Oxandrolone for growth hormone‐treated girls aged up to 18 years with Turner syndrome

Abstract

Background

The final adult height of untreated girls aged up to 18 years with Turner syndrome (TS) is approximately 20 cm shorter compared with healthy females. Treatment with growth hormone (GH) increases the adult height of people with TS. The effects of adding the androgen, oxandrolone, in addition to GH are unclear. Therefore, we conducted this systematic review to investigate the benefits and harms of oxandrolone as an adjuvant therapy for people with TS treated with GH.

Objectives

To assess the effects of oxandrolone on growth hormone‐treated girls aged up to 18 years with Turner syndrome.

Search methods

We searched CENTRAL, MEDLINE, Embase, the ICTRP Search Portal and ClinicalTrials.gov. The date of the last search was October 2018. We applied no language restrictions.

Selection criteria

We included randomised controlled clinical trials (RCTs) that enrolled girls aged up to 18 years with TS who were treated with GH and oxandrolone compared with GH only treatment.

Data collection and analysis

Three review authors independently screened titles and abstracts for relevance, selected trials, extracted data and assessed risk of bias. We resolved disagreements by consensus, or by consultation with a fourth review author. We assessed trials for overall certainty of the evidence using the GRADE instrument.

Main results

We included six trials with 498 participants with TS, 267 participants were randomised to oxandrolone plus GH treatment and 231 participants were randomised to GH only treatment. The individual trial sample size ranged between 22 and 133 participants. The included trials were conducted in 65 different paediatric endocrinology healthcare facilities including clinics, centres, hospitals and academia in the USA and Europe. The duration of interventions ranged between 3 and 7.6 years. The mean age of participants at start of therapy ranged from 9 to 12 years. Overall, we judged only one trial at low risk of bias in all domains and another trial at high risk of bias in most domains. We downgraded the level of evidence mainly because of imprecision (low number of trials, low number of participants or both).

Comparing oxandrolone plus GH with GH only for final adult height showed a mean difference (MD) of 2.7 cm in favour of oxandrolone plus GH treatment (95% confidence interval (CI) 1.3 to 4.1; P < 0.001; 5 trials, 270 participants; moderate‐quality evidence). The 95% prediction interval ranged between 0.3 cm and 5.1 cm. For adverse events, we based our main analysis on reliable date from two trials with overall low risk of bias. There was no evidence of a difference between oxandrolone plus GH and GH for adverse events (RR 1.81, 95% CI 0.83 to 3.96; P = 0.14; 2 trials, 170 participants; low‐quality evidence). Six out of 86 (18.6%) participants receiving oxandrolone plus GH compared with 8/84 (9.5%) participants receiving GH only reported adverse events, mainly signs of virilisation (e.g. deepening of the voice). One trial each investigated the effects of treatments on speech (voice frequency; 88 participants), cognition (51 participants) and psychological status (106 participants). The overall results for these comparisons were inconclusive (very low‐quality evidence). No trial reported on health‐related quality of life or all‐cause mortality.

Authors' conclusions

Addition of oxandrolone to the GH therapy led to a modest increase in the final adult height of girls aged up to 18 years with TS. Adverse effects identified included virilising effects such as deepening of the voice, but reporting was inadequate in some trials.

Plain language summary

Oxandrolone for growth hormone‐treated girls aged up to 18 years with Turner syndrome

Review question

What are the effects of giving oxandrolone to growth hormone‐treated girls aged up to 18 years with Turner syndrome?

Background

Turner syndrome is a genetic disorder of girls with a chromosomal anomaly where all or part of one of the two X chromosomes is missing or changed. Turner syndrome shows various signs and symptoms, among them short stature. Growth failure in childhood leading to short final adult height contributes to social and emotional impairment. Without treatment, girls with Turner syndrome are about 20 cm shorter than healthy youngsters. Treatment of girls with Turner syndrome with growth hormone increases adult height. We wanted to find out if the addition of oxandrolone would further improve the final adult height and the effects of this combination on other symptoms. Oxandrolone is an androgen. Androgens are essential male sex hormones and are important for females as well.

Study characteristics

We included six randomised controlled trials (clinical trials where people are randomly put into one of two or more treatment groups). The duration of the treatments ranged between 3 and 7.6 years. Study authors allocated 498 participants to treatment groups, 267 participants to oxandrolone plus growth hormone treatment and 231 participants to growth hormone only treatment. The average age of the children at begin of treatment ranged from 9 to 12 years.

This evidence is up to date as of October 2018.

Key results

When comparing oxandrolone plus growth hormone to growth hormone only, the final adult height was on average 2.7 cm higher in favour of oxandrolone plus growth hormone therapy. Only two studies provided reliable data on side effects: 6 out of 86 (19%) participants given oxandrolone plus growth hormone compared with 8 out of 84 (10%) participants given growth hormone only reported side effects, mainly signs of development of male physical characteristics (for example deepening of the voice). One study investigated the effects of treatments on speech, the process of acquiring knowledge and understanding (cognition), and mental and emotional (psychological) status. The overall results for these were inconclusive. No trial measured people's satisfaction with their life and health or death from any cause.

Quality of the evidence

For side effects and effects on speech, cognition and psychological status, we are uncertain or very uncertain, mainly because the number of studies and participants was low and results were vague. For final adult height, we think that further research is likely to have an important impact on our confidence in the results and may change the results.

Summary of findings

Summary of findings for the main comparison. Oxandrolone for growth hormone‐treated girls aged up to 18 years with Turner syndrome.

Oxandrolone for growth hormone‐treated girls aged up to 18 yearswith Turner syndrome
Patient: girls aged up to 18 years with Turner syndrome Settings: mostly paediatric endocrine centres Intervention: oxandrolone + growth hormone Comparison: growth hormone alone
Outcomes	Growth hormone	Oxandrolone + growth hormone	Relative effect (95% CI)	No of participants (trials)	Certainty of the evidence (GRADE)	Comments
Improvement in final adult height (cm) Follow‐up: 3–10 years	The mean final adult height ranged across control groups from 149.6 cm to 155.6 cm	The mean final adult height in the intervention groups was 2.7 cm higher (1.3 cm higher to 4.1 cm higher)	—	270 (5)	⊕⊕⊕⊝ Moderatea	95% prediction interval 0.3–5.1 cm
Adverse events (N) Follow‐up: 3–10 years	95 per 1000	172 per 1000 (79 to 377)	RR 1.81 (0.83 to 3.96)	170 (2)	⊕⊕⊝⊝ Lowb	Data based on 2/4 trials with overall low risk of bias for adverse events.
Health‐related quality of life	Not reported					Follow‐up study with < 50% of initially randomised participants showed inconclusive results comparing oxandrolone + GH for total and mean scores (3 questionnaires).
Effects on speech (voice frequency) Follow‐up: 6 years	See comment	88 (1)	⊕⊝⊝⊝ Very lowc	Most voice frequencies remained within the normal range, but became occasionally lower, especially with oxandrolone + GH 0.06 mg/kg/day; results were inconclusive for oxandrolone + GH 0.03 mg/kg/day.
Effects on cognition (WISC‐R questionnaire) Follow‐up: 2 years	See comment			51 (1)	⊕⊝⊝⊝ Very lowc	Summary scores for working memory, spatial cognition, executive function and verbal abilities using the WISC‐R; after 2 years, comparison of oxandrolone + GH vs GH showed inconclusive results.
Effects on psychological status (several questionnaires) Follow‐up: 6 years	See comment			106 (1)	⊕⊝⊝⊝ Very lowc	There were no evident psychological virilising effects in the area of behaviour, aggression, romantic and sexual interest, mood and gender role.
All‐cause mortality	Not reported
*The basis for the assumed risk (e.g. the median control group risk across studies) is provided in footnotes. The corresponding risk (and its 95% confidence interval) is based on the assumed risk in the comparison group and the relative effect of the intervention (and its 95% CI). CI: confidence interval; GH: growth hormone; RR: risk ratio; WISC‐R: Wechsler Intelligence Scale for Children‐Revised.
GRADE Working Group grades of evidence High quality: further research is very unlikely to change our confidence in the estimate of effect. Moderate quality: further research is likely to have an important impact on our confidence in the estimate of effect and may change the estimate. Low quality: further research is very likely to have an important impact on our confidence in the estimate of effect and is likely to change the estimate. Very low quality: we are very uncertain about the estimate.

^aDowngraded one level because of imprecision (low median sample size) – for details, see Appendix 15.
 ^bDowngraded two levels because of serious imprecision (low number of trials and low median number of participants, CI for the pooled estimate consistent with benefit and harm) – for details, see Appendix 15.
 ^cDowngraded one level because of inconsistency (inconclusive results) and two levels because of serious imprecision (single trial with low number of participants).

Background

Turner syndrome (TS) is one of the common chromosomal abnormalities in girls. TS was initially described by Otto Ullrich in 1930 and later in 1938 by Henry Turner who described this syndrome in seven females with short stature, sexual infantilism, webbed neck and cubitus valgus (Turner 1938; Ullrich 1930). The incidence of TS is 1 in 2000 to 1 in 2500 female live births (Nielsen 1991; Pinsker 2012; Stochholm 2006). Most of the pregnancies with TS result in abortion. Gravholt and colleagues studied the prevalence of TS karyotypes among antenatally tested fetuses and TS among liveborn infants in Denmark from 1970 to 1993 (Gravholt 1996). Their data showed that among female fetuses tested by amniocentesis, the prevalence of TS karyotypes was 176/100,000 while the prevalence was 392/100,000 among female fetuses tested by chorion villus sampling. These data proved that the prevalence of TS is not only influenced by testing during pregnancy or after birth but also depends on the method of antenatal testing (Gravholt 1996).

Description of the condition

TS is characterised by complete or partial loss of the second X chromosome (Nielsen 1991). Complete loss of one X chromosome (45 X) results in classical features of the disease while mosaics present with varying and generally lesser degrees of manifestation. Variable aberrations of the second X chromosome such as deletion, ring chromosome and isochromosome, cause phenotypic features of TS in people with 46 XX (Mohamed 2015a). The most frequent chromosomal abnormality in TS is 45 X (Nielsen 1991; Pinsker 2012; Stochholm 2006). The Danish TS registry showed that the 45 X karyotype was found in half of the cohort, followed by mosaics (Gravholt 1996).

TS is a multisystem disorder with the cardiovascular, skeletal, endocrine and reproductive systems mostly affected (Stochholm 2006). Lymphoedema of the hand and foot is an early sign of TS. Other dysmorphic features include widely spaced nipples, cubitus valgus, short webbed neck, low posterior hairline and narrow hyperconvex nails (Baxter 2007; Nielsen 1991; Pinsker 2012; Stochholm 2006). Congenital heart diseases are more frequent in people with TS compared to the general population (Stochholm 2006). One Italian study group for TS reported on cardiac anomalies in 594 people with the disease (Mazzanti 1998). The prevalence of cardiac malformations in this cohort was 23%, with bicuspid aortic valve (12.5%), aortic coarctation (6.9%) and aortic valve disease (3.2%). Growth failure in childhood leading to short final adult height is the cardinal sign in girls with TS (Pinsker 2012; Stochholm 2006). While their birth weight is slightly impaired in utero, girls with TS may present early in childhood with failure to thrive (Davenport 1999). The height of girls with TS, when plotted on growth curves specific for this disorder, shows that growth velocity declines below the reference growth curve for females often as early as two to four years of age (Saenger 1999). The adult height (defined as the height at which the epiphyses are closed or height velocity is less than 1 cm/year) of people with TS is approximately 20 cm below that of the average female population. Naeraa and Nielsen studied growth parameters in 78 females with TS (Naeraa 1990; Nielsen 1991). They found that though there was no pubertal growth spurt, the steady decrease of height velocity (defined as increment in height per year in centimetres) was interrupted at the age of nine years. Height velocity was then constant until 12 years of age, thereafter it slowly decreased. The mean final height of this cohort was 146.8 cm (standard deviation (SD) 5.8 cm, 76 females) compared to 166.8 cm in the general female population (Naeraa 1990). Therefore, short final adult height in TS is explained by poor childhood growth in addition to the lack of a pubertal growth spurt.

Hypergonadotropic hypogonadism resulting from ovarian failure is a cardinal feature of TS. Primary infertility and lack of pubertal signs are seen in most girls with TS (Bondy 2007a). Spontaneous puberty and rarely pregnancy are reported in some women with TS, most of them carrying mosaic karyotype (Hadnott 2011; Mohamed 2015b). Assisted reproduction remains an option to achieve pregnancy for women with primary ovarian failure including those with TS (Hadnott 2011).

Characteristic neurocognitive features of TS include normal verbal function and impaired visual‐spatial and perceptual abilities, attention, working memory and spatially dependent executive function (Ross 2006). However, global developmental delay is uncommon in TS (Nielsen 1991; Pinsker 2012; Ross 2009; Stochholm 2006). Short stature, delayed puberty and the presence of dysmorphic features contribute to the social and emotional impairment observed in TS (Ross 2006). Failure to treat growth and puberty stigmata may lead to further psychological and psychiatric problems (Bondy 2007a; Carel 2006; Menke 2010).

Description of the intervention

Growth hormone

Growth hormone (GH) is a peptide hormone secreted by the anterior pituitary gland. GH promotes linear growth by stimulating production of insulin‐like growth factor 1 that acts at the growth plate by enhancing differentiation of prechondrocytes and the expansion of osteoblasts (Reh CS 2010).

GH is currently used as a once‐daily subcutaneous injection, typically given late in the evening to mimic the physiological secretory pattern of endogenous GH (Ranke 1999; Reh CS 2010). There is a wide range of recommended dosing of GH for the treatment of GH deficiency. The standard dose of classical GH deficiency ranges between 0.5 IU/kg/week and 0.7 IU/kg/week (0.17 mg/kg/week and 0.23 mg/kg/week) in most countries (Tanaka 1999). As GH is usually not deficient in people with TS, higher doses have been used for augmenting growth in TS (Bell 2010; Reh CS 2010; Takeda 2010; Tanaka 1999).

GH is generally safe with adverse events, such as benign intracranial hypertension, slipped femoral epiphysis, worsening of scoliosis and impaired glucose homeostasis, infrequently observed (Bell 2010).

Recombinant human GH was approved by the US Food and Drug Administration (FDA) for use in children with GH deficiency in 1985 and for girls with TS in 1996. Since the late 1980s, multiple studies have confirmed the efficacy and safety of GH therapy (Bell 2010; Reh CS 2010; Takeda 2010; Tanaka 1999).

Currently, GH therapy is recommended for all girls aged up to 18 years with TS as the evidence supports this intervention (Baxter 2007; Canadian Growth Hormone Advisory Committee 2005; Takeda 2010). Standard management of TS entails starting GH in childhood to maximise height gain and to improve the final adult height (Bondy 2007a; Davenport 2007; Ranke 2007; Stephure 2005). Furthermore, treatment with GH positively affects body composition by increasing muscle mass and decreasing fat mass (Gravholt 2002). One Cochrane systematic review investigated the effects of GH in girls aged up to 18 years with TS (Baxter 2007). It included four randomised controlled trials (RCTs) with 365 participants receiving GH for at least one year. This review concluded that GH increased short‐term growth in girls with TS by approximately 3 cm in the first year of treatment and 2 cm in the second year. In one trial, treatment increased final height by approximately 6 cm compared with the height in an untreated control group (Baxter 2007). The optimal age for initiation of GH therapy for young girls has not been established. Davenport and colleagues showed that early GH treatment can correct growth failure and normalise height in infants and toddlers with TS (Davenport 2007).

Carel and colleagues carried out a population‐based cohort study of health‐related quality of life determinants in 568 young women with TS using the Medical Outcome Study Short Form 36 (SF‐36) score and the General Health Questionnaire 12 score (Carel 2005; Carel 2006). This trial concluded that health‐related quality of life was normal and unaffected by height in young adults with TS treated with GH. Taback and colleagues found similar results in a follow‐up study on the health‐related quality of life of young adults from a long‐term controlled trial of GH treatment in participants with TS (Taback 2011). The trial found no benefit or adverse effects on health‐related quality of life either from receiving or not receiving GH injections.

Most people with TS have delayed puberty leading to the lack of a pubertal growth spurt, which compromises the final adult height. Therefore, oestrogen is commonly started with GH to induce puberty and maximise final adult height (Bondy 2007a; Van Pareren 2003).

The age of starting oestrogen in girls aged up to 18 years with TS, as well as the dose and the duration of therapy, varies considerably among physicians (Bondy 2007a; Pinsker 2012; Van Pareren 2003).

Oxandrolone

Oxandrolone is an anabolic steroid (a synthetic derivative of testosterone). It is a weak androgen (Menke 2010), and is available in 2.5 mg and 10 mg tablets. The dose for children is based on bodyweight (0.1 mg/kg); however, studies have used a variable oxandrolone dose (mostly 0.03 mg/kg/day to 0.06 mg/kg/day) to promotion growth in TS (Bareille 1997; Freriks 2012; Freriks 2015; Gault 2011; Job 1991; Joss 1984; Menke 2010; Nilsson 1996; Rosenfeld 1986; Rosenfeld 1987; Rosenfeld 1988; Rosenfeld 1989; Rosenfeld 1990; Rosenfeld 1992; Stahnke 1992; Stahnke 2002; Zeger 2011). A stable and validated liquid formulation of oxandrolone has been developed. This formulation uses commercially available oxandrolone tablets which are crushed and dispersed in syrup (Garg 2011). Historically, oxandrolone was initially approved in 1964 for treatment of wasting associated with conditions such as chronic infection, severe trauma and after extensive surgery (Mann 1999). Since the late 1990s, oxandrolone has been used as an adjuvant therapy to promote healing in severe burns (Hart 2001; Sheffield‐Moore 1999).

In order to improve growth, oxandrolone is usually started a few years before the appropriate age for induction of puberty in girls with TS. Both GH and oxandrolone are discontinued when the adult final height has been achieved (Bondy 2007a; Sas 2014; Stochholm 2006).

