KR20220015466A - Screening Method for Diagnosing Growth Hormone Deficiency in Pediatric Patients Using Masimorelin - Google Patents

Abstract

The present invention relates to a screening method for diagnosing growth hormone deficiency in a pediatric patient using masimorelin. The present invention further provides a masimorelin substance for use in diagnosing growth hormone deficiency in a pediatric patient. The method comprises the steps of providing one or more blood samples obtained from a subject within a range of about 15 to about 100 minutes after administering an amount of masimorelin effective to induce growth hormone secretion, the growth hormone levels in each blood sample determining whether the subject has growth hormone deficiency diagnosing whether or not The method of the present invention is a stand-alone test, does not need to be repeated, and does not require an alternative growth hormone stimulation test to reliably diagnose a pediatric patient.

Description

Screening Method for Diagnosing Growth Hormone Deficiency in Pediatric Patients Using Masimorelin

Growth hormone (GH) is a major body metabolic hormone that regulates protein, lipid, and carbohydrate homeostasis, and is necessary for the growth, development and maintenance of body and mind. GH is produced in the anterior pituitary by stimulation by growth hormone-releasing hormone (GHRH) in the hypothalamus. GH is secreted by the pituitary gland in a pulsating manner of about 6 to 10 random bursts over a 24-hour cycle.

Growth hormone deficiency (GHD) can be divided into four broad categories based on the cause of GH deficiency: 1) pituitary or "traditional" GHD, 2) hypothalamic GHD, 3) functional GHD, and 4) idiopathic GHD. . GHD may become clinically apparent in children or adults. In the United States, it is estimated that the incidence of GHD in children is between 1 in 4,000 and 1 in 10,000. More than 50,000 adults in the United States have GHD, and 6,000 new cases are reported each year, including children with GHD who convert to adult GHD (Human Growth Foundation www.hgfound.org).

GHD is a disease in children characterized by decreased developmental parameters, such as stunted growth and short stature. GHD in children can be congenital or acquired, and can be a solitary symptom or a complex symptom with other pituitary hormonal defects. There are many distinct causes of GHD in children, but the causes are often unknown (idiopathic GHD). Definitions of the various types of GHD and other causes of short stature are given in the International Classification of Endocrine Diagnostics in Children (ICPED). If left untreated, childhood-onset GHD leads to permanent short stature. For children, the diagnosis of GHD includes detailed medical history, clinical features, growth (development) analysis, biochemical examination of components of the GH-IGF axis (GH=growth hormone; IGF=insulin-like growth factor), and skeletal maturity using MRI and Based on radiological evaluation of pituitary structures (GHRS, 2000).

Similar to the case in adults, the diagnosis of GHD in children relies on biochemical tests, which are based on the growth hormone stimulation test (GHST), which determines GH levels that can be induced with agents known to stimulate the secretion of GH. Currently used GHSTs, such as insulin resistance testing (ITT) or glucagon stimulation testing (GST), have not been specifically developed or approved for this purpose, and have been adjusted from other indicators, and therefore may not have a performance characteristic, e.g. specificity, There are limitations on sensitivity, safety, or validity (Molitch et al., 2011; Cook et al., 2009).

ITT has been considered as the optimal criterion for the evaluation of GHD. Intravenous administration of insulin is used to induce hypoglycemia, which in turn leads to GH secretion. However, because of the potential risks associated with hypoglycemia associated with symptoms such as tremors, somnolence, and tachycardia, intensive medical monitoring of the patient is required, so this test is labor intensive. Dangerous side effects are often reported. Moreover, ITT is contraindicated for patients with seizure disorders and ischemic heart disease (Yuen 2011; Yuen 2013). Therefore, because of its inconvenience and safety concerns, ITT is not widely used.

GST is an alternative that has grown in use (Molitch 2011; Yuen 2011; Yuen 2013). Common side effects of GST include nausea, vomiting, and headache. Limited examination time (3 to 4 hours) and the need for intramuscular injection. There remains a realistic unmet medical need for safe and reliable alternative testing.

A test for diagnosing GHD based on masimorelin, an orally ingestible peptidomimetic ghrelin receptor agonist with growth hormone secretion promoting (GHS) activity, is disclosed in WO 2007/093820 A1 by Larsen.

Ghrelin potently stimulates GH secretion (Kojima 1999). The GH-secreting effect of ghrelin is believed to be mediated by specific receptors primarily at the pituitary and hypothalamic levels (Nogueiras 2006). In membrane specimens containing GHS receptors derived from the human hypothalamus and pituitary, it has been demonstrated that masimorelin exhibits similar binding affinity to the natural ligand ghrelin to the human GHS receptor (Broglio 2002). Masimorelin is readily absorbed from the gastrointestinal tract and is believed to act in the same way as ghrelin.

Based on the ability of masimorelin to secrete GH pulses immediately after oral administration to healthy subjects, masimorelin has been developed as an oral diagnostic agent for GH deficiency in adults.

Masimorelin as a compound and its therapeutic use of GHD are disclosed in WO 01/96300 A1 by Martinez et al.

Masimorelin GHST for adult GHD (AGHD) is disclosed by Garcia et al. (J Clin Endocrinol Metab. 2013-1157, J Clin Endocrinol Metab. 2018-00665, and titles published at the 99th Annual Meeting of the 2017 Endocrinology Society) Poster of "Validation of Macimorelin As a Diagnostic Test for Adult Growth Hormone Deficiency (AGHD): A Phase 3 Study in Comparison with the Insulin Tolerance test (ITT)").

WO 2019/121762 A1 discloses a method for measuring the level of growth hormone in a human or animal subject. The method comprises the steps of orally administering to a subject a composition containing masimorelin, collecting one, two, or three post-dose samples from the subject within the range of 25 to 95 minutes after administration, and 1, 2, or comparing the growth hormone levels of the three samples to a single threshold, wherein the single threshold is 2.8 ng/mL, and the “single threshold” is a threshold growth hormone level indicating a sufficient response to masimorelin stimulation. means

The role of GH stimulation tests in pediatric patients is controversial because of the frequent occurrence of low GH levels in provocation tests and concerns about the efficacy and reproducibility of GHST.

Thus, guidelines for diagnosing GHD in children usually require the results of two GHSTs to conclude a diagnosis of GHD, unless there is a general brain defect that necessitates the use of only one GHST (GHRS, 2000, GHRS 2019).

The consensus guidelines of the Society for GH Research published in 2000 (GHRS, 2000), also the guidelines of the American Society of Clinical Endocrinologists published in 2003 (Gharib, 2003), and national guidelines (Binder, 2014) provide standardized GHST protocols. Therefore, it is recommended that a limited number of GHST preparations be used after an overnight fast. These include arginine (ARG), clonidine, glucagon, insulin, and L-dopa.

The consensus guidelines of the GH Research Society states that the combination of GHRH and ARG as GHST is considered useful for diagnosing GHD in children and adults, if appropriate endpoints are applied (GHRS, 2000). This combination has been shown to have high sensitivity and high specificity for children and adolescents (Maghnie, 2002) and for late adolescents and young adults (Corneli, 2007). In the latter experiment, endpoints are established only for lean patients. Given that GH secretion is a function of body weight and obesity (Colao, 2009), the appropriate endpoint (taking into account age, BMI, and waist circumference) for overweight and obese pediatric patients remains to be elucidated.

For children on the GHD clinical criteria, a peak GH concentration of less than 10 ng/mL has traditionally been used as an endpoint to support the diagnosis. In other studies, the sensitivity, specificity, and GH endpoints used for different GHSTs have recently been reviewed (van Vught, 2009).

Against this background, various GHSTs are being used in clinical practice, and the requirement of two GHSTs as part of standard diagnostic procedures is variably met by repetition of the same GHST or successive performance of two different GHSTs.

These two GHSTs are performed on the same day or two consecutive days, and they require 4 to 6 blood samples per test. Especially for children, the amount of blood obtained is considered a safety concern, and safe dose limits should be observed, eg as recommended by WHO (Howie 2011). In addition to this safety issue, it should be noted that performing two tests is time- and resource-consuming, and the burden associated with the child and their parents, as well as the pediatric endocrinologist, is great.

Therefore, instead of two tests, there is a confirmed need for a single test that has proven safety, tolerability, easy application, strong test properties with respect to sensitivity and specificity, and reliable reproducibility. A brief report on the present invention was published in 2020, announcing positive results in a pediatric study investigating the dosing of marshmorelin.

In one aspect, the present invention provides a screening method for diagnosing growth hormone deficiency in a pediatric patient using masimorelin, comprising:

(a) providing 1 to 5 blood samples obtained from the subject within the range of about 15 to about 100 minutes after administration of an amount of masimorelin effective to induce growth hormone secretion;

(b) measuring the growth hormone level in each blood sample provided in step (a);

(c) comparing the measured growth hormone level obtained in step (b) with a single threshold, wherein the single threshold is greater than or equal to 10.0 ng/mL;

(d) determining as having growth hormone deficiency the subject having the highest growth hormone level in the blood sample obtained in step (b) below said single threshold, wherein the highest growth hormone level in the blood sample obtained in step (b) is determining a subject not lower than the single threshold as not having growth hormone deficiency.

In another aspect, the present invention relates to a masimorelin substance for use in diagnosing growth hormone deficiency in a pediatric patient, comprising:

(a) providing 1 to 5 blood samples obtained from the subject within the range of about 15 to about 100 minutes after administration of an amount of masimorelin effective to induce growth hormone secretion;

(b) measuring the growth hormone level in each blood sample provided in step (a);

(c) comparing the measured growth hormone level obtained in step (b) with a single threshold, wherein the single threshold is greater than or equal to 10.0 ng/mL;

(d) determining as having growth hormone deficiency the subject having the highest growth hormone level in the blood sample obtained in step (b) below said single threshold, wherein the highest growth hormone level in the blood sample obtained in step (b) is determining a subject not lower than the single threshold as not having growth hormone deficiency.

In another aspect, the present invention relates to a screening method for diagnosing growth hormone deficiency in a pediatric patient using masimorelin, comprising:

(a) providing one or more blood samples obtained from the subject within the range of about 15 to about 100 minutes after administering an amount of masimorelin effective to induce growth hormone secretion;

(b) measuring the growth hormone level in each blood sample;

(c) comparing each measured growth hormone level to a single threshold;

(d) diagnosing whether the subject has growth hormone deficiency based on comparing the growth hormone levels measured in step (b) of the one or more blood samples to the single threshold value, wherein the subject Determining whether you have this growth hormone deficiency is a step based solely on induction of growth hormone levels by a single masimorelin administration.

