Saizen - Pharmaceutical Information, Clinical Trials, Detailed Pharmacology, Toxicology.
Россия
  • Россия
  • Украина

Saizen - Scientific Information

Manufacture: EMD Serono, Inc
Country: Canada
Condition: Adult Human Growth Hormone Deficiency, Pediatric Growth Hormone Deficiency, Prader-Willi Syndrome, Short Stature for Age, Turner's Syndrome
Class: Growth hormones, Hormones
Form: Subcutaneous (SC), Powder
Ingredients: Mannitol, disodium hydrogen phosphate dihydrate, sodium dihydrogen phosphate monohydrate, sodium chloride, sucrose, phosphoric acid and sodium hydroxide.

Pharmaceutical Information

Drug Substance

Proper name: somatropin for injection
Chemical name: recombinant human Growth Hormone for Injection (r-hGH)
Molecular Formula: C990 H1528 N262 O300 S7
Structural Formula: -PHE-PRO-THR-ILE-PRO-LEU-SER-ARG-LEU-PHE
-ASP-ASN-ALA-MET-LEU-ARG-ALA-HIS-ARG-LEU-HIS-GLN-LEU-ALA
-PHE-ASP-THR-TYR-GLN-GLU-PHE-GLU-GLU-ALA-TYR-ILE-PRO
-LYS-GLU-GLN-LYS-TYR-SER-PHE-LEU-GLN-ASN-PRO-GLN-THR
-SER-LEU-CYS-PHE-SER-GLU-SER-ILE-PRO-THR-PRO-SER-ASN
-ARG-GLU-GLU-THR-GLN-GLN-LYS-SER-ASN-LEU-GLU-LEU-LEU
-ARG-ILE-SER-LEU-LEU-LEU-ILE-GLN-SER-TRP-LEU-GLU-PRO
-VAL-GLN-PHE-LEU-ARG-SER-VAL-PHE-ALA-ASN-SER-LEU-VAL
-TYR-GLY-ALA-SER-ASP-SER-ASN-VAL-TYR-ASP-LEU-LEU-LYS
-ASP-LEU-GLU-GLU-GLY-ILE-GLN-THR-LEU-MET-GLY-ARG-LEU
-GLU-ASP-GLY-SER-PRO-ARG-THR-GLY-GLN-ILE-PHE-LYS-GLN
-THR-TYR-SER-LYS-PHE-ASP-THR-ASN-SER-HIS-ASN-ASP-ASP
-ALA-LEU-LEU-LYS-ASN-TYR-GLY-LEU-LEU-TYR-CYS-PHE-ARG
-LYS-ASP-MET-ASP-LYS-VAL-GLU-THR-PHE-LEU-ARG-ILE-VAL
-GLN-CYS-ARG-SER-VAL-GLU-GLY-SER-CYS-GLY-PHE
Molecular Weight: 22, 125 daltons
Description of Drug Substance: Somatropin is a polypeptide hormone consisting of 191 amino acid residues and its structure is identical to that of growth hormone extracted from human pituitary glands. A large loop is formed by a disulfide bond between Cys53 and Cys165. A second, smaller loop is formed by a disulfide bond near the carboxyl-terminal between Cys182 and Cys189. The solution is a slightly opalescent liquid.
Biological Activity: The biological activity of growth hormone is approximately 3.0 international units/1mg

Clinical Trials

SAIZEN (somatropin for injection) is a polypeptide hormone of recombinant DNA origin which is composed of 191 amino acid residues in the identical sequence and configuration as human pituitary growth hormone. It is indicated as classical endocrine replacement therapy for the long-term treatment of patients with growth failure due to inadequate secretion of normal endogenous growth hormone. In vitro, preclinical and clinical studies have demonstrated that SAIZEN is therapeutically equivalent to human growth hormone (hGH) of pituitary origin.

Study demographics and trial design
Study demographics and design of clinical trials performed with SAIZEN in registered indications
Clinical
indication
Study
Number
Trial
design
Dosage, route of administration
and duration
Study
subjects
number
Age
range
(years)
Pediatric GHD 1. GF 2078
2. GF 2376
3. GF 2386
4. GF 2415
5. GF 2537
Each study: open-label, multicentre, Phase III 0.20 mg/kg/week, 3-6 injections
Subcutaneous administration, 2 years
304 0.0-19.0
Turner syndrome GF 3152 Open-label phase III randomised XO group: r-hGH 18 IU/m2/week, 1 year, then r-hGH 24 IU/m2/week, 1 year. XM group: r-hGH 18 IU/m2/week, 2 years and oxandrolone 0.100 mg/kg/day the 1st year, then oxandrolone 0.05 mg/kg/day the 2nd year.
Subcutaneous administration.
0/91 5.7-15.4
GF 5413 (continuation of study GF 3152) Open-label phase III randomised r-hGH 24 IU/m2/week, with or without oxandrolone 0.05 mg/kg/day, up to a total of 6 years over studies GF 3152 & 5413.
Subcutaneous administration
0/91 5.7-15.4
CRF GF 4941 Open-label, Phase III 28 IU/m2, 3 years, then 36IU/m2 if unsatisfactory growth.
Subcutaneous administration
81 1.7, 16.4
SGA GF 4001 Open-label, randomised, multicentre, Phase III r-hGH 0.067 mg/kg/day (0.2 IU/kg/day), 3 years.
Subcutaneous administration
101 1.9-8.1
GF 6283 Openlabelled, randomised, multicentre, Phase III r-hGH 0.067 mg/kg/day (0.2 IU/kg/day), 2 years.
Subcutaneous administration
58 2.0-5.0
Adult GHD GF 7364 Doubleblind, placebocontrolled, randomised, phase III (1) 0.005 mg/kg/day, 4 weeks, then 0.010 mg/kg/day, 5 months or (2) placebo, 6 months.
(1) and (2) followed by 12-30 months open-label: 0.005 mg/kg/day, 4 weeks, then 0.010 mg/kg/day.
Subcutaneous administration.
115 20.2-69.7

*: information missing in the clinical study report.

Paediatric Indications

Inadequate endogenous growth hormone secretion

Efficacy and safety of SAIZEN has been studied in five pivotal studies using pretreatment growth measurements compared with treatment growth measured as a method of control.
Of the total patients enrolled in these studies, 70.3% with at least 12 months treatment have been analyzed for efficacy. All patients were prepubertal or pubertal children with classic growth hormone deficiency with or without previous growth hormone treatment. The patients in the different studies treated with SAIZEN were assessed for the occurrence of adverse events and laboratory abnormalities and were tested regularly for the presence of antibodies against hGH and against proteins of the host cells (C-127 mouse cells).

Study Results

In addition to the treatment related adverse events reported above, two patients developed anti-hGH antibodies. In both cases, the antibodies did not have any growth inhibiting effect. None of the patients developed antibodies to host cell protein. Three transfer patients who had anti-hGH antibodies prior to treatment became negative within 6 months of treatment with SAIZEN. Hypothyroidism was seen in several patients. One patient died of recurrent craniopharyngioma and one patient experienced lipoatrophy.

