**2. Subjects and methods**

#### **2.1 Patients**

Eighteen patients participated in the study. At diagnosis, all of the patients had a history of increased growth velocity, breast development of Tanner stage 2 or more, and a bone age that was more than 1 yr above their chronological age. Ten girls had idiopathic central precocious puberty (ICPP), and 8 girls had idiopathic central early puberty (ICEP). The diagnosis of ICPP was made based upon the onset of breast development before the age of 7 yr and 6 mo, the generation of pubic hair before the age of 9 yr, or the onset of menses before the age of 10 yr and 6 mo, according to the diagnostic criteria currently used in Japan. ICEP was defined as the appearance of pubertal signs between 8 -10 yr of age. Furthermore, neither set of patients showed any evidence of hypothalamo-pituitary lesions on magnetic resonance imaging or additional conditions that might have affected their body mass index (BMI).

The median age at the start of treatment was 8.3 yr (range: 6 to 11). All patients received leuprolide acetate (LUPRON DEPOT, Takeda, Osaka, Japan) at an initial dose of 30 g/kg, which was administered subcutaneously every 4 weeks according to the standard gonadotropin-suppressive therapy currently used in Japan.

#### **2.2 Methods**

Standard anthropometric measurements were taken at the baseline and during the 2-year GnRHa treatment period. BMI was calculated as weight (kg)/height (m2), compared with age- and sex-matched reference values, and expressed as a standard deviation score (SDS) according to the method of Inokuchi (Inokuchi, 2009). The percentage of overweight (POW) was calculated as 100 x (the measured weight – normal weight)/normal weight (%). Normal weight data were derived from the 1990 Ministry of Health and Welfare data (Yamazaki et al., 1994). A POW of ≥ 20% was considered to indicate obesity (Asayama et al., 2003). Pubertal development was determined according to the method of Tanner (Tanner & Whitehouse, 1976).

Pituitary-gonadal axis function was considered to be adequately suppressed during treatment if the concentrations of LH and E2 were maintained within the prepubertal normal ranges of our laboratory; i.e., if a) the basal serum LH level was below 0.5 mIU/ml and b) the basal serum E2 level was below 10 pg/ml. In the patients that demonstrated incomplete suppression, the dose of leuprolide acetate was increased to 150 g/kg. Bone age (BA) was assessed by one investigator using an x-ray of the left hand, according to the method of Greulich and Pyle (Greulich & Pyle, 1959).