Justice

Terms specified for the description of the method of the present invention always have the following meanings, unless otherwise specified in the specification or claims.

As used herein, a “subject” or “pediatric patient” is a human child of any gender (male or female) between about 2 years of age and less than 18 years of age. For example, the subject can be about 3 to less than 18 years old, about 4 to less than 18 years old, about 5 to less than 18 years old, about 6 to less than 18 years old, about 7 to less than 18 years old, about 8 to less than 18 years old. less than 18 years old, about 9 to less than 18 years old, about 10 to less than 18 years old, about 11 to less than 18 years old, about 12 to less than 18 years old, about 2 to less than 17 years old, about 2 to less than 16 years old less than, about 2 to less than 15, about 2 to less than 14, about 2 to less than 13, about 2 to less than 12, about 2 to less than 10, about 2 to less than 9, or about 2 to less than 8 years of age.

As used herein, "masimorelin" refers to a peptidomimetic compound that acts as a ghrelin receptor agonist with growth hormone secretion promoting (GHS) activity. Its chemical structure and therapeutic use of GHD are disclosed in US Pat. No. 6,861,409, WO 01/96300, and WO 2007/093820.

As used herein, the term “effective amount” means an amount of a given substance that is quantitatively sufficient to achieve the desired effect. For example, an effective amount of masimorelin for inducing growth hormone secretion in a recipient is that amount of the compound that, when administered to a subject, is capable of achieving a detectable increase in growth hormone secretion.

As used herein, the term “testing” refers to the act of clarifying a person suspected of having a particular disease based on a subject's symptoms and leading to identification of the presence or absence of the disease. In other words, “testing” for a disease encompasses the identification or exclusion of the disease.

As used herein, the term “treatment” refers to an act that leads to the elimination, reduction, alleviation, recovery, or prevention or delay of the onset or recurrence of any symptoms of a related disease.

As used herein, the term “blood sample” encompasses a portion of whole blood, such as a serum or plasma sample, as well as a whole blood sample. When two or more blood samples are used in the same method of testing, these blood samples are always of the same type. For example, if the first sample is serum, the second and all subsequent samples are also serum. Also, if more than one blood sample is provided, the term “blood sample” means a blood sample obtained at different time points following administration of an effective amount of masimorelin to induce hormone secretion. The two blood samples may be sampled at, for example, about 30±10 minutes and about 45±10 minutes or about 30±10 minutes and about 60±10 minutes after administration of an effective amount of masimorelin to induce hormone secretion. may mean a blood sample obtained. The three blood samples may be administered, for example, at about 30±10 minutes, about 45±10 minutes, and about 60±10 minutes or preferably, about may mean a blood sample obtained at 30±10 minutes, at about 45±10 minutes, and at about 90±10 minutes, or alternatively at about 30±10 minutes, at about 60±10 minutes, and at about 90±10 minutes. have.

As used herein, the term “about” means a range of ±10% of a reference value. For example, “about 10” defines a range from 9 to 11.

The term “single threshold” relates to a threshold level of growth hormone that exhibits a sufficient response to masimorelin stimulation: instead of the threshold of 2.8 ng/mL normally used for adult testing, the threshold used in the method of the present invention is about 10.0 ng/mL or greater, such as about 10.0 to 25.0 ng/mL, about 10.0 to 20.0 ng/mL, about 10.1 to 19.5 ng/mL, about 10.2 to 19.0 ng/mL, about 10.3 to 18.5 ng/mL , about 10.4 to 18.0 ng/mL, about 10.5 to 17.5 ng/mL, about 10.6 to 17.0 ng/mL, about 11.0 to 16.5 ng/mL, about 12.0 to 16.0 ng/mL, about 13.0 to 15.5 ng/mL, about 14.0 to 15.0 ng/mL, about 15.0 to 16.0 ng/mL, about 15.5 to 18.0 ng/mL, about 16.0 to 18.0 ng/mL, about 16.5 to 18.0 ng/mL, about 17.0 to 18.0 ng/mL, about 17.5 to 18.5 ng/mL, about 18.0 to 19.0 ng/mL, about 18.5 to 19.5 ng/mL, about 19.0 to 20.0 ng/mL, about 20.0 to 21.0 ng/mL, or a higher range of about 25.0 to 30.0 ng/mL . A single threshold used in the method of the present invention may also be a single value, e.g., 10.5 ng/mL, 11.0 ng/mL, 11.5 ng/mL, 12.0 ng/mL, 12.5 ng/mL, 13.0 ng/mL, 13.5 ng/mL, 14.0ng/mL, 14.5ng/mL, 15.0ng/mL, 15.5ng/mL, 16.0ng/mL, 16.5ng/mL, 17.0ng/mL, 17.5ng/mL, 18.0ng/mL, 18.5 ng/mL, 19.0 ng/mL, 20.0 ng/mL, or 25.0 ng/mL. Also, single threshold means ng per mL of whole blood or serum/plasma. Most preferably, a single threshold means ng per mL of serum. The terms “single threshold” and “endpoint” are used interchangeably.

Weight percent, weight %, % w/w, etc. are synonyms meaning the concentration of a substance divided by the weight of the composition and multiplied by 100.

All patents, patent applications, and other publications cited in this application are incorporated by reference in their entirety for all purposes.

1 shows individual masimorelin concentrations versus time, ie 0.25 mg/kg body weight, linear scale (Pharmacokinetic Analysis Set (PKS), Number of Patients (N)=24) in Cohort 1 for each individual.
2 shows individual masimorelin concentrations versus time, ie, 0.5 mg/kg body weight, linear scale (PKS, N=24) in Cohort 2;
3 shows individual masimorelin concentrations versus time, ie, 1.0 mg/kg body weight, linear scale (PKS, N=24) in Cohort 3;
4 shows individual GH concentrations versus time, ie, 0.25 mg/kg body weight, linear scale (Pharmacodynamic Analysis Set (PDS), N=24) after masimorelin GHST in Cohort 1.
5 shows individual GH concentrations versus time, ie, 0.5 mg/kg body weight, linear scale (PDS, N=24) after masimorelin GHST in Cohort 2.
6 shows individual GH concentrations versus time, ie, 1.0 mg/kg body weight, linear scale (PDS, N=24) after masimorelin GHST in Cohort 3.
7 shows a recipient response characteristic (ROC) analysis of masimorelin GHST in Cohort 1, ie, 0.25 mg/kg body weight (PDS, N=24).
8 shows a recipient response characteristic (ROC) analysis of masimorelin GHST in Cohort 2, ie, 0.5 mg/kg body weight (PDS, N=24).
9 shows a recipient response characteristic (ROC) analysis of masimorelin GHST in Cohort 3, ie, 1.0 mg/kg body weight (PDS, N=24).

It is an object of the present invention to provide a unique method for measuring the level of growth hormone in a pediatric patient and detecting GHD in a pediatric patient. The above object is to develop a new method that should reduce the test time and the number of blood samples, thereby reducing the burden on the test manager and the test subject, and providing safe, reliable, and excellent diagnostic performance.

The object of the present invention is surprisingly solved in one aspect by providing a screening method for diagnosing growth hormone deficiency in a pediatric patient using masimorelin, comprising:

(a) providing 1 to 5 blood samples obtained from the subject within the range of about 15 to about 100 minutes after administration of an amount of masimorelin effective to induce growth hormone secretion;

(b) measuring the growth hormone level in each blood sample provided in step (a);

(c) comparing the measured growth hormone level obtained in step (b) with a single threshold, wherein the single threshold is greater than or equal to 10.0 ng/mL;

(d) determining as having growth hormone deficiency the subject having the highest growth hormone level in the blood sample obtained in step (b) below said single threshold, wherein the highest growth hormone level in the blood sample obtained in step (b) is determining a subject not lower than the single threshold as not having growth hormone deficiency.

Therefore, the 1 to 5 blood samples were obtained by administering an effective amount of masimorelin for inducing growth hormone secretion, not earlier than about 15 minutes after administration of an effective amount of masimorelin for inducing growth hormone secretion. Then, no later than about 100 minutes, it is obtained from the subject.

For example, if the level of a single growth hormone in the blood sample is not lower than the single threshold, it is determined that the subject does not have growth hormone deficiency. Instead, if the growth hormone levels of all blood samples are below the single threshold, the subject is determined to have growth hormone deficiency.

In a preferred embodiment, the screening method for diagnosing growth hormone deficiency in a pediatric patient using masimorelin is an in vitro screening method for diagnosing growth hormone deficiency in a pediatric patient using masimorelin.

In a preferred embodiment, the single threshold of growth hormone is from about 10.0 to about 25.0 ng/mL, preferably from about 10.2 to about 20.0 ng/mL, more preferably from about 12.0 to about 19.0 ng/mL, more preferably is in the range of from about 14.0 to about 18.0 ng/mL, more preferably from about 16.0 to about 18.0 ng/mL, and most preferably from about 17.0 to about 18.0 ng/mL.

In a preferred embodiment, 1 to 4 blood samples are provided in step (a), preferably 1 to 3 blood samples are provided in step (a), more preferably 2 or 3 blood samples are provided in step (a) A blood sample is provided from a dog. As noted above, if more than one blood sample is provided, these blood samples are obtained at different time points after administration of an effective amount of masimorelin to induce hormone secretion.

In another preferred embodiment, the blood sample in step (a) is in the range of about 20 to about 100 minutes, preferably about 25 to about 100 minutes after administration of an effective amount of masimorelin for inducing growth hormone secretion. minutes, more preferably in the range of about 25 to about 95 minutes, and most preferably in the range of about 30 to about 90 minutes. Thus, the blood sample is administered, for example, no later than about 20 minutes but no later than about 100 minutes, preferably no earlier than about 25 minutes, after administration of an effective amount of masimorelin to induce growth hormone secretion. no later than 100 minutes, more preferably no later than about 25 minutes but no later than about 95 minutes, and most preferably no earlier than about 30 minutes but no later than about 90 minutes.

