 1.00 and 4.00 hours is at least 1 SEM (preferably referring to two experimental points or at least 2 SEM if referring to one experimental point) when it is less than 8.0 ng / ml for men or 14 mg / ml for women. In addition, if prolactin at the time of day is too high and substantial clinical benefit would be achieved by adjusting daily prolactin time levels, administration of the prolactin stimulator will in most cases not be required (although it may be desirable).

In general, the time at which an inhibitor composition other than bromocriptine or a stimulator composition other than metoclopramide is to be administered to a patient can be determined by determining the time between administration of the composition and the time at which the composition exerts its maximum biological (i.e. stimulating or inhibiting) effect. The time at which the stimulator has its maximum stimulating effect (or where the inhibitor has its maximum inhibitory effect) can be determined by administering the stimulating or inhibiting drug to a patient, where the drug has a known prolactin profile, and then the time elapsing between drug administration and exerting a maximal effect on shifting (either inhibiting or stimulating) the (known) patient prolactin profile.

Fine-tuning the timing of administration of a drug to a subject should take into account the reflected effect that administration of a prolactin inhibitor during the day may have at sleep time on the subject's prolactin levels. Conversely, the reflected effect that administration of a prolactin stimulator may have on prolactin levels during waking hours of a subject should also be considered. In this manner and using the methodology of the above guidance, the time of administration for a particular inhibitor or stimulator can be determined using routine experimental procedures.

The exact time of administration of the modulator, which will result in the most effective results with respect to the efficacy of the treatment of the patients will depend on the activity, pharmacokinetics and bioavailability of the particular modulator, the physiological condition of the patient (including age, type and disease state, physical condition and response). dose and modulator), routes of administration, and the like. The above guidance can be used as a basis for determining optimal administration time.

The above data is applicable to adjust the initial therapy regimen. Generally, the patient will receive from about 3 to about 100 mg of bromocriptine per kilogram of body weight per day, and preferably from about 10 to 40 mg per kg of body weight per day. The exact dose of a prolactin inhibitor (or prolactin stimulator) required to achieve optimal prolactin effect should be adjusted for each patient based on the patient's sensitivity to the drug (ie drug responses), age, type and condition of the disease as well as physical state. The patient is periodically re-evaluated by measuring prolactin levels at predetermined intervals over a 24-hour period (or a shorter period of time if necessary), initially such re-evaluation is usually performed at the end of four weeks after initiation of therapy and subsequent re-evaluation performed every 4 weeks to every 8 weeks during therapy and thereafter every 3 months. Typical daily doses of bromocriptine for humans are from 0.2 to 15 mg, preferably from 0.8 to 8 mg.

Prolactin stimulators are usually administered for approximately one hour before bedtime for a patient with a normal sleep period.

Adjustment of the amount of drug (or drugs) administered and, optionally, the duration of administration may be made as described above based on repeated evaluations.

In general, adjustment of the administration time and amount of the drug (or drugs) is not considered necessary if the prolactin peak at sleep time during therapy is higher than normal if the prolactin peak occurs at the right time and peak peak is sharp (with normal values on each side of the normal peak) ).

The efficacy of a particular regimen in an individual patient and the required adjustment (in terms of dose and timing), if so, can be determined by comparing the patient's reassessment of the prolactin profile or the reassessment of key prolactin levels to the normalized profile (or healthy key levels).

In the treatment of vertebrates, generally, doses of the inhibitor of accelerated release prolactin (bromocriptine) and / or stimulator (metoclopramide) are always given, generally once a day, usually within a time period of about 10 to about 180 days, but treatment may continue indefinitely ( months) or even years. A preferred prolactin inhibitor (fast-released bromocriptine) is administered daily at dosage levels ranging from about 3 to about 100 pg, preferably from about 10 to about 40 pg per kg body weight, and the prolactin stimulator (metoclopramide) is administered daily at dose levels in the range of from about 5 to about 200 pg, preferably from about 5 to about 100 pg per kg body weight per day, to adjust or change the prolactin profile (or key prolactin level) and continue for a time sufficient to regain the prolactin daily rhythm in plasma, at which time treatment may be discontinued. If the subject has a relapse, the treatment may be reintroduced.