The effectiveness of growth hormone treatment on growth was assessed primarily by changes in height velocity:

  1. Height velocity in cm/year, as a change from baseline.
  2. Height velocity as a change in standard deviation with reference to mean value for chronological age [Standard Deviation Score for Chronological Age (SDS CA)].
  3. Height velocity as a change in the standard deviation with reference to mean values for bone age [Standard Deviation Score for Bone Age (SDS BA)].

In contrast to the absolute data in cm/year, the calculation in SDS takes the different normal height velocities of different age groups into account.

In the above multicentre studies, 54.7% of the subjects were naive patients treated for 12 months. The efficacy results are summarized in the table below.

Height Velocities in Naive Patients Treated with SAIZEN for 12 Months in Comparison with 12 Month Literature Data for pit-hGH
Growth velocity SAIZEN pit-hGH
Before During 12 months Before During 12 months
cm/year
Germany 3.60 ± 1.22 (n=27) 9.54 ± 2.76 (n=27)
USA 3.49 ± 1.10 (n=50) 8.56 ± 1.65 (n=50)
Italy 3.22 ± 1.39 (n=26) 8.54 ± 2.45 (n=26) 3.68 ± 1.07 (n=25) 7.66 ± 2.37 (n=36)
United Kingdom 3.77 ± 1.75 (n=12) 10.02 ± 2.08 (n=12)
France 3.88 ± 1.07 (n=25) 8.03 ± 1.59 (n=25)
SDS CA
Germany -2.69 ± 1.28 (n=26) +3.44±2.81 (n=26) -2.45 ± 0.84 (n=?) +2.29 ± 2.49 (n=36)
USA -2.82 ± 1.27 (n=50) +3.26±2.38 (n=50)
SDS BA
Germany -2.92 ± 1.14 (n=25) ++2.19 ± 1.2 (n=25) -3.0 ± 0.99 (n=22) +0.71 ± 1.93 (n=35)
USA -3.08 ± 1.09 (n=50) +2.48 ± 2.41 (n=50)

Turner’s Syndrome

Turner’s Syndrome is caused by the apparent complete or partial absence of one of the X chromosomes or by other chromosomal abnormalities involving the second X chromosome. It occurs with an incidence of about 1 in 1000 neonates. The majority of Turner’s syndrome patients experience poor growth and gonadal dysfunction.

Short stature is nearly a constant clinical feature of girls with Turner’s Syndrome. Although there is considerable variation in phenotypic expression, the observed height velocities and final height of girls with TS are usually significantly lower than the normal average. The effectiveness of any therapeutic agent in the treatment of short stature in girls with TS is judged by the change in height velocity and the effect on predicted and/or final height.

The study described here is a pivotal, Phase III trial in patients with Turner’s Syndrome. The purpose of this study was to assess the long term efficacy as well as the safety of SAIZEN alone and in combination with oxandrolone in the treatment of growth retarded girls with Turner’s Syndrome.

This was an open, comparative, randomised multicentre study where a total of 91 girls (aged 10.3 ± 2.3 years) with Turner’s Syndrome were randomly allocated to receive either SAIZEN alone (XO group) or SAIZEN in combination with oxandrolone (XM group). The study medication dosing regimen is presented below.

Dosage Regimen

XO (initial n = 47) XM (initial n = 44)
1st Year SAIZEN 18 IU/m2/week (~0.029mg/kg/day) 18 IU/m2/week (~0.029 mg/kg/day)
Oxandrolone - 0.1 mg/kg/day
2nd Year SAIZEN 24 IU/m2/week (~0.038 mg/kg/day) 18 IU/m2/week (~0.029 mg/kg/day)
Oxandrolone - 0.05 mg/kg/day
XO (n=21) XOM (n=26) XM (n=33) XMO (n=11)
3rd Year and subsequent SAIZEN 24 IU/m2/week (~0.038mg/kg/day) 24 IU/m2/week (~0.038mg/kg/ day) 24IU/m2/week (~0.038mg/kg/ day) 24 IU/m2/week (~0.038 mg/kg/day)
Oxandrolone - 0.05 mg/kg/day 0.05 mg/kg/day -

After the second year, oxandrolone 0.05 mg/kg/day was also recommended for the XO group if HV was less than +2SD above the respective mean HV for age of girls with TS (group XOM). If subjects of the XM group had to stop oxandrolone, but continue SAIZEN, the dose of SAIZEN was increased to 24 IU/m2/week (group XMO).

Study Results

Changes in Height Velocity (cm/year)

The mean HV increased significantly over baseline in both groups (XO and XM) during the first 12 months of treatment. The difference between the mean gains in HV of the two groups, 2.4 ± 1.3 (XO) vs. 4.6 ± 1.8 (XM) cm/year, was statistically significant (P<0.0001). During the second year of treatment, the mean HV was maintained at a higher level compared with baseline in both groups. The TS girls in the XO group grew at a rate of 5.5 ± 1.1 cm/year (+1.5 ± 1.1 cm/year over baseline) significantly more than before treatment (P<0.0001). In the XM group, a mean HV of 6.5 ± 1.4 cm/year was achieved with a +2.4 ± 1.9 cm/year gain over baseline (P<0.0001). The difference in HV between the XO versus XM groups over the first two years of treatment was also statistically significant (P<0.05).

Height

Heights at the start of treatment showed a normal distribution around the 50th centile curve for a population of untreated girls with TS. After following 2 to 7.5 years of treatment, the majority of the TS patients were more than 1 SD above the mean for age, and had achieved a height greater than 150 cm. The best responses were achieved in the youngest patients.

Predicted Final Height

Changes in final height predictions (FHP) were analysed in the 35 TS girls who completed 6 years of treatment using the Bayley-Pinneau method based on bone age. The mean change in predicted adult height after 6 years ranged between 9 - 11 cm for the 4 groups. This suggests that final height will have improved by approximately 1.7 SD since the start of therapy.

Final Height

Twenty-six of the original 91 TS girls have reached final height defined as a height velocity = 0.5 cm/year over the last year of observation or treatment. For the group as a whole, the mean (±SD) final height was 150.6 ± 5.5 cm. The results are consistent with those reported in the literature for other TS girls treated with GH. When normalized for age, the improvement for the group of patients who reached final height as a whole was 1.1 SD. It should be noted that the age at attainment of final height was 18.2 years compared to 15.7 years for those who had not yet reached final height and on average were treated for a shorter period of time, 4.9 versus 5.3 years.

Factors influencing response to treatment

In order to assess the influence of certain factors on the response to SAIZEN therapy, we examined the effect of age at start of therapy and the duration of therapy on the change in height standard deviation score (HSDS). There was a highly significant inverse relationship between age at start and response to therapy (r=-0.49, p<0.0001), suggesting that the earlier the treatment is initiated, the better final height is likely to be. There was also a highly significant positive relationship between the duration of treatment and the improvement in HSDS (r=0.44, p<0.0001), suggesting that the final height will indeed be influenced by the duration of treatment.