In another preferred embodiment, if more than one blood sample is provided, the blood sample is obtained from the subject at about 10 to about 60 minute intervals, preferably about 15 to about 30 minute intervals. Alternatively, the blood sample may be obtained at any time interval deemed appropriate by the attending physician. For example, the blood sample may be about 5 minutes, about 10 minutes, about 15 minutes, about 20 minutes, about 25 minutes, about 30 minutes, about 35 minutes, about 40 minutes, about 45 minutes, about 50 minutes, about 45 minutes. minutes, or about 60 minutes apart.

In a more preferred embodiment, said blood sample in step (a) is a whole blood sample, a serum sample, or a plasma sample. Preferably, said blood sample in step (a) is a serum sample or a plasma sample. If more than one blood sample is obtained, the two or more blood samples are of the same type, and therefore the blood samples are either a whole blood sample, a serum sample, or a plasma sample. Most preferably, the blood sample is a serum sample.

In a more preferred embodiment, in step (a) about 0.8 mg to about 1.2 mg of masimorelin per kg of subject's body weight is administered, preferably in step (a) about 1.0 mg of masimorelin per kg of subject's body weight is administered . In general, an effective amount of masimorelin is any one of, or a minimum amount, in the range of, as a minimum, from about 0.8 to about 0.9 mg/kg of body weight, as high as about 1.0, from about 1.1 to about 1.2 mg/kg of body weight of a subject. Within the range defined by any one of, for example, may be about 0.9 to about 1.1 mg / kg body weight. The effective amount of marshmorelin may also be a single value, for example, about 0.8 mg/kg, about 0.9 mg/kg, about 1.0 mg/kg, about 1.1 mg/kg, or about 1.2 mg/kg body weight. Body weight (reported in kg) may preferably be rounded to the nearest whole number. Most preferably, in step (a), about 1.0 mg of masimorelin per kg body weight of the subject is administered.

For example, one unit dose consists of a composition containing 1817.2 mg masimorelin for preparation as an oral suspension. Generally, the prepared suspension contains 0.5 mg of marshmorelin per mL of suspension. Pediatric patients are administered in aliquots of the reconstituted suspension adjusted for body weight so that the dose for the child is 1.0 mg/kg body weight. The unit dose is defined as masimorelin calculated as the free base with a content of 100%. The mass of masimorelin free base or its free base equivalent in the unit dose may be adjusted according to the content.

In a preferred embodiment, in step (a), masimorelin is administered as a suitable pharmaceutical salt in the form of a composition comprising masimorelin, wherein the appropriate pharmaceutical salt is preferably an acetate salt of masimorelin, masimorelin trifluoro acetate salt of, or a combination thereof.

In a more preferred embodiment, the administration of masimorelin in step (a) is oral administration.

If the administration of masimorelin is oral administration, the masimorelin may be prepared as an oral suspension. The suspension may be administered within about 90 minutes, preferably within about 60 minutes, more preferably within about 30 minutes after preparation of the oral suspension. Preferably, the oral suspension is drunk for a time of about 1 minute or less, preferably about 30 seconds or less.

In a more preferred embodiment, the subject fasts for about 10 hours, preferably about 9 hours, more preferably about 8 hours prior to administration of masimorelin. More preferably, in step (a) the subject fasts for about 100 minutes after administration of masimorelin, and in step (a) the subject fasts for about 100 minutes, about 95 minutes, or about 90 minutes after administration of masimorelin means you can

In a more preferred embodiment, one blood sample obtained from said subject is provided about 60±30 minutes after administration of masimorelin in step (a). One blood sample may alternatively be administered at about 30±10 minutes, about 40±10 minutes, about 45±10 minutes, about 50±10 minutes, about 60±10 minutes, about 70±10 minutes, can be obtained from the subject at about 80±10 minutes, about 90±10 minutes, or about 100±10 minutes. One blood sample may be obtained at any point within the range of about 15 to about 100 minutes after administration of masimorelin as deemed appropriate by the attending physician.

In another more preferred embodiment, two blood samples obtained from said subject at about 30±10 minutes and about 45±10 minutes after administration of masimorelin in step (a) are provided. The two blood samples may alternatively be administered at about 20±10 minutes, about 30±10 minutes, about 40±10 minutes, about 45±10 minutes, about 50±10 minutes, about 60±10 minutes, can be obtained from the subject at about 70±10 minutes, about 80±10 minutes, about 90±10 minutes, or about 100±10 minutes. Two blood samples may be obtained at any time in the range of about 15 to about 100 minutes after administration of masimorelin as deemed appropriate by the attending physician.

In another preferred embodiment, two blood samples obtained from said subject at about 30±10 minutes and about 60±10 minutes after administration of masimorelin in step (a) are provided.

In another preferred embodiment, three blood samples obtained from said subject are provided at about 30±10 minutes, about 45±10 minutes, and about 60±10 minutes after administration of masimorelin in step (a), or at step (a) ), at about 30±10 min, about 45±10 min, and about 90±10 min after administration of masimorelin, three blood samples obtained from the subject are provided, or at about 30±10 min after administration of masimorelin in step (a) Three blood samples obtained from the subject are provided at ±10 min, at about 60±10 min, and at about 90±10 min. The three blood samples may alternatively be administered at about 20±10 minutes, about 30±10 minutes, about 40±10 minutes, about 45±10 minutes, about 50±10 minutes, about 60±10 minutes, can be obtained from the subject at about 70±10 minutes, about 80±10 minutes, about 90±10 minutes, or about 100±10 minutes. Three blood samples may be obtained at any time in the range of about 15 to about 100 minutes after administration of masimorelin as deemed appropriate by the attending physician.

If more than three blood samples are provided in step (a), these blood samples may be obtained at any time within the range of about 15 to about 100 minutes after administration of masimorelin as deemed appropriate by the attending physician. can get For example, suitable time points are about 20±10 minutes, about 30±10 minutes, about 40±10 minutes, about 45±10 minutes, about 50±10 minutes, about 60±10 minutes, about 70 minutes after administration of massimorelin. ±10 min, about 80±10 min, about 90±10 min, or about 100±10 min.

In a particularly preferred method, obtained from the subject at a time selected from the group consisting of about 30±10 minutes, about 45±10 minutes, about 60±10 minutes, and about 90±10 minutes after the administration of masimorelin in step (a) , 1 to 4, more preferably 1 to 3, more preferably 2 or 3 blood samples are provided.

In a more preferred embodiment, in step (a), masimorelin is administered in the form of a composition further comprising masimorelin and optionally a pharmaceutically acceptable excipient such as a carrier material. Preferably, the masimorelin is administered in the form of a composition comprising masimorelin and a sweetener. A suitable sweetener is, for example, saccharin. Advantageously, saccharin has been found to be a suitable taste masking agent for masimorelin.

In a more preferred embodiment, in step (a) about 3.5% (w/w) of masimorelin (calculated as free base), about 93.1% (w/w) of spray dried lactose monohydrate, about 2.0% ( w/w) crospovidone A, about 0.1% (w/w) colloidal silicon dioxide, about 1.0% (w/w) sodium stearyl fumarate, and about 0.3% (w/w) saccharin Massimorelin in the form of a composition comprising sodium dihydrate is administered.

In a more preferred embodiment, the subject is a human child from 2 to less than 18 years of age, preferably said subject is a human child from 2 to less than 17 years of age, more preferably said subject is from 2 to less than 16 years of age is a human child of

In a more preferred embodiment, the method is a standalone test, does not need to be repeated, and does not require an alternative growth hormone stimulation test to reliably diagnose growth hormone deficiency in a pediatric patient.

In a more preferred embodiment, the determining according to step (d) whether the subject has growth hormone deficiency is based solely on induction of growth hormone levels by a single masimorelin administration.

Surprisingly, it has been found that the method of the present invention is suitable as a stand-alone test because it does not require additional GHST to reliably diagnose GHD in pediatric patients.

The object of the present invention is surprisingly solved, in another aspect, by providing a masimorelin substance for use in diagnosing growth hormone deficiency in a pediatric patient, comprising:

(a) providing 1 to 5 blood samples obtained from the subject within the range of about 15 to about 100 minutes after administration of an amount of masimorelin effective to induce growth hormone secretion;

(b) measuring the growth hormone level in each blood sample provided in step (a);

(c) comparing the measured growth hormone level obtained in step (b) with a single threshold, wherein the single threshold is greater than or equal to 10.0 ng/mL;

(d) determining as having growth hormone deficiency the subject having the highest growth hormone level in the blood sample obtained in step (b) below said single threshold, wherein the highest growth hormone level in the blood sample obtained in step (b) is determining a subject not lower than the single threshold as not having growth hormone deficiency.

The 1 to 5 blood samples were administered in less than about 15 minutes after administration of an effective amount of masimorelin for inducing growth hormone secretion, and after administration of an effective amount of masimorelin for inducing growth hormone secretion, No later than approximately 100 minutes, obtained from the subject.

In a preferred embodiment, the masimorelin substance for use in diagnosing growth hormone deficiency in a pediatric patient is a masimorelin substance for use in diagnosing growth hormone deficiency in vitro in a pediatric patient.

In a preferred embodiment, the single threshold of growth hormone is from about 10.0 to about 25.0 ng/mL, preferably from about 10.2 to about 20.0 ng/mL, more preferably from about 12.0 to about 19.0 ng/mL, more preferably is in the range of from about 14.0 to about 18.0 ng/mL, more preferably from about 16.0 to about 18.0 ng/mL, and most preferably from about 17.0 to about 18.0 ng/mL.

In a preferred embodiment, 1 to 4 blood samples are provided in step (a), preferably 1 to 3 blood samples are provided in step (a), more preferably 2 or 3 blood samples are provided in step (a) A blood sample is provided from a dog. If more than one blood sample is provided, the blood samples are obtained at different time points after administration of an effective amount of masimorelin to induce hormone secretion.

In another preferred embodiment, the blood sample in step (a) is in the range of about 20 to about 100 minutes, preferably about 25 to about 100 minutes after administration of an effective amount of masimorelin for inducing growth hormone secretion. minutes, more preferably in the range of about 25 to about 95 minutes, and most preferably in the range of about 30 to about 90 minutes. Thus, the blood sample may be administered, for example, no later than about 20 minutes but no later than about 100 minutes, preferably no earlier than about 25 minutes, after administration of an effective amount of masimorelin to induce growth hormone secretion. no later than 100 minutes, more preferably no later than about 25 minutes but no later than about 95 minutes, and most preferably no earlier than about 30 minutes but no later than about 90 minutes.