In the treatment of humans, in particular, a prolactin inhibitor (accelerated release bromocriptine) is generally administered daily at dosage levels ranging from about 3 to about 100 pg, preferably from about 10 to about 40 pg per kg body weight. Metoclopramide as a prolactin stimulator is generally administered daily at a dosage level ranging from about 5 to about 50 pg, preferably from about 5 to about 20 pg per kg body weight. (For a person, the daily dosage amount of metoclopramide is usually from 0.5 to 5.0 mg, preferably from 0.5 to 2.0 mg.) Such treatment (using one or both types of drug) usually continues for a period of time from about 10 usually up to 180 days, as a result of the adjustment and recovery of the lipid and glucose metabolism in a patient who is a lean (i.e. normal) healthy person, at which time treatment may be discontinued. For some patients (for example patients with particularly poor physical condition or elderly patients), it may not be possible to restore their prolactin rhythm within the aforementioned periods of time and such patients may require longer or equally continuous treatment with prolactin stimulators and / or inhibitors. The dose and temporal administration information set forth above is for bromocriptine and metoclopramide and will be varied for the other means used in the dosing methodology and timing disclosed herein.

In a practical embodiment of the invention, the prolactin inhibiting compound and the prolactin stimulator are administered daily to a subject, preferably orally or subcutaneously, intravenously or intramuscularly. Dermal release systems, such as skin patches, as well as lace and other well known drug delivery systems, may also be used.

The deposition of body fat, including deposition on the adipose, arterial wall and plasma fat, is reduced in obese persons, remains at the same reduced height and is generally maintained (after treatment according to the invention is discontinued) as in normal (lean) persons after prolonged time period. A subject who exhibits the effects of insulin resistance, hyperlipidemia, hyperinsulinemia and / or hyperglycaemia, or both insulin resistance and hyperinsulinemia and / or hyperglycemia, treated with a prolactin inhibitor and / or a prolactin stimulator at the appropriate time of day, elaborated above, becomes more sensitive to insulin (i.e., it will have lower insulin resistance) and the effects of hyperinsulinemia and / or hyperglycemia and related abnormal metabolic values will decrease on a long-term basis. Treatment usually ends between about 10 and about 180 days on average in humans. Thus, administration of a prolactin inhibitor (or dopamine agonist) and / or a prolactin stimulator in this manner will restore the phase relationships of the two neural oscillations and their different daily manifestations to changing metabolism on a long-term basis (e.g. several years) if not sustained. In other words, the result of a time daily dosing of a prolactin inhibitor (or dopamine agonist) and / or a prolactin stimulator will be reversible in the long term for the major part of the pathologies generally associated with the development of type II diabetes. Using the methods of the present invention, body fat levels, plasma insulin concentrations (including in cases of patients with oral hypoglycaemic medication), insulin resistance, hyperglycemia, and elevated blood pressure, or all of these pathological conditions, may be reduced in the long term by such treatments or treatments, from higher levels, which often occur in obese, hyperinsulinemic, hyperlipidemic and / or hyperglycemic persons to adjust or approach much lower and much more convenient levels that occur in normal persons with normal insulin levels.

DETAILED DESCRIPTION OF THE INVENTION

The following are working examples of diagnosis, assays, and therapy of the present invention, which do not constitute a limitation of the invention.

Study 1 (130 010)

Subject: male, age 52, weight 132 kg, height 167 cm

Pathological condition before treatment:

Obesity. 191% of ideal body weight (based on a standardized table, such as the one published by Metropolitan Life Insurance Co., New York, NY, USA, available from that company)

(b) Type II diabetes: fasting plasma glucose: 9.18 mmol / l, glycosylated hemoglobin: 9.4% (measured by affinity chromatography), normal fasting plasma glucose: between 4.48 and 6.72 mmol / l l.