Adverse effects of the intervention

Adverse effects associated with oxandrolone include signs of virilisation such as clitoral enlargement, acne, lowering of the voice and more rapid skeletal maturation (Bondy 2007b; Menke 2011a; Menke 2011b; Rosenfeld 1998).

How the intervention might work

Although oxandrolone is a known anabolic steroid, the exact mechanism by which oxandrolone improves growth is unknown. It may act directly on the growth plate (Haeusler 1996; Sheffield‐Moore 1999; Wilson 1988). The advantage of oxandrolone over other growth‐promoting androgens is that it may improve the final adult height while not promoting bone maturation that leads to early fusion of the epiphysis (Bareille 1997; Ferrández 1989; Gault 2011; Haeusler 1996; Job 1991; Joss 1984; Joss 1997; Menke 2010; Nilsson 1996; Rosenfeld 1992; Sheanon 2015; Stahnke 2002; Zeger 2011).

Why it is important to do this review

The effects of oxandrolone as an adjuvant therapy to GH aiming to maximise the final adult height is unclear. Therefore, we conducted this systematic review to evaluate the effects and safety of oxandrolone in girls with TS already treated with GH. This will help physicians and health policy developers to make evidence‐based recommendations regarding the role of oxandrolone in TS.

Objectives

To assess the effects of oxandrolone on growth hormone‐treated girls aged up to 18 years with Turner syndrome.

Methods

Criteria for considering studies for this review

Types of studies

We included RCTs.

Types of participants

GH‐treated girls aged up to 18 years with TS.

Diagnostic criteria

As some people with TS lack the clinical phenotype suggestive of the disease, all participants in this review had their diagnosis of TS confirmed by a chromosomal study. This included complete (45 X) or partial loss of the second X chromosome (such as deletion, ring chromosome and isochromosome). Mosaics were also included. We included trials that enrolled some participants with TS who had an abnormal Y chromosome.

Types of interventions

All participants received GH treatment for at least one year. All other concurrent therapies were comparable between the intervention and comparator groups.

Intervention

• Oxandrolone plus GH treatment.

Comparator

• GH treatment only.

Minimum duration of intervention

Minimum duration of intervention was one year.

Summary of specific exclusion criteria

We excluded trials including participants diagnosed on clinical grounds without confirmation from chromosomal studies.

Types of outcome measures

We did not exclude a trial if it failed to report one or several of our primary or secondary outcome measures. If the trial reported none of our primary or secondary outcomes, we planned not to include the trial but wanted to provide some basic information in the 'Characteristics of awaiting classification' table.

Primary outcomes

• Improvement in final adult height.

• Health‐related quality of life.

• Adverse events.

Secondary outcomes

• Increase in height velocity.

• All‐cause mortality.

• Bone maturation.

• Effects on cognition.

• Effects on psychological status.

• Effects on speech.

• Socioeconomic effects.

Method and timing of outcome measurement

• Improvement in final adult height: defined as the height at which epiphyses were closed or height velocity was less than 2 cm per year (Bondy 2007a; Stochholm 2006), and measured after the expected age of normal female puberty.

• Health‐related quality of life: evaluated by a validated instrument such as the SF‐36 (Carel 2005; Carel 2006), and measured after the completion of the RCT.

• Adverse events: such as virilisation effects on hair distribution, deepening of voice, clitromegaly and measured during or after completion of the RCT.

• Increase in height velocity (in girls who had not reached their final adult height): height velocity is defined as increment in height per year in centimetres and measured before participants achieved their final adult height.

• All‐cause mortality: defined as death from any cause and measured during and at the end of the RCT.

• Bone maturation: assessed by X‐ray of the left hand for bone age and closure of the epiphysis and measured in years during and at the end of the RCT.

• Effects on cognition: measured using a validated instrument such as the Wechsler Intelligence Scale for Children Third Edition (WISC) (Wechsler 1991).

• Effects on psychological status: measured by a validated instrument such as the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (DSM‐IV) (Conway 2012), and measured after completion of the RCT.

• Effects on speech: assessed by speaking fundamental frequency (Andersson‐Wallgren 2008), and measured during or after completion of the RCT.

• Socioeconomic effects: defined as the impact of the disease on social and economic parameters such as education, marriage, employment and income, housing, equity and overall social well‐being. Socioeconomic effects were measured after completion of the RCT.

Search methods for identification of studies

Electronic searches

We searched the following sources from inception of each database to the specified date and placed no restrictions on the language of publication (Lefebvre 2011).

• Cochrane Central Register of Controlled Trials (CENTRAL; 2018, Issue 9; searched 10 October 2018).

• Ovid MEDLINE(R) and Epub Ahead of Print, In‐Process & Other Non‐Indexed Citations and Daily (from 1946 to 9 October 2018; searched 10 October 2018).

• Embase (from 1974 to 2015 week 32; searched 13 August 2015*).

• ClinicalTrials.gov (searched 10 October 2018).

• World Health Organization (WHO) International Clinical Trials Registry Platform (ICTRP) (www.who.int/trialsearch/; searched 10 October 2018).

*RCTs indexed in Embase are now prospectively added to CENTRAL via a highly sensitive screening process (Cochrane 2018) (used for Embase after August 2015).

For detailed search strategies, see Appendix 1. We continuously applied an email alert service for MEDLINE via OvidSP to identify newly published trials using the search strategy detailed in Appendix 1. If we detected additional relevant key words during any of the electronic or other searches, we modified the electronic search strategies to incorporate these terms and documented the changes. We placed no restrictions on the language of publication when searching the electronic databases or reviewing reference lists of identified trials.

Searching other resources

We tried to identify other potentially eligible trials or ancillary publications by searching the reference lists of included trials, systematic reviews, meta‐analyses and health technology assessment reports.

We defined grey literature as records detected in ClinicalTrials.gov or WHO ICTRP. In addition, we searched manufacturers' websites and databases from regulatory agencies (European Medicines Agency (EMA) and US FDA) (Hart 2001; Schroll 2015).

We did not use abstracts or conference proceedings for data extraction unless full data were available from trial authors because this information source does not fulfil the CONSORT requirements which consist of "an evidence‐based, minimum set of recommendations for reporting randomized trials" (CONSORT 2016; Scherer 2007). We planned to present information on abstracts or conference proceedings in the 'Characteristics of studies awaiting classification' table.

Data collection and analysis

Selection of studies

Three review authors (SM, YA, KA) independently scanned the abstract, title, or both, of every record retrieved by the literature searches, to determine which trials we should assessed further. We investigated the full‐text of all potentially relevant records. We resolved disagreements through consensus or by recourse to a fourth review author (YSM). If resolving disagreement was not possible, we categorised the trial as a 'Study awaiting classification' and we contacted trial authors for clarification. We presented an adapted PRISMA flow diagram to show the process of trial selection (Liberati 2009). We listed all articles excluded after full‐text assessment in a 'Characteristics of excluded studies' table and provided the reasons for exclusion.

Data extraction and management

For trials that fulfilled the inclusion criteria, three review authors (SM, HA, YSA) independently extracted key participant and intervention characteristics. We reported data on efficacy outcomes and adverse events using standard data extraction forms from the Cochrane Metabolic and Endocrine Disorders (CMED) Group. We resolved any disagreements by discussion, or, if required, by a fourth review author (YA) (for details see Characteristics of included studies table; Table 2; Appendix 2; Appendix 3; Appendix 4; Appendix 5; Appendix 6; Appendix 7; Appendix 8; Appendix 9; Appendix 10; Appendix 11; Appendix 12; Appendix 13; Appendix 14; Appendix 15).

1. Overview of trial populations.

Study ID	Intervention(s) and comparator(s)	Description of power and sample size calculation	Screened/eligible (N)	Randomised (N)	Analysed (N)	Finishing trial (N)	Randomised finishing trial (%)	Follow‐up (extended follow‐up)a
Zeger 2011b (parallel RCT)	I: oxandrolone + GH	Sample size was estimated for height velocity: 60 girls (30 per group) yielded a > 95% probability to detect a statistically significant difference in height velocity between the GH + oxandrolone and GH + placebo groups at year 2. 70 girls completed the initial 2 years, thereafter 35 girls receiving oxandrolone and 31 girls receiving placebo chose to continue in the 2‐year double‐blind extension study (47 girls completed this phase and were evaluated at the end of year 4).	NR/76	40	37	37 (24 extension period)	93 (60 extension period)	2 years (2 years)
	C: GH + placebo			36	33	33 (23 extension period)	92 (64 extension period)
	Total:			76	70	70 (47 extension period)	92 (62 extension period)
Gault 2011c (parallel factorial RCT)	I: oxandrolone + GH	Assuming an SD of 5 cm, 50 girls were needed in each group to detect a difference between groups in mean final height (defined as height velocity < 1 cm/year and bone age ≥ 15.5 years) of 2.8 cm with 80% power at 5% significance; total of 100 participants.	NR/106	51	NA	NA	NA	Unclear (15 years)
	C: GH + placebo			55	NA	NA	NA
	Total:			106	NA	NA	NA
Menke 2010d (parallel RCT)	I1: oxandrolone 0.03 mg/kg/day + GH	15 girls per dosage and age group were needed to achieve a power of 80% to detect a difference (P = 0.05, 2 sided) in first‐year height velocity of 2 cm with an assumed SD of 2.6. ITT analyses were performed and differences in adult height gain were also assessed by a PP analysis.	NR/184	46	42 (ITT) 30 (PP)	NA	NA	1 year (2 years)
	I2: oxandrolone 0.06 mg/kg/day + GH			39	36 (ITT) 22 (PP)	NA	NA
	C: GH + placebo			48	42 (ITT) 30 (PP)	NA	NA
	Total:			133	120 (ITT) 82 (PP)	NA	NA
Stahnke 2002e (parallel RCT)	I1: oxandrolone + GH	—	NR/91	44 (33)	NA	NA	NA	Initial phase: 12 months (2 years; 4 years; 5 years)
	I2: transient oxandrolone + GH			0 (38)	NA	NA	NA
	C: GH			47 (20)	NA	NA	NA
	Total:			91 (53)	NA	NA	NA
Rosenfeld 1998f (parallel RCT/CCT after 1–2 years)	I1: oxandrolone	—	NR/70e	18	NA	NA	NA	12–24 months (3–6 years)
	I2: oxandrolone + GH			17
	C1: GH			17
	C2: no treatment			18
	total:			70	NA	NA	NA
Job 1991 (parallel RCT)	I: oxandrolone + GH	—	NR/22	12	12	12	100	3 years
	C: GH			10	10	10	100
	Total:			22	22	22	100
Overall total	All interventions All comparators All interventions and comparators			267	—
				231
				498

^aFollow‐up under randomised conditions until end of trial (= duration of intervention + follow‐up postintervention or identical to duration of intervention); extended follow‐up refers to follow‐up of participants once the original trial was terminated as specified in the power calculation.
 ^bTwo years' double‐blind extension study, original trial duration 2 years.
 ^cTwo by two factorial design: participants were randomised to oxandrolone or placebo from 9 years of age until final height; those with evidence of ovarian failure at 12 years were further randomised to oral ethinylestradiol or placebo; participants who received placebo and those recruited after the age of 12.3 years started ethinylestradiol at age 14; "in 2004 and 2008, the sole European manufacturer of oxandrolone 2.5 mg ceased production, resulting in 34 participants in 2004 and 11 in 2008 temporarily suspending active oxandrolone treatment (mean duration: 2004, 52 days; 2008, 163 days). In 2008, the decision was taken to terminate the treatment arm forthwith, resulting in eight participants stopping oxandrolone treatment prematurely."
 ^d68 women (GH plus placebo: 23; oxandrolone 0.03 mg plus GH: 27; oxandrolone 0.06 mg plus GH: 18) participated in a follow‐up study (all participants and investigators remained blinded for the study medication, mean age was 24 years, mean time since stopping oxandrolone plus GH was 8.7 years); see Freriks 2012 under Menke 2010.
 ^eFirst 12 months: oxandrolone plus GH versus GH only therapy.
 ^fInitial 6 months observation period before randomisation; 70 girls admitted to trial, 67 girls completed the first trial year, 65 girls completed 3 years, 62 girls completed 3–6 years; at the end of an initial period of 12–20 months, girls in the original control and oxandrolone groups were given combination therapy of GH plus oxandrolone.

C: comparator; CCT: controlled clinical trial; GH: growth hormone; I: intervention; ITT: intention‐to‐treat; N: number; NA: not applicable; NR: not reported; PP: per protocol; RCT: randomised controlled trial; SD: standard deviation.

We provided information, including trial identifier for potentially relevant ongoing trials in the Characteristics of ongoing studies table and Appendix 7 ('Matrix of trial endpoints (publications and trial documents)'). We tried to find the protocol of each included trial and reported in Appendix 7 primary, secondary and other outcomes in comparison with data in publications.

We emailed all authors of included trials to enquire whether they were willing to answer questions regarding their trials. Appendix 14 shows the results of this survey. We sought relevant missing information on the trial from the primary trial authors, if required.

Dealing with duplicate and companion publications

In the event of duplicate publications, companion documents or multiple reports of a primary trial, we maximised the information yield by collating all available data and used the most complete data set aggregated across all known publications. We listed duplicate publications, companion documents, multiple reports of a primary trial, and trial documents of included trials (such as trial registry information) as secondary references under the study ID of the included trial.

Data from clinical trials registers

If data from included trials were available as study results in clinical trial registries, such as ClinicalTrials.gov or similar sources, we made full use of this information and extracted the data. If there was also a full publication of the trial, we collated and critically appraised all available data. If an included trial was marked as a completed trial in a clinical trial registry but no additional information (study results, publication, or both) was available, we planned to ad this trial to the 'Characteristics of studies awaiting classification' table.

Assessment of risk of bias in included studies

Three review authors (SM, HA, YSA) independently assessed the risk of bias of each included trial. We resolved disagreements by consensus or by consulting a fourth review author (YA). In cases of disagreement, we consulted the remainder of the review author team and made a judgement based on consensus. If adequate information was unavailable from the publications, trial protocols or other sources, we contacted the trial authors for more detail to request missing data on 'Risk of bias' items.

We used the Cochrane 'Risk of bias' assessment tool assigning assessments of low, high, or unclear risk of bias (see Appendix 2; Appendix 3; Higgins 2011a; Higgins 2017). We evaluated individual bias items as described in the Cochrane Handbook for Systematic Reviews of Interventions, according to the criteria and associated categorisations contained therein(Higgins 2017).

Summary assessment of risk of bias

We presented a 'Risk of bias' graph and a 'Risk of bias' summary figure.

We distinguished between self‐reported and investigator‐assessed and adjudicated outcome measures.

We considered the following self‐reported outcomes.

• Health‐related quality of life.

• Adverse events.

• Effects on cognition.

• Effect on psychological status.

• Effects on speech.

We considered the following endpoints to be investigator‐assessed.

• Increment in height velocity and final adult height.

• Adverse events.

• All‐cause mortality.

• Bone maturation.

• Effects on cognition.

• Effect on psychological status.

• Effects on speech.

• Socioeconomic effects.

Risk of bias for a trial across outcomes

Some risk of bias domains such as selection bias (sequence generation and allocation sequence concealment) affect the risk of bias across all outcome measures in a trial. In case of high risk of selection bias, all endpoints investigated in the associated trial were marked as 'high' risk. Otherwise, we did not perform a summary assessment of the risk of bias across all outcomes for a trial.

Risk of bias for an outcome within a trial and across domains

We assessed the risk of bias for an outcome measure including all of the entries relevant to that outcome (i.e. both trial‐level entries and outcome‐specific entries). 'Low' risk of bias was defined as low risk of bias for all key domains, 'unclear' risk of bias as unclear risk of bias for one or more key domains and 'high' risk of bias as high risk of bias for one or more key domains.

Risk of bias for an outcome across trials and across domains

These are our main summary assessments that were incorporated in our judgements about the certainty of the evidence in the 'Summary of finding' tables. 'Low' risk of bias was defined as most information coming from trials at low risk of bias, 'unclear' risk of bias as most information coming from trials at low or unclear risk of bias and 'high' risk of bias as sufficient proportion of information coming from trials at high risk of bias.

Measures of treatment effect

When there was at least two included trials for a comparison of a given outcome, we tried to express dichotomous data as odds ratio (OR) or risk ratios (RR) with 95% confidence intervals (CIs). For continuous outcomes measured on the same scale (e.g. weight loss in kilograms), we estimated the intervention effect using the mean difference (MD) with 95% CIs. For continuous outcomes measuring the same underlying concept (e.g. health‐related quality of life) but using different measurement scales, we planned to calculate the standardised mean difference (SMD) with 95% CIs. We planned to express time‐to‐event data as hazard ratios (HR) with 95% CIs.

Unit of analysis issues

We took into account the level at which randomisation occurred, such as cross‐over trials, cluster‐randomised trials and multiple observations for the same outcome. If more than one comparison from the same trial was eligible for inclusion in the same meta‐analysis, we either combined groups to create a single pair‐wise comparison, or we appropriately reduced the sample size so that the same participants did not contribute data to the meta‐analysis more than once (splitting the 'shared' group into two or more groups). Although the latter approach offers some solution for adjusting the precision of the comparison, it does not account for correlations arising from inclusion of the same set of participants being in multiple comparisons (Higgins 2011b).

We attempted to re‐analyse cluster‐RCTs that did not appropriately adjust for potential clustering of participants within clusters in their analysis. Variance of the intervention effects were inflated by a design effect. Calculation of a design effect involves estimation of an intracluster correlation coefficient (ICC). We obtained estimates of ICCs by contacting trial authors, or by imputing ICC values using either estimates from other included trials that reported ICCs or external estimates from empirical research (e.g. Bell 2013). We planned to examine the impact of clustering by performing sensitivity analyses.

Dealing with missing data

If possible, we obtained missing data from the authors of included trials. We carefully evaluated important numerical data such as screened, randomly assigned participants, as well as intention‐to‐treat, as‐treated and per‐protocol populations. We investigated attrition rates (e.g. dropouts, losses to follow‐up, withdrawals), and we critically appraised issues concerning missing data and imputation methods (e.g. last observation carried forward).