In another preferred embodiment, if more than one blood sample is provided, the blood sample is obtained from the subject at about 10 to about 60 minute intervals, preferably about 15 to about 30 minute intervals. Alternatively, the blood sample may be obtained at any time interval deemed appropriate by the attending physician. For example, the blood sample may be about 5 minutes, about 10 minutes, about 15 minutes, about 20 minutes, about 25 minutes, about 30 minutes, about 35 minutes, about 40 minutes, about 45 minutes, about 50 minutes, about 45 minutes. minutes, or about 60 minutes apart.

In a more preferred embodiment, said blood sample in step (a) is a whole blood sample, a serum sample, or a plasma sample. Preferably, said blood sample in step (a) is a serum sample or a plasma sample. If more than one blood sample is obtained, the two or more blood samples are of the same type, and therefore the blood samples are either a whole blood sample, a serum sample, or a plasma sample. Most preferably, the blood sample is a serum sample.

In a more preferred embodiment, in step (a) about 0.8 mg to about 1.2 mg of masimorelin per kg of subject's body weight is administered, preferably in step (a) about 1.0 mg of masimorelin per kg of subject's body weight is administered . In general, an effective amount of masimorelin is any one of, or a minimum amount, in the range of, as a minimum, from about 0.8 to about 0.9 mg/kg of body weight, as high as about 1.0, from about 1.1 to about 1.2 mg/kg of body weight of a subject. Within the range defined by any one of, for example, may be about 0.9 to about 1.1 mg / kg body weight. The effective amount of marshmorelin may also be a single value, for example, about 0.8 mg/kg, about 0.9 mg/kg, about 1.0 mg/kg, about 1.1 mg/kg, or about 1.2 mg/kg body weight. Body weight (reported in kg) may preferably be rounded to the nearest whole number.

For example, one unit dose consists of a composition containing 1817.2 mg masimorelin for preparation as an oral suspension. Generally, the prepared suspension contains 0.5 mg of marshmorelin per mL of suspension. Pediatric patients are administered in aliquots of the reconstituted suspension adjusted for body weight so that the dose for the child is 1.0 mg/kg body weight. The unit dose is defined as masimorelin calculated as free base in a content of 100%. The mass of masimorelin free base or its free base equivalent in the unit dose may be adjusted according to the content.

In a preferred embodiment, in step (a), masimorelin is administered as a suitable pharmaceutical salt in the form of a composition comprising masimorelin, wherein the appropriate pharmaceutical salt is preferably an acetate salt of masimorelin, masimorelin trifluoro acetate salt of, or a combination thereof.

In a more preferred embodiment, the administration of masimorelin in step (a) is oral administration.

If the administration of masimorelin is oral administration, the masimorelin may be prepared as an oral suspension. The suspension may be administered within about 90 minutes, preferably within about 60 minutes, more preferably within about 30 minutes after preparation of the oral suspension. Preferably, the oral suspension is drunk for a time of about 1 minute or less, preferably about 30 seconds or less.

In a more preferred embodiment, the subject fasts for about 10 hours, preferably about 9 hours, more preferably about 8 hours prior to administration of masimorelin. More preferably, in step (a) the subject fasts for about 100 minutes after administration of masimorelin, and in step (a) the subject fasts for about 100 minutes, about 95 minutes, or about 90 minutes after administration of masimorelin means you can

In a more preferred embodiment, one blood sample obtained from said subject is provided about 60±30 minutes after administration of masimorelin in step (a). One blood sample may alternatively be administered at about 30±10 minutes, about 40±10 minutes, about 45±10 minutes, about 50±10 minutes, about 60±10 minutes, about 70±10 minutes, can be obtained from the subject at about 80±10 minutes, about 90±10 minutes, about 100±10 minutes. One blood sample may be obtained at any point within the range of about 15 to about 100 minutes after administration of masimorelin as deemed appropriate by the attending physician.

In another more preferred embodiment, two blood samples obtained from said subject at about 30±10 minutes and about 45±10 minutes after administration of masimorelin in step (a) are provided. The two blood samples may alternatively be administered at about 20±10 minutes, about 30±10 minutes, about 40±10 minutes, about 45±10 minutes, about 50±10 minutes, about 60±10 minutes, can be obtained from the subject at about 70±10 minutes, about 80±10 minutes, about 90±10 minutes, or about 100±10 minutes. Two blood samples may be obtained at any time in the range of about 15 to about 100 minutes after administration of masimorelin as deemed appropriate by the attending physician.

In another preferred embodiment, two blood samples obtained from said subject at about 30±10 minutes and about 60±10 minutes after administration of masimorelin in step (a) are provided.

In another preferred embodiment, three blood samples obtained from said subject are provided at about 30±10 minutes, about 45±10 minutes, and about 60±10 minutes after administration of masimorelin in step (a), or at step (a) ), at about 30±10 min, about 45±10 min, and about 90±10 min after administration of masimorelin, three blood samples obtained from the subject are provided, or at about 30±10 min after administration of masimorelin in step (a) Three blood samples obtained from the subject are provided at ±10 min, at about 60±10 min, and at about 90±10 min. The three blood samples may alternatively be administered at about 20±10 minutes, about 30±10 minutes, about 40±10 minutes, about 45±10 minutes, about 50±10 minutes, about 60±10 minutes, can be obtained from the subject at about 70±10 minutes, about 80±10 minutes, about 90±10 minutes, or about 100±10 minutes. Three blood samples may be obtained at any time in the range of about 15 to about 100 minutes after administration of masimorelin as deemed appropriate by the attending physician.

If more than three blood samples are provided, these blood samples may be obtained at any time within the range of about 15 to about 100 minutes after administration of masimorelin as deemed appropriate by the attending physician. For example, suitable time points are about 20±10 minutes, about 30±10 minutes, about 40±10 minutes, about 45±10 minutes, about 50±10 minutes, about 60±10 minutes, about 70 minutes after administration of massimorelin. ±10 min, about 80±10 min, about 90±10 min, or about 100±10 min.

In a particularly preferred embodiment, from the subject at a time selected from the group consisting of about 30±10 minutes, about 45±10 minutes, about 60±10 minutes, and about 90±10 minutes after the administration of masimorelin in step (a). The obtained, 1 to 4, more preferably 1 to 3, more preferably 2 or 3 blood samples are provided.

In a more preferred embodiment, in step (a), masimorelin is administered in the form of a composition further comprising masimorelin and optionally a pharmaceutically acceptable excipient such as a carrier material. Preferably, the masimorelin is administered in the form of a composition comprising masimorelin and a sweetener. A suitable sweetener is, for example, saccharin. It has been found that saccharin is a suitable taste masking agent for marshmorelin.

In a more preferred embodiment, in step (a) about 3.5% (w/w) of masimorelin (calculated as free base), about 93.1% (w/w) of spray dried lactose monohydrate, about 2.0% ( w/w) crospovidone A, about 0.1% (w/w) colloidal silicon dioxide, about 1.0% (w/w) sodium stearyl fumarate, and about 0.3% (w/w) saccharin Massimorelin in the form of a composition comprising sodium dihydrate is administered.

In a more preferred embodiment, the subject is a human child from 2 to less than 18 years of age, preferably said subject is a human child from 2 to less than 17 years of age, more preferably said subject is from 2 to less than 16 years of age is a human child of

In a more preferred embodiment, the substance is used in a stand-alone test, does not need to be repeated, and does not require an alternative growth hormone stimulation test to reliably diagnose growth hormone deficiency in pediatric patients.

In a more preferred embodiment, the determining according to step (d) whether the subject has growth hormone deficiency is based solely on induction of growth hormone levels by a single masimorelin administration.

Surprisingly, it has been found that the method of the present invention is suitable as a stand-alone test because it does not require additional GHST to reliably diagnose GHD in pediatric patients.

The object of the present invention is surprisingly solved in another aspect by providing a screening method for diagnosing growth hormone deficiency in a pediatric patient using masimorelin, comprising:

(a) providing one or more blood samples obtained from the subject within the range of about 15 to about 100 minutes after administering an amount of masimorelin effective to induce growth hormone secretion;

(b) measuring the growth hormone level in each blood sample;

(c) comparing each measured growth hormone level to a single threshold;

(d) diagnosing whether the subject has growth hormone deficiency based on comparing the growth hormone levels measured in step (b) of the one or more blood samples to the single threshold value, wherein the subject Determining whether you have this growth hormone deficiency is a step based solely on induction of growth hormone levels by a single masimorelin administration.

In a preferred embodiment, the screening method for diagnosing growth hormone deficiency in a pediatric patient using masimorelin is an in vitro screening method for diagnosing growth hormone deficiency in a pediatric patient using masimorelin.

In a preferred method, obtained from the subject at a time selected from the group consisting of about 30±10 minutes, about 45±10 minutes, about 60±10 minutes, and about 90±10 minutes after administration of masimorelin in step (a), 1 to 4, more preferably 1 to 3, more preferably 2 or 3 blood samples are provided.

In a preferred embodiment, in step (d), it is determined that the subject having the highest growth hormone level measured in step (b) is lower than said single threshold has growth hormone deficiency, and wherein the highest growth hormone level measured in step (b) is determined A subject whose growth hormone level is not lower than this single threshold is determined not to have growth hormone deficiency.

In another preferred embodiment, the single threshold is from about 10.0 to about 25.0 ng/mL, preferably from about 10.2 to about 20.0 ng/mL, more preferably from about 12.0 to about 19.0 ng/mL, more preferably about 14.0 to about 18.0 ng/mL, more preferably about 16.0 to about 18.0 ng/mL, and most preferably about 17.0 to about 18.0 ng/mL.

In another preferred embodiment, about 0.8 mg to about 1.2 mg of masimorelin is administered per kg of subject's body weight, preferably about 1.0 mg of masimorelin is administered per kg of subject's body weight.