In order to separate the effects of age at start and duration of treatment on the response to therapy, the effect of age at start on HSDS was analyzed by holding the duration of treatment constant. Highly significant negative correlations were again evident: r=-0.54 (p<0.0001) for the 56 girls treated for 5 years, and r=-0.56 (p<0.0005) for the 35 girls treated for 6 years.

Chronic Renal Failure

A primary intent to treat analysis is not available for the published literature in this area. Growth failure is common and serious sequelae of chronic renal failure in childhood with profound consequences for a child’s psychological development and social integration. Even when managed optimally, patients with CRF continue to lose height over time, both relative to their normally growing peers and in terms of final height.

The study described here is a pivotal, multicenter study where a total of 81 children with chronic renal failure (17 post -transplant, 27 on dialysis and 37 compensated CRF) were evaluated to assess the safety and efficacy of SAIZEN in the treatment of growth failure in children with chronic renal failure (CRF).

SAIZEN was administered subcutaneously at a weekly dose of 0.35 mg/kg per week (28 IU/m2/ week) for the first 3 years of treatment.

Study Results

For the group as a whole, laboratory changes were similar to those reported in patients receiving GH for growth hormone deficiency. This suggests that the dose used in this study is, in effect, an appropriately physiological one for children with CRF.

Changes in Height Velocity (HV)

After 12 months: Of the 63 patients who were available for analysis, 59 (94%) experienced an increase over baseline in HV. Mean HV (SD) increased by 4.4 ± 4.0 cm/yr. (p <0.001).
After 24 months: In the 44 children available for analysis, 39 (89%) experienced a sustained increase over baseline HV and the mean HV for this cohort was 7.5 ± 2.9 cm/yr., an increase of 3.0 ± 3.6 cm/yr. over baseline (p <0.001).

Changes in Height Standard Deviation Score (H SDS)

After 12 months: Of the 63 children available for analysis, 55 (87%) experienced an increase over baseline in H SDS. The percentage of children with a normal H SDS increased from 1% (1 of 81) to 17% (11 of 63). For the group as a whole, H SDS increased by +0.7 ± 0.7 (p<0.001).
After 24 months: In the 44 children available for analysis, 38 (86%) experienced a sustained increase over baseline in H SDS. The percentage of children achieving a normal H SDS increased to 43% (19 of 44).

The mean HV SDS remained significantly higher than at baseline and was greater than zero, consistent with ongoing catch-up growth.

Mean Height Velocity (cm/yr) and Mean Height SDS during the First Two Years of the Study
12 Months (n=63) 24 Months (n=44)
HV 9.0 ± 3.6 7.5 ± 2.9
HV from Baseline + 4.4 ± 4.0* +3.0 ± 3.6*
H SDS -3.0 ± 1.7 -2.5 ± 1.5
H SDS from Baseline +0.7 ± 0.7* + 1.2 ± 1.2*

Final Height

The change in the HA/BA (Height Age/Bone Age) ratio was also quantified during the 2 years of the study. The change in linear growth relative to the change in skeletal maturation, a measure of the preservation or loss of potential final height, was examined by estimating the HA/ BA ratio. A ratio of 1.0 indicates proportional gains in height and skeletal maturation and preservation of potential final height: a value above 1.0 suggests potential improvement in final height while a value below 1.0 indicates a loss of final height.

At baseline, for the 80 patients in which they could both be obtained, the mean HA was 5.1 and the mean BA was 5.7. For the 56 children with data at one year, the HA/ BA was 1.6 - 2.2, significantly greater than unity (p=0.040) and suggestive of improvement in predicted final height. For the cohort of children treated for 2 years, HA/ BA remained 1.0 during the second year, consistent with ongoing preservation of final adult height.

Analysis of Efficacy Subgroups

A secondary analysis, stratifying the study participants according to whether they had entered the study with compensated CRF, end stage renal disease (on dialysis) or whether they had undergone transplantation, was also performed. HV and H SDS rose in all three groups at one year of treatment. In the second year HV remained elevated above baseline levels in both the compensated and transplant groups and H SDS improved in all three groups.

Small for Gestational Age

A child born SGA is defined as a neonate whose birth weight or birth crown-heel length is at least 2 standard deviations (SD) below the mean ( -2 SD is equivalent to the 2.3 percentile) for the infant’s gestational age, based on data derived from a reference population. Most children born SGA catch up in growth during their first years of life, but between 8% and 15% of all children born SGA do not.

Two randomized, controlled Phase III clinical studies GF 4001 (n=101) and GF 6283 (n=58) were designed to assess efficacy and safety of treatment with SAIZEN in children born SGA who did not catch up in growth during their first years of life. Their design and patient demographics are summarized in the following table:

Study Demographics and Trial Design

Study
Number
Trial Design Dosage
(kg/day)
Route of
Administration
Administration
Scheme
Duration
(years)
Study
Subjects
Number
Mean
Age
(range)
(years)
Gender
GF 4001 Open, Randomised (Saizen group + No treatment control group) 0.067 mg (0.2 IU) Subcutaneous injection Continuous Study: 4
Treatment: 3
101 4.5 (2-8) M 51
F 49
GF 6283 Open, Randomised (Saizen group + No treatment control group) 0.067 mg (0.2 IU) Subcutaneous injection Intermittent Study: 4
Treatment: 2
58 3.3 (2-5) M 28
F 30

Study GF 4001 was initiated in 1990 and a dose of 0.2 IU/kg/day (corresponding to 0.067 mg/kg/day) was selected for use based on the results from the EMD Serono study GF 2773 and on published information (Albertsson-Wikland et al., 1989). The aim of the treatment was to induce catch-up growth and, because failure to catch up is thought to be related to a relative resistance to GH, a dose higher than that used to treat children with GH deficiency was administered. The study included a group treated from study initiation (Group T), and a control group (Group C) that did not receive any treatment during the first year but did receive SAIZEN in subsequent years. During the course of the study all children received SAIZEN treatment for three years and were followed for five years after treatment.

Study GF 6283 was initiated in 1993 and examined the same SAIZEN dose as study GF 4001. The 4-year study used two different regimens, continuous treatment for two years with follow-up for two additional years (Treatment-Treatment-Observation-Observation, TTOO) or discontinuous treatment (treatment during the first and third study years and observation during the second and fourth study years, TOTO). The TOTO regimen was chosen to investigate whether discontinuous treatment would result in a better effect on catch-up growth than continuous treatment (TTOO) since it was well known that the effect of SAIZEN on height velocity is greater during the first year of treatment than during subsequent years.

The two pivotal studies used similar inclusion and exclusion criteria: birth weight less than the 10th percentile of the gestational age-related standards, height SD score (H-SDS) ≤ –3.0 for chronological age and sex, and height velocity (HV) less than +0.5 SD for chronological age and sex. At study entry, children were aged between 2 and 8 years in study GF 4001 and between 2 and 5 years in study GF 6283. An additional inclusion criterion in study GF 6283 was a parental height ≥1.48 m for women and ≥1.60 m for men.