For trials in which the SD of the outcome was not available at follow‐up or we could not recreate it, we planned to standardise it using the mean of the pooled baseline SD from trials that reported this information.

When included trials did not report means and SDs for outcomes, and we did not receive requested information from trial authors, we planned to impute these values by estimating the mean and the variance from the median, range, and size of the sample (Hozo 2005).

We planned to investigate the impact of imputation on meta‐analyses by performing sensitivity analyses, and we reported for every outcome which trials had imputed SDs.

Assessment of heterogeneity

In the event of substantial clinical, methodological or statistical heterogeneity, we did not report trial results as the pooled effect estimate in a meta‐analysis.

We identified heterogeneity by visual inspection of the forest plots and by using a standard Chi² test with a significance level of α = 0.1 (Deeks 2017). In view of the low power of this test, we considered the I² statistic, which quantifies inconsistency across trials, to assess the impact of heterogeneity on the meta‐analysis (Higgins 2002; Higgins 2003).

Had we found heterogeneity, we planned to determine possible reasons for this by examining individual trial and subgroup characteristics.

Assessment of reporting biases

If we had included 10 or more trials investigating a particular outcome, we planned to use funnel plots to assess small‐trial effects. Several explanations may account for funnel plot asymmetry, including true heterogeneity of effect with respect to trial size, poor methodological design (and hence bias of small trials) and publication bias (Sterne 2017). Therefore, we interpreted results carefully (Sterne 2011).

Data synthesis

We planned to undertake (or display) a meta‐analysis only if we judged participants, interventions, comparisons and outcomes to be sufficiently similar to ensure an answer that was clinically meaningful. Unless good evidence showed homogeneous effects across trials of different methodological quality, we primarily summarised low risk of bias data using a random‐effects model (Wood 2008). We interpreted random‐effects meta‐analyses with due consideration for the whole distribution of effects and presented a prediction interval (Borenstein 2017a; Borenstein 2017b; Higgins 2009). A prediction interval requires at least three trials to be calculated and specifies a predicted range for the true treatment effect in an individual trial (Riley 2011). For rare events, such as event rates below 1%, we planned to use the Peto OR method, provided there was no substantial imbalance between intervention and comparator group sizes, and intervention effects were not exceptionally large. In addition, we performed statistical analyses according to the statistical guidelines presented in the Cochrane Handbook for Systematic Reviews of Interventions (Deeks 2017).

Subgroup analysis and investigation of heterogeneity

We expected the following characteristics to introduce clinical heterogeneity, and planned to carry out the following subgroup analyses including investigation of interactions.

• Participants who reached their final adult height at the end of the trial versus those who did not reach their final height.

• Participants who received intervention or comparator for three years versus less than three years.

• Age of participants.

• Participants who received low‐dose oxandrolone (0.03 mg/kg/day) versus those who received high‐dose oxandrolone (0.06 mg/kg/day).

Sensitivity analysis

We planned to perform sensitivity analyses to explore the influence of the following factors (when applicable) on effect sizes by restricting analysis to the following.

• Published studies.

• Taking into account risk of bias, as specified in the Assessment of risk of bias in included studies section.

• Very long or large studies to establish the extent to which they dominated the results.

• Use of the following filters: diagnostic criteria, imputation, language of publication, source of funding (industry versus other) or country.

We planned to test the robustness of results by repeating analyses using different measures of effect size (i.e. RR, OR, etc.) and different statistical models (fixed‐effect and random‐effects models).

Certainty of the evidence

We presented the overall certainty of the evidence for each outcome specified below, according to the GRADE approach, which takes into account issues related to internal validity (risk of bias, inconsistency, imprecision, publication bias) and external validity (such as directness of results). Three review authors (SM, HA, YSA) independently rated the certainty of the evidence for each outcome. We resolved differences in assessment by discussion or by consultation with a fourth review author (YA).

We included an appendix entitled 'Checklist to aid consistency and reproducibility of GRADE assessments' (Appendix 15), to help with standardisation of the 'Summary of findings' table (Meader 2014). Alternatively, we planned to use GRADEpro Guideline Development Tool (GDT) software and would have presented evidence profile tables as an appendix (GRADEpro GDT). We presented results for outcomes as described in the Types of outcome measures section. If meta‐analysis was not possible, we presented the results in a narrative format in the 'Summary of findings' table. We justified all decisions to downgrade the certainty of the evidence using footnotes, and we made comments to aid the reader's understanding of the Cochrane Review when necessary.

'Summary of findings' table

We presented a summary of the evidence in a 'Summary of findings' table. This provided key information about the best estimate of the magnitude of effect, in relative terms and as absolute differences for each relevant comparison of alternative management strategies, numbers of participants and trials addressing each important outcome, and a rating of overall confidence in effect estimates for each outcome. We created the 'Summary of findings' table using the methods described in the Cochrane Handbook for Systematic Reviews of Interventions (Schünemann 2017), along with Review Manager 5 (Review Manager 2014).

Interventions presented in the 'Summary of findings' table were oxandrolone plus GH and comparators were GH treatment only.

We reported the following outcomes, listed according to priority.

• Improvement in final adult height.

• Adverse events.

• Health‐related quality of life.

• Effects on speech.

• Effects on cognition.

• Effects on psychological status.

• All‐cause mortality.

Results

Description of studies

For a detailed description of trials, see the Characteristics of included studies, Characteristics of excluded studies, and Characteristics of ongoing studies tables.

Results of the search

The search identified 300 records. After removal of duplicates, we screened the title and abstracts of 199 records. We identified 164 of these as clearly irrelevant. The remaining 35 records underwent full‐text review (see Figure 1). We excluded five trials for the following reasons: no control group (Nilsson 1996), short follow‐up period (Ferrández 1989), and not an RCT design (Joss 1997; Sas 2014; Sheanon 2015). We included six trials (30 records) in the review (Gault 2011; Job 1991; Menke 2010; Rosenfeld 1998; Stahnke 2002; Zeger 2011).

1.

Study flow diagram. RCT: randomised controlled trial.

Included studies

A detailed description of the characteristics of included trials is presented elsewhere (see Characteristics of included studies table; Appendix 4; Appendix 5; Appendix 6; Appendix 7; Appendix 9; Table 2). The following is a succinct overview.

Source of data

We obtained all included trials from published literature.

Comparisons

In general, trials compared oxandrolone plus GH treatment to GH treatment alone. The dose of oxandrolone and GH treatments varied across trials, see Appendix 4.

Overview of trial populations

Trial authors randomised 498 participants, 267 participants to oxandrolone plus GH treatment and 231 participants to comparator groups. Individual trial sample size ranged from 22 to 133 participants (see Table 2).

Trial design

Five trials were multicentre (Gault 2011; Job 1991; Menke 2010; Rosenfeld 1998; Zeger 2011). The number of centres ranged from 2 to 36. Three trials were blinded for participants, personnel and outcome assessors (Gault 2011; Menke 2010; Zeger 2011). The trials were performed from 1983 to 2011. The duration of interventions ranged from 3 to 7.6 years. All six trials had a run‐in period. One trial was terminated early due to unavailability of oxandrolone because the sole European manufacturer of oxandrolone stopped production in 2004 and 2008 (Gault 2011).

Settings

Five trials were conducted in paediatric clinics or paediatric endocrinology clinics (Gault 2011; Job 1991; Menke 2010; Stahnke 2002; Zeger 2011). One trial was in medical centres (Rosenfeld 1998).

Participants

All participants were aged up to 18 years with TS. All participants came from high‐income countries. Most trials did not specify ethnic groups. The mean age of participants at start of therapy ranged from 9 to 12 years. All trials reported comorbidities, co‐interventions and comedications used by participants. Major exclusion criteria included major illnesses and previous use of hormonal treatments (see Appendix 5; Appendix 6).

Diagnosis

All trials include females with TS, with specific referral to diagnosis criteria.

Interventions

None of the trials reported treatment before enrolment. The intervention (oxandrolone) was given as oral medication. The total dose for oxandrolone ranged from a daily dose of 0.03 mg/kg to 0.125 mg/kg (0.125 mg/kg (Rosenfeld 1998), 0.1 mg/kg (Stahnke 2002), 0.06 mg/kg (Job 1991; Menke 2010; Zeger 2011), 0.05 mg/kg (Gault 2011; Stahnke 2002), 0.03 mg/kg (Menke 2010)).

Duration of treatment ranged from 3 to 13 years. All trials used interventions and comparators (two trials used two comparator arms which we did not include in the final analysis; Rosenfeld 1998; Stahnke 2002; Appendix 4).

Outcomes

Two trials explicitly stated primary and secondary outcomes in the publication (Gault 2011; Menke 2010). Three trials had information in trial registers (Gault 2011; Menke 2010; Zeger 2011), and one trial published a paper on the design of the study (Stahnke 2002) (see Appendix 7). Where available, there were no major differences between defined primary outcomes in the publication and trial registers. All trials collected a median of one (one to six) outcomes. Five trials reported adverse events (Gault 2011; Menke 2010; Rosenfeld 1998; Stahnke 2002; Zeger 2011). Only one trial investigated health‐related quality of life (Menke 2010). The included trials had variable definitions of our primary outcome measurement improvement in final adult height (see Appendix 9).

Excluded studies

We excluded five trials that did not fulfil our inclusion criteria. One trial had no comparator group (Nilsson 1996), one had a short follow‐up period (six months only; Ferrández 1989), and three trials were not RCTs (Joss 1997; Sas 2014; Sheanon 2015). A short description of the excluded trials can be found in the Characteristics of excluded studies table.

Risk of bias in included studies

For details on the risk of bias of the included trials, see the Characteristics of included studies table.

For an overview of review authors' judgements about each risk of bias item for individual trials and across all trials, see Figure 2 and Figure 3.

2.

Risk of bias graph: review authors' judgements about each risk of bias item presented as percentages across all included studies (blank cells indicate that the particular outcome was not measured in some studies).

3.

Risk of bias summary: review authors' judgements about each risk of bias item for each included study (blank cells indicate that the study did not measure that particular outcome).

Allocation

Two trials reported adequately on random sequence generation and allocation concealment (low risk of bias; Gault 2011; Menke 2010). The remaining trials were at unclear risk of selection bias.

Blinding

Three trials were double‐blinded for participants and personnel and blinded on all reported outcomes for outcome assessors (Gault 2011; Menke 2010; Zeger 2011). Three trials had no sufficient information on blinding procedures (unclear risk; Job 1991; Rosenfeld 1998; Stahnke 2002). We judged two trials at high risk of performance bias and detection bias for the outcome adverse events (Rosenfeld 1998; Stahnke 2002).

Incomplete outcome data

Three trials (Job 1991; Menke 2010; Zeger 2011) performed an ITT analysis (low risk of bias). We judged one trial at high risk of attrition bias for the outcome measures adverse events and improvement in final height/increase in height velocity (Stahnke 2002).

Selective reporting

Four trials were at low risk of reporting bias (Gault 2011; Menke 2010; Rosenfeld 1998; Zeger 2011). Two trials were at high risk of reporting bias.

Other potential sources of bias

Five trials were at low risk of bias (Gault 2011; Job 1991; Menke 2010; Rosenfeld 1998; Zeger 2011). One trial was at high risk if other bias because participants were switched between groups to create a third new group after randomisation that could have negatively affected the randomisation process (Stahnke 2002).

Effects of interventions

See: Table 1

Baseline characteristics

For details of baseline characteristics, see Appendix 5 and Appendix 6.

Oxandrolone plus growth hormone versus growth hormone only

Primary outcomes

Improvement in final adult height

Oxandrolone plus GH compared with GH only showed an improvement in final height (MD 2.7 cm, 95% CI 1.3 to 4.1; P < 0.001; 5 trials, 270 participants; Analysis 1.1; moderate‐quality evidence). The 95% prediction interval ranged between 0.3 cm and 5.1 cm.

1.1. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 1 Final adult height.

Several additional analyses did not change this result substantially (e.g. excluding low‐quality trials, Analysis 1.2, and the subgroup of participants who reached final adult height by excluding trials measuring near adult height and predicted height, Analysis 1.3). For illustrative purposes, we provided supplementary data with very limited amount of information reporting various height measures and usually consisting of one trial only (Analysis 1.4; Analysis 1.5; Analysis 1.6; Analysis 1.7; Analysis 1.8; Analysis 1.9; Analysis 1.10; Analysis 1.11; Analysis 1.12; Analysis 1.13; Analysis 1.14; Analysis 1.15; Analysis 1.16; Analysis 1.17; Analysis 1.18; Analysis 1.19; Analysis 1.20; Analysis 1.21; Analysis 1.22). Data on adult height could not be used from one trial because authors did not provide SDs (Zeger 2011).

1.2. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 2 Final adult height: excluding low‐quality trials.

1.3. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 3 Final adult height: trials reporting strictly adult height (excluding trials measuring near adult height and predicted height).

1.4. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 4 Final adult height: trials reporting strictly adult height (excluding low‐quality trials).

1.5. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 5 Final height/near adult height reported.

1.6. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 6 Predictable adult height.

1.7. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 7 Maximum height.

1.8. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 8 Adult height gain.

1.9. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 9 Final adult height SDS (Turner reference).

1.10. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 10 Final adult height SDS (healthy reference).

1.11. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 11 Final adult height delta height SDS (Turner reference).

1.12. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 12 Final adult height delta height SDS (healthy reference).

1.13. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 13 Final adult height minus predicted adult height (height gain).

1.14. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 14 Difference in height gain vs placebo (final height – placebo final height).

1.15. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 15 Near adult height.

1.16. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 16 Near adult height SDS (Turner reference).

1.17. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 17 Near adult height SDS (healthy reference).

1.18. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 18 Near adult height delta height SDS (Turner reference).

1.19. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 19 Near adult height delta height SDS (healthy reference).

1.20. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 20 Increase in near adult height from baseline.

1.21. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 21 Near adult height minus predicted adult height.

1.22. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 22 Difference in height gain over placebo (near adult height – placebo final height).

Health‐related quality of life

One trial investigated health‐related quality of life (Menke 2010).

The follow‐up study of Menke 2010, where only 68 women (133 children and adolescents were initially randomised) participated in the double‐blinded follow‐up, with a mean age of 24 years and mean time since stopping oxandrolone plus GH of 8.7 years, reported health‐related quality of life using three questionnaires (Beck Depression Inventory – Second Edition Netherlands (BDI‐II‐NL); Symptom Checklist‐90‐revised (SCL‐90‐R); SF‐36). The comparison of oxandrolone plus GH versus GH only in 46 participants for total and mean scores showed inconclusive results (Analysis 1.23; Analysis 1.24; Analysis 1.25).

1.23. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 23 Health‐related quality of life (BDI‐II‐NL).

1.24. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 24 Health‐related quality of life (SCL‐90‐R).

1.25. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 25 Health‐related quality of life (SF‐36).

Adverse events

The random‐effects RR for adverse events comparing oxandrolone plus GH with GH only was 3.27 (95% CI 0.75 to 14.22; P = 0.11; 4 trials, 283 participants; Analysis 1.26). Thirty‐five out of 162 (21.6%) participants receiving oxandrolone plus GH compared with 8/121 (6.6%) participants receiving GH only reported adverse events. Fixed‐effect meta‐analysis showed a RR of 3.43 (95% CI 1.64 to 7.17). Evidence‐synthesis in the case of very few trials is difficult (Bender 2018). Elimination of the trials with high or unclear risk of performance bias, detection bias and attrition bias for the outcome measure adverse events (Rosenfeld 1998; Stahnke 2002) resulted in a RR of 1.81 (95% CI 0.83 to 3.96; P = 0.42; 2 trials, 170 participants; low‐quality evidence). In the remaining two trials, 16/86 (18.6%) participants receiving oxandrolone plus GH compared with 8/84 (9.5%) participants receiving GH only reported adverse events (Menke 2010; Zeger 2011). Adverse events comprised signs of virilisation (e.g. deepening of the voice, hirsutism), hypothyroidism, impaired glucose tolerance and increased liver enzymes. For full details, see Appendix 10; Appendix 11; Appendix 12.

1.26. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 26 Adverse events.

Secondary outcomes

Increase in height velocity

Oxandrolone plus GH compared with GH only for height velocity showed different results depending on whether a random‐effects model or fixed effect model was used. Trials used several methods to report the increment in height per year (Analysis 1.27; Analysis 1.28; Analysis 1.29; Analysis 1.30; Analysis 1.31; Analysis 1.32; Analysis 1.33; Analysis 1.34; Analysis 1.35). Taking into account the small number of trials and most effect estimates pointing in the same direction, we used a qualitative evidence synthesis approach (Bender 2018).

1.27. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 27 Height growth (velocity) year 1.

1.28. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 28 Height growth (velocity) SDS year 1 (Turner reference).

1.29. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 29 Height velocity year 2.

1.30. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 30 Height velocity, SDS year 2 (Turner reference).

1.31. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 31 Height velocity, SDS year 2 (healthy reference).

1.32. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 32 Height velocity year 3.

1.33. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 33 Height velocity SDS year 3 (Turner reference).

1.34. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 34 Height velocity year 4.

1.35. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 35 Height velocity SDS year 4 (Turner reference).

• Two trials (59 participants) measured height velocity at one year and showed an MD in favour of oxandrolone plus GH of 1.8 cm and 3.8 cm (Analysis 1.27).

• Three trials (98 participants) measured height velocity at two years and showed an MD in favour of oxandrolone plus GH of 0.3 cm, 1.5 cm and 1.9 cm (Analysis 1.29).

• Two trials (57 participants) measured height velocity at three years and showed an MD in favour of oxandrolone plus GH of 0.7 cm and 2.1 cm (Analysis 1.32).

• Two trials (107 participants) measured height velocity at four years and showed a disadvantage and advantage of oxandrolone plus GH of –0.2 cm and 4 cm (Analysis 1.34).

All‐cause mortality

None of the trials reported all‐cause mortality.

Bone maturation

Oxandrolone plus GH compared with GH only showed an MD for bone age of 0.16 years (95% CI –0.46 to 0.78; P = 0.23; 3 trials, 91 participants; Analysis 1.36).

1.36. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 36 Bone maturation, bone age in years.

