At a Glance

Androgen insensitivity syndrome (AIS), previously named testicular feminization syndrome, is an X-linked recessive disorder causing failure of external male genitalia development in a patient with a 46,XY genotype. The mechanism of the disorder is androgen resistance, rather than androgen deficiency, hence the name change.

Androgens are hormones required in male development. The principle androgens in humans are testosterone and its more potent metabolite, dihydrotestosterone (DHT). The function of androgen is mediated by the androgen receptor (AR). Loss-of-function mutations in the AR gene cause different degrees of impairment of androgen function, resulting in variable phenotypes of AIS. Approximately 750 different AR mutations that cause various forms of AIS have been reported. There are 3 clinical phenotypes of AIS in humans: complete (CAIS), partial (PAIS), and mild (MAIS). In all cases, the affected patients have normal or remnants of testes, normal testosterone production, and normal conversion of testosterone to DHT. However, the post receptor events do not occur.

CAIS prevents the development of the penis, descent of the testicles, and masculinization. At birth, a child with CAIS appears to be a typical girl. Patients typically present during teenage years with abnormal pubertal changes. Although breast development occurs, little pubic hair or other androgenic hair appears. The discrepancy of breast and hair development is sometimes the clue for further investigations. Delayed menarche, primary amenorrhea, difficulty with intercourse, and infertility can all be presenting symptoms. Occasionally, an inguinal lump is noticed and confirmed as a testis. Remnants of testicular tissue may also be discovered as an inguinal hernia without an obvious lump. Abdominal surgery performed for other reasons may reveal the presence of testes or lack of ovaries and uterus. In rare situations, a karyotype performed for unrelated reasons reveals a 46,XY karyotype in a phenotypic female to the surprise of all.

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PAIS is usually diagnosed during early childhood, since such children may have both male and female physical appearance. The degree of sexual ambiguity varies widely in persons with PAIS. Many have partial closing of the outer vaginal lips, an enlarged clitoris, and a short vagina. Some show undervirilized male external genitalia, such as hypospadias or diminished penile size.

MAIS patients usually have male genitalia. Impaired pubertal virilization, infertility, and gynecomastia in puberty are the presenting symptoms.

What Tests Should I Request to Confirm My Clinical Dx? In addition, what follow-up tests might be useful?

Measurement of serum testosterone or DHT levels is the first step. Total testosterone levels are usually elevated in the range of normal males. In normal females, testosterone is approximately 5-10% of male levels. Luteinizing hormone (LH) is abnormally high because of diminished negative feedback. Estradiol, from the conversion of high levels of testosterone, is also elevated in these patients, despite having no ovaries. (Table 1)

Table 1.
Serum Testosterone Serum LH Karyotype of a phenotypical female
Increased Increased 46, XY

A low level of testosterone is suggestive of hypogonadism and excludes the possibility of AIS. An elevated ratio of testosterone to DHT suggests a patient has 5-alpha reductase deficiency, instead of PAIS.

Karyotype or fluorescence in-situ hybridization (FISH) analysis using Y chromosomal specific probe on peripheral blood or buccal mucosa cells is important to determine a patient’s genetic makeup. Pelvic ultrasound is also used to find testes in the abdomen or inguinal area and the absence of fallopian tubes, a uterus, or a proximal vagina. Diagnosis of CAIS is confirmed by 46,XY karotype and lack of uterus and cervix by ultrasound in a phenotypic female.

Currently, mutation analysis by sequencing of AR is available commercially. Mutations of AR are found in 95% of CAIS, less than 50% of PAIS, and even less for MAIS. It is usually not necessary for diagnosis in most cases; however, it is very important for genetic counseling.

Are There Any Factors That Might Affect the Lab Results? In particular, does your patient take any medications – OTC drugs or Herbals – that might affect the lab results?

The testosterone assay is most commonly performed using immunoassay. The immunoassay has lower sensitivity, which may not detect low levels of testosterone. Immunoassay is also prone to interference by patients’ endogenous compounds, especially in young children. Variation among different immunoassays is high, which inhibits standardization of the testosterone test.

Liquid chromatography tandem mass spectrometry (LC-MS/MS) has become the most accurate method for steroid measurement. Although the variation is still relatively high, the specificity and sensitivity of steroid measurement is significantly improved. LC-MS/MS methodology requires expensive instruments and experienced technologists and is only used in large medical centers or reference laboratories. If there is discrepancy between a test result and clinical suspicion, LC-MS/MS can be used to confirm the testosterone level. In addition, the lack of specificity is more critical in the newborn and early infant periods. In our hospital, all the testosterone levels in patients younger than 1 month old are performed using LC-MS/MS.

What Lab Results Are Absolutely Confirmatory?

Approximately 750 different mutations have been found in the AR gene that affects the function of the androgen receptor. Finding known mutations in the AR gene is absolutely confirmatory. The sensitivity of mutation analysis by sequencing is high for CAIS (~95%) but low for PAIS (<50%) and MAIS (percentage unknown).

What Tests Should I Request to Confirm My Clinical Dx? In addition, what follow-up tests might be useful?

In a phenotypic female patient with increased serum testosterone and LH, a follow-up cytogenetic analysis of chromosome should be performed. If the patient has a karyotype of 46,XY, the diagnosis of AIS is confirmed.

Counseling is the most important first step in the treatment of these patients. Patients and parents usually have difficulty accepting the diagnosis and may benefit from referral to the Androgen Insensitivity Syndrome Support Group (AISSG).

For CAIS, treatment usually includes estrogen replacement therapy starting in late adolescence. Surgical removing of testes is also done after puberty to prevent malignant transformation. Vaginal construction can be accomplished by dilation or surgical creation.

For PAIS, treatment is similar to CAIS for patients with female gender. However, for PAIS patients with male gender identities (undervirilized male external genitalia), they may be treated with supra-physiological level of testosterone and/or DHT.

Are There Any Factors That Might Affect the Lab Results? In particular, does your patient take any medications – OTC drugs or Herbals – that might affect the lab results?

As mentioned, for young infants, testosterone level should be tested using LC-MS/MS to improve the sensitivity and specificity.

Reference ranges vary with age and sexual developmental stages of the child. Using age and Tanner stage specific reference ranges is very important. Testosterone levels are low at birth, rise within weeks, and remain at normal male pubertal levels for about 2 months before declining to the low, barely detectable childhood levels. Testosterone has a diurnal variation, with peak serum levels at 0400-0800 and minimum levels at 1600–2000. Errors in diagnosis could occur without using the appropriate reference ranges.

If a patient takes or abuses exogenous testosterone, the serum testosterone level will be increased. However, if a patient has normal function of AR, the negative feedback of hypothalamus-pituitary-gonad axis will cause decreased serum LH level, instead of increased LH level seen in AIS patients.