Similar numbers of boys and girls were included in each of two treatment groups for Study GF 4001. In Study GF 6283 distribution of boys and girls in the two treatment groups were slightly skewed (the two treatment groups contained 28 and 30 patients with respectively 11 males/17 females in TTOO and 17 males/13 females in TOTO). Taken together, the children enrolled in the two studies were representative of short children born SGA. They had not demonstrated any catch-up growth and no child had GH deficiency as determined by standard GH stimulation tests. Mean parental heights were somewhat below the French population average in both studies.

Study Results

The greatest effect on growth was observed during the first year of r- hGH treatment. However, during continued treatment, HV-SDS values demonstrated that growth rate remained consistently above the mean for age and sex and that catch-up growth was maintained (see table below). Mean HV values were 7.62 and 7.46 cm/year during the second year of SAIZEN therapy in the T and C groups, respectively, and 6.62 and 6.86 cm/year during the third year in study GF 4001 (compared with 5.83 and 6.16 cm/year at baseline in the two treatment groups, respectively). The corresponding mean HV-SDS values were 1.97 and 2.05 after 2 years of SAIZEN therapy and 1.07 and 1.33 after 3 years in study GF 4001.

Mean HV during the second year in the group that received continuous treatment in study GF 6283 was 7.85 cm/year (compared with 7.40 cm/year at baseline). This corresponded to HV-SDS of 1.55 after 2 years of continuous treatment in study GF 6283.
As a result of the accelerated growth, the children’s mean H-SDS CA improved to –1.67 and – 1.73 after 2 years, and –1.43 and –1.41 after 3 years in study GF 4001, and to –1.55 after 2 years of continuous treatment in study GF 6283. Thus, 2 years of treatment with SAIZEN yielded a net height increase of 1.5, 1.7 and 2.0 SD (studies GF 4001 and GF 6283) and 3 years of continuous treatment yielded an increase in height of 1.9 SD (study GF 4001).

Growth Results in Patients Treated with SAIZEN from Studies GF4001 and GF6283
Values presented as group means (SD)
Study Treatment Arm Number
Enrolled/
Completed
Height velocity SDS
(based on chronological age)
Height SDS
(based on chronological age)
Height SDS
(based on bone age)
At start of
treatment
1 year 2 years 3 years 4 years At start of
treatment
1 year 2 years 3 years 4 years At start of
treatment
1 year 2 years 3 years 4 years
GF 4001 Group T,
0.067 mg/kg/day
50/44 -1.42 (1.23) 4.00 (1.68) 1.97 (1.56) 1.07 (1.59) -3.34 (0.64) -2.21 (0.81) -1.67 (0.91) -1.43 (0.93) -1.04 (2.02) -0.54 (1.73) -0.43 (1.56) -0.50 (1.56)
Group C,
0.067 mg/kg/day
50/39 -0.40 (1.11) 4.01 (2.11) 2.05 (1.68) 1.33 (1.12) -3.24 (0.85) -2.22 (0.89) -1.73 (1.03) -1.41 (1.11) -1.37 (1.40) -0.89 (1.47) -1.01 (1.52) -1.01 (1.52)
GF 6283 Group TTOO,
0.067 mg/kg/day
28/25 -0.97 (1.01) 3.76 (1.31) 1.55 (1.82) -2.28 (1.67)* -1.82 (1.25)* -3.55 (0.60) -2.18 (0.62) -1.55 (0.82) -1.80 (0.78)* -1.99 (0.81)* -0.66 (2.35) -0.78 (1.28) -0.61 (1.24) -0.88 (1.35)* -1.21 (1.28)*
Group TOTO,
0.067 mg/kg/day
30/28 -1.31 (1.44) 3.15 (1.50) -1.78 (1.06)* 2.56 (1.69) -2.41 (1.19)* -3.43 (0.74) -2.16 (0.88) -2.35 (0.90)* -1.68 (1.05) -2.00 (1.01)* -1.04 (2.79) -0.93 (1.93) -1.51 (1.63)* -1.22 (1.73) -1.68 (1.61)*

* Denotes values obtained during observation;

Growth and bone maturation

Bone maturation was followed during the pivotal studies with SAIZEN by assessing the ratio of bone age to chronological age (BA/CA), the ratio of the rate of change in bone age to the rate of change in chronological age (ΔBA/ΔCA), and H-SDS based on bone age.
BA/CA ratios increased during r-hGH treatment but remained below unity at all times. The increase of the BA/CA ratio is consistent with the increased height velocity during treatment and suggests that adequate catch-up growth is achieved during treatment with SAIZEN.

Effects of discontinuation of treatment

Study GF 6283 provide follow-up information for 2 years after treatment and study GF 4001 provides such information for 5 years after treatment cessation.

In study GF 4001, mean H-SDS CA values of the combined treatment groups were –1.38 at the end of treatment, and –1.70, –1.83, –1.96, –2.16 and –2.23 after 1, 2, 3, 4 and 5 years of follow-up, respectively (pre-treatment values were –3.34 and –3.24 for the two groups, respectively). This means that over 5 years patients may have lost an average improvement in height of 0.85 SDS. Thus, the net effect of treatment for 3 years over a total of 8 years was a gain in height of approximately 1 SDS. These data indicate clearly that a substantial part of the effect on growth was lost following discontinuation of treatment.

Similarly, after 2 years of follow-up in study GF 6283, mean H-SDS CA was –1.99 (compared to a pre-treatment value of –3.55) and in study GF 6018 it was –1.4 (pre-treatment value of –2.6). Thus, the net effect of treatment for 2 years during a total of 4 years was a gain in height of approximately 1.2 to 1.5 SD.

Hence, discontinuation of treatment is accompanied by a substantial loss of the benefit on growth. The data support the notion that treatment should be given continuously until final height.

The impact of ethnic factors has not been evaluated in SGA clinical trials.

Adult Indications

Adult Growth Hormone Deficiency

The pathogenesis and disease history leading to adult GHD differs depending on whether the deficiency has existed since childhood or has been acquired during adult life.

Hypopituitarism acquired in adult life is often caused by pituitary or peripituitary tumours and/or their associated therapy. It is estimated that acquired hypopituitarism with GHD annually affects 10 people per million. The evolution of GHD after radiation therapy of a pituitary adenoma or craniopharyngioma has been investigated during ten years following radiotherapy. The time taken for pronounced GHD to develop was between one and four years and was dependent upon the GH status before radiotherapy.

The safety and efficacy of SAIZEN replacement therapy in adults was evaluated in a pivotal study. This was a randomized, double -blind, placebo -controlled (DBPC) study involving 115 patients. Sixty patients received 0.005 mg/kg/day for one month and then 0.01 mg/kg/day for five months, and 55 patients received corresponding placebo, administered by daily subcutaneous injection. This was followed by 12 to 30 months open-label treatment for all patients.