Effects on cognition

Zeger 2011 reported summary scores for working memory, spatial cognition, executive function and verbal abilities using the Wechsler Intelligence Scale for Children‐Revised (WISC‐R). After two years, comparison of oxandrolone plus GH versus GH only in 51 participants showed inconclusive results (very low‐quality evidence; Analysis 1.37).

1.37. Analysis.

Comparison 1 Oxandrolone plus growth hormone (GH) versus GH only, Outcome 37 Effect on cognition at year 2.

Effects on psychological status

One trial in 106 participants investigated psychological and behavioural characteristics using the Child Behavior Checklist (CBCL), the Junior Dutch Personality Questionnaire (DPQ‐J), the state‐subscale of the Spielberger's State‐Trait Anger Scale, the Romantic and Sexual Interest Questionnaire, the Mood Questionnaire and the Gender Role Questionnaire (Menke 2010). There were no evident psychological virilising effects in the areas of behaviour, aggression, romantic and sexual interest, mood and gender role (very low‐quality evidence).

Effects on speech

One trial including 88 participants assessed whether oxandrolone treatment caused voice deepening (Menke 2010). GH‐treated girls had relatively high‐pitched voices. Oxandrolone plus GH decreased voice frequency in a dose‐dependent way. Most voice frequencies remained within the normal range, but became occasionally lower, especially with oxandrolone plus GH 0.06 mg/kg/day treatment. Results were inconclusive for oxandrolone plus GH 0.03 mg/kg/day (very low‐quality evidence).

Socioeconomic effects

None of the trials reported on details of socioeconomic effects. However, one trial comparing oxandrolone plus GH with GH plus placebo reported costs of treatment (Menke 2010). Trial authors stated: "When corrected for bone age at starting GH, the duration of GH therapy was shorter on GH+Ox [growth hormone plus oxandrolone] 0.03 and 0.06 (–0.4 yr [years] and –0.8 yr, P = 0.06 and P = 0.001, respectively), and the cumulative costs of GH were lower (–10,100 ± 6,100 and –13,500 ± 6,300 euro, P = 0.1 and P = 0.03, respectively) than on GH+Pl [growth hormone plus placebo] (mean cumulative costs, 161,200 ± 59,500 euro)".

Subgroup analyses

With the exception of the subgroup of participants who reached final adult height by excluding trials measuring near adult height and predicted height, we did not perform subgroups analyses because there were not enough trials.

Sensitivity analyses

With the exception of low‐quality versus high‐quality trials, we did not perform sensitivity analyses because there were not enough trials.

Assessment of reporting bias

We did not draw funnel plots due to limited number of trials.

Ongoing trials

We found no ongoing trials. All trials registered in different phases were completed and published.

Discussion

Summary of main results

This Cochrane systematic review intended to examine the effects of oxandrolone therapy on different outcomes in girls with TS. Six RCTs met our inclusion criteria and all provided data. All trials compared a combination of oxandrolone and GH, at different dosing regimens, to GH alone or combined with placebo. The intervention period lasted from 3 to 7.6 years. Trials were carried out from 1983 to 2011 and enrolled 498 participants from fairly comparable patient populations.

The findings of our review led to the following main conclusions: overall, oxandrolone had a positive effect on adult height, height gain and height velocity when used in combination with GH. There were numerically more adverse events reported in the oxandrolone treatment arms. However, results did not clearly show an advantage or disadvantage of the intervention or comparator group, which may be caused by including only two trials and a small number of participants with events.

Overall completeness and applicability of evidence

This review provides the most up‐to‐date assessment of the efficacy and safety of the combination therapy of oxandrolone and GH on adult height in girls with TS. The six included trials used a combination of oxandrolone and GH, no trial used oxandrolone alone for the whole intervention period. All trials compared the combination therapy to GH alone.

This review provides reliable information about the impact of oxandrolone in combination with GH on a range of adult height outcomes. Adult height was the most commonly reported outcome among the included trials. Other commonly reported outcomes were adult height gain, height velocity, bone maturation and overall presence of adverse events. Limited evidence due to small number of studies was available for the outcomes bone maturation, socioeconomic effects, cognition, psychological status and speech. No trial reported all‐cause mortality or health‐related quality of life (one follow‐up study with less than 50% of initially randomised participants showed inconclusive results comparing oxandrolone plus GH for total and mean scores in three different questionnaires).

Quality of the evidence

We assessed the certainty of the evidence for our prespecified outcomes using the GRADE approach (Appendix 15). Most trials did not report study methodology in adequate detail. For example, we judged that only two trials sufficiently reported both allocation concealment and random sequence generation (Gault 2011; Menke 2010). Contact with trial authors did not provide any additional information.

The major reason for downgrading the level of certainty of the evidence was imprecision due to the low number of trials, low number of participants or both.

Potential biases in the review process

We followed the Cochrane search strategy which includes searching trials registers as well as handsearching any relevant articles or review papers. Therefore, we believe it is unlikely that we missed relevant trials. However, there is a possibility that we did not detect unpublished data. Due to the small number of trials and limited amount of data, we could only perform very limited subgroup and sensitivity analyses. The doses used for the interventions and comparisons varied across trials and we were unable to analyse each dose regimen separately due to the limited number of trials. In addition, treatment durations and follow‐up periods varied across trials. There was incomplete outcome reporting in some trials. Although we attempted to locate all relevant data, we were unable to reach the original trials investigators due to incomplete correspondence with trial investigators.

Agreements and disagreements with other studies or reviews

Our review and meta‐analysis supports the conclusion of three recent reviews. The review published by Sheanon and Backeljauw (Sheanon 2015), included four published, randomised, placebo‐controlled, double‐blinded trials. The four mentioned trials were also included in our meta‐analysis and they revealed that adding oxandrolone to GH‐treated girls with TS can result in an increase in height velocity and lead to a mild increase in their adult heights. The review published by Li, Cheng and Xiu (Li 2018) included 10 published, randomised trials (10 trials and 11 records; Davenport 2007; Gault 2011; Job 1991; Menke 2010; Quigley 2002; Rosenfeld 1986; Rosenfeld 1998; Ross 2011; Stahnke 2002; Stephure 2005; Zeger 2011). We excluded some of these records as they were out of the scope of our review, and included the remainder. Li and colleagues concluded that adding oxandrolone to the therapy of GH‐treated girls with TS resulted in an increase in height velocity and led to a mild increase in their adult heights by 2.46 cm (Li 2018). Another narrative review was published by a group of authors investigating RCTs addressing the effects of oxandrolone in GH‐treated girls with TS (Sas 2014). This review found that addition of oxandrolone increased the final adult height by 2.3 cm to 4.6 cm. Gault and colleagues published a conference abstract in 2015 (Gault 2015), which updated the interim analysis of the 2011 paper (Gault 2011), and added 20 participants with data that could not be included in our meta‐analysis as there were no comparison groups and different outcomes (i.e. delayed pubertal induction) (Beller 2013).

Authors' conclusions

Implications for practice.

The available data support the use of the combination therapy of oxandrolone and growth hormone (GH) in comparison to GH only for improving adult height in girls aged up to 18 years with Turner syndrome. Adverse events appeared comparable between the combination therapy and GH treatment alone. However, only two trials provided reliable data. No trial reported all‐cause mortality or health‐related quality of life (one follow‐up study with less than 50% of initially randomised participants showed inconclusive results comparing oxandrolone plus GH for total and mean scores in three different questionnaires). Comparisons for the outcomes cognition, psychological status and speech (voice frequency) showed inconclusive results. The doses and durations of oxandrolone therapy across trials varied, making it difficult to draw conclusions on the optimal regimen.

Implications for research.

The effect of oxandrolone on girls with Turner syndrome needs to be further investigated. In particular, future trials are needed to confirm the optimal dosing regimen and duration of oxandrolone therapy. Further research is also required to get a better insight into patient‐important outcomes such as health‐related quality of life, mortality, cognition, psychological status and speech. Finally, we advise investigators planning to study oxandrolone therapy in girls with Turner syndrome to use standardised definitions for study outcomes, provide detailed information on the intervention used including any other co‐interventions, and report on all possible adverse events with oxandrolone and comparator therapy.

Notes

Parts of the background and methods sections, the appendices, additional tables and figures 1 to 3 of this review are based on a standard template established by the Cochrane Metabolic and Endocrine Disorders Group.

Appendices

Appendix 1. Search strategies

Cochrane Library

1. [mh "Turner Syndrome"] 2. ((turner* or bonnevie or ullrich) near/3 syndrom*):ti,ab,kw 3. ("gonadal dysgenesis" or "monosomy X"):ti,ab,kw 4. {or #1‐#3} 5. [mh Oxandrolone] 6. (oxandrolone* or oxandrin or anavar or "SC 11585" or SC11585 ):ti,ab,kw 7. #5 or #6 8. #4 and #7

MEDLINE (OvidSP)

1. Turner Syndrome/ 2. ((turner* or bonnevie or ullrich) adj3 syndrome*).tw. 3. (gonadal dysgenesis or monosomy X).tw. 4. or/1‐3 5. Oxandrolone/ 6. (oxandrolone* or oxandrin or anavar or "SC 11585" or SC11585 ).tw. 7. or/5‐6 8. 4 and 7

Embase (OvidSP)

1. Turner syndrome/ 2. ((turner* or bonnevie or ullrich) adj3 syndrome*).tw. 3. (gonadal dysgenesis or monosomy X).tw. 4. or/1‐3 5. oxandrolone/ 6. (oxandrolone* or oxandrin or anavar or "SC 11585" or SC11585 ).tw. 7. or/5‐6 8. 4 and 7 [9:Wong 2006"sound treatment studies" filter – BS version] 9. random*.tw. or clinical trial*.mp. or exp health care quality/ 10. 8 and 9

WHO ICTRP Search Portal (standard search)

turner* AND oxandrolone* OR turner* AND oxandrin OR turner* AND anavar OR turner* AND SC11585 OR bonnevie AND oxandrolone* OR bonnevie AND oxandrin OR bonnevie AND anavar OR bonnevie AND SC11585 OR ullrich AND oxandrolone* OR ullrich AND oxandrin OR ullrich AND anavar OR ullrich AND SC11585

ClinicalTrials.gov (advanced search)

Search Terms: ("Turner syndrome" OR "Turners syndrome" OR "ullrich syndrome" OR " gonadal dysgenesis" OR "monosomy X") AND (oxandrolone OR oxandrin OR anavar OR "SC 11585" OR SC11585)

Manufacturers' websites, FDA and EMA websites

oxandrolone

Appendix 2. Assessment of risk of bias

Risk of bias domains

Random sequence generation (selection bias due to inadequate generation of a randomised sequence) For each included trial, we described the method used to generate the allocation sequence in sufficient detail to allow an assessment of whether it should produce comparable groups. Low risk of bias: the trial authors achieved sequence generation using computer‐generated random numbers or a random numbers table. Drawing of lots, tossing a coin, shuffling cards or envelopes and throwing dice were adequate if an independent person performed this who was not otherwise involved in the trial. We considered the use of the minimisation technique as equivalent to being random. Unclear risk of bias: insufficient information about the sequence generation process. High risk of bias: the sequence generation method was non‐random or quasi‐random (e.g. sequence generated by odd or even date of birth; sequence generated by some rule based on date (or day) of admission; sequence generated by some rule based on hospital or clinic record number; allocation by judgement of the clinician; allocation by preference of the participant; allocation based on the results of a laboratory test or a series of tests; or allocation by availability of the intervention). Allocation concealment (selection bias due to inadequate concealment of allocation prior to assignment) We described for each included trial the method used to conceal allocation to interventions prior to assignment and we assessed whether intervention allocation could have been foreseen in advance of or during recruitment, or changed after assignment. Low risk of bias: central allocation (including telephone, interactive voice‐recorder, web‐based and pharmacy‐controlled randomisation); sequentially numbered drug containers of identical appearance; sequentially numbered, opaque, sealed envelopes. Unclear risk of bias: insufficient information about the allocation concealment. High risk of bias: used an open random allocation schedule (e.g. a list of random numbers); assignment envelopes used without appropriate safeguards; alternation or rotation; date of birth; case record number; any other explicitly unconcealed procedure. We also evaluated trial baseline data to incorporate assessment of baseline imbalance into the 'Risk of bias' judgement for selection bias (Corbett 2014). Chance imbalances may also affect judgements on the risk of attrition bias. In the case of unadjusted analyses, we distinguished between trials that we rated at low risk of bias on the basis of both randomisation methods and baseline similarity, and trials that we judged at low risk of bias on the basis of baseline similarity alone (Corbett 2014). We reclassified judgements of unclear, low or high risk of selection bias as specified in Appendix 3. Blinding of participants and study personnel (performance bias due to knowledge of the allocated interventions by participants and personnel during the trial) We evaluated the risk of detection bias separately for each outcome (Hróbjartsson 2013). We noted whether endpoints were self‐reported, investigator‐assessed or adjudicated outcome measures (see below). Low risk of bias: blinding of participants and key study personnel was ensured, and it was unlikely that the blinding could have been broken; no blinding or incomplete blinding, but we judged that the outcome was unlikely to have been influenced by lack of blinding. Unclear risk of bias: insufficient information about the blinding of participants and study personnel; the trial did not address this outcome. High risk of bias: no blinding or incomplete blinding, and the outcome was likely to have been influenced by lack of blinding; blinding of trial participants and key personnel attempted, but likely that the blinding could have been broken, and the outcome was likely to have been influenced by lack of blinding. Blinding of outcome assessment (detection bias due to knowledge of the allocated interventions by outcome assessment) We evaluated the risk of detection bias separately for each outcome (Hróbjartsson 2013). We noted whether endpoints were self‐reported, investigator‐assessed or adjudicated outcome measures (see below). Low risk of bias: blinding of outcome assessment was ensured, and it was unlikely that the blinding could have been broken; no blinding of outcome assessment, but we judged that the outcome measurement was unlikely to have been influenced by lack of blinding. Unclear risk of bias: insufficient information about the blinding of outcome assessors; the trial did not address this outcome. High risk of bias: no blinding of outcome assessment, and the outcome measurement was likely to have been influenced by lack of blinding; blinding of outcome assessment, but likely that the blinding could have been broken, and the outcome measurement was likely to have been influenced by lack of blinding. Incomplete outcome data (attrition bias due to amount, nature or handling of incomplete outcome data) For each included trial or each outcome (or both), we described the completeness of data, including attrition and exclusions from the analyses. We stated whether the trial reported attrition and exclusions, and reported the number of participants included in the analysis at each stage (compared with the number of randomised participants per intervention/comparator group). We also noted if the trial reported the reasons for attrition or exclusion and whether missing data were balanced across groups or were related to outcomes. We considered the implications of missing outcome data per outcome such as high dropout rates (e.g. above 15%) or disparate attrition rates (e.g. difference of 10% or more between trial arms). Low risk of bias: no missing outcome data; reasons for missing outcome data unlikely to be related to true outcome (for survival data, censoring unlikely to introduce bias); missing outcome data balanced in numbers across intervention groups, with similar reasons for missing data across groups; for dichotomous outcome data, the proportion of missing outcomes compared with observed event risk was not enough to have a clinically relevant impact on the intervention effect estimate; for continuous outcome data, plausible effect size (mean difference or standardised mean difference) among missing outcomes was not enough to have a clinically relevant impact on observed effect size; appropriate methods, such as multiple imputation, were used to handle missing data. Unclear risk of bias: insufficient information to assess whether missing data in combination with the method used to handle missing data were likely to induce bias; the trial did not address this outcome. High risk of bias: reason for missing outcome data was likely to be related to true outcome, with either imbalance in numbers or reasons for missing data across intervention groups; for dichotomous outcome data, the proportion of missing outcomes compared with observed event risk enough to induce clinically relevant bias in the intervention effect estimate; for continuous outcome data, plausible effect size (mean difference or standardised mean difference) among missing outcomes enough to induce clinically relevant bias in observed effect size; 'as‐treated' or similar analysis done with substantial departure of the intervention received from that assigned at randomisation; potentially inappropriate application of simple imputation. Selective reporting (reporting bias due to selective outcome reporting) We assessed outcome reporting bias by integrating the results of the appendix 'Matrix of trial endpoints (publications and trial documents)' (Boutron 2014; Mathieu 2009), with those of the appendix 'High risk of outcome reporting bias according to the Outcome Reporting Bias In Trials (ORBIT) classification' (Kirkham 2010). This analysis formed the basis for the judgement of selective reporting. Low risk of bias: the trial protocol was available and all the trial's prespecified (primary and secondary) outcomes that were of interest to this review were reported in the prespecified way; the study protocol was unavailable, but it was clear that the published reports included all expected outcomes (ORBIT classification). Unclear risk of bias: insufficient information about selective reporting. High risk of bias: not all the trial's prespecified primary outcomes were reported; one or more primary outcomes were reported using measurements, analysis methods or subsets of the data (e.g. subscales) that were not prespecified; one or more reported primary outcomes were not prespecified (unless clear justification for their reporting was provided, such as an unexpected adverse effect); one or more outcomes of interest in the Cochrane Review were reported incompletely so that we could not enter them in a meta‐analysis; the trial report failed to include results for a key outcome that we would expect to have been reported for such a trial (ORBIT classification). Other bias Low risk of bias: the trial appeared free from other sources of bias. Unclear risk of bias: there was insufficient information to assess whether an important risk of bias existed; insufficient rationale or evidence that an identified problem introduced bias. High risk of bias: the trial had a potential source of bias related to the specific trial design used; the trial was claimed to be fraudulent; or the trial had some other serious problem.