The subject's prolactin profile, based on 24-hour follow-up (pre-therapeutic examination), is shown graphically as a dashed line in FIG. 1. The subject's prolactin appears to be too high during the day and early in the evening. At 7.00 am 13.2 ng / ml, at 8.00 am 11.8 ng / ml, at 16.00 am 9.9 ng / ml and at 19.00 am 12.2 ng / ml. The subject is administered accelerated release bromocriptine as follows:

Weeks 1 and 2: 1.6 mg at 5.00, weeks 3 and 4: 0.8 mg at 5.00 and additional 0.8 mg at 10.00, weeks 5 to 18: 1.6 mg at 5.00. The reassessment of the prolactin profile is performed on the subject after 18 weeks and is graphically depicted in FIG. 2 as a faint black line. Key prolactin values are: o 7.00 am 11.6 ng / ml, o 8.00 am 9.5 ng / ml, o 16.00 am 3.3 ng / ml, and o 7.00 pm 6.5 o / ml.

The improvement observed in this patient at prolactin levels after 12 weeks of treatment includes significantly lower values during most of the day and early evening.

The clinical effect of treating this patient involves a reduction in glycosylated hemoglobin from a baseline of 9.4% to 7.6% after 18 weeks of treatment. The patient lost 8.7 kg of body fat over 18 weeks, plasma cholesterol decreased from 5.95 mmol / l to 4.68 mmol / l, triglycerides decreased from 1.86 mmol / l to 1.22 mmol / l. 1, glucose cheaply decreased from 9.18 mmol / l to 7.49 mmol / l and oral glucose tolerance test results improved during the treatment period.

Study 2 (130 007)

Subject: male, age 53, weight 91 kg, height: 178 cm.

Pathological condition before treatment:

a) Obesity 130% of ideal body weight.

b) Type II diabetes: fasting plasma glucose: 9.41 mmol / l, glycosylated hemoglobin: 7.5%.

According to the initial assessment, the subject has a prolactin profile, shown in dashed line in FIG. 3. Daily levels (especially 7.00 am) are too high: 18.00 ng / ml at 7.00, 15.0 ng / ml at 8.00, 8.1 ng / ml at 16.00 and 19.00 at 16.00 hour is 8.8 ng / ml. The night peak is also slightly shifted for later. The patient is given fast-released bromocriptine as follows: Weeks 1 and 2: 1.6 mg at 8.30 hours, weeks 3 and 4: 1.6 mg at 8.00 hours and weeks 5 and 6: 0.8 mg at 8.30 hours, weeks 7 to 7 18: 1.6 mg at 5.00 hours.

The second profile (thin black line) of prolactin plotted after 18 weeks of treatment shows that the subject's prolactin profile is very sensitive to bromocriptine. Daily time levels are greatly reduced, but night time levels are also reduced below normal levels. Prolactin levels are: at 7.00 hrs 1.9 ng / ml, at 8.00 hrs 1.7 ng / ml, at 16.00 hrs 0.9 ng / ml and at 19.00 hrs 1.5 ng / ml.

The clinical effect of treating this patient involves a reduction in glycosylated hemoglobin from a baseline of 7.5% to 6.1%. Patient lost weight by 10.3 kg body fat, patients OGTT test shows significant improvement, with area under glucose curve decreasing by approximately 30% and fasting glucose decreased from 9.41 mmol / l to 7.45 mmol / l during the treatment period .

The data show that metabolic conditions are regulated at least to a clinically significant degree in part of the daily neuroendocrine rhythm interactions. This suggestion suggests that the daily rhythms of cortisol and prolactin are individually expressed from two separate daily systems and that the phase relationships of the two systems can be restored. Thus, in a hamster model, it was found that the 0-hour relationship of restoring daily oscillations in a pattern that maintains an insulin-sensitive condition in a poor individual and a 12-hour relationship allows retention of patterns that maintain an obese insulin-resistant condition. Another important addition to the present study is that the effects of injections of a prolactin-incubating dopamine agonist incubating agent or other prolactin-inhibiting compound at certain times are long lasting. Apparently as soon as the phase relationship of the two daily oscillations equals, it tends to maintain its changed pattern.

Changes in the phase relationships of two daily neuroendocrine oscillations are evidenced by changes in the phase relationships of their daily expressions. This expectation is fulfilled by the appropriate plasma glucocorticosteroid and prolactin rhythm. In several investigated species, the phase relationships of the two hormonal rhythms differ in poor and fat animals.