Study Results

Lean body mass (DEXA)

There was a statistically significant increase (p<0.0001) after 6 months of r-hGH treatment compared to placebo (r-hGH: baseline 49.1 ± 11.7 kg, N = 59, 6 months 49.6 ± 12.0 kg, N = 52); (placebo: baseline 53.7 ± 12.2 kg, N = 54, 6 months 53.9 ± 11.9 kg, N = 52). After adjusting for centre and baseline lean body mass, the estimated overall treatment difference was an increase of 2.21 kg (95% CI 1.27–3.15) compared to placebo. This increase was sustained throughout the 30 months of follow-up treatment. A subsidiary, subgroup analysis by gender shows a treatment difference of 2.91 ± 0.47 kg compared to placebo, in males, while the difference is 0.80 ± 0.70 kg in females.

Treadmill exercise test (Weber protocol)

There was a slightly greater increase in VO2max in the r-hGH group compared to placebo, but the difference was not statistically significant (r-hGH: baseline 21.21 ± 7.71 mL/kg/min N = 36, 6-months 25.50 ± 7.78 mL/kg/min, N = 26; placebo: baseline 23.36 ± 6.98 mL/kg/min, N = 35, 6 months 26.47 ± 8.58 mL/kg/min, N = 31). No statistically significant differences were noted for anaerobic threshold (r-hGH: baseline 13.13 ± 3.80, N = 35, 6-months 16.29 ± 4.41 mL/kg/min, N = 26; placebo: baseline 14.69 ± 4.29, N = 34; 6 months 16.38 ± 6.00 mL/kg/min, N = 31). The changes observed during the follow- up phase VO2max and anaerobic threshold paralleled study duration and were thus related to the time on study rather than to time on r-hGH treatment.

Other body composition results

DEXA assessments demonstrated a statistically significant reduction of total fat mass (p<0.0001) in the r-hGH group compared to placebo (r-hGH: baseline 27.73 ± 10.72 kg, N = 59, 6 months 23.82 ± 9.65 kg, N = 52; placebo: baseline 28.90 ± 14.83 kg, N= 54, 6 months 29.12 ± 15.33 kg, N = 52). The fat mass remained relatively stable throughout 30 months of follow-up treatment. The changes in total bone mineral content or density were not statistically different between the treatment groups or during the follow-up.

Of the BIA assessments, only the change in total body water was subject to statistical analysis, and no significant difference was found between treatment groups. Anthropometry demonstrated no statistically significant between-group differences for skinfolds, waist/hip ratio or body weight and was maintained throughout the 30 month follow-up. The sum of circumferences decreased significantly in the r-hGH group relative to placebo (p<0.017). Body weight and BMI were stable throughout r-hGH treatment.

Bone turnover markers

Intact parathyreoid hormone decreased significantly in the r-hGH treated group (p=0.0071), whereas bone specific alkaline phospatase, C-terminal propeptide, osteocalcin levels (p<0.0001 for each parameter) and urinary excretion of deoxyproline (p<0.001) rose significantly during r-hGH treatment compared to placebo. The changes were noted during the first 6 months of treatment and were followed by relatively little further change.

Perceived well-being

The results from the Nottingham Health Profile (NHP) questionnaire demonstrated a statistically significant difference between the treatment groups during the DBPC phase for the domain Emotional Reactions (p<0.017). The domains for Social Isolation, Energy and Sleep were also found to be statistically significant for the first 6 months of treatment for the follow-up phase of the study and were maintained or further improved during this phase. No other significant differences were found in NHP or the General Well-Being Index.

Handgrip strength

No statistically significant differences were found in the assessments of dominant or non-dominant hand-grip strength.

Mid-thigh cross-sectional MRI

There were no statistically significant differences in the assessments of percentages of fat, muscle or bone (carried out in only 1 centre); however the decrease in fat, increase in muscle and stability of bone content echo the trends in body composition.

Cardiac function

Two-dimensional echocardiography showed statistically significant differences between the treatment groups for ejection fraction percentage (increase in the r-hGH group, p<0.048; r-hGH: baseline 54.90 ± 11.21 %, N = 52, 6 months 60.89 ± 9.47 %, N = 48; placebo: baseline 54.41 ± 12.91 %, N = 50, 6 months 57.30 ± 8.61 %, N = 49) and remained significantly higher than baseline throughout 30 months. Left ventricular posterior wall thickness also increased significantly during the first 6 months of treatment and remained stable and greater than baseline values although significantly only after 12 months.

Comparative Bioavailability Studies

Somatropin reconstituted with Bacteriostatic Water for Injection, 0.3% w/v metacresol has been determined to be bioequivalent to Somatropin reconstituted with Bacteriostatic Water for Injection, 0.9% Benzyl Alcohol based on the statistical evaluation for Cmax, AUClast and AUC(0-∞). Summaries of the pharmacokinetic parameters are presented below.

Summary of the pharmacokinetic parameters of Somatropin in healthy volunteers

PK metrics Cmax
(mcg/L)
AUClast
(hr*mcg/L)
AUC(infin)
(hr*mcg/L)
AUC(infin)-
%extrapol.
tmax (h)
Median
values
t½ (h)
Mean values
Somatropin 8.8 mg Benzyl alcohol Mean (Min-Max) 39.0 (20.8-61.5) 280 (140-411) 291 (145-418) 4.11 (1.25-10.5) 4.5 (3-7) 2.6 (1.6-4.1)
Somatropin 8.8 mg metacresol Mean (Min-Max) 45.1 (21.5-69.2) 306 (191-458) 320 (205-495) 4.23 (1.84-13.5) 4.0 (2-7) 2.7 (1.2-5.8)

Summary of average bioequivalence without correction for potency

Bioequivalence of Somatropin Diluents
Meta-cresol (T) vs. Benzyl alcohol (R)
Parameter Estimated
Ratio (T/R) of
Relative
Means
Estimated
Intra-subject
CV(%)
90% C.I. (%) Standards Decision
Cmax 115.6% 19.8 105.2 - 127.0 Mean Ratio within 80% - 125% BE
AUCT 110.3% 10.8 104.8 - 116.0 90% C.I. within 80% - 125% BE

T = Test, R = Reference, BE = Bioequivalent

Median hGH Serum Concentration versus Time Profiles after Subcutaneous Administration of 4 mg r-hGH as Somatropin benzyl alcohol (Reference) or Somatropin meta-cresol (Test)

Detailed Pharmacology

Animal Pharmacology

Two pharmacokinetic studies in primates (Cynomolgus monkeys) were conducted with SAIZEN, consisting of one single dose study using two routes of administration, subcutaneous and intravenous, and one repeat dose study by subcutaneous administration.

In the single dose study, the animals were dosed with 0.16 mg (in 0.5 mL)/kg SAIZEN (r-hGH) and 0.5 IU (in 0.5 mL)/kg Asellacrin [natural hGH or pituitary extraction] for comparison, and the pharmacokinetic profiles were compared. Eight monkeys were treated in a cross-over design with all four treatments with at least 1 week between subsequent treatments. Blood samples were drawn from a forelimb vein of the fasted animals immediately before and up to 48 hours after administration.

For both SAIZEN and Asellacrin, the half-life was significantly longer after subcutaneous administration than after intravenous administration. The relative bioavailability of SAIZEN was not significantly different from that for Asellacrin.