Appendix 3. Selection bias decisions

Selection bias decisions for trials reporting unadjusted analyses – comparison of results obtained using method details alone with results using method details and trial baseline informationa
Reported randomisation and allocation concealment methods	Risk of bias judgement using methods reporting	Information gained from study characteristics data	Risk of bias using baseline information and methods reporting
Unclear methods	Unclear risk	Baseline imbalances present for important prognostic variable(s)	High risk
		Groups appeared similar at baseline for all important prognostic variables	Low risk
		Limited or no baseline details	Unclear risk
Would generate a truly random sample, with robust allocation concealment	Low risk	Baseline imbalances present for important prognostic variable(s)	Unclear riskb
		Groups appeared similar at baseline for all important prognostic variables	Low risk
		Limited baseline details, showing balance in some important prognostic variablesc	Low risk
		No baseline details	Unclear risk
Sequence was not truly random, or allocation concealment was inadequate	High risk	Baseline imbalances present for important prognostic variable(s)	High risk
		Groups appeared similar at baseline for all important prognostic variables	Low risk
		Limited baseline details, showing balance in some important prognostic variablesc	Unclear risk
		No baseline details	High risk
aTaken from Corbett 2014; judgements highlighted in bold indicate situations in which the addition of baseline assessments would change the judgement about risk of selection bias, compared with using methods reporting alone. bImbalance identified that appeared likely to be due to chance. cDetails for the remaining important prognostic variables were not reported.

Appendix 4. Description of interventions

Study ID		Intervention(s) and comparator Description (mean number of engagements per week, frequency of use)
Zeger 2011	I:	Oxandrolone 0.06 mg/kg/day PO + GH 0.05 mg/kg daily, SC for 2 years Participants who choose to continue were started on ethinylestradiol 50 ng/kg/day during year 3 and 100 ng/kg/day during year 4 (2‐year extension)
	C:	Placebo + GH 0.05 mg/kg daily, SC for 2 years Participants who choose to continue were started on ethinylestradiol 50 ng/kg/day during year 3 and 100 ng/kg/day during year 4 (2‐year extension)
Gault 2011	I:	Oxandrolone 0.05 mg/kg/day PO from 9 years of age + GH 10 mg/m²/week, daily SC I: and C: 2 × 2 factorial design: girls with evidence of ovarian failure at 12 years were further randomised to oral ethinylestradiol (year 1: 2 μg/day, year 2: 4 μg/day, year 3: 4 months each of 6 μg/day, 8 μg/day and 10 μg daily) or placebo, the placebo group subsequently started ethinylestradiol at 14 years. The code for randomisation 2 was broken at 15 years to allow introduction of progesterone therapy: oral norethisterone 5 mg/day for 5 days each month
	C:	Placebo from 9 years of age + GH 10 mg/m²/week, daily SC
Menke 2010	I1:	Oxandrolone 0.03 mg/kg/day, PO at bedtime, from 8 years of age + GH 1.33 mg/m², SC, OD In the absence of spontaneous puberty, oestrogen therapy was started between the age of 12.0 and 12.99 (after a number of full years of GH therapy) in age groups 1 (2.00–7.99 years) and 2 (8.00–11.99 years) and at inclusion (i.e. between the age of 12 and 16 years) in age group 3 (12.00–15.99 years) Cyclic progesterone was added after ≥ 2 years of oestrogen therapy. Doses were adjusted every 6 months, and interventions were stopped when height velocity was less than 1 cm per 6 months or when participants decided to stop because they were satisfied with their height. Thereafter participants were followed up for 2 subsequent year visits to measure growth after discontinuing interventions
	I2:	Oxandrolone 0.06 mg/kg/day PO at bedtime, from 8 years of age + GH 1.33 mg/m², SC, OD In the absence of spontaneous puberty, oestrogen therapy was started between the age of 12.0 and 12.99 (after a number of full years of GH therapy) in age groups 1 (2.00–7.99 years) and 2 (8.00–11.99 years) and at inclusion (i.e. between the age of 12 and 16 years) in age group 3 (12.00–15.99 years) Cyclic progesterone was added after ≥ 2 years of oestrogen therapy. Doses were adjusted every 6 months, and interventions were stopped when height velocity was < 1 cm per 6 months or when participants decided to stop because they were satisfied with their height. Thereafter participants were followed up for 2 subsequent year visits to measure growth after discontinuing interventions
	C:	Placebo, PO, OD from 8 years of age + GH 1.33 mg/m² daily, SC at bedtime In the absence of spontaneous puberty, oestrogen therapy was started between the age of 12.0 and 12.99 (after a number of full years of GH therapy) in age groups 1 (2.00–7.99 years) and 2 (8.00–11.99 years) and at inclusion (i.e. between the age of 12 and 16 years) in age group 3 (12.00–15.99 years) Cyclic progesterone was added after ≥ 2 years of oestrogen therapy. Doses were adjusted every 6 months, and interventions were stopped when height velocity was < 1 cm per 6 months or when participants decided to stop because they were satisfied with their height. Thereafter participants were followed up for 2 subsequent year visits to measure growth after discontinuing interventions
Stahnke 2002	I1:	Oxandrolone 0.1 mg/kg/day + GH 2.6 IU/m² daily (18 IU/m²/week), SC; during year 2: oxandrolone was reduced to 0.05 mg/kg and after 2 years GH was increased to 24–28 IU/m²/week
	I2:	Transient oxandrolone + GH Some of the girls were taken off oxandrolone due to virilising adverse effects of oxandrolone. Oestrogen was initiated at a mean age of 14.9 years
	C1:	GH 2.6 IU/m² daily (18 IU/m²/week), SC; during year 2: 3.4 IU/m² (24 IU/m²/week) and later 28 IU/m²/week
	C2:	Transient oxandrolone + GH Some of the girls were later given oxandrolone in addition to GH because of waning growth velocity. Oestrogen was initiated at a mean age of 14.9 years
Rosenfeld 1998	I1:	Oxandrolone 0.125 mg/kg/day + GH 0.125 mg/kg 3 times per week At the end of the initial period of 12–24 months, oxandrolone was lowered to 0.0625 mg/kg/day. Oestrogen treatment started ≥ 14 years
	I2:	Oxandrolone 0.125 mg/kg/day At the end of the initial period of 12–24 months, girls in the original control and oxandrolone groups were given a combination of GH and oxandrolone (lowered to 0.0625 mg/kg/day). Oestrogen treatment started at age ≥ 14 years
	C1:	GH 0.125 mg/kg 3 times per week After 3–4 years, GH total weekly dose of 0.375 mg/kg was divided into 7 daily administrations. Oestrogen treatment started ≥ 14 years
	C2:	No treatment ("control") At the end of the initial period of 12–24 months, girls in the original control and oxandrolone groups were given a combination of GH and oxandrolone (lowered to 0.0625 mg/kg/day). Oestrogen treatment started at age ≥ 14 years
Job 1991	I:	Oxandrolone 0.06 mg/kg/day, PO + GH 0.01 U/kg, SC at night, OD
	C:	GH 0.01 U/kg daily, SC at night
C: comparator; GH: growth hormone; I: intervention; OD: once daily: PO: orally; SC: subcutaneous.

Appendix 5. Baseline characteristics (I)

Study ID	Intervention(s) and comparator(s)	Duration of intervention (years)	Description of participants	Study period (year to year)	Country	Setting	Ethnic groups (%)
Zeger 2011	I: oxandrolone + GH	4	Girls with TS aged 10–15 years at trial entry	1989–1999	USA	University and developmental endocrinology branch	White (not reported in the trial; however, based on location of trial)
	C: GH + placebo
Gault 2011	I: oxandrolone + GH	5	Girls with TS at or after 9 years of age at trial entry	1999–2008	UK	36 paediatric endocrinology departments in UK hospitals	White (not reported in the trial; however, based on location of trial)
	C: GH + placebo
Menke 2010	I1: oxandrolone 0.03 mg/kg/day + GH	6	Girls with TS aged 2–16 years at trial entry	1991–2003	Netherlands	10 paediatric endocrine centres	White (not reported in the trial; however, based on location of trial)
	I2: oxandrolone 0.06 mg/kg/day + GH
	C: GH + placebo
Stahnke 2002	I1: oxandrolone + GH	5	Girls with TS aged 3–16 years at trial entry	1987–1999	Germany	Pediatric department	White (not reported in the trial; however, based on location of trial)
	I2: transient oxandrolone + GH
	C: GH
Rosenfeld 1998	I: oxandrolone alone for the first 12–24 months then continued with GH for the rest of the intervention period	6.1	Girls with TS aged 5–12 years at trial entry	1983–1994	USA	11 medical centres	White (not reported in the trial; however, based on location of trial)
	C: GH	7.6
Job 1991	I: oxandrolone + GH	3	Girls with TS aged ≥ 4 years at trial entry	1986–1989	France	7 paediatric endocrinology clinics	White (not reported in the trial; however, based on location of trial)
	C: GH
C: comparator; GH: growth hormone; I: intervention; TS; Turner syndrome.

Appendix 6. Baseline characteristics (II)

Study ID	Intervention(s) and comparator(s)	Age when therapy was started (mean years (SD), range)	Comedications/co‐interventions	Comorbidities
Zeger 2011	I: oxandrolone + GH	10–14.9	—	—
	C: GH + placebo	10–14.9
Gault 2011	I: oxandrolone + GH	9.0	—	—
	C: GH + placebo	9.0
Menke 2010	I1: oxandrolone 0.03 mg/kg/day + GH	10.2 (2.5)	—	—
	I2: oxandrolone 0.06 mg/kg/day + GH	10.2 (2.2)
	C: GH + placebo	10.9 (2.3)
Stahnke 2002	I1: oxandrolone + GH	10.2 (2.2)	—	—
	I2: transient oxandrolone + GH	‐
	C: GH	10.5 (2.5)
Rosenfeld 1998	I: oxandrolone	9.9 (2.3)	—	—
	C: GH	9.1 (2.1)
Job 1991	I: oxandrolone + GH	11.4 (1.6)	—	—
	C: GH	10.1 (3.1)
—: denotes not reported. C: comparator; GH: growth hormone; I: intervention; SD: standard deviation.

Appendix 7. Matrix of trial endpoints (publications and trial documents)

Zeger 2011	Endpoints quoted in trial document(s) (ClinicalTrials.gov, FDA/EMA document, manufacturer's website, published design paper)a,c
	Source: NCT00004275 (clinicaltrials.gov/ct2/show/NCT00004275)/NCT00029159 (clinicaltrials.gov/ct2/show/NCT00029159) (extension study; during the 2‐year extension participants received oestrogen replacement as ethinylestradiol, 50 ng/kg/day during year 3 and 100 ng/kg/day during year 4) Primary outcome measure(s): NCT00029159 (extension study): cognitive function (time frame: 4 years) Secondary outcome measure(s): — Other outcome measure(s): — Trial results available in trial register: no
	Endpoints quoted in publication(s)b,c
	Primary outcome measure(s): — Secondary outcome measure(s): — Other outcome measure(s): height, weight, waist and hip circumference, breast and pubic hair Tanner staging, hormone measurements, bone age, mid‐parental height, predicted adult height, near‐adult height, bone mineral content, adverse events
	Endpoints quoted in abstract of publication(s)b,c
	Primary outcome measure(s): — Secondary outcome measure(s): — Other outcome measure(s): auxological data, breast and pubic hair Tanner stages, hormone and lipid levels
Gault 2011	Endpoints quoted in trial document(s) (ClinicalTrials.gov, FDA/EMA document, manufacturer's website, published design paper)a,c
	Source: ISRCTN50343149 (apps.who.int/trialsearch/Trial2.aspx?TrialID=ISRCTN50343149) Primary outcome measure(s): final adult height (cm), defined as height velocity < 1 cm/year and bone age ≤ 15.5 years Secondary outcome measure(s): maximum height (the most recently available height), age of attaining final height, 3 summary growth parameters (size, tempo, velocity) Other outcome measure(s): — Trial results available in trial register: no
	Endpoints quoted in publication(s)b,c
	Primary outcome measure(s): final height Secondary outcome measure(s): maximum height (most recently available height), age of attaining final height, 3 summary growth parameters Other outcome measure(s): —
	Endpoints quoted in abstract of publication(s)b,c
	Primary outcome measure(s): final height Secondary outcome measure(s): — Other outcome measure(s): —
Menke 2010	Endpoints quoted in trial document(s) (ClinicalTrials.gov, FDA/EMA document, manufacturer's website, published design paper)a,c
	Source: NTR708 (apps.who.int/trialsearch/Trial2.aspx?TrialID=NTR708)/NTR1934 (apps.who.int/trialsearch/Trial2.aspx?TrialID=NTR1934) (observational follow‐up study) Primary outcome measure(s): clinical, auxological and biochemical parameters; (changes in) characteristics of the voice; psychological parameters (follow‐up study: final adult height) Secondary outcome measure(s): carbohydrate metabolism; abnormalities of liver function; thyroxine binding globulin levels; masculinising effects (follow‐up study: body proportions and composition; metabolic parameters; virilising effects; cardiovascular, liver and thyroid function; (neuro)psychological function; quality of life) Other outcome measure(s): — Trial results available in trial register: no
	Endpoints quoted in publication(s)b,c
	Primary outcome measure(s): adult height gain, defined as adult height minus predicted adult height Secondary outcome measure(s): influence of age group on the effect of oxandrolone; effect of oxandrolone on: short‐term height gain, adult height gain adjusted for bone age at start, safety parameters, pubertal development, bone maturation, duration and costs of GH therapy Other outcome measure(s): adverse events
	Endpoints quoted in abstract of publication(s)b,c
	Primary outcome measure(s): adult height gain (adult height minus predicted adult height) Secondary outcome measure(s): — Other outcome measure(s): safety parameters
Stahnke 2002	Endpoints quoted in trial document(s) (ClinicalTrials.gov, FDA/EMA document, manufacturer's website, published design paper)a,c
	Source: published design paper Primary outcome measure(s): — Secondary outcome measure(s): — Other outcome measure(s): bone age, cortical thickness and bone diameter
	Endpoints quoted in publication(s)b,c
	Primary outcome measure(s): — Secondary outcome measure(s): — Other outcome measure(s): bone age, height, projected adult height, recent height minus projected adult height, glucose levels, HbA1c, lipid profile, liver enzymes
	Endpoints quoted in abstract of publication(s)b,c
	Primary outcome measure(s): — Secondary outcome measure(s): — Other outcome measure(s): height, projected adult height
Rosenfeld 1998	Endpoints quoted in trial document(s) (ClinicalTrials.gov, FDA/EMA document, manufacturer's website, published design paper)a,c
	Source: NT
	Endpoints quoted in publication(s)b,c
	Primary outcome measure(s): — Secondary outcome measure(s): — Other outcome measure(s): height, recent height minus projected adult height, most recent height of each girl relative to each girl's projected adult height, adverse events
	Endpoints quoted in abstract of publication(s)b,c
	Primary outcome measure(s): — Secondary outcome measure(s): — Other outcome measure(s): height, recent height minus projected adult height
Job 1991	Endpoints quoted in trial document(s) (ClinicalTrials.gov, FDA/EMA document, manufacturer's website, published design paper)a,c
	Source: NT
	Endpoints quoted in publication(s)b,c
	Primary outcome measure(s): — Secondary outcome measure(s): — Other outcome measure(s): bone age maturation, annual growth rate, height and weight, predictable adult height, biochemical parameters (triglycerides, cholesterol, HbA1c), anti‐GH antibodies
	Endpoints quoted in abstract of publication(s)b,c
	Primary outcome measure(s): — Secondary outcome measure(s): — Other outcome measure(s): growth rate, growth velocity, predictable adult height, metabolic parameters
—: denotes not reported. aTrial document(s) refers to all available information from published design papers and sources other than regular publications (e.g. FDA/EMA documents, manufacturer's websites, trial registers). bPublication(s) refers to trial information published in scientific journals (primary reference, duplicate publications, companion documents or multiple reports of a primary trial). cPrimary and secondary outcomes refer to verbatim specifications in publication/records. Unspecified outcome measures refer to all outcomes not described as primary or secondary outcome measures. EMA: European Medicines Agency; FDA: Food and Drug Administration (US); HbA1c: glycosylated haemoglobin A1c; NT: no trial document available

Appendix 8. High risk of outcome reporting bias according to ORBIT classification

Study ID	Outcome	High risk of bias (category A)a	High risk of bias (category D)b	High risk of bias (category E)c	High risk of bias (category G)d
Zeger 2011	ND
Gault 2011	ND
Menke 2010	ND
Stahnke 2002	ND
Rosenfeld 1998	ND
Job 1991	Increase in height velocity	Yes	No	No	No
	Adverse events	No	No	No	Yes
aClear that outcome was measured and analysed; trial report stated that outcome was analysed but reported only that result was not significant (Classification 'A', table 2, Kirkham 2010). bClear that outcome was measured and analysed; trial report stated that outcome was analysed but reported no results (Classification 'D', table 2, Kirkham 2010). cClear that outcome was measured but was not necessarily analysed; judgement said likely to have been analysed but not reported because of non‐significant results (Classification 'E', table 2, Kirkham 2010). dUnclear whether outcome was measured; not mentioned, but clinical judgement said likely to have been measured and analysed but not reported on the basis of non‐significant results (Classification 'G', table 2, Kirkham 2010). ND: none detected; ORBIT: Outcome Reporting Bias In Trials.