The phase relationship between the daily rhythm of plasma insulin concentration and the rhythm of lipogenic insulin sensitivity appears to be different in poor and fat animals. While the daily lipogenic susceptibility interval remains almost invisible initially, the phase of insulin rhythm changes markedly. For example, peak insulin concentrations occur near an indistinct onset in obese hamster females maintained for a short day. That is, the daily peaks of the lipogenic stimulus (i.e., insulin) and the lipogenic response to insulin coincide in fat animals and not in poor animals.

It is to be understood that various modifications and changes may be made without departing from the spirit and scope of the invention.

Claims (31)

A method for determining abnormalities in the daily rhythm of a prolactin of a first subject, which is a vertebrate, with a daily rhythm of prolactin, comprising the steps of: (a) measuring the blood level of the first subject's prolactin at intervals spaced independently during the first subject's waking hours and during the first subject's sleep time, for a period of about 24 hours, to form a first subject's prolactin profile; (b) comparing the prolactin profile with a previous determination of a normalized prolactin profile for healthy subjects of the same species; and (c) identifying the first subject as having an abnormal daily prolactin rhythm if the prolactin level measured in the first subject is (i) greater than 1 SEM above the mean prolactin level of these healthy subjects at two time points during waking hours; or (ii) less than 1 SEM below the mean prolactin level of these healthy subjects at two time points during sleep. The method of claim 1, wherein in step c) the first subject prolactin level is a level of prolactin that was measured in step a) and is / (i) greater than 2 SEM above the mean prolactin level of these healthy subjects during waking hours; or (ii) less than 2 SEM below the mean prolactin level of these healthy subjects at bedtime. 3. A method of determining abnormalities in the daily rhythm of prolactin of a first human subject comprising the steps of: (a) measuring the blood level of the first subject's prolactin at intervals spaced independently over the first subject's waking hours and during the first subject's sleep period of about 24 hours to form the first subject's prolactin profile; (b) a comparison of the prolactin profile with a previous determination of a normalized prolactin profile for healthy subjects of the same sex; and (c) identifying the first subject as having an abnormal daily prolactin rhythm if the prolactin level measured in the first subject is (i) greater than 1 SEM above the mean prolactin level of these healthy subjects at two time points during waking hours; or (ii) less than 1 SEM below the mean prolactin level of these healthy subjects at two time points during sleep. The method of claim 3, wherein in step c) the level of prolactin of the first subject is the level of prolactin that was measured in step a) and is (i) greater than 1 SEM above the mean prolactin level of these healthy subjects during waking hours; or (ii) less than 2 SEM below the mean prolactin level of these healthy subjects at bedtime. The method of claim 1, wherein the predetermined normalization values are generated for a statistically significant number of healthy subjects of the same sex as said first subject. 6. A method of determining abnormalities in the daily rhythm of prolactin of human subjects to be tested, comprising the steps of: (a) a comparison of the prolactin profile of these subjects to be tested that meet a predetermined normalized prolactin profile for healthy human subjects over a predetermined period of about 24 hours; and (b) identifying those members of this group who have a blood prolactin level of (i) greater than 1 SEM above the level of prolactin of these healthy subjects at two different times during waking hours; or (ii) less than 1 SEM below the level of prolactin of these healthy subjects at two different times at bedtime. 7. The method of claim 6 wherein in step b) the prolactin level of said members is a blood prolactin level and is (i) at any time during the waking hours, at least 2 SEM higher than the prolactin level of these healthy subjects; or (ii) at any time during sleep time at least 2 SEM lower than the prolactin level of these healthy subjects. 8. A method of determining abnormalities in the daily rhythm of a prolactin of a human subject to be tested comprising the steps of: (a) a comparison of the subject's prolactin profile for testing, wherein the profile is determined over a period of approximately 24 hours, with a prior determination of a normalized prolactin profile for healthy human subjects; and (b) determining whether the subject to be tested has an abnormal daily prolactin rhythm based on whether the subject's prolactin level is a blood prolactin level; and (i) is greater than 1 SEM above the level of prolactin in healthy subjects at two different times during waking hours; or (ii) is less than 1 SEM below the level of prolactin in healthy subjects at two different times at bedtime. The method of claim 8, wherein in step b) the level of prolactin of the subject to be tested is the level of prolactin in the blood and is (i) at any time greater than 2 SEM above the prolactin level of healthy subjects during waking hours; or (ii) at any time during sleep less than 2 SEM below the prolactin level during sleep and healthy subjects. 