Mean Half-Life (T½ Values)
Subcutaneous vs Intravenous
Subcutaneous T½ (hours) Intravenous T½ (hours)
r-hGH 5.1 r-hGH 3.2
n-hGH 4.7 n-hGH 2.4

In the repeat dose study, the pharmacokinetic profiles of SAIZEN and Asellacrin were compared by subcutaneous administration alone. Animals were treated at 24 hour intervals for 8 consecutive days, with either 0.16 mg/kg of SAIZEN or 0.5 IU/kg of Asellacrin, after being fasted for 16 hours. Blood samples were taken from a forelimb vein of each animal immediately prior to and 1 hour after each administration (meantime for highest serum concentration after administration as shown in the single dose pharmacokinetic study), on study days 1-7, and prior to and up to 48 hours after treatment on day 8.

Neither increasing nor decreasing trends could be detected in the serum concentration values obtained prior to or one hour after each daily administration. No accumulation nor induction phenomena occurred in this study. No significant difference for the elimination half life was detected for Asellacrin when compared with the single dose study. However, a significant difference (near to the 5% significance level) was seen for SAIZEN. It was concluded that the serum kinetic profile of hGH is not significantly altered by repeated administration. No significant difference was seen between the values of the main pharmacokinetic parameters calculated after the last administration in monkeys.

Human Pharmacology

Human growth hormone is physiologically released in response to pulses of the hypothalamic growth hormone releasing factor (GRF) at night during sleep. Endogenously liberated hGH has a short half-life. At least part of the actual growth-promoting effect takes place through stimulation of the production of insulin-like growth factors or somatomedins, a group of peptide hormones with a long half- life which, via a negative feedback mechanism, influence the synthesis and release of hGH.

A single-blind, randomized, placebo-controlled study in 16 normal-weight, healthy male volunteers was conducted to evaluate the pharmacology of SAIZEN. SAIZEN 0.067 mg/kg or placebo was administered over a period of 14 days. There was no difference between the treatment groups regarding blood pressure, headache, queasiness, and itching and local side effects (redness, swelling, soreness, and sensitivity to touch which occurred in both groups) which was caused by the high osmolarity of the administered solution.

A single-blind, placebo-controlled, partly randomized cross-over phase I study was performed in six healthy male volunteers, under standard diet. Single doses of r-hGH were administered: 5 mg as a 6-hour iv infusion, 6 mg and 20 mg by sc route.

During the iv infusion, GH concentrations rapidly reached a plateau followed by a a rapid decay, while the GH concentration curve rose and decreased smoothly after the sc injection, the absorption representing the rate limiting step. A moderate degree of non-linearity was observed between the 6 and 20 mg sc doses, the latter displaying a greater bioavailability and a prolonged mean absorption time. The 3 doses of r-hGH elicited similar side effects, though in varying magnitudes (transient weight gain, a rise in IGF-1, IGFBP-3, non-esterified fatty acids and glycerol, and a decrease in blood urea, urinary nitrogen, sodium and potassium excretions). These effects were more striking after the 6 mg sc dose than after the 5 mg iv infusion, although both doses produced similar AUCs of GH. The biological changes induced by the 20 mg dose were not much higher than after the 6 mg dose, suggesting a relative saturation of the effects of r-hGH at high doses. This was not the case for the glycemia and the C-peptide urinary excretion, which were significantly elevated only after the 20 mg dose.

Variable Treatment
vs time
interaction
Effect of
5mg iv vs
placebo
Effect of
6mg sc vs
placebo
Effect of 20
mg sc vs
placebo
Effect of 6mg
sc vs 20 mg sc
Weight gain 0.08 24h-72 h 24h-96h 24h-72h 72h-96h
IGF-1 <0.0001 8h-48h 8h-72h 8h-96h 16h-48h
IGFBP-3 0.014 12h-48h 16h-48h 16h-72h 72h-96h
NEFA 0.0002 4h-12h 4h-12h 4h-24h --
Glycerol 0.01 - 24h 8h, 24h, 48h 8h
Blood urea <0.0001 8h-24h 8h-36h 12h-48h 48h
Urinary nitrogen 0.05 8h-24h 4h-24h 12h-24h 4h-8h
Urinary sodium 0.006 4h-24h 4h-24h 0h-96h 4h-12h
Urinary potassium <0.0001 4h-24h 4h-24h 0h-96h 4h-12h

The above table list the time periods (hours) over which the variable differs significantly (p<0.5) from the placebo (or 6 mg sc). The values are calculated from a 3 way analysis of variance for repeated measures and randomized block design.

Apart from its growth-promoting effect, growth hormone influences the carbohydrate, the lipid, and the protein metabolism.

Insulin-like Growth Factor-I (IGF-1)

Reduced IGF-1 serum concentrations are often found in children with hypothalamic-hypophyseal hGH deficiency. Recent investigations show that plasma IGF-1 levels do not (although they generally increase on hGH treatment) directly correlate with the therapeutic success of hGH, because other factors (nutrition, binding protein concentration) modulate the plasma concentrations, and because IGF-1 formation in the tissues is not directly reflected in the plasma levels. In addition, some animal experiments show that not all growth-promoting effects are necessarily mediated through IGF-1.
SAIZEN (1.33 mg Somatropin/m2) administration to 12 healthy adults (6 males and 6 females), showed increased mean serum concentrations of IGF-1 within 24 hours. SAIZEN (0.067 mg/kg) administration to 8 healthy male volunteers every 48 hours over a period of 14 days, resulted in an increased mean serum concentration.

Contrary to earlier reports, other authors have found no correlation between the growth velocity and short- term or long-term changes in the IGF-1 serum levels. In the U.S. multicentre study for SAIZEN, the mean IGF-1 serum level (n = 50) rose from a pre-treatment level of 8.12 ± 6.36 nmol/L to 16.95 ± 9.82 nmol/L within 12 months of treatment, whereas the other SAIZEN multicentre studies confirmed that IGF-1 cannot be used as a parameter for evaluating the efficacy of growth hormone. The reason for this might be that IGF-1 is produced by different tissues and that the effect on the target cells is carried out in a paracrine or autocrine manner. Furthermore, IGF-1 (as determined by RIA), is only one of the growth factors responsible for the stimulation of growth.