Appendix 9. Definition of endpoint measurement^a

Study ID	Endpoints	Definition
Zeger 2011	Final adult height	Predicted adult height was determined using the Bayley‐pinneau method. Near‐adult height was defined as height obtained when bone age was > 13.5 years (IO)
	Increase in height velocity	Change in height and height SDS from baseline to year 1, 2, 3 and 4 (IO)
	Health‐related quality of life	NR
	Bone maturation	Greulich and Pyle method (IO)
	Effects on cognition	Cognition: working memory, spatial cognition, executive functions, verbal abilities
	Effects on psychological status	NR
	Effects on speech	NR
	Socioeconomic effects	NR
	Severe/serious hypoglycaemia	Participants were evaluated for adverse effects at each visit (IO; SO)
Gault 2011	Final adult height	Final height, defined as height velocity < 1 cm/year and bone age ≥ 15.5 years. Maximum height (most recently available height). Final height, height SDS, change in height SDS were measured (IO)
	Increase in height velocity	ND
	Health‐related quality of life	NR
	Bone maturation	Tanner Whitehouse II method (IO)
	Effects on cognition	NR
	Effects on psychological status	NR
	Effects on speech	NR
	Socioeconomic effects	NR
	Severe/serious hypoglycaemia	Adverse effects classified as serious adverse events, when they involved admission to hospital for procedures including appendicectomy and ear, nose and throat surgery (IO; SO)
Menke 2010	Final adult height	Adult height measured in cm. Projected adult height according to the Lyon method. Adult height gain as adult height (the last measured height after discontinuing interventions) minus predicted adult height, calculated using the modified projected adult height method (IO)
	Increase in height velocity	ND
	Health‐related quality of life	BDI‐II‐NL; SCL‐90‐R; RAND scales
	Bone maturation	Tanner and Whitehouse radius, ulna, short‐bones score (IO)
	Effects on cognition	RAND; WAIS III‐Verbal IQ and WAIS III‐Performance IQ
	Effects on psychological status	CBCL, Junior DPQ‐J, romantic, sexual interest and mood questionnaires
	Effects on speech	NR
	Socioeconomic effects	NR
	Severe/serious hypoglycaemia	All adverse events reported by the participant, parent or medical doctor were registered (IO; SO)
Stahnke 2002	Final adult height	Projected adult height calculated using the Lyon method. Adult or near adult final height defined as: height velocity at ≤ 0.5 cm/year or bone age > 13.5 or chronological age > 17 years or participant satisfied with height (IO)
	Increase in height velocity	Height velocity of year 1 and 2 (IO)
	Health‐related quality of life	NR
	Bone maturation	Greulich and Pyle method (IO)
	Effects on cognition	NR
	Effects on psychological status	NR
	Effects on speech	NR
	Socioeconomic effects	NR
	Severe/serious hypoglycaemia	(IO; SO)
Rosenfeld 1998	Final adult height	Treatment stopped (final height) when height velocity was < 2.5 cm/year or bone age ≥ 14 years. Final adult height measured in cm (IO)
	Increase in height velocity	Height velocity of year 1–2 (IO)
	Health‐related quality of life	NR
	Bone maturation	Greulich and Pyle method (IO)
	Effects on cognition	NR
	Effects on psychological status	NR
	Effects on speech	NR
	Socioeconomic effects	NR
	Severe/serious hypoglycaemia	(IO; SO)
Job 1991	Final adult height	Predicted adult height was determined using the Bayley‐Pinneau method (IO)
	Increase in height velocity	Growth velocity in year 1, 2 and 3 (IO)
	Health‐related quality of life	NR
	Bone maturation	Greulich and Pyle method (IO)
	Effects on cognition	NR
	Effects on psychological status	NR
	Effects on speech	NR
	Socioeconomic effects	NR
	Severe/serious hypoglycaemia	(IO; SO)
aIn addition to definition of endpoint measurement, description who measured the outcome. AO: adjudicated outcome measurement; BDI‐II‐NL: Beck Depression Inventory – Second Edition Netherlands; CBCL: Child Behavior Check List; DPQ‐J: Junior Dutch Personality Questionnaire; IO: investigator‐assessed outcome measurement; IQ: intelligence quotient; ND: not defined; NR: not reported; RAND: Measure of Health‐Related Quality of Life; SCL‐90‐R: Symptom Checklist‐90‐Revised; SDS: standard deviation score; SO: self‐reported outcome measurement WAIS: Wechsler Adult Intelligence Scale.

Appendix 10. Adverse events (I)

Study ID	Intervention(s) and comparator(s)	Participants included in analysis (N)	Deaths (N)	Deaths (%)	Participants with ≥ 1 adverse event (N)	Participants with ≥ 1 adverse event (%)	Participants with ≥ 1 severe/serious adverse event (N)	Participants with ≥ 1 severe/serious adverse event (%)
Zeger 2011	I: oxandrolone + GH	40	—	—	20	50	10	25
	C: GH + placebo	36	—	—	9	25	6	16.7
Gault 2011	I: oxandrolone + GH	43	—	—	197 (did not specify per group)	—	—	—
	C: GH + placebo	49	—	—		—	—	—
Menke 2010	I1: oxandrolone 0.03 mg/kg/day + GH	46	—	—	6	13	0	0
	I2: oxandrolone 0.06 mg/kg/day + GH	39	—	—	15	38.5	0	0
	C: GH + placebo	48	—	—	2	4.2	0	0
Stahnke 2002	I1: oxandrolone + GH	33	—	—	19	57.6	0	0
	I2: transient oxandrolone + GH	38	—	—	0	0	0	0
	C: GH	20	—	—	0	0	0	0
Rosenfeld 1998	I: oxandrolone	43	—	—	0	0	0	0
	C: GH	17	—	—	0	0	0	0
Job 1991	I: oxandrolone + GH	12	—	—	—	—	—	—
	C: GH	10	—	—	—	—	—	—
—: denotes not reported. C: comparator; GH: growth hormone; I: intervention; N: number of participants.

Appendix 11. Adverse events (II)

Study ID	Intervention(s) and comparator(s)	Participants included in analysis (N)	Participants discontinuing trial due to an adverse event (N)	Participants discontinuing trial due to an adverse event (%)	Participants with ≥ 1 hospitalisation (N)	Participants with ≥ 1 hospitalisation (%)	Participants with ≥ 1 outpatient treatment (N)	Participants with ≥ 1 outpatient treatment (%)
Zeger 2011	I: oxandrolone + GH	40	0	0	10	25	10	25
	C: GH + placebo	36	0	0	6	16.7	3	8.3
Gault 2011	I1: oxandrolone + GH	51	—	—	10 (did not specify per group		—	—
	C1: GH + placebo	55	—	—			—	—
Menke 2010	I1: oxandrolone 0.03 mg/kg/day + GH	46	2	4.5	—	—	—	—
	I2: oxandrolone 0.06 mg/kg/day + GH	39	7	17.5	—	—	—	—
	C: GH + placebo	48	0	0	—	—	—	—
Stahnke 2002	I1: oxandrolone + GH	33	12	27.3	0	0	0	0
	I2: transient oxandrolone + GH	38			0	0	0	0
	C: GH	20	2	4.3	0	0	0	0
Rosenfeld 1998	I: oxandrolone	43	0	0	0	0	0	0
	C: GH	17	0	0	0	0	0	0
Job 1991	I: oxandrolone + GH	12	—	—	—	—	—	—
	C: GH	10	—	—	—	—	—	—
—: denotes not reported. C: comparator; GH: growth hormone; I: intervention; N: number of participants.

Appendix 12. Adverse events (III)

Study ID	Intervention(s) and comparator(s)	Participants included in analysis (N)	Participants with a specific adverse event (description)	Participants with ≥ 1 specific adverse events (N)	Participants with ≥ 1 specific adverse event (%)
Zeger 2011	I: oxandrolone + GH	40	(1) Increased liver enzymes (2) Hypertension (3) Virilisation (subjective clitoris changes) (4) Acne (5) Hirsutism	(1) 3 (2) 1 (3) 2 (4) 1 (5) 3	(1) 7.5 (2) 2.5 (3) 5.5 (4) 2.5 (5) 7.5
	C: GH + placebo	36	(1) Increased liver enzymes (2) Virilisation (subjective clitoris changes)	(1) 1 (2) 2	(1) 2.8 (2) 5.6
Gault 2011	I: oxandrolone + GH	51	(1) Acne (2) Hypertension (3) Abnormal liver function	(1) 1 (2) 1 (3) 1	(1) 2 (2) 2 (3) 2
	C: GH + placebo	55	(1) Hot flushes	(1) 1	(1) 1.8
Menke 2010	I1: oxandrolone 0.03 mg/kg/day + GH	46	(1) Impaired glucose tolerance (2) Hypothyroidism (3) Virilisation (voice deepening) (4) Clitromegaly (5) Hirsutism	(1) 3 (2) 12 (3) 3 (4) 4 (5) 5	(1) 6.5 (2) 26.1 (3) 6.5 (4) 8.7 (5) 10.9
	I2: oxandrolone 0.06 mg/kg/day + GH	39	(1) Impaired glucose tolerance (2) Hypothyroidism (3) Virilisation (voice deepening) (4) Clitromegaly (5) Hirsutism	(1) 2 (2) 26 (3) 9 (4) 12 (5) 5	(1) 5.1 (2) 66.7 (3) 23.1 (4) 30.8 (5) 12.8
	C: GH + placebo	48	(1) Elevated liver enzymes (2) Hirsutism (3) Voice deepening	(1) 3 (2) 1 (3) 1	(1) 6.3 (2) 2.1 (3) 2.1
Stahnke 2002	I1: oxandrolone + GH	33	(1) Enlargement of clitoris (2) Carbohydrate intolerance (3) Voice deepening	(1) 19 (2) 1 (3) 7	(1) 26.8 (2) 1.4 (3) 9.9
	I2: transient oxandrolone + GH	38
	C: GH	20	(1) Carbohydrate intolerance	(1) 2	(1) 10
Rosenfeld 1998	I: oxandrolone	43	(1) Aortic stenosis with cerebrovascular accident (2) Slipped femoral epiphysis	(1) 1 (2) 1	(1) 1.7 (2) 1.7
	C: GH	17	—	—	—
Job 1991	I: oxandrolone + GH	12	—	—	—
	C: GH	10	—	—	—
—: denotes not reported. C: comparator; GH: growth hormone; I: intervention; N: number of participants.

Appendix 13. Health‐related quality of life: instruments

Study ID	Instrument	Dimensions (subscales) (number of items)	Validated instrument	Answer options	Scores	Minimum score Maximum score	Weighting of scores	Direction of scales	Minimal important difference
Menke 2010 (follow‐up study)	SF‐36 (G)	Physical functioning (PF) (10) Role‐physical (RP) (4) Bodily pain (BP) (2) General health (GH) (5) Vitality (VT) (4) Social functioning (SF) (2) role‐emotional (RE) (3) Mental health (MH) (5) Reported health transition (RHT) (1)	Yes	3, 5 and 6‐point Likert‐scale	Scores for dimensions Physical Component Summary (PCS) Mental Component Summary (MCS)	Minimum scores: scores for dimensions/PCS/MCS: norm‐based scale Maximum scores: scores for dimensions/PCS/MCS: norm‐based scale	No	Higher values mean better assessment	PCS: 2–3 points MCS: 3 points Dimensions: PF/BT/VT: 2 points, if score < 40; 3 points, if score ≥ 40 RP: 2 points SF/MH: 3 points RE: 4 points
	RAND 36	Physical functioning, social functioning, limitations due to physical problems, limitations due to emotional problems, mental health, vitality, bodily pain and general health	Yes	0–100	For all subscales	0 100	Yes	Higher values mean better assessment	0.010–0.048
	SCL‐90‐R	General psychological, somatic and cognitive well being	Yes	5‐point scale	8 subscales are defined as agoraphobia, somatisation, anger‐hostility, depression, interpersonal sensitivity, paranoid ideation, anxiety, cognitive performance difficulty and sleep disturbance	ND	ND	ND	ND
	BDI‐II‐NL	Cognitive, somatic and affective	Yes	0–3	21 items divided into 3 categories: cognitive, somatic and affective	0 63	No	Higher values mean worse assessment	16
BDI‐II‐NL: Beck Depression Inventory – Second Edition Netherlands; ND: not defined; RAND: Measure of Health‐Related Quality of Life; SCL‐90‐R: Symptom Checklist‐90‐Revised; SF‐36: 36‐item Short‐Form health survey.

Appendix 14. Survey of trial investigators providing information on included trials

Study ID	Date trial author contacted	Date trial author replied	Trial author asked for additional information (short summary)	Trial author provided data (short summary)
Zeger 2011	20 March 2014	No answer	NA	NA
Gault 2011	20 March 2014	20 March 2014	No	NA
Menke 2010	20 March 2014	No answer	NA	NA
Stahnke 2002	20 March 2014	23 March 2014	No	NA
Rosenfeld 1998	20 March 2014	Deceased	NA	NA
Job 1991	20 March 2014	20 March 2014	No	NA
NA: not applicable.

Appendix 15. Checklist to aid consistency and reproducibility of GRADE assessments

Item		(1) Final adult height	(2) Adverse events	(3) Health‐related quality of life	(4) Effects on speech	(5) Effects on cognition	(6) Effects on psychological status	(7) All‐cause mortality
Trial limitations (risk of bias)a	Was random sequence generation used (i.e. no potential for selection bias)?	Unclear	Unclear	Not reported	NA	NA	NA	Not reported
	Was allocation concealment used (i.e. no potential for selection bias)?	Unclear	Unclear
	Was there blinding of participants and personnel (i.e. no potential for performance bias) or outcome not likely to be influenced by lack of blinding?	Yes	Yes
	Was there blinding of outcome assessment (i.e. no potential for detection bias) or was outcome measurement not likely to be influenced by lack of blinding?	Yes	Yes
	Was an objective outcome used?	Yes	Yes
	Were > 80% of participants enrolled in trials included in the analysis (i.e. no potential reporting bias)?b	Unclear	Unclear
	Were data reported consistently for the outcome of interest (i.e. no potential selective reporting)?	Yes	Yes
	No other biases reported (i.e. no potential of other bias)?	Yes	Yes
	Did the trials end up as scheduled (i.e. not stopped early)?	Yes	Yes
Inconsistencyc	Point estimates did not vary widely?	Yes	Yes
	To what extent did confidence intervals overlap (substantial: all confidence intervals overlap ≥ 1 of the included studies point estimate; some: confidence intervals overlap but not all overlap ≥ 1 point estimate; no: ≥ 1 outlier: where the confidence interval of some of the studies do not overlap with those of most included studies)?	Substantial	Some
	Was the direction of effect consistent?	Yes	Yes
	What was the magnitude of statistical heterogeneity (as measured by I²) – low (I² < 40%), moderate (I² 40–60%), high I² > 60%)?	Low	High
	Was the test for heterogeneity statistically significant (P < 0.1)?	Not statistically significant	Yes statistically significant
Indirectness	Were the populations in included studies applicable to the decision context?	Highly applicable	Highly applicable
	Were the interventions in the included studies applicable to the decision context?	Highly applicable	Highly applicable
	Was the included outcome not a surrogate outcome?	Yes	Yes
	Was the outcome timeframe sufficient?	Sufficient	Sufficient
	Were the conclusions based on direct comparisons?	Yes	Yes
Imprecisiond	Was the confidence interval for the pooled estimate not consistent with benefit and harm?	Yes	Yes
	What is the magnitude of the median sample size (high: > 300 participants, intermediate: 100–300 participants, low: < 100 participants)?e	Low (↓)	Low (↓)
	What was the magnitude of the number of included studies (large: > 10 studies, moderate: 5–10 studies, small: < 5 studies)?b	Moderate	Small (↓)
	Was the outcome a common event (e.g. occurs > 1/100)?	NA	Yes
Publication biasde	Was a comprehensive search conducted?	Yes	Yes
	Was grey literature searched?	Yes	Yes
	Were no restrictions applied to study selection on the basis of language?	Yes	Yes
	There was no industry influence on studies included in the review?	Yes	Yes
	There was no evidence of funnel plot asymmetry?	Unclear	Unclear
	There was no discrepancy in findings between published and unpublished trials?	Unclear	Unclear
aQuestions on risk of bias are answered in relation to the majority of the aggregated evidence in the meta‐analysis rather than to individual trials. bDepends on the context of the systematic review area. cQuestions on inconsistency are primarily based on visual assessment of forest plots and the statistical quantification of heterogeneity based on I² statistic. dWhen judging the width of the confidence interval it is recommended to use a clinical decision threshold to assess whether the imprecision is clinically meaningful. eQuestions address comprehensiveness of the search strategy, industry influence, funnel plot asymmetry and discrepancies between published and unpublished trials. (↓): key item for potential downgrading the certainty of the evidence (GRADE) as shown in the footnotes of the 'Summary of finding' table; NA: not applicable.

Data and analyses

Comparison 1. Oxandrolone plus growth hormone (GH) versus GH only.

Outcome or subgroup title	No. of studies	No. of participants	Statistical method	Effect size
1 Final adult height	5	270	Mean Difference (IV, Random, 95% CI)	2.71 [1.32, 4.11]
2 Final adult height: excluding low‐quality trials	3	229	Mean Difference (IV, Random, 95% CI)	2.87 [‐0.56, 6.30]
3 Final adult height: trials reporting strictly adult height (excluding trials measuring near adult height and predicted height)	3	226	Mean Difference (IV, Random, 95% CI)	2.54 [0.31, 4.78]
4 Final adult height: trials reporting strictly adult height (excluding low‐quality trials)	2	166	Mean Difference (IV, Random, 95% CI)	2.87 [‐0.56, 6.30]
5 Final height/near adult height reported	2		Mean Difference (IV, Random, 95% CI)	Subtotals only
6 Predictable adult height	2	44	Mean Difference (IV, Random, 95% CI)	0.29 [‐2.78, 3.36]
7 Maximum height	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
8 Adult height gain	4	246	Mean Difference (IV, Random, 95% CI)	3.15 [1.93, 4.37]
9 Final adult height SDS (Turner reference)	2	106	Mean Difference (IV, Random, 95% CI)	0.10 [‐0.31, 0.51]
10 Final adult height SDS (healthy reference)	2	166	Mean Difference (IV, Random, 95% CI)	‐0.30 [‐1.28, 0.68]
11 Final adult height delta height SDS (Turner reference)	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
12 Final adult height delta height SDS (healthy reference)	2	166	Mean Difference (IV, Random, 95% CI)	0.54 [0.24, 0.83]
13 Final adult height minus predicted adult height (height gain)	2	106	Mean Difference (IV, Random, 95% CI)	3.04 [1.15, 4.94]
14 Difference in height gain vs placebo (final height – placebo final height)	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
15 Near adult height	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
16 Near adult height SDS (Turner reference)	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
17 Near adult height SDS (healthy reference)	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
18 Near adult height delta height SDS (Turner reference)	1		Mean Difference (IV, Random, 95% CI)	Totals not selected
19 Near adult height delta height SDS (healthy reference)	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
20 Increase in near adult height from baseline	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
21 Near adult height minus predicted adult height	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
22 Difference in height gain over placebo (near adult height – placebo final height)	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
23 Health‐related quality of life (BDI‐II‐NL)	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
24 Health‐related quality of life (SCL‐90‐R)	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
25 Health‐related quality of life (SF‐36)	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
26 Adverse events	4	283	Risk Ratio (M‐H, Random, 95% CI)	3.27 [0.75, 14.22]
27 Height growth (velocity) year 1	2		Mean Difference (IV, Random, 95% CI)	Totals not selected
28 Height growth (velocity) SDS year 1 (Turner reference)	2	58	Mean Difference (IV, Random, 95% CI)	1.52 [0.72, 2.33]
29 Height velocity year 2	3		Mean Difference (IV, Random, 95% CI)	Totals not selected
30 Height velocity, SDS year 2 (Turner reference)	2		Mean Difference (IV, Random, 95% CI)	Totals not selected
31 Height velocity, SDS year 2 (healthy reference)	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
32 Height velocity year 3	2		Mean Difference (IV, Random, 95% CI)	Totals not selected
33 Height velocity SDS year 3 (Turner reference)	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
34 Height velocity year 4	2		Mean Difference (IV, Random, 95% CI)	Totals not selected
35 Height velocity SDS year 4 (Turner reference)	2		Mean Difference (IV, Random, 95% CI)	Totals not selected
36 Bone maturation, bone age in years	3	91	Mean Difference (IV, Random, 95% CI)	0.16 [‐0.46, 0.78]
37 Effect on cognition at year 2	1		Mean Difference (IV, Fixed, 95% CI)	Totals not selected
37.1 Working memory	1		Mean Difference (IV, Fixed, 95% CI)	0.0 [0.0, 0.0]
37.2 Spatial cognition	1		Mean Difference (IV, Fixed, 95% CI)	0.0 [0.0, 0.0]
37.3 Executive function	1		Mean Difference (IV, Fixed, 95% CI)	0.0 [0.0, 0.0]
37.4 Verbal abilities	1		Mean Difference (IV, Fixed, 95% CI)	0.0 [0.0, 0.0]

Characteristics of studies

Characteristics of included studies [ordered by study ID]

Gault 2011.