10. A method for determining abnormalities in the daily rhythm of a prolactin of a first vertebrate subject with a daily prolactin rhythm, comprising the steps of: a) measuring the blood level of the first subject's prolactin at intervals spaced independently at least twice during the early morning and at least twice during the early evening, to generate key levels of prolactin of the first subject, (b) comparing each of the key levels of prolactin with the corresponding previous determination of normalized prolactin levels for healthy subjects of the same species at the same time points; and (c) identifying the first subject as having an abnormal daily prolactin rhythm if at least one key first subject prolactin level is greater than 1 SEM above the corresponding normalized prolactin level of said healthy subjects during the waking hours. 11. A method of determining abnormalities in the daily rhythm of prolactin of a first human subject comprising the steps of: a) measuring the blood level of the first subject's prolactin at intervals spaced independently at least twice during the early morning and at least twice during the early evening, to generate key levels of prolactin of the first subject, (b) comparing each of the key levels of prolactin with the corresponding previous determination of normalized prolactin levels of healthy subjects of the same sex at the same time points; and c) identifying the first subject as having an abnormal daily prolactin rhythm if at least one key prolactin level of the first subject during waking hours is greater than 1 SEM above the corresponding normalized prolactin level of said healthy subjects. The method of claim 10, wherein the predetermined normalization values are generated for a statistically significant number of healthy subjects of the same sex as the first subject. 13. A method of determining abnormalities in the daily rhythm of prolactin of a group of human subjects to be tested, comprising the steps of: (a) comparing a set of key prolactin levels of these subjects to be tested that meet a predetermined period of about 24 hours with a set of predetermined normalized prolactin levels for healthy human subjects at the same time points as the key prolactin levels of said subjects to be tested; (b) identifying those members of the group who have: (i) at least one key prolactin level that is greater than 1 SEM above the corresponding normalized prolactin level of these healthy subjects, during waking hours; or (ii) at least one nightly key prolactin level that is less than 1 SEM below the corresponding normalized prolactin level of these healthy subjects. subjects. 14. A method of determining abnormalities in the daily rhythm of a prolactin of a human subject to be tested, comprising the steps of: (a) comparing a set of key prolactin levels of the subject to be tested with a set of corresponding previous determinations of normalized prolactin levels for healthy human subjects at the same time points as the key prolactin levels; and (b) determining whether the subject to be tested has an abnormal daily prolactin rhythm based on whether. (i) at least one key prolactin level during the waking hours of the subject to be tested is greater than 1 SEM per corresponding normalized prolactin level in healthy subjects; or (ii) at least one night key prolactin level of that subject is less than 1 SEM below the corresponding normalized prolactin level in healthy subjects . 15. A method for normalizing the daily prolactin profile of a first subject having an abnormal daily prolactin rhythm comprising the steps of: - comparing a series of key prolactin levels of said subject to a corresponding series of prolactin levels of healthy subjects of the same species at the same time points as the key levels, and adjusting key prolactin levels of the first subject to generally align or approximate corresponding prolactin levels to these healthy subjects. The method of claim 15, wherein the treatment step comprises at least one of - administering to said first subject a prolactin inhibitor if at least one key prolactin level of said first subject during waking hours is greater than 1 SEM above the corresponding level of the healthy subject's prolactin, and administering to said first subject a prolactin stimulator if at least one nocturnal key prolactin level of said first subject is less than 1 SEM below the corresponding prolactin level of a healthy subject. 