In a more preferred embodiment, the subject is a human child from 2 to less than 18 years of age, preferably said subject is a human child from 2 to less than 17 years of age, more preferably said subject is from 2 to less than 16 years of age is a human child of

One key feature of the method of this invention is that it is a single stimulation test format, as opposed to the standard two-test format currently used by medical professionals. Instead of having two separate tests performed more than a day apart and involving as many as 8 to 12 blood draws, this new method provides a reliable diagnosis with accuracy, specificity, and sensitivity for detecting growth hormone deficiency. To achieve performance, only 1 test and only 1 to 5, preferably 2 to 4 blood draws are required, thus greatly reducing the burden of testing and potential harm to the child being tested. Advantageously, these blood samples can be collected within a short period of time, eg, after administration of masimorelin, at intervals of about 15 minutes, up to about 30 minutes, in a total time of about 90 minutes.

Surprisingly, the method of the present invention achieved significant improvement by using a higher threshold for diagnosing growth hormone deficiency. Generally, when testing GHD in adult patients, a single threshold of about 2-3 ng/mL is used, and when testing GHD in pediatric patients, a single threshold of less than 10 ng/mL is used. We unexpectedly found that better diagnostic performance can be achieved for pediatric patients when a higher single threshold of 10.0 ng/mL or greater is used. For example, a single threshold of about 16.0 to 19.0 ng/mL, preferably about 17.0 to 18.0 ng/mL, has been found to be very effective in representing GHD in the proprietary method of this invention.

In addition, the method of the present invention achieves a significant improvement by using a higher masimorelin dose for diagnosing growth hormone deficiency. A typical masimorelin dose used in current practice is 0.5 mg/kg patient body weight, especially when testing GHD for adult patients, but we found that higher masimorelin doses in a growth hormone stimulation test for pediatric patients It has been unexpectedly found that better diagnostic performance can be achieved when a dose is used. For example, it has been found that a masimorelin dose of about 0.8 to about 1.2 mg/kg, preferably about 1.0 mg/kg of body weight, exhibits very effectively GHD in the single test method of the present invention.

The present invention is

- As a standalone test (requires one test method and single GH stimulation only),

- for example, after administration of masimorelin, 2 to 4 blood samples are collected, at intervals of about 15 minutes, up to about 30 minutes, for a total time of about 90 minutes,

- at least about 10.0 ng/mL, preferably from about 10.0 ng/mL to about 25.0 ng/mL, more preferably from about 10.2 ng/mL to about 20 ng/mL, most preferably from about 17.0 to about 18.0 ng/mL having a GH endpoint in the range of

Provided is a method of measuring growth hormone levels in a human child, comprising a method of assessing pituitary-associated GH deficiency in the human child following a single oral administration of masimorelin to the child.

The following examples are provided for illustrative purposes only, and do not limit the invention. Those of ordinary skill in the art can readily recognize several non-critical parameters that can be changed or adjusted to obtain essentially the same or similar results.

(Example)

Growth hormone deficiency (GHD) in children is a rare, etiologically diverse disease that causes growth failure and short stature. An insufficient response to two different growth hormone stimulation tests (GHST) is required for the diagnosis of GHD. Masimorelin acetate, a potent orally administered growth hormone (GH) secretagogue, is approved by the FDA and EMA for the diagnosis of GHD (AGHD) in adults. This study (AEZS-130-P01) was designed to study masimorelin acetate as a diagnostic test in children suspected of having GHD.

This is an open-label, group comparison, dose escalation trial, and the safety, tolerability, and safety, tolerability, to study pharmacokinetics, and pharmacodynamics. Masimorelin GHST was administered between two standard GHST doses, performed according to local clinical practice, with a 7-28 day recovery period between tests. Blood samples were collected predose (±15 min) and at 15 min, 30 min, 45 min, 60 min, 90 min, 120 min, and 360 min post-masimorelin acetate infusion.

Overall, 24 pediatric patients (8 per cohort [C1, C2, C3]) were included in the pharmacokinetic/pharmacodynamic (PK/PD) analysis. 5 males and 3 females were observed in C1 and C2, and 7 males and 1 female were observed in C3. In all three cohorts, at least three subjects were in Tanner stage I or II. All 24 subjects (100%) were Caucasian, had a median age of 9.8, 9.0, and 10.5 years (range 4-15 years), and had a median body mass index of 16.1 kg/m ² (12.4-21.4 kg) at screening. /m ² ). Overall, 88 adverse events were reported, many of which were related to standard GHST; There was nothing related to the marshmorelin test. The highest plasma concentrations of marshmorelin were observed mainly between 30 and 45 minutes. The mean C _max values were 3.46 ng/mL, 8.13 ng/mL, and 12.87 ng/mL for C1, C2, and C3, respectively. AUC increased with dose; The mean AUC0-6 values were 6.69 h*ng/mL, 18.02 h*ng/mL, and 30.92 h*ng/mL. The mean excretion half-lives were 1.22 h, 1.61 h, and 1.71 h, respectively. The PK and PD profiles of all three cohorts were similar, with peak GH levels observed predominantly within 30-60 minutes after masimorelin infusion.

Masimorelin acetate was safe and well tolerated in all dosing cohorts. Dose-dependent increases in masimorelin C _max and AUC in children and adolescents correlated well with data from adult subjects. A strong dose proportional GH response was also achieved. The PD results indicated that the GH response was similar in all administration groups, and that there was a slight change in t _max earlier as the massimorelin dose increased.

Moreover, the results of the masimorelin GHST showed a surprisingly high agreement with the results of the two standard GHSTs, as well as the final diagnosis evaluated by the principal investigator. At C3, GH secretion was clearly stimulated, for all 8 patients. Finally, the results of masimorelin GHST applied as a single test showed agreement with the results of the combination of two sGHSTs as well as the PI assessment for 7 out of 8 subjects.

Example 1: Massimorelin-containing composition for CGHD diagnosis

The massimorelin-containing composition includes the following ingredients listed in Table 1.

One unit dose consists of a composition containing 1817.2 mg masimorelin for preparation as an oral suspension. Generally, the prepared suspension contains 0.5 mg of marshmorelin per mL of suspension.

Small portions of the reconstituted suspension adjusted for body weight are administered to pediatric subjects to a dose of 1.0 mg/kg body weight for the child. It should be noted that aliquots of the reconstituted suspension adjusted for body weight are administered to adults so that the dose for adult subjects is 0.5 mg/kg body weight.

The unit dose is defined as masimorelin calculated as free base in a content of 100%. The mass of masimorelin free base or its free base equivalent in the unit dose may be adjusted according to the content.

Masimorelin, as a suitable pharmaceutical salt, may be included in the unit dosage. Examples of suitable pharmaceutical salts are acetate salts and trifluoro acetate salts.

The unit dose may be contained in an appropriate container for easy use of the GHD test. Examples of suitable containers are bags or containers of suitable size made of glass or plastic.

For a 60 kg subject, the container contains 63.6 mg of masimorelin, 1691.8 mg of spray dried lactose monohydrate, 36.3 mg of crospovidone type A, 1.8 mg of colloidal silicon dioxide, 18.2 mg of sodium stearyl fumarate, and 5.5 mg of saccharin sodium dihydrate. When reconstituted in 120 mL of water, 2 mL provides 1.0 mg of marshmorelin.

Example 2 : Use of saccharin in a composition containing massimorelin to mask unpleasant taste

To determine the diagnostic efficacy of masimorelin for AGHD, in a multicenter, randomized, interactive, crossover study, 100 subjects with confirmed adult growth hormone deficiency (AGHD) were given GHRH+L-arginine and masimorelin as GHSTs. did

This study was conducted in two parts. In the first part, the masimorelin-containing composition described in Example 1 was used at a dose of 0.5 mg/kg, except without saccharin, mild to moderate in 12 of 52 subjects (21%). An unpleasant taste of

While the study was discontinued, it was found that saccharin was a suitable taste masking agent, although it still did not completely mask the bitter taste. In the second part of the study, the masimorelin-containing composition (with saccharin) described in Example 1 was used, and only one (2%) of 48 test subjects reported an unpleasant taste of mild intensity.

Example 3: Pharmacokinetic, pharmacodynamic, and exploratory test properties of Masimorelin as a diagnostic test

Study AEZS-130-P01 is an open-label, group comparison, dose escalation trial, for pediatric patients with suspected GHD, after a single oral dose at 0.25 mg/kg, 0.5 mg/kg, and 1 mg/kg, Masimorel To study the safety, tolerability, pharmacokinetics (PK) and pharmacodynamics (PD) of lean acetate.

Objectives are defined as follows:

Primary:

To study the safety and tolerability of Masimorelin Acetate following escalation of a single oral dose of Masimorelin in pediatric patients suspected of having GHD.

Secondary:

ㆍTo study the PK of masimorelin acetate in pediatric patients suspected of having GHD.

ㆍTo study the PD of masimorelin acetate as measured by growth hormone (GH) secretion in pediatric patients suspected of having GHD.

To analyze the PK/PD relationship following administration of a single oral dose of masimorelin acetate in pediatric patients suspected of having GHD.

methodology:

Plasma concentrations of masimorelin and serum concentrations of GH were analyzed in a central laboratory.

Massimorelin Plasma Concentrations: Analysis of plasma samples for massimorelin concentrations was performed at a central laboratory, Prolytic GmbH, Germany, using effective liquid chromatography-mass spectrometry (LCMS/MS) with a detection limit of 0.2 ng/mL. used (Franz, 2005).

Preliminary pharmacokinetics (PK) were determined by the peak concentration (C _max ) of the masimorelin plasma concentration and the time to measure the peak concentration (t _max ) within the sampling cycle.

GH Serum Concentration: Analysis of serum samples for GH concentration was performed in a central laboratory by effective immunochemiluminescence assay (IDS-iSYS Human Growth Hormone (hGH), Immunodiagnostic Systems Ltd [UK]) (Manolopoulou et al. , 2012). This assay is standardized in the recombinant growth hormone calibration standard WHO 98/574 and follows the recommendations for assay standardization described by Clemmons (Clemmons et al., 2011).

Analytical laboratories that received GH were Central Laboratory Synevo Lodz, Krakusa Str. 28, 93-515 Lodz. It was Poland. The lower limit of quantification was <0.05 ng/mL.

number of patients

Overall, masimorelin was administered as a single oral dose to 24 pediatric patients suspected of having GHD, 8 patients per dosing group. Of these 8 patients, at least 3 patients per dosing group were prepubertal (Tanner stage I) and pubertal (Tanner stage II-IV), respectively.