Methods	Design: parallel, factorial, randomised controlled trial
Participants	Inclusion criteria: TS confirmed by karyotyping (all karyotypes were eligible including mosaic), aged 7–13 years, no previous GH treatment, or previous GH treatment of 8.3–11.7 mg/m²/week in 5–7 injections a week, no previous oxandrolone and oestrogen therapy, and open epiphyses Exclusion criteria: major systemic illness that might affect growth and social or psychological difficulties likely to seriously impair concordance Diagnostic criteria: karyotyping Setting: paediatric endocrinology departments in UK hospitals Age group: children/adolescents Gender distribution: females Country where trial was performed: UK
Interventions	Intervention: oxandrolone 0.05 mg/kg/day PO from 9 years of age + GH 10 mg/m²/week, daily SC Comparator: placebo from 9 years of age + GH 10 mg/m²/week, daily SC Duration of intervention: 5 years Number of study centres: 36 paediatric endocrinology departments in UK hospitals Treatment before study: no previous GH treatment or previous GH treatment of 8.3–11.7 mg/m²/week in 5–7 injections a week
Outcomes	Composite outcome measures reported: no
Study details	Trial identifier: ISRCTN50343149 Trial terminated early: yes, due to unavailability of oxandrolone because the sole European manufacturer of oxandrolone stopped production in 2004 and 2008
Publication details	Language of publication: English Funding: non‐commercial funding was provided by Scottish Executive Chief Scientist Office (1999–2004) and thereafter by the British Society of Paediatric Endocrinology and Diabetes, with a contribution to funding of pharmacy staff from the Child Growth Foundation Publication status: peer‐reviewed journal/full article
Stated aim for study	Quote: "to examine the effect of oxandrolone and the timing of pubertal induction on final height in girls with Turner's syndrome receiving a standard dose of growth hormone."
Notes	2 × 2 factorial design: girls with evidence of ovarian failure at 12 years were further randomised to ethinylestradiol PO (year 1: 2 μg daily, year 2: 4 μg daily, year 3: 4 months each of 6, 8 and 10 μg daily) or placebo. The placebo group subsequently started ethinylestradiol at 14 years. The code for randomisation 2 was broken at 15 years to allow introduction of progesterone therapy: norethisterone 5 mg PO daily for 5 days each month.
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	Low risk	Quote: methods: randomisation and masking: "the British Society for Paediatric Endocrinology (BSPE) and Diabetes Clinical Trial Unit (DCTU) (Cambridge, UK) used minimization by weighted randomisation for the study's two randomizations." (page 2) Comment: no selection bias
Allocation concealment (selection bias)	Low risk	Quote: methods: treatment: "white coated, flat bevelled edge placebo tablets (7 and 5 mm in diameter) were specially manufactured to match the size, weight and appearance of oxandrolone 2.5 mg and ethynylestradiol 2 mcg tablets respectively. The study's central pharmacy supplied all tablets in a double blind, placebo‐controlled fashion; only staff at the BSPE, DCTU and the central distributing pharmacy were not blinded to treatment allocations." (page 3) Comment: no allocation bias.
Blinding of participants and personnel (performance bias) adverse events	Low risk	Quote:abstract: design: "randomised, double blind, placebo‐controlled trial." Comment: investigator‐assessed, both doctors and participants were blinded.
Blinding of participants and personnel (performance bias) bone maturation	Low risk	Quote: methods: study procedures: "radiographs of the left wrist were obtained annually, and a single observer (WFP) analysed them for bone age estimation according to the Tanner Whitehouse II method. Biochemical analyses followed local protocols, and the results were reported centrally." (page 3) Comment: investigator‐assessed, blinding was documented for bone maturation.
Blinding of participants and personnel (performance bias) improvement in final adult height/increase in height velocity	Low risk	Quote: methods: study procedures: "at clinic visits every four to six months, height was measured with a Harpenden stadiometer to the last completed 1 mm and converted to an SD score by using the British 1990 reference. Radiographs of the left wrist were obtained annually, and a single observer (WFP) analysed them for bone age estimation according to the Tanner Whitehouse II method. Biochemical analyses followed local protocols, and the results were reported centrally." (page 3) Comment: investigator‐assessed. Blinding was documented for height velocity.
Blinding of outcome assessment (detection bias) adverse events	Low risk	Quote: abstract: design: "randomized, double blind, placebo‐controlled trial;" Methods: Study procedures: "standardised pharmacovigilance procedures were followed, with recording of adverse events or reactions at each visit and subsequent review by the Data and Safety Monitoring Group. Data collection ceased if participants withdrew consent, although we included previously collected data in the analysis." (page 3) Comment: investigator‐assessed. Both doctors and participants were blinded.
Blinding of outcome assessment (detection bias) bone maturation	Low risk	Quote: methods: study procedures: "at clinic visits every four to six months, height was measured with a Harpenden stadiometer to the last completed 1 mm and converted to an SD score by using the British 1990 reference. Radiographs of the left wrist were obtained annually, and a single observer (WFP) analysed them for bone age estimation according to the Tanner Whitehouse II method. Biochemical analyses followed local protocols, and the results were reported centrally." (page 3) Comment: investigator‐assessed. Blinding was documented for bone maturation.
Blinding of outcome assessment (detection bias) improvement in final adult height/increase in height velocity	Low risk	Quote: methods: study procedures: "at clinic visits every four to six months, height was measured with a Harpenden stadiometer to the last completed 1 mm and converted to an SD score by using the British 1990 reference. Radiographs of the left wrist were obtained annually, and a single observer (WFP) analysed them for bone age estimation according to the Tanner Whitehouse II method. Biochemical analyses followed local protocols, and the results were reported centrally." (page 3) Comment: investigator‐assessed. Blinding was documented for height velocity.
Incomplete outcome data (attrition bias) adverse events	Unclear risk	Quote: "14 withdrew (4 of them because of early organizational errors)." Comment: no intention‐to‐treat analysis done.
Incomplete outcome data (attrition bias) bone maturation	Unclear risk	Quote: "14 withdrew (4 of them because of early organizational errors)." Comment: no intention‐to‐treat analysis done
Incomplete outcome data (attrition bias) improvement in final adult height/increase in height velocity	Unclear risk	Quote: "14 withdrew (4 of them because of early organizational errors)." Comment: no intention‐to‐treat analysis done
Selective reporting (reporting bias)	Low risk	Comment: no inconsistencies between protocol and publication
Other bias	Low risk	Comment: none detected

Job 1991.

Methods	Design: parallel, randomised controlled trial
Participants	Inclusion criteria: individual informed written consent, chronological age at least 4 years, bone age ≤ 11 years according to Greulich and Pyle, height ≤ –2 SD, growth velocity < –1 SD according to French usual charts, Turner phenotype without abnormality of external genitalia or associated malformations or diseases Exclusion criteria: lack (or well‐defined abnormality) of 1 X chromosome (karyotyping), no steroid or other hormonal treatment during the past years Diagnostic criteria: karyotyping Setting: paediatric endocrinology clinics Age group: children/adolescents Gender distribution: females Country where trial was performed: France
Interventions	Intervention: oxandrolone 0.06 mg/kg/day, PO, + GH 0.01 U/kg, SC at night, OD Comparator: GH 0.01 U/kg daily, SC at night Duration of intervention: 3 years Number of study centres: 7 paediatric endocrine clinics Treatment before study: none
Outcomes	Annual growth rate, height and weight, bone maturation and predictable adult height Composite outcome measures reported: no
Study details	Trial identifier: not reported Trial terminated early: no
Publication details	Language of publication: English Funding: funding not documented Publication status: peer‐reviewed journal/full article
Stated aim for study	Quote: "however, very few results after 2–3 years of treatment are presently known, apart from those of the American multicenter study where the schedule of treatment has been changed for certain groups of patients along time and the bone age changed are not mentioned." (page 229) "This choice relates to the aim of the work, which is not to compare the data with those from untreated Turner syndrome, but to give some insight in the way allowing the Turner girls to feel closer to the normal." (page 231) Comment: the aim was not clearly stated but it could be understood and extracted from the publication, that trial authors investigated the short‐term effects of GH with or without oxandrolone on height, velocity and predicted adult height.
Notes
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	Unclear risk	Quote: patients and methods: "the patients were randomised in two groups: A, to be treated with recombinant human Growth Hormone (hGH) 191 aa (Genotonorm), 0.1 IU/kg daily, injected subcutaneously at night, and B, to receive the same daily injection of Genotonorm plus 0.06 mg/kg/day of oxandrolone per os." (page 229) Comment: method of random sequence generation not clearly described.
Allocation concealment (selection bias)	Unclear risk	Quote: patients and methods: "the patients were randomised in two groups: A, to be treated with recombinant human Growth Hormone (hGH) (Genotonorm), 0.1 IU/kg daily, injected subcutaneously at night, and B, to receive the same daily injection of Genotonorm plus 0.06 mg/kg/day of oxandrolone per os." (page 229) Comment: method of concealment not clearly described.
Blinding of participants and personnel (performance bias) adverse events	Unclear risk	Comment: investigator‐assessed. The blinding of participants was not clearly documented.
Blinding of participants and personnel (performance bias) bone maturation	Low risk	Quote: "evaluation of bone age according to Greulich and Pyne was done centrally by two skilled paediatricians in a blind fashion." (page 230) Comment: investigator‐assessed. The blinding of participants was not clearly documented. Outcome unlikely influenced by lack of participant blinding.
Blinding of participants and personnel (performance bias) improvement in final adult height/increase in height velocity	Unclear risk	Quote: "evaluation of bone age according to Greulich and Pyne was done centrally by two skilled paediatricians in a blind fashion." (page 230). Also, Table (2) Effects of treatment on growth, and Figure (1) Total height gain (cm/3 years) plotted versus age at the onset of treatment. Comment: investigator‐assessed.
Blinding of outcome assessment (detection bias) adverse events	Unclear risk	Comment: not reported.
Blinding of outcome assessment (detection bias) bone maturation	Low risk	Quote: "evaluation of bone age according to Greulich and Pyne was done centrally by two skilled paediatricians in a blind fashion." (page 230). Also, Table (2) Effects of treatment on growth, and Figure (1) Total height gain (cm/3 years) plotted versus age at the onset of treatment. Comment: investigator‐assessed.
Blinding of outcome assessment (detection bias) improvement in final adult height/increase in height velocity	Low risk	Quote: "blinding was done centrally by two paediatricians to evaluate bone age but blinding of participants was not clearly documented," and table (2) and figure (1). Comment: investigator‐assessed; outcome unlikely influenced by potential lack of blinding.
Incomplete outcome data (attrition bias) adverse events	Low risk	Quote: "table 3" Comment: no missing data.
Incomplete outcome data (attrition bias) bone maturation	Low risk	Quote: "table 2" Comment: no missing data.
Incomplete outcome data (attrition bias) improvement in final adult height/increase in height velocity	Low risk	Quote: "table 2 and Fig 1" Comment: no missing data.
Selective reporting (reporting bias)	High risk	Comment: evidence of outcome reporting bias for adverse events and increase in height velocity (see Appendix 8).
Other bias	Low risk	Comment: none detected.

Menke 2010.

Methods	Design: parallel, randomised controlled trial
Participants	Inclusion criteria: karyotype associated with TS (except for cytogenetical evidence of Y chromosomal material), calendar age 2–15.99 years, and bone age < 12.00 years Exclusion criteria: bone age > 12 years; Y chromosomal material; growth failure due to other causes; use of drugs that could interfere with growth; and previous GH, sex hormone or androgen therapy; and refusal to participate Diagnostic criteria: karyotyping Setting: paediatric endocrine centres Age group: children/adolescents Gender distribution: females Country where trial was performed: Netherlands
Interventions	Intervention 1: oxandrolone 0.03 mg/kg/day + GH Intervention 2: oxandrolone 0.06 mg/kg/day + GH Comparator: GH + placebo Duration of intervention: 6 years Run‐in period: 11 years and 6 months Number of study centres: 10 paediatric endocrine centres in the Netherlands Treatment before study: none
Outcomes	Primary outcome: adult height gain = adult height – predicted height Composite outcome measures reported: no
Study details	Trial identifier: not reported Trial terminated early: no
Publication details	Language of publication: English Funding: supported by commercial funding by Pfizer and Eli Lilly Publication status: peer‐reviewed journal//full article
Stated aim for study	Quote(article 2010 (# i): abstract): "in a randomised, placebo‐controlled, double‐blind study, we assessed the effect of oxandrolone at a dosage of either 0.06 or 0.03 mg/kg/day on glucose metabolism in 133 Growth Hormone (GH)‐treated girls with TS." (page 115)
Notes
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	Low risk	Quote: article 2010 (# i): materials and methods: "after stratification for calendar age and height standard deviation (SD) score (SDS), they were randomised by a computer‐generated schedule with a block size of 6 and blindly assigned to receive, orally at bedtime after reaching the age of 8 years, oxandrolone 0.03 mg/kg/d, oxandrolone 0.06 mg/kg/d, or a similar‐appearing placebo (Pl)." (page 1152) Comment: random sequence generation documented.
Allocation concealment (selection bias)	Low risk	Quote:article 2010 (# i): participants and methods: treatment: "the capsules were manufactured and distributed by one hospital pharmacy. All patients and doctors were blinded for the allocation of the patients and will remain so until the last patient will finish the study." (page 1152) Comment: allocation concealment clearly documented.
Blinding of participants and personnel (performance bias) adverse events	Low risk	Quote: article 2010 (# i): participants and methods:treatment: "the capsules were manufactured and distributed by one hospital pharmacy. All patients and doctors were blinded for the allocation of the patients and will remain so until the last patient will finish the study." (page 1152) "Only the independent pharmacist, the statistician, and data analyst saw unblended data, but none of them had any contact with the participants." Comment: investigator‐assessed. Participants and doctors were blinded.
Blinding of participants and personnel (performance bias) bone maturation	Low risk	Quote: article 2010 (# i): participants and methods: assessments: "to assess bone maturation (Delta‐bone age/ Delta calendar age), one trained, and up until 2008, blinded investigator determined bone age of the yearly made hand x‐rays retrospectively and chronologically according to the Tanner and Whitehouse radius, ulna, short‐bones score." (page 1153) Comment: investigator‐assessed. Participants and doctors were blinded.
Blinding of participants and personnel (performance bias) effects on cognition/psychological status/speech	Low risk	Quote: article 2010 (# i): participants and methods: treatment: "the capsules were manufactured and distributed by one hospital pharmacy. All patients and doctors were blinded for the allocation of the patients and will remain so until the last patient will finish the study. Only the independent pharmacist, the statistician, and data analyst saw unblended data, but none of them had any contact with the participants." (page 1152) Comment: investigator‐assessed. Participants and doctors were blinded.
Blinding of participants and personnel (performance bias) health‐related quality of life	Low risk	Quote:article 2010 (# i): participants and methods: treatment: "the capsules were manufactured and distributed by one hospital pharmacy. All patients and doctors were blinded for the allocation of the patients and will remain so until the last patient will finish the study. Only the independent pharmacist, the statistician, and data analyst saw unblended data, but none of them had any contact with the participants." (page 1152) Comment: investigator‐assessed. Participants and doctors were blinded.
Blinding of participants and personnel (performance bias) improvement in final adult height/increase in height velocity	Low risk	Quote: article 2010 (# i): participants and methods:treatment: "the capsules were manufactured and distributed by one hospital pharmacy. All participants and doctors were blinded for the allocation of the patients and will remain so until the last patient will finish the study." (page 1152) Quote: article 2010 (# i): participants and methods: assessments: "two trained observers performed all half‐yearly measurements during the total study period." Comment: patients and doctors were blinded.
Blinding of participants and personnel (performance bias) socioeconomic effects	Low risk	Quote: article 2010 (# i): participants and methods:treatment: "the capsules were manufactured and distributed by one hospital pharmacy. All patients and doctors were blinded for the allocation of the patients and will remain so until the last patient will finish the study." (page 1152) Quote: article 2010 (# i): patients and methods: assessments: "secondary outcomes included the influence of age group on the effect of oxandrolone, and the effect of oxandrolone on: short‐term height gain, adult height gain adjusted for bone age at start, safety parameters, pubertal development, bone maturation, and duration and costs of GH therapy." (page 1153) Comment: investigator‐assessed. The cumulative cost of GH therapy was calculated on page 1153 but the impact on the social status was not elaborated.
Blinding of outcome assessment (detection bias) adverse events	Low risk	Quote:article 2010 (# i) participants and methods: treatment: "the capsules were manufactured and distributed by one hospital pharmacy. All patients and doctors were blinded for the allocation of the patients and will remain so until the last patient will finish the study." (page 1152) "Only the independent pharmacist, the statistician, and data analyst saw unblended data, but none of them had any contact with the participants." Comment: investigator‐assessed. Participants and doctors were blinded.
Blinding of outcome assessment (detection bias) bone maturation	Low risk	Quote: article 2010 (# i): participants and methods: assessments: "to assess bone maturation (Delta‐bone age/ Delta calendar age), one trained, and up until 2008, blinded investigator determined bone age of the yearly made hand x‐rays retrospectively and chronologically according to the Tanner and Whitehouse radius, ulna, short‐bones score." (page 1153) Comment: investigator‐assessed. Participants and doctors were blinded.
Blinding of outcome assessment (detection bias) effects on cognition/psychological status/speech	Low risk	Quote: article 2010 (# i): methods: gender role behavior and preferences: "the favourite profession was scored as "masculine" vs. "neutral or feminine" by one blinded observer. To test inter‐observer agreement, the same was done by a second observer." (page 299) Quote: article 2010 (# i): participants and methods:treatment: "the capsules were manufactured and distributed by one hospital pharmacy. All patients and doctors were blinded for the allocation of the patients and will remain so until the last patient will finish the study." (page 1152) Comment: investigator‐assessed. Participants and doctors were blinded.
Blinding of outcome assessment (detection bias) health‐related quality of life	Low risk	Quote: article 2015: methods: participants and previous treatment: "for the current study all patients and investigators remained blinded for the study medication." (page 60) Comment: investigator‐assessed. Participants and doctors were blinded.
Blinding of outcome assessment (detection bias) improvement in final adult height/increase in height velocity	Low risk	Quote: article 2010 (# i): participants and methods:treatment: "the capsules were manufactured and distributed by one hospital pharmacy. All patients and doctors were blinded for the allocation of the patients and will remain so until the last patient will finish the study." (page 1152) Quote: article 2010 (# i): participants and methods: assessments: "two trained observers performed all half‐yearly measurements during the total study period." Comment: investigator‐assessed. Participants and doctors were blinded.
Blinding of outcome assessment (detection bias) socioeconomic effects	Low risk	Quote: article 2010 (# i): participants and methods:treatment: "the capsules were manufactured and distributed by one hospital pharmacy. All patients and doctors were blinded for the allocation of the participants and will remain so until the last patient will finish the study." (page 1152) Quote: article (# i) 2010: patients and methods: assessments: "secondary outcomes included the influence of age group on the effect of oxandrolone, and the effect of oxandrolone on: short‐term height gain, adult height gain adjusted for bone age at start, safety parameters, pubertal development, bone maturation, and duration and costs of GH therapy." (page 1153) Comment: investigator‐assessed. Participants and doctors were blinded.
Incomplete outcome data (attrition bias) adverse events	Low risk	Quote: article 2010 (# i): statistical analysis: "intention‐to‐treat analyses (ITT) were performed." (page 1154) "All adverse events reported by the patient, parent, or medical doctor were registered. Virilizing adverse events included voice deepening, clitoral enlargement, or an increase in body hair, all being previously identified as oxandrolone‐related virilization." (page 1153)
Incomplete outcome data (attrition bias) bone maturation	Low risk	Quote: article 2010 (# i): statistical analysis: "intention‐to‐treat analyses (ITT) were performed." (page 1154)
Incomplete outcome data (attrition bias) effects on cognition/psychological status/speech	Low risk	Quote:article 2010 (# i): statistical analysis: "intention‐to‐treat analyses (ITT) were performed." (page 1154)
Incomplete outcome data (attrition bias) health‐related quality of life	Low risk
Incomplete outcome data (attrition bias) improvement in final adult height/increase in height velocity	Low risk	Quote: article 2010 (# i): statistical analysis: "intention‐to‐treat analyses (ITT) were performed." (page 1154)
Incomplete outcome data (attrition bias) socioeconomic effects	Low risk	Quote: article 2010 (# i): statistical analysis: "intention‐to‐treat analyses (ITT) were performed." (page 1154)
Selective reporting (reporting bias)	Low risk	Comment: none detected.
Other bias	Low risk	Comment: none detected.