Carbohydrate Metabolism

By inhibiting glucose uptake in the tissue, hGH has an anti-insulinogenic effect. At the same time, hGH increases the release of pancreatic insulin. SAIZEN (0.067 mg/kg) administration to 8 male volunteers every 48 hours over a period of 14 days resulted in a greater increase in glucose and a higher insulin secretion during the OGTT than before treatment whereas a single intramuscular or subcutaneous injection (1.33 mg SOMATROPIN/m2) had no significant effect on the blood sugar and C-peptide levels within the first 4 hours. The mean basal insulin level was increased after 4 hours (p=0.0738). Results of these two studies are summarized:

Glucose and Insulin levels in male patients receiving SC injections of r-hGH every 48hr for 14 days [mean (se)]
GLUCOSE mg/dL INSULIN IU/mL
Occasion HGH-rDNA Placebo HGH-rDNA Placebo
0 min premedication 80.19 (1.70) 80.86 (3.92) 6.35 (1.27) 9.28 (2.26)
stage II Day 7 92.58 (3.23) 84.02 (1.67) 17.28 (2.96) 9.26 (1.52)
stage II Day 15 89.87 (2.98) 80.86 (2.00) 16.03 (1.54) 10.26 (1.48)
30 min premedication 126.82 (6.08) 130.64 (9.74) 83.16 (15.82) 91.14 (23.64)
stage II Day 7 168.49(12.50) 151.59 (8.38) 100.64 (28.45) 97.7 (21.96)
stage II Day 15 168.71(12.00) 144.61 (8.69) 124.46 (20.15) 102.87 (26.19)
60 min premedication 88.3 (6.67) 98.66 (11.38) 47.78 (11.7) 49.81 (8.36)
stage II Day 7 196.64 (11.75) 145.74 (14.08) 176 (33.26) 123.14 (36.59)
stage II Day 15 173.67 (18.31) 133.8 (12.39) 169.99 (23.78) 114.45 (17.08)
90 min premedication 73.21 (3.94) 89.2 (10.30) 22.45 (5.31) 42.42 (9.99)
stage II Day 7 187.86 (15.30) 113.3 (8.15) 216.91 (26.80) 94.61 (26.57)
stage II Day 15 152.94 (15.30) 110.37 (8.41) 154.5 (30.96) 101.38 (17.60)
120 min premedication 65.10 (4.18) 77.94 (10.44) 11.71 (2.23) 32.81 (10.58)
stage II Day 7 149.12 (12.99) 105.19 (7.62) 177.94 (33.99)) 81.62 (24.20)
stage II Day 15 118.03 (10.97) 88.75 (8.37) 138.92 (24.24) 53.41 (6.57)
150 min premedication 67.12 (6.26) 68.25 (6.74) 6.4 (1.95) 13 (4.91)
stage II Day 7 116.9 (13.81) 88.97 (7.10) 95.51 (21.69) 38.17 (8.49)
stage II Day 15 86.5 (10.22) 69.6 (8.77) 76.45 (21.10) 34.06 (8.93)
180 min premedication 70.5 (5.18) 72.53 (6.65) 4.86 (0.91) 9.19 (2.25)
stage II Day 7 98.43 (11.26) 76.36 (4.50) 59.04 (17.99) 22.89 (5.08)
stage II Day 15 70.05 (5.62) 77.04 (2.90) 29.45 (7.50) 15.5 (2.01)
Glucose, C-peptide and Insulin levels in patients receiving a single SC or IM injection of r-hGH [mean (sd)]
time (h) male after IM
injection
female after IM
injection
male after SC
injection
female after SC
injection
GLUCOSE (m/dL)
0 85.33 (5.57) 84.67 (4.80) 84.20 (4.82) 82.33 (4.80)
1 83.17 (5.12) 82.17 (8.66) 84.80 (5.54) 80.67 (4.84)
2 83.00 (8.53) 82.33 (7.92) 81.80 (6.42) 81.50 (5.32)
3 87.50 (7.37) 86.83 (5.19) 86.00 (6.28) 79.33 (7.42)
4 91.40 (6.02) 84.67 (7.37) 83.40 (4.28) 81.67 (8.52)
C-PEPTIDE (IU/mL)
0 83.17 (26.55) 66.83 (6.37) 65.40 (18.31) 68.17 (13.75)
1 62.83 (16.83) 61.83 (8.47) 61.20 (15.27) 65.83 (17.75)
2 61.50 (20.50) 63.17 (5.98) 70.50 (18.31) 65.83 (11.13)
3 60.83 (22.05) 64.50 (13.38) 68.60 (15.27) 61.00 (13.36)
4 71.00 (20.25) 68.00 (10.24) 68.80 (17.48) 65.00 (14.21)
INSULIN (IU/mL)
0 12.23 (2.77) 12.62 (2.12) 11.00 (2.61) 12.44 (2.20)
1 10.33 (2.95) 11.08 (1.86) 11.56 (3.34) 11.32 (1.39)
2 10.25 (4.09) 11.80 (1.67) 9.78 (2.40) 11.22(3.37)
3 10.25 (4.09) 12.32(3.95) 11.20 (2.61) 9.66 (2.34)
4 12.94 (5.97) 13.36 (1.86) 14.74 (6.50) 11.30 (2.03)

Lipid Metabolism

Due to an activation of hormone- sensitive lipases in the fatty tissue, the parenteral application of hGH leads to an increase in FFA in the serum with a maximum between 3 - 6 hours, which lasts for several hours. Following subcutaneous and intramuscular injections of SAIZEN (1.33 mg Somatropin/m2), a significant increase in the mean FFA levels could be demonstrated after 4 hours. A drop of serum cholesterol after administration of high doses of SAIZEN was also found in 8 male volunteers.

Protein Metabolism

In hGH deficient children, the administration of hGH produces a marked retention of nitrogen. This was the reason for Prader et. al. to develop the nitrogen retention test for the differential diagnosis of short stature. Such metabolic tests have subsequently been simplified and refined using the stable isotope 15N. Recently, GH-deficient patients have been shown to respond to a recombinant hGH preparation in the same way as to pit-hGH. The effect on protein metabolism is closely connected with the growth-promoting effect. The growing organism requires more protein, and this requirement is then met by stimulation of protein synthesis. SAIZEN administered subcutaneously to 8 male volunteers every 48 hours over a period of 14 days (0.2 IU/kg [approximately 0.067 mg/kg]) resulted in a statistically significant drop of the mean urea blood level from 34.4 ± 2.5 mg/dL to 24.8 ± 1.5 mg/dL.

Pharmacokinetics

An open, non-comparative study was conducted in 12 healthy adults (6 males and 6 females). SAIZEN (1.33 mg Somatropin/m2) was injected intramuscularly and subcutaneously in equal doses, with at least one week between each injection. To determine the Cmax and Tmax, blood samples were taken at 1, 2, 3, 4, 6, 9, 12, 15 (3 mg only) and 24 hours; hGH was determined by specific radioimmunoassay. While the mean peak serum concentration of circulating hGH (36.9 ng/mL) occurred 3 hours after intramuscular injection, subcutaneous injections resulted in a more sustained elevation at lower levels, reaching a mean peak serum concentration between 4 hours (mean 16.4 ng/mL) and 6 hours (mean 16.3 ng/mL). The areas under the curves (AUCs) were very similar after subcutaneous and intramuscular injections. Comparable results were obtained after subcutaneous administration of the same amount of SAIZEN in a higher concentrated solution (3.33 mg/mL instead of 1.33 mg/mL). The Tmax values reported for intramuscular injection of pit-hGH in the literature are between 2 and 4 hours which correlates well with the values found for SAIZEN.


Toxicology

A series of toxicology studies including acute, subacute, subchronic and long-term studies were conducted with SAIZEN. The animal species used for these studies included mice, rats and monkeys.

Single Dose Studies

Six acute toxicity studies were conducted in rats, mice, rats and monkeys.