Key criteria for persons included as patients:

Subjects must meet all of the following criteria to be eligible to participate in the trial.

1. Male or female pediatric patients aged 2 to less than 18 years;

2. Suspected GHD based on developmental and clinical criteria;

3. Indicators for the performance of the induced growth hormone stimulation test (GHST).

Subjects who have had sex hormone injections prior to GHST as part of standard diagnostic procedures should also receive sex hormone injections for masimorelin GHST.

Investigational drug (IMP) Masimorelin

A disposable aluminum pouch (syn.: bag) contained 63.6 mg of marshmorelin, and when dissolved in 120 mL of water, provides 0.5 mg/mL of marshmorelin.

A sequential cohort of trial participants received masimorelin in increments of a single oral dose: 0.25 mg/kg body weight in Cohort 1 (C1), 0.5 mg/kg body weight in Cohort 2 (C2), and Cohort 3 ( C3) at 1 mg/kg body weight.

To determine masimorelin PK/PD, blood samples were obtained pre-dose and at 15 min, 30 min, 45 min, 60 min, 90 min, 120 min, and 360 min post-massmorelin administration.

As a test, a single oral dose of Masimorelin Acetate was administered to the patient on the day the Masimorelin GHST was performed.

Masimorelin GHST Preparation

Masimorelin oral suspensions were prepared by test personnel and administered according to the instructions below (steps #1 to #5), taking the dose of cohort C3 (ie, 1.0 mg/kg) as an example.

1. Weigh the patient and determine the number of pouches/bags required (one pouch may be needed for the patient). Weight (reported in kg) may be rounded to the nearest whole number;

2. In a suitable clear glass or polypropylene container, dissolve the entire contents of the pouch in 120 mL of water (i.e., once applied, one pouch is 120 mL, two pouches are 240 mL), and gently agitate for at least 3 minutes (a small amount of dissolved particles may remain);

3. Based on the masimorelin dose of 1.0 mg/kg, determine the required volume of suspension corresponding to the patient's body weight, i.e. the volume of suspension required is 2 mL/kg (e.g., 1.0 mg/kg of A 30 kg patient requiring a marshmorelin dose may require 60 mL of the prepared suspension);

4. Using a graduated syringe, measure the patient's required volume and transfer it to a glass cup (to minimize the risk of loss due to incomplete drainage, volumes less than 20 mL should be administered by oral syringe; see instructions below);

5. The suspension should be used within 30 minutes of preparation.

Fasting before GHST

Patients were required to fast for 8 hours prior to initiation and throughout the sampling period of the marshmorelin stimulation test.

administration

Administration of the marshmorelin oral suspension was under the supervision of test personnel. Following step #4 of the specific cohort dosing instructions as described above, the entire contents of the prepared vials were allowed to drink by the patient - for a period of not more than 30 seconds.

blood sampling

Blood samples were collected at the following time points: pre-dose (sampling time frame: +/- 15 min), and 15 min, 30 min, 45 min, 60 min, 90 min, 120 min (+/- 5 min) after masimorelin administration. minute time zone), and 360 minutes (+/- 10 minute time zone). Serum concentrations of GH and plasma concentrations of masimorelin were analyzed in a central laboratory.

The test masimorelin GHST was performed after the first sGHST was completed. In order to avoid carryover effects or interference between subsequent GHSTs and to provide an adequate follow-up period for the observation of possible drug-related adverse events by previously used inducers, a recovery period of at least 1 week and up to 4 weeks is introduced between GHSTs. .

Standard GHST used in P01 study

Two standard growth hormone tests (sGHST) were to be performed on patients, according to local practice. The sGHST formulation was considered as a 'background' and not as an IMP.

The following pharmacological agents were accepted as sGHST: insulin (insulin tolerance test (ITT)), arginine, arginine/growth hormone releasing hormone (GHRH), clonidine, glucagon, L-dopa.

A single dose of the sGHST formulation is administered by intramuscular injection (i.m.), intravenous injection (i.v.), subcutaneous injection (s.c.), or orally (depending on the formulation) on the day sGHST is performed. Batch numbers were recorded on-site in the patient record and in the ‘Standard GHST Patient Responsibility Record’.

Evaluation standard

test endpoint

Safety and Tolerability

• Patient tolerability (including taste receptivity and effects on sleep, appetite, and gastrointestinal symptoms), adverse events (AEs);

• Determination of changes in laboratory parameters relevant to safety;

ㆍEffects on biological parameters (pulse rate, blood pressure, ECG).

Pharmacokinetics

• Concentration-time profile of masimorelin;

ㆍTarget parameters: AUC, C _max , T _max , T _1/2 .

pharmacodynamics

• Concentration-time profile of GH;

ㆍTarget parameters: C _max , T _max ;

• Preliminary PK/PD: T _max of Masimorelin vs. T _max of GH; C _max of marshmorelin vs. C _max of GH .

Etc

• Establishment of recommended doses for diagnostic purposes in pediatric patients suspected of having GHD;

• Analysis of appropriate GH endpoints for follow-up examination, to establish a diagnosis of GHD in pediatric patients.

Statistical method:

All statistical analyzes are considered exploratory in nature. Data sets were analyzed using SAS version 9.3 or later.

In general, summary statistics of quantitative variables (n, arithmetic mean, standard deviation, median, minimum, and maximum) and frequency tables of qualitative data were presented by administration group.

Masimorelin PK: PK parameters were analyzed for the PK analysis set (PKS), by n (number of measurements), arithmetic mean, standard deviation and coefficient of variation (CV), median, minimum, maximum, and also (T _max excluded) by geometric mean, geometric standard deviation, and geometric CV. For T _max , additional median, minimum, and maximum values, as well as frequencies, were shown.

Masimorelin PD: Regardless of the validity of the PK data, GH concentration data were analyzed for the PD Analysis Set (PDS). The GH peak concentration was correlated with the outcome of the clinical diagnostic method (diagnosing whether or not GHD was confirmed).

PK/PD analysis: Plasma concentrations of masimorelin in individual patients were correlated with each GH concentration at the same time, as well as outcomes of clinical diagnostic methods (diagnosing whether GHD was confirmed).

Results and Conclusions:

Of a total of 27 screened patients, 24 patients received the masimorelin test, and there were 8 patients in each of the 3 dosing cohorts (C1, C2, and C3).

Therefore, the PK analysis set (PKS), PD analysis set (PDS), and PK/PD set, as well as the safety population (SAF) consisted of 24 patients.

Baseline characteristics:

Overall, 17 patients (70.8%) were male, 7 patients (29.2%) were female, and 100% were Caucasian. At screening, the median parameters for all three dosing cohorts were: age 10.5 years (range: 4-15 years), height 123.35 cm (range: 46.0-152.5 cm), body weight 25.5 kg (range: 12-43 kg), and body mass The index (BMI) was 16.1 kg/m ² (range: 12.4 to 21.4 kg/m ² ).

Tanner status was distributed as follows: in C3 as well as in C1, 4 patients exhibited Tanner I, 4 patients exhibited Tanner II, 5 patients in C2 exhibited Tanner I, and 3 patients exhibited Tanner I. Shown Tanner II. Sex hormone injection was applied by intramuscular injection administration of testosterone depot to 2 male patients in C3.

As for the baseline history, another pituitary axis deficiency (ie, thyroid deficiency) was reported for only 2 patients in C2. As part of a standard 'workup', IGF-1 and IGF-BP3 values were found on electronic case records (eCRFs) collected according to local diagnostic criteria.

IGF-1 values were present in 7 patients in C1 and 8 patients each in C2 and C3, with median values of 88.00 μg/L in C1 (SD 68.72), 100.00 μg/L in C2 (SD 97.90), and C3. at 119.50 μg/L (SD 68.88). IGF-BP3 values were obtained for one patient in C3.

Bone age showed a median value of 102.2 months (range: 24-156 months). As part of the developmental parameters, the median renal SDS was -2.35 (range: -3.2 to 1.7), the median BMI SDS was -0.60 (range: -2.1 to 2.0), and the median annual renal rate SDS was -1.50 (range: -3.3 to 0.5).

SGHST: Overall, ITT was administered to 22 patients (ie, 5 patients (20.8%) at visit 1 (V1) and 17 patients (70.8%) at visit 3 (V3)) and arginine in 8 patients at V1 Patients (33.3%) were administered and clonidine was administered to 16 patients (ie, 11 patients (45.8%) in V1 and 5 patients (20.8%) in V3). Glucagon was administered to only one patient and no L-dopa was administered.

Treatment compliance to marshmorelin was 100% in all three cohorts.

Pharmacokinetic and Pharmacodynamic Results:

Plasma Concentration Data

In general, masimorelin plasma concentrations displayed a dose-dependent increase ( FIG. 1 ), with high inter-individual variability. Plasma concentrations after administration of masimorelin showed a sharp increase with the highest level observed between 0.25 and 2 hours. At the time of the last sampling, 6 hours post-dose, plasma concentrations were strongly reduced.

Pharmacokinetics:

In general, masimorelin plasma concentrations displayed a dose-dependent increase ( FIG. 1 ), with high inter-individual variability. Plasma concentrations after administration of masimorelin showed a sharp increase with the highest level observed between 0.25 and 2 hours. At the time of the last sampling, 6 hours post-dose, plasma concentrations were strongly reduced.

AUC and C _max of masimorelin show a dose-dependent increase, with an arithmetic mean AUC0-6 of 6.69 h*ng/mL at C1, 18.02 h*ng/mL at C2, and 30.92 h*ng/mL at C3, The arithmetic mean C _max is 3.46 ng/mL for C1, 8.13 ng/mL for C2, and 12.87 ng/mL for C3 (Table 2).

The mean T _max was similar among all three groups, and the arithmetic mean was 45.5 min at C1, 40.6 min at C2, and 31.9 min at C3. The mean T _1/2 shows a slight increase with higher doses, ie, 73.18 min at C1, 96.31 min at C2, and 102.85 min at C3.

Pharmacodynamics:

As shown in FIG. 2 , the GH concentration is increasing after masimorelin administration, and the value tends to become higher as the dose increases. Large inter-individual variability is expected in the observed population with persons suspected of having GHD.