Rosenfeld 1998.

Methods	Design: parallel, randomised controlled trial (after 1–2 years controlled clinical trial)
Participants	Inclusion criteria: TS karyotype 45X, structurally abnormal X chromosome or mosaicism chronological age = 4.7–12.4 years, maximal bone age 1.2 years Exclusion criteria: any demonstrable Y chromosome material, no history of diabetes or endocrine disease, except for hypothyroidism treated with appropriate replacement therapy Diagnostic criteria: karyotyping Setting: medical centres Age group: children/adolescents Gender distribution: females Country where trial was performed: USA
Interventions	Intervention: oxandrolone alone for the first 12–24 months then continued with GH for the rest of the intervention period Comparator(s): GH Duration of intervention: 6.1 years Number of study centres: 11 medical centres Treatment before study: none
Outcomes	Composite outcome measures reported: no
Study details	Trial identifier: not reported Trial terminated early: no
Publication details	Language of publication: English Funding: supported in part by Genentech Inc. (financial and editorial) Publication status: peer‐reviewed journal/full article/other
Stated aim for study	Quote: "to carry out a multicenter, prospective, randomised trial of human growth hormone, alone or in combination with oxandrolone in patients with Turner syndrome."
Notes
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	Unclear risk	Quote:article 1998: methods: "on completion of the observation period subjects were randomly assigned to one of the four study groups." (page 320) Comment: method of random sequence generations not clearly mentioned.
Allocation concealment (selection bias)	Unclear risk	Quote: article 1998: methods: "on completion of the observation period subjects were randomly assigned to one of the four study groups", "subjects receiving growth Hormone alone (group 3) continued to receive this treatment whereas all subjects (groups 1, 2, and 4) received a combination of growth hormone plus oxandrolone, although the latter at a reduced dose of 0.0625 mg/kg/day." (page 320) Comment: no details on allocation concealment.
Blinding of participants and personnel (performance bias) adverse events	High risk	Comment: investigator‐assessed and patient‐reported outcome measure; participants and doctors were not blinded; outcome measure likely influenced by lack of blinding.
Blinding of participants and personnel (performance bias) bone maturation	Low risk	Quote:article 1998: "bone ages were analysed in a blinded manner at the Fels Institute, Yellow Springs, Ohio, according to the method of Greulich and Pyle." (page 320) Comment: investigator‐assessed; outcome measure unlikely influence by lack of blinding.
Blinding of participants and personnel (performance bias) improvement in final adult height/increase in height velocity	Low risk	Comment: investigator‐assessed, no blinding; outcome measure unlikely influenced by lack of blinding.
Blinding of outcome assessment (detection bias) adverse events	High risk	Comment: investigator‐assessed and patient‐reported, participants and doctors were not blinded; outcome measure likely influenced by lack of blinding.
Blinding of outcome assessment (detection bias) bone maturation	Low risk	Quote: article 1998: "bone ages were analysed in a blinded manner at the Fels Institute, Yellow Springs, Ohio, according to the method of Greulich and Pyle." (page 320) Comment: investigator‐assessed.
Blinding of outcome assessment (detection bias) improvement in final adult height/increase in height velocity	Low risk	Comment: investigator‐assessed, participants and doctors were not blinded; outcome measure unlikely to be influenced by lack of blinding.
Incomplete outcome data (attrition bias) adverse events	Unclear risk	Comment: missing data were not clearly reported, intention‐to‐treat analysis not done.
Incomplete outcome data (attrition bias) bone maturation	Low risk	Comment: missing data were reported with reasons.
Incomplete outcome data (attrition bias) improvement in final adult height/increase in height velocity	Unclear risk	Quote:article 1998: "of the 70 subjects enrolled in the study between 1983 and 1984, 65 completed the first 3 years of the protocol. Three subjects were subsequently discontinued from the study because of their poor compliance." (page 320) Comment: missing data of dropouts were not reported and intention‐to‐treat analysis was not done.
Selective reporting (reporting bias)	Low risk	Comment: none detected.
Other bias	Low risk	Comment: none detected.

Stahnke 2002.

Methods	Design: parallel, randomised controlled trial
Participants	Inclusion criteria: girls with clinical characteristics of TS verified by karyotype analysis, with monosomy XO, with mosaicism, with structurally abnormal X chromosome, chronological age (CA) > 5 years and bone age (BA) < 11 years. Exclusion criteria: any demonstrable Y chromosome material, any endocrine or metabolic disorder (i.e. GH deficiency or carbohydrate intolerance) Diagnostic criteria: karyotyping Setting: paediatric department Age group: children/adolescents Gender distribution: females Country where trial was performed: Germany
Interventions	Intervention: oxandrolone + GH Comparator: GH Duration of intervention: 5 years Run‐in period: 5 years Number of study centres: not mentioned Treatment before study: none
Outcomes	Composite outcome measures reported: no
Study details	Trial identifier: not reported Trial terminated early: no
Publication details	Language of publication: English Funding: not clearly mentioned Publication status: peer‐reviewed journal/full article and journal supplement
Stated aim for study	Quote: introduction: "the aim of this study was to find out if even lower doses of rhGH would improve final heights in girls with Turner Syndrome when combined with oxandrolone" and "2002‐Abstract: the objective of this study was to find out whether moderate doses of growth hormone (GH) in combination with oxandrolone and late initiation of puberty could improve adult height even in relatively old patients with Ultrich‐Turner syndrome (UTS)."
Notes
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	Unclear risk	Quote: article 1992: materials and methods: "patients were randomly assigned to receive either recombinant human growth hormone (rhGH) 18 IU/m²/week administered by daily subcutaneous injection or the same dose of rh growth hormone plus oxandrolone 0.1 mg/kg/day orally for the first year of therapy." (page 38) Comment: method of random sequence generation not clearly mentioned.
Allocation concealment (selection bias)	Unclear risk	Comment: allocation concealment not mentioned.
Blinding of participants and personnel (performance bias) adverse events	High risk	Comment: no blinding; outcome measure likely influenced by lack of blinding.
Blinding of participants and personnel (performance bias) improvement in final adult height/increase in height velocity	Low risk	Comment: no blinding; outcome measure unlikely influenced by lack of blinding.
Blinding of outcome assessment (detection bias) adverse events	High risk	Comment: no blinding; outcome measure likely influenced by lack of blinding.
Blinding of outcome assessment (detection bias) improvement in final adult height/increase in height velocity	Low risk	Comment: no blinding; outcome measure unlikely influenced by lack of blinding.
Incomplete outcome data (attrition bias) adverse events	High risk	Comment: investigator‐assessed and self‐reported outcome measurement. Comment: 23/91 (25%) participants left the trial (oxandrolone) due to adverse effects and their details were not reported.
Incomplete outcome data (attrition bias) improvement in final adult height/increase in height velocity	High risk	Comment: missing data not clearly reported, no intention‐to‐treat analysis. Comment: data of 6 participants were not included in the final analysis because they either showed no beneficial response to GH administration or only for a short period up to 2 years after starting treatment. (page 131, article 2002)
Selective reporting (reporting bias)	High risk	Comment: general reporting of data not adequate.
Other bias	High risk	Comment: participants were switched between groups to create a third new group after randomisation that could have negatively affected the randomisation process. Authors performed a per‐protocol analysis but did not do an intention‐to‐treat analysis.

Zeger 2011.

Methods	Design: parallel, randomised controlled trial
Participants	Inclusion criteria: karyotype diagnosis of TS, no prior treatment with oestrogen or androgen, no prior treatment with GH exceeding 12 months or in the preceding 3 months, chronological age 10.0–14.9 years and bone age ≤ 12 years) Exclusion criteria: Y chromosome component in the peripheral karyotype (page 39) Diagnostic criteria: karyotyping Setting: university and developmental endocrinology branch Age group: children/adolescents Gender distribution: females Country where trial was performed: USA
Interventions	Intervention: oxandrolone + GH Comparator: GH + placebo Duration of intervention: 4 years Number of study centres: 2; Thomas Jefferson University and Developmental Endocrinology Branch of the National Institute of Child Health and Human Development, USA Treatment before study: none
Outcomes	Composite outcome measures reported: no
Study details	Trial identifier: NCT00029159 Trial terminated early: no
Publication details	Language of publication: English Commercial funding: GH was donated by Eli Lilly & Co., Indianapolis, IN, USA – oxandrolone and placebo were supplied by Bio‐Technology general Corporation, Iselin, NJ, USA Publication status: peer‐reviewed journal/full article
Stated aim for study	Quote:abstract: "to investigate the benefit of adding oxandrolone to GH in a long‐term, randomised, placebo (Pl)‐controlled prospective trial to near adult height in TS."
Notes
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	Unclear risk	Quote: "subjects were randomised (by the NIH [National Institutes of Health] Clinical Center Research Pharmacy) at baseline to receive daily treatment for 2 years …" (page 39) Comment: no details
Allocation concealment (selection bias)	Unclear risk	Comment: no details
Blinding of participants and personnel (performance bias) adverse events	Low risk	Quote: methods: "subjects and investigators remained blinded to treatment group." (page 39)
Blinding of participants and personnel (performance bias) bone maturation	Low risk	Quote: "bone ages (by X‐ray) were assessed by the method of Greulich and Pyle [19] and were read by a single‐blinded investigator (J.L.R.)." (page 40) Comment: investigator‐assessed.
Blinding of participants and personnel (performance bias) improvement in final adult height/increase in height velocity	Low risk	Quote: methods: "Subjects and investigators remained blinded to treatment group." (page 39) Comment: investigator‐assessed.
Blinding of outcome assessment (detection bias) adverse events	Low risk	Comment: investigator‐assessed. History and examination was performed by a physician blinded to the treatment group and all laboratory test were read by a single‐blinded investigator. (page 40)
Blinding of outcome assessment (detection bias) bone maturation	Low risk	Quote: "study participants were monitored at baseline and every 6 months by obtaining a history and performing physical examination, measurement of height, weight, waist and hip circumference, breast and pubic hair Tanner staging (determined by a physician blinded to treatment group), and hormone measurements [thyroxine, thyroid‐stimulating hormone, insulin, luteinizing hormone, follicle‐stimulating hormone (FSH), estradiol, free testosterone, insulin‐like‐growth factor 1 (IGF‐1) and insulin‐like growth factor binding protein 3 (IGFBP‐3)]. Bone ages (by X‐ray) were assessed by the method of Greulich and Pyle and were read by a single‐blinded investigator (J.L.R.)." (page 40) Comment: investigator‐assessed. History and examination was performed by a physician blinded to the treatment group and all laboratory test were read by a single‐blinded investigator. (page 40)
Blinding of outcome assessment (detection bias) improvement in final adult height/increase in height velocity	Low risk	Comment: investigator‐assessed. History and examination was performed by a physician blinded to the treatment group and all laboratory test were read by a single‐blinded investigator. (page 40)
Incomplete outcome data (attrition bias) adverse events	Low risk	Comment: missing data reported and reasons explained. (pages 39 and 40)
Incomplete outcome data (attrition bias) bone maturation	Low risk	Comment: missing data reported and reasons explained. (pages 39 and 40)
Incomplete outcome data (attrition bias) improvement in final adult height/increase in height velocity	Low risk	Comment: missing data reported and reasons explained. (pages 39 and 40)
Selective reporting (reporting bias)	Low risk	Comment: none detected.
Other bias	Low risk	Comment: none detected.

Note: blank cells in risk of bias domains with 'unclear risk of bias' indicate that the associated outcome was not investigated.

GH: growth hormone; OD: once daily; PO: per oral; SC: subcutaneous; SD: standard deviation; TS: Turner syndrome.

Characteristics of excluded studies [ordered by study ID]

Study	Reason for exclusion
Ferrández 1989	Duration of trial 6 months.
Joss 1997	Not a randomised trial, authors compared 3 modalities of treatment, high growth hormone, low growth hormone and oxandrolone.
Nilsson 1996	All 3 treatment groups were given oxandrolone and growth hormone in different combinations.
Sas 2014	Narrative review of 3 randomised controlled trials investigating the role of oxandrolone in growth hormone‐treated girls with Turner syndrome.
Sheanon 2015	Systematic review/meta‐analysis of randomised controlled trials of girls with Turner syndrome who received oxandrolone or growth hormone, or both.

Differences between protocol and review

We also included trials that enrolled some participants with Turner syndrome who had an abnormal Y chromosome. We had not mentioned the population of people with Turner syndrome phenotype and abnormal Y chromosome as an inclusion or exclusion criterion in our review protocol (Mohamed 2013).

Two review authors (HA, YSA) joined the group and participated in data extraction, data analysis, quality assessment, data interpretation and review writing.

We did not use funnel plots to assess small‐study effects as there were too few included studies.

Contributions of authors

SM: acquiring trial reports, trial selection, assessment of the risk of bias, data extraction, data analysis and data interpretation.

HA: data extraction, data analysis and data interpretation.

YA: acquiring trial reports, trial selection and data extraction.

YSA: assessment of the risk of bias, data extraction, data analysis and data interpretation.

KA: acquiring trial reports, trial selection, data analysis and data interpretation.

All review authors contributed to writing, read and approved the final review.

Sources of support

Internal sources

• Celiac Disease Research Chair, College of Medicine, King Saud University, Riyadh, Saudi Arabia, Saudi Arabia.

  Provide access to journals and references, computer and secretarial assistance

External sources

• Metabolic and Endocrine Group, Cochrane, Germany.

  Editing and reviewing protocol and helping with search strategy

Declarations of interest

SM: none.

HA: none.

YA: none.

YSA: none.

KA: none.

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