Species Route of Administration Dose
Rats and mice Subcutaneous 13.3mg/kg
Rats and mice Intravenous 13.3mg/kg
Rats Subcutaneous
Oral
83.33mg/kg
Rats Oral 1.67, 3.33, 6.67, 13.3 mg/kg
Monkeys Subcutaneous 1.67, 3.33, 6.67 mg/kg
Mice Oral 1.67, 3.33, 6.67, 13.3 mg/kg

No effects were observed except for minor histological changes (vacuole in hepatocytes and a hyaline droplet in renal epithelium) in one female monkey of the 6.67 mg/kg dose group.

Repeated Dose Studies

Six studies were conducted: 2 four-week subacute studies (one in rats and the other in monkeys), two thirteen-week studies (one in rats and one in monkeys) and 2 fifty-two-week studies (rats and monkeys) by subcutaneous route.

Four-Week Studies
RATS (4 week study) MONKEYS (4 week study)
# Animals 15 animals/sex/group 28/sex
Dosage Daily injections of 0, 0.067, 0.33, 1.6 and 3.33 mg/kg in 0.9% NaCl Daily injections of 0, 0.067, 0.33 and 1.6 mg/kg of Somatropin or 0.2 or 0.5 IU/kg Asellacrin in 0.9% NaCl
Observations Clinical signs, mortality, ophtalmoscopic examinations, body weight, laboratory analyses and postmortem examination (autopsy, organ weight, histology)
Results The drug was well tolerated up to 3.33 mg/kg. Small number of slight hematological, biochemical and morphological changes, most notably in 1.6 and 3.33 mg/kg groups. These were for the most part reversible, and none appeared detrimental to the health of the animals. Local tolerability was satisfactory. The drug was well tolerated up to 1.6 mg/kg. None of the observations appeared to be treatment-related and no clearly identifiable antibodies were observed.
Thirteen-week Studies
RATS (13 week study) MONKEYS (13 week study)
# Animals 15/sex/group 12/sex/group
Dosage Daily injections of 0, 0.067, 0.33 and 3.33 mg/kg Somatropin, or 0.2 and 10 IU/kg Asellacrin in 0.9% NaCl Daily injections of 0.067, 0.33 and 1.67 mg/kg/day SAIZEN
Observations Clinical signs, mortality, body weight, ophtalmoscopic examinations, laboratory analyses and postmortem examination (autopsy, organ weight, histology)
Results No overt signs of toxicity, number of slight changes were observed due mainly to the biological activity of a heterologous hormone administered for a prolonged period of time. Higher values of GOT, GPT, -GTP and LAP found in males of the 1.67 mg group were the only drug-related changes.
Fifty-two-week Studies
RATS (52 week study) MONKEYS (52 week study)
# Animals 100 males and 100 females 16 males and 16 females
Dose 0, 0.067, 0.2 and 0.6 mg/kg 0, 0.067, 0.2 and 0.6 mg/kg
Observations Clinical signs, mortality, body weight, ophtalmoscopic examinations, laboratory analyses and post-mortem examination (autopsy, organ weight, histology)
Results No treatment-related deaths. Ten rats died from incidental causes or spontaneous incidental pathology. No treatment-related clinical signs. No clinical changes at the injection site. Body weight and food consumption unaffected. No treatmentrelated eye abnormalities were seen. None of the hematology, blood chemistry or urinalysis value modifications were related to the dose levels. All rats developed high levels of antibodies by week 12, still present 8 weeks after end of study. Slight tendency towards increase of adrenal gland and spleen mean absolute weights without dosecorrelation possible partial correlation with the slight increase in body weight. No drug-related modifications in gross pathology and histology. No treatment-related deaths. One male (0.2 mg/kg) died on day 245 (accidental trauma). No clinical or laboratory modifications attributable to the drug. No gross alternations at injection sites. No anti-hGH detected. Post-mortem examination showed no changes
 attributable to the drug.

Other Studies

Two local tolerance studies were conducted in rats and rabbits to compare the drug reconstituted in bacteriostatic saline, in metacresol, bacteriostatic saline alone and metacresol alone. In rats, which were treated with either the drug reconstituted in bacteriostatic saline, or bacteriostatic saline alone, the slight changes observed were considered mainly due to the needle trauma and potentiated by the vasodilator activity of benzyl alcohol 0.9%. Both intramuscular and subcutaneous injections were well-tolerated. The changes observed were related to the mild activity of low concentration benzyl alcohol. In rabbits which were injected with the drug reconstituted with 0.3% metacresol at the concentrations of 5 and 10 mg/mL, the 5 mg/mL concentration caused minimal changes that were similar in type and degree to the negative control (physiological saline). The 10 mg/mL solution was slightly more irritant.

The potential of somatropin, bacteriostatic saline and metacresol for sensitization were assessed in the guinea pig maximization test. None of the animals showed a positive reaction when challenged with an occlusive skin patch. In the group treated with drug reconstituted in bacteriostatic saline, 40% and 50% of the animals showed positive at the first and second intradermal challenges respectively. When reconstituted in metacresol, the drug caused severe anaphylaxis after intradermal challenge, due to the high dose used in the study. In the study using bacteriostatic saline as the diluent, lower doses were injected and only a moderate skin reaction due to sensitization was observed in guinea pigs. The drug appears to be a moderate sensitizer in guinea pigs, which is understandable since the human protein present in the drug is heterologous to this species. One study investigating the ocular and dermal irritation potentials of the drug in rabbits showed no abnormalities.

Mutagenicity

A series of mutagenicity studies, consisting of Ames Test, Gene Conversion Test in S.cerevisiae, Unscheduled DNA Synthesis in Cultured HeLa Cells, Chromosome Aberration in Human Lymphocytes Cultured In Vitro, Micronucleus Test, were conducted.

No mutagenic activity was observed with the drug for any of the mutagenicity tests listed above.

Reproductive Toxicology

In three independent pre-clinical reproductive toxicology (fertility, teratology, and pre- and post-natal) studies involving rats, SAIZEN was administered by SC route at the dosages of 0, 0.033, 0.33 and 3.33 mg/kg/day. SAIZEN was administered SC to males during pre-mating and mating, and females during pre-mating, mating, gestation and lactation periods. In these studies, no compound related embryotoxic or teratogenic effects were observed in any experimental group. In addition, no interference with the F0 and F1 reproductive performance was found. SAIZEN administration did have the effect of increasing body weight and food and water intakes at the dosage of 3.33 mg/kg/day and to a lesser extent at the dosage of 0.33 mg/kg/day. Since the increase in body weight noted in parents, offspring and fetuses is related to the pharmacological activity of SAIZEN (r-hGH), the non-toxic effect dose is considered to be 3.33 mg/kg/day for both parents and their progeny.

In a reproductive (teratology) study in female rabbits, SAIZEN was administered by SC route at the dosages of 0, 0.033, 0.33 and 3.33 mg/kg/day for 13 days during the organogenesis period. Administration of SAIZEN during pregnancy did not induce embryotoxic or teratogenic effects, and the autopsy of females at the scheduled killing did not reveal any pathological changes. The no-effect level is considered to be 3.33 mg/kg/day for does and fetuses.