After masimorelin administration, peak GH levels are within 0.5 to 1 hour at C1 (mean T _max is 52.5 minutes (SD 11.3)) and 0.25 to 1 hour at C2 (mean T _max is 37.5 minutes (SD 13.9)), and within 0.5 to 0.75 h at C3 (mean T _max is 37.5 min (SD 8.0)) (see Table 3).

Exploratory analysis for GH endpoints:

Peak GH values by GHD diagnosis were compared based on GHST results and the investigator's evaluation. The diagnostic properties of the tested GH values as endpoints (i.e., sensitivity, specificity, and Yuden index (unweighted, weighted)) were listed as the most definitive representations of the diagnostic properties, with a peak GH in C1 of 10.03 ng/mL, Peak GH at C2 was 10.43 ng/mL and peak GH at C3 was 17.13 ng/mL.

The diagnostic results of GHST are shown in Table 4. In this table, the diagnostic result of sGHST is considered ‘confirmed’ if two sGHSTs are provided, and both sGHSTs have a peak GH≤7ng/mL, and ‘unconfirmed’ if one or more peaks are greater than 7ng/mL. Results 'unconfirmed' are classified as 'excluded' if two sGHST results are provided and a GH peak > 7 ng/mL, and 'unclear' if none of the above are met. The Investigator's evaluation was based on local diagnostic standard practice. Masimorelin GHST was tested against endpoints calculated from individual peak GH values.

Based on the above considerations, Table 4 shows the agreement between the PI's assessment and the results of both sGHSTs: in 21 (87.5%) patients (i.e., 8 confirmed and 13 unconfirmed). , there was agreement between the investigator's assessment and the sGHST results. For 3 (12.5%) patients, the investigator concluded with GHD, but sGHST excluded the diagnosis (1 patient) or was unclear (2 patients).

Furthermore, the diagnostic results can be summarized as follows (Table 5): Masimorelin GHST was 1 in C2 (9.09%) of 11 total patients assessed as 'GHD' by the investigator in all 3 cohorts. 'GHD unconfirmed', only for patients of .

Of a total of 13 patients assessed by the investigator as 'unconfirmed' to have GHD, masimorelin GHST was administered in 3 (23.08%) patients in C1 and 1 (7.69%) in C3 patients, respectively. GHD was checked.

Considering the data presented above, the strongest test characteristic of the masimorelin test is observed at C3: GH secretion was clearly stimulated in all eight patients. Finally, the results of masimorelin GHST applied as a single test showed agreement with the results of the combination of two sGHSTs as well as the PI assessment for 7 out of 8 subjects.

Receptor Response Characteristics (ROC) Analysis

For all GH endpoints tested, the ROC curve of C1 showed the lowest sensitivity and specificity compared to C2 and C3 ( FIG. 3 ). The relevant area under the curve (AUC) increases with increasing dose.

Comparing the characteristics of the GH endpoints between the three cohorts, the GH endpoint of 17.130 ng/mL of C3 and the sensitivity of 1.0, specificity of 0.8, Yuden index ≥ 0.80, and ROC AUC of 0.93 show the strongest test characteristics. (See Table 6).

Sensitivity analysis was performed by observing the development of ROC AUC based on test results of sGHST classified as ‘confirmed’ versus ‘unconfirmed’. Again, compared to C2 (sensitivity 0.75, specificity 0.75, ROC AUC 0.563) and C1 (sensitivity 1.0, specificity 0.71, ROC AUC 0.714), the strongest test characteristic, e.g., a sensitivity of 1.00, a specificity of 0.80 , and ROC AUC of 0.933 were shown in C3.

PK and PD Summary

Overall, masimorelin PK and PD of C1, C2, and C3 show similar profiles:

• Marsimorelin T _max was similar in all three groups, with mean T _max values of about 0.5 to 0.75 hours;

• mean masimorelin C _max shows a dose proportional increase;

• AUC increases with masimorelin dose;

ㆍMaximum GH secretion is observed 0.25 to 2 hours after masimorelin administration, and the mean T _max value is about 0.5 to 1 hour.

At C3 upon administration of masimorelin at 1.0 mg/kg body weight, the maximum values of AUC and C _max are observed. Moreover, the sensitivity analysis supports dosing at C3, which has the strongest assay characteristics exhibited by an approximate 17 ng/mL GH endpoint and a specificity of 0.80, a sensitivity of 1.00, a Yuden index of 0.80, and a ROC AUC of 0.933.

Safety Results:

Overall, 88 AEs were observed in 23 patients with SAF, i.e., 27 events in 8 patients in C1, 28 events in 8 patients in C2, and 33 events in 7 patients in C3. .

A total of 70 treatment-induced adverse events (TEAEs) were observed in 21 patients with SAF, i.e., 22 events in 8 patients in C1, 24 events in 6 patients in C2, and 24 events in 7 patients in C3. Suggestions were observed.

reported to be related to the marshmorelin test There was no TEAE .

During the course of this trial, there were no reported SAEs or severe TEAEs . No AEs or TEAEs leading to patient discontinuation were reported.

Most AEs were related to ITT, ie, 62 events (70.5%) were related in 21 (91.3%) patients. Clonidine-related AEs (13 (14.8%)) were observed in 7 (30.4%) patients, and arginine-related AEs were reported in 1 (4.3%) patient. It should be noted that ITT was administered to 22 patients, arginine was administered to 8 (33.3%) patients, clonidine was administered to 16 patients, and glucagon was administered to only 1 patient.

As AEs, infectious disease, a single case of anal fissure, and adverse events of the test formulation used for sGHST have been reported.

AEs were mainly of mild to moderate intensity. ITT-related AEs included symptoms of hypoglycemia (eg, tremor, sweating) that were the clinical endpoints of this sGHST. With regard to clonidine, hypotension-related symptoms known as side effects of this sGHST agent have been reported.

As a TEAE of one patient in Cohort 1, a severe severity was reported: 5 AEs (ie, palpitations, tachycardia, hunger, hyperhidrosis, and tremor) as part of expected hypoglycemia, HU01-01 patient during ITT occurred from

Clinical laboratory, vital signs, physical examination, ECG:

No clinically significant changes were observed in safety clinical laboratory parameters, vital signs, and physical examination. Clinically significant abnormalities and significant changes in ECG parameters are not described.

Tolerability questionnaire:

The GHST tolerability questionnaire was completed by the patient or parent/legal guardian.

Overall, there was a majority or strong agreement for the pre-determined phrase for masimorelin in all three dosing groups.

In C1, 1 case of 'acceptable taste' and 1 case of disagreement regarding 'good stomach condition the next day' were recorded. 1 case of strong disagreement with 'a bowel movement the next day' was checked.

No strong disagreement or disagreement was recorded on any of the C2 massimorelin test-related questionnaires.

In C3, there was one non-consent to 'acceptable taste'.

In the handwritten evaluation, two patients reported a 'bitter taste' after masimorelin GHST. These assessments were not evaluated as AEs by the investigator.

Overall, the feedback provided by the patient or parent/guardian after the masimorelin test demonstrates good tolerability and safety of the test.

conclusion:

This trial was conducted to study the safety, tolerability, PK and PD of masimorelin acetate following single oral doses of 0.25 mg/kg, 0.5 mg/kg, and 1.0 mg/kg in pediatric patients suspected of having GHD. carried out In addition, this trial provides for the identification of appropriate masimorelin doses for further testing within the test efficacy trial, and also the analysis of test GH endpoints.

In all three dosing cohorts, the C _max and T _max for plasma levels of masimorelin were found to be within the expected ranges from the adult development program.

In the observed pediatric population, a dose-dependent increase in masimorelin C _max (3.46 vs. 8.13 vs. 12.87 ng/mL) was observed as well as mean AUC0-6 (6.69 vs. 18.02 vs. 30.92 h*ng/mL). The elimination half-life t _1/2 was in the range of 1.25 to 1.75 h.

In general, PK parameters in the pediatric population were within a range similar to those of adults.

Masimorelin dose (C1) of 0.25 mg/kg did not result in maximal stimulation of GH secretion in children, which resulted in a comparison of the consistency of the analyzed GH endpoints to the results of the PI assessment and sGHST, as well as the PK/PD It became clear through a review of the data.

A dose of 0.5 mg/kg (C2) showed strong GH secretion, a ROC AUC of 0.80, as well as a strong level of agreement between the principal investigator (PI) assessment based on masimorelin GHST versus sGHST. However, a dose of 1.0 mg/kg (C3) appears to lead to more sustained and stronger GH stimulation, probably due to sufficiently strong masimorelin exposure for all subjects.

Finally, the sensitivity analysis supports dosing at C3, which has the strongest assay characteristics, with an endpoint of approximately 17 ng/mL GH and specificity of 0.80, sensitivity of 1.00, Yuden index of 0.80, and ROC AUC of 0.93. .

Moreover, the results of the masimorelin GHST showed a surprisingly high agreement with the results of the two standard GHSTs, as well as the final diagnosis evaluated by the principal investigator. At C3, GH secretion was clearly stimulated, for all 8 patients. Finally, the results of masimorelin GHST applied as a single test showed agreement with the results of the combination of two sGHSTs as well as the PI assessment for 7 out of 8 subjects.