17. A method of determining adjustments that cause an abnormal daily rhythm of a patient's prolactin to adapt to or approach the daily rhythm of a prolactin in a healthy subject, comprising: (a) comparing a series of key prolactin levels of said patient with corresponding normalized prolactin levels at the same time points for healthy human subjects of the same sex as said patient; (b) determining whether said patient is to be administered a prolactin inhibitor if at least one key level of said patient's prolactin exceeds the corresponding prolactin level of healthy subjects at the same time by more than 1 SEM; c) selecting at least one time for administration of said prolactin inhibitor, which is between 6.00 and 12.00 hours; and d) selecting an amount of said prolactin inhibitor to be administered to said subject such that at least one key prolactin level during said waking hours of said subject adapts or approaches the corresponding prolactin level of healthy subjects. 18. A method of determining an adjustment that induces an abnormal daily rhythm of a patient's prolactin to adapt to or approach the daily rhythm of a prolactin of a healthy subject, comprising: (a) a comparison of a number of key prolactin levels of this first subject with a predetermined normalized prolactin profile of healthy subjects; (b) determining whether said subject needs a prolactin stimulator by determining whether at least one key prolactin level of said first subject during sleep is at least 1 SEM lower than the normal prolactin level of said healthy subjects during sleep; (c) selecting a time period for administration to said subject prior to a time at which the level of prolactin at bedtime peaks in healthy subjects; (d) a selected amount of said prolactin stimulator to be administered to said subject such that at least one key prolactin level of said subject during sleep is not less than 1 SEM below the corresponding prolactin level of said healthy subjects at the same time. 19. A method of determining abnormalities in the daily rhythm of prolactin of a first human subject comprising the steps of: (a) collection of blood samples of the first subject for at least two blood samples collected at predetermined time intervals separated from each other during the early morning and at least two blood samples collected during the early evening; (b) measuring the prolactin content of each of the said samples; (c) plotting the prolactin content of each of said samples versus the time at which said sample was captured to create experimental points of the key prolactin level; (d) comparing these experimental prolactin key points to a predetermined normal prolactin profile for healthy subjects; and (e) identifying the first subject as having a abnormal daily prolactin rhythm, based on whether at least one experimental point of the key prolactin level of said subject is greater than 1 SEM above the corresponding point on the normal prolactin profile of healthy subjects. The method of claim 10, further comprising - in step a) measuring the blood level of the first subject's prolactin at least once at night to produce at least one nocturnal key prolactin level of said first subject, - in step b) comparing said at least one nocturnal key prolactin level with a corresponding predetermined normalized prolactin level of healthy subjects at the same time, and - in step c), identifying the first subject as having an abnormal daily prolactin rhythm, since this at least one nocturnal key prolactin level is less than 1 SEM below the corresponding normalized prolactin level. The method of claim 11, further comprising - in step a) measuring the blood level of the first subject's prolactin at least once at night to produce at least one nocturnal key prolactin level of said first subject, - in step b) comparing the at least one nocturnal key prolactin level with the corresponding predetermined normalized prolactin level of healthy subjects at the same time, and - in step c), identifying the first subject as having an abnormal daily prolactin rhythm. Because this at least one nocturnal key prolactin level is less than 1 SEM below the corresponding normalized prolactin level. The method of claim 10, wherein the prolactin inhibitor is bromocriptine and the subject is a human. The method of claim 22, wherein the bromocriptine is administered at least once between 6.00 and 12.00 hours. The method of claim 23, wherein the bromocriptine is administered in a total daily amount ranging from 0.8 to 8.0 mg. 25. The method of claim 24, wherein at least one portion of bromocriptine is given at 6.00 hrs. 26. The method of claim 16, wherein the prolactin inhibitor is bromocriptine and the subject is a human. The method of claim 26, wherein the bromocriptine is administered at least once between 6.00 and 13.00 hours. The method of claim 27, wherein the bromocriptine is administered in a total daily amount ranging from 0.8 to 8.0 mg. The method of claim 28, wherein at least one portion of bromocriptine is administered at 6.00 hrs. The method of claim 10, wherein, with step c), the key level of prolactin of the first subject (i) is greater than 1 SEM above the corresponding normalized prolactin level of said healthy subjects at two time points during waking hours. The method of claim 10, wherein with step c) the key level of the first subject's prolactin (i) is greater than 2 SEM above the corresponding normalized prolactin level of said healthy subjects during the waking hours. 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