In conclusion, in all three dosing cohorts, masimorelin showed good safety and tolerability, with no reported AEs in the observed population. PK and PD profiles are within the expected range from an adult development program. The overall properties of masimorelin in the first pediatric trial supported the selection of a masimorelin dose of 1.0 mg/kg in the study of a phase 3 trial of trial efficacy.

reference list

Claims (39)

A screening method for diagnosing growth hormone deficiency in a pediatric patient using masimorelin, comprising:
(a) providing 1 to 5 blood samples obtained from the subject within the range of about 15 to about 100 minutes after administration of an amount of masimorelin effective to induce growth hormone secretion;
(b) measuring the growth hormone level in each blood sample provided in step (a);
(c) comparing the measured growth hormone level obtained in step (b) with a single threshold, wherein the single threshold is greater than or equal to 10.0 ng/mL;
(d) determining as having growth hormone deficiency the subject having the highest growth hormone level in the blood sample obtained in step (b) below said single threshold, wherein the highest growth hormone level in the blood sample obtained in step (b) is determining a subject not lower than the single threshold as not having growth hormone deficiency. The method of claim 1,
The single threshold is from about 10.0 to about 25.0 ng/mL, preferably from about 10.2 to about 20.0 ng/mL, more preferably from about 12.0 to about 19.0 ng/mL, more preferably from about 14.0 to about 18.0 ng/mL. mL, more preferably from about 16.0 to about 18.0 ng/mL, and most preferably from about 17.0 to about 18.0 ng/mL. 3. The method of claim 1 or 2,
1 to 4 blood samples are provided in step (a), preferably 1 to 3 blood samples are provided in step (a), more preferably 2 or 3 blood samples are provided in step (a) how to be 4. The method according to any one of claims 1 to 3,
In step (a), the blood sample is administered within a range of about 20 to about 100 minutes, preferably within a range of about 25 to about 100 minutes, more Preferably within the range of about 25 to about 95 minutes, and most preferably within the range of about 30 to about 90 minutes. 5. The method according to any one of claims 1 to 4,
Where more than one blood sample is provided, said blood sample is obtained from said subject at about 10 to about 60 minute intervals, preferably about 15 to about 30 minute intervals. 6. The method according to any one of claims 1 to 5,
The method in step (a) wherein the blood sample is a serum sample or a plasma sample. 7. The method according to any one of claims 1 to 6,
A method wherein in step (a) about 0.8 mg to about 1.2 mg of masimorelin per kg of subject's body weight is administered, and preferably, in step (a), about 1.0 mg of masimorelin per kg of subject's body weight is administered. 8. The method according to any one of claims 1 to 7,
In step (a), the administration of masimorelin is oral administration. 9. The method according to any one of claims 1 to 8,
A method wherein one blood sample obtained from said subject is provided about 60±30 minutes after administration of masimorelin in step (a). 9. The method according to any one of claims 1 to 8,
wherein two blood samples obtained from said subject are provided at about 30±10 minutes and about 45±10 minutes after administration of masimorelin in step (a). 9. The method according to any one of claims 1 to 8,
wherein two blood samples obtained from said subject are provided at about 30±10 minutes and about 60±10 minutes after administration of masimorelin in step (a). 9. The method according to any one of claims 1 to 8,
Three blood samples obtained from the subject are provided at about 30±10 minutes, about 45±10 minutes, and about 60±10 minutes after the administration of masimorelin in step (a), or administration of masimorelin in step (a) Three blood samples obtained from the subject are provided at about 30±10 minutes, about 45±10 minutes, and about 90±10 minutes after, or about 30±10 minutes, about 60 minutes after masimorelin administration in step (a) A method in which three blood samples obtained from the subject are provided at ±10 minutes, and at about 90 ±10 minutes. 13. The method according to any one of claims 1 to 12,
Administering masimorelin in the form of a composition further comprising masimorelin and optionally a pharmaceutically acceptable additive in step (a), preferably comprising masimorelin and optionally saccharin in step (a) A method of administering marshmorelin in the form of a composition. 14. The method according to any one of claims 1 to 13,
Masimorelin calculated as about 3.5% (w/w) free base in step (a), about 93.1% (w/w) spray dried lactose monohydrate, about 2.0% (w/w) type A cross in the form of a composition comprising povidone, about 0.1% (w/w) colloidal silicon dioxide, about 1.0% (w/w) sodium stearyl fumarate, and about 0.3% (w/w) saccharin sodium dihydrate How to administer Masimorelin. 15. The method according to any one of claims 1 to 14,
wherein said subject is a human child between the ages of 2 and less than 18 years of age. 16. The method according to any one of claims 1 to 15,
The method is a stand-alone test, does not need to be repeated, and does not require an alternative growth hormone stimulation test for reliably diagnosing growth hormone deficiency in pediatric patients. 16. The method according to any one of claims 1 to 15,
A method wherein the determining according to step (d) whether the subject has growth hormone deficiency is based solely on induction of growth hormone levels by a single masimorelin administration. Masimorelin substances for use in diagnosing growth hormone deficiency in pediatric patients, comprising:
(a) providing 1 to 5 blood samples obtained from the subject within the range of about 15 to about 100 minutes after administration of an amount of masimorelin effective to induce growth hormone secretion;
(b) measuring the growth hormone level in each blood sample provided in step (a);
(c) comparing the measured growth hormone level obtained in step (b) with a single threshold, wherein the single threshold is greater than or equal to 10.0 ng/mL;
(d) determining as having growth hormone deficiency the subject having the highest growth hormone level in the blood sample obtained in step (b) below said single threshold, wherein the highest growth hormone level in the blood sample obtained in step (b) is determining a subject not lower than the single threshold as not having growth hormone deficiency. 19. The method of claim 18,
The single threshold is from about 10.0 to about 25.0 ng/mL, preferably from about 10.2 to about 20.0 ng/mL, more preferably from about 12.0 to about 19.0 ng/mL, more preferably from about 14.0 to about 18.0 ng/mL. mL, more preferably from about 16.0 to about 18.0 ng/mL, and most preferably from about 17.0 to about 18.0 ng/mL. 20. The method according to claim 18 or 19,
1 to 4 blood samples are provided in step (a), preferably 1 to 3 blood samples are provided in step (a), more preferably 2 or 3 blood samples are provided in step (a) Masimorelin Substances for Uses 21. The method according to any one of claims 18 to 20,
In step (a), the blood sample is administered within a range of about 20 to about 100 minutes, preferably within a range of about 25 to about 100 minutes, more Masimorelin material for use obtained from a subject, preferably within the range of about 25 to about 95 minutes, and most preferably within the range of about 30 to about 90 minutes. 22. The method according to any one of claims 18 to 21,
Masimorelin material for use, if more than one blood sample is provided, the blood sample is obtained from the subject at about 10 to about 60 minute intervals, preferably about 15 to about 30 minute intervals. 23. The method according to any one of claims 18 to 22,
Massimorelin material for use in step (a), wherein said blood sample is a serum sample or a plasma sample. 24. The method according to any one of claims 18 to 23,
Masimorelin for use in which about 0.8 mg to about 1.2 mg of masimorelin per kg of subject's body weight is administered in step (a), preferably about 1.0 mg of masimorelin per kg of subject's body weight is administered in step (a) matter. 25. The method according to any one of claims 18 to 24,
Massimorelin substance for use wherein the administration of masimorelin in step (a) is oral administration. 26. The method according to any one of claims 18 to 25,
A masimorelin substance for use, wherein one blood sample obtained from said subject is provided about 60±30 minutes after administration of masimorelin in step (a). 26. The method according to any one of claims 18 to 25,
A masimorelin material for use, wherein two blood samples obtained from the subject are provided at about 30±10 minutes and about 45±10 minutes after the administration of masimorelin in step (a). 26. The method according to any one of claims 18 to 25,
A masimorelin material for use, wherein two blood samples obtained from said subject are provided at about 30±10 minutes and about 60±10 minutes after administration of masimorelin in step (a). 26. The method according to any one of claims 18 to 25,
Three blood samples obtained from the subject are provided at about 30±10 minutes, about 45±10 minutes, and about 60±10 minutes after the administration of masimorelin in step (a), or administration of masimorelin in step (a) Three blood samples obtained from the subject are provided at about 30±10 minutes, about 45±10 minutes, and about 90±10 minutes after, or about 30±10 minutes, about 60 minutes after masimorelin administration in step (a) A masimorelin material for use in which three blood samples obtained from said subject are provided at ±10 minutes, and at about 90 ±10 minutes. 30. The method according to any one of claims 18 to 29,
Administering masimorelin in the form of a composition further comprising masimorelin and optionally a pharmaceutically acceptable additive in step (a), preferably comprising masimorelin and optionally saccharin in step (a) A masimorelin substance for use in administering masimorelin in the form of a composition. 30. The method according to any one of claims 18 to 29,
Massimorelin calculated as about 3.5% (w/w) free base in step (a), about 93.1% (w/w) spray dried lactose monohydrate, about 2.0% (w/w) type A cross in the form of a composition comprising povidone, about 0.1% (w/w) colloidal silicon dioxide, about 1.0% (w/w) sodium stearyl fumarate, and about 0.3% (w/w) saccharin sodium dihydrate A masimorelin substance for use in administering masimorelin. 32. The method according to any one of claims 18 to 31,
A masimorelin substance for use, wherein said subject is a human child between the ages of 2 and less than 18 years of age. 33. The method according to any one of claims 18 to 32,
A masimorelin substance for use in which the substance is used in a stand-alone test, does not need to be repeated, and does not require an alternative growth hormone stimulation test for reliably diagnosing growth hormone deficiency in pediatric patients. 34. The method according to any one of claims 18 to 33,
A masimorelin substance for use, wherein the determining according to step (d) whether the subject has growth hormone deficiency is based solely on induction of growth hormone levels by a single masimorelin administration. A screening method for diagnosing growth hormone deficiency in a pediatric patient using masimorelin, comprising:
(a) providing one or more blood samples obtained from the subject within the range of about 15 to about 100 minutes after administering an amount of masimorelin effective to induce growth hormone secretion;
(b) measuring the growth hormone level in each blood sample;
(c) comparing each measured growth hormone level to a single threshold;
(d) diagnosing whether the subject has growth hormone deficiency based on comparing the growth hormone level measured in step (b) of the one or more blood samples to the single threshold value, wherein the subject has a growth hormone deficiency. Determining whether you have this growth hormone deficiency is a step based solely on induction of growth hormone levels by a single masimorelin administration. 36. The method of claim 35,
In step (d), it is determined that the subject having the highest growth hormone level measured in step (b) is lower than said single threshold has growth hormone deficiency, wherein the highest growth hormone level measured in step (b) is said A method of determining a subject not lower than a single threshold as not having growth hormone deficiency. 37. The method of claim 35 or 36,
The single threshold is from about 10.0 to about 25.0 ng/mL, preferably from about 10.2 to about 20.0 ng/mL, more preferably from about 12.0 to about 19.0 ng/mL, more preferably from about 14.0 to about 18.0 ng/mL. mL, more preferably from about 16.0 to about 18.0 ng/mL, and most preferably from about 17.0 to about 18.0 ng/mL. 38. The method according to any one of claims 35 to 37,
A method in which about 0.8 mg to about 1.2 mg of masimorelin per kg of subject's body weight is administered, preferably about 1.0 mg of masimorelin per kg of subject's body weight is administered. 39. The method according to any one of claims 35 to 38,
wherein said subject is a human child between the ages of 2 and less than 18 years of age.

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