Genetic errors that give rise to ambiguous genitalia

article source:


Genetic errors that give rise to ambiguous genitalia 
John Paul's parents were happy to see their newborn son complete with ten fingers and toes. While confident there was nothing wrong, the parents took their pediatrician's advise to subject John Paul to newborn screening. This blood test detects five congenital metabolic diseases-congenital hypothyroidism (CH), congenital adrenal hyperplasia (CAH), glucose 6 phosphate dehydrogenase (G6PD) deficiency, galactosemia, and phenylketonuria (PKU)-in the hope of catching them early to avoid serious and life-threatening symptoms.

A few drops of John Paul's blood from a heel prick were blotted on special filter paper, air-dried, and sent to the Newborn Screening Laboratory at the National Institutes of Health. Result: John Paul was positive for CAH. The pediatrician was concerned that the baby might experience salt-wasting and dehydration-a fatal condition common in 75 percent of CAH patients. If left untreated it could lead to hypotension and shock. A confirmatory test showed John Paul did not have salt-wasting nephropathy. But the newborn's problems did not end there. Karyotyping-a blood test that maps all 46 of a person's chromosomes-revealed John Paul had two X chromosomes, making him genetically female.

What Lies Beneath

Among patients positive for CAH, genital ambiguity at birth is fairly common. Dr. Sylvia Estrada, clinical assistant professor of pediatrics and endocrinology at UP-PGH, says there are many causes of ambiguous genitalia and "we lump these all into what we call the intersex disorders."

Estrada says that cases of ambiguous genitalia involving newborns to teenagers get referred to her clinic. Investigating the cause of the ambiguous genitalia is just the tip of the iceberg. Says she: "When faced with a patient [who has ambiguous genitalia] I ask [the parents or patient] what the primary concern is. Is it because of these anomalies or is it because the parents are now thinking the child can become the opposite sex. As the physician you would like to know the concerns and what is causing all of these before we address the problem."

Karyotyping can tell whether the baby is genetically male or female, has a complete set of chromosomes, or has an extra, missing, or abnormally shaped chromosome. But because it is expensive, it is not routinely done. "Unfortunately not everyone can afford endocrine tests. But definitely if you are presented with an ambiguous genitalia you have to do karyotyping which [normally] costs PhP2,500."

Under the Influence

Of the 23 pairs of chromosomes in humans, one pair determines gender. Males have an X and a Y chromosome (XY) while females have two X chromosomes (XX). Extra or missing chromosomes cause clinical disorder. For instance trisomy, a condition where three chromosomes of a kind are present in an individual has been implicated in syndromes like Down's (where the patient has three chromosome 21), Edwards (trisomy 18), and Patau (trisomy 13).

At seven weeks of embryonic life, gonadal tissues that become the sex organs have not yet differentiated into either male or female.

An area in the short arm of the Y chromosome called the sex-determining region of the Y chromosome (SRY) houses the genes for testis determining factor (TDF). Under the influence of TDF gonadal tissue in the fetus develops into the testes, producing testosterone and mullerian inhibiting substance (MIS) or antimullerian hormone (AMH). Testosterone and MIS/AMH induce completion of the male internal genital ducts or obliteration of tissues that give rise to female internal genital ducts. Testosterone conversion into dihydrotestosterone (DHT) will ensure development of the male external genitalia.

When TDF is absent, the gonadal tissue remains in the abdomen and cells that give rise to male internal genital ducts regress. The gonadal tissue then proceeds to develop into the ovaries and uterus that soon secrete estrogens stimulating the completion of the female internal genital ducts and ensuring development of the vagina, labia, and clitoris.

Teratogens, chromosomal aberrations, or alterations in the genes' coding for certain hormones can alter the normal course of gonadal development in the fetus, causing ambiguous genitalia. Points out Estrada: "Chromosomal abnormalities can cause ambiguous genitalia. It can also happen in mosaics where you have two cell lines in the same person. You can also have a true hermaphrodite or gonadal dysgenesis. We have all these cases in the Philippines."
Klinefelter syndrome, a condition where males have an extra X chromosome (XXY), affects approximately one in 1,000 newborn males. The extra X chromosome is maternal in origin in 54 percent of cases and paternal in 46 percent. Klinefelter variants have been reported to have as many as four X chromosomes (XXXXY). Affected individuals suffer from some degree of mental retardation. During puberty testicular growth and secondary male sexual characteristics like deepening of the voice or facial hair fail to manifest. The testes and phallus tend to be small for age. In some cases the testes are not in the scrotum but remain in the abdomen (cryptorchidism). The urethral meatus may not be at the tip of the penis but lie somewhere below the shaft (hypospadias).

In other cases, ambiguous genitalia is present. Androgen deficiency is usually detected in this condition with about 80 percent having enlarged breasts (gynecomastia). Varicose veins and breast cancer are also likely to occur.

Mosaic individuals have two or more types of cells containing different numbers of chromosomes. Although many mosaic individuals with ambiguous genitalia have cells with a normal number of chromosomes, some of their cells have two X chromosomes (XX) while others have one X and Y (XY). This could be perplexing to the individual who is programmed to express both male and female sex organs. Many true hermaphrodites are mosaic individuals whose gonadal findings may be any combination of ovary, testis, or ovotestis.

Other individuals suffer from a condition called mixed gonadal dysgenesis (MGD). Patients with this condition may present with undeveloped male and female internal genitalia. Chromosomally some cells will have an XY while others will have only one X chromosome (XO).

Gonadoblastoma, seminoma, and embryonal cell carcinomas develop. This can be circumvented by gonadectomy prior to adulthood when the tumors most likely appear. Estrogen and progesterone support is usually indicated especially if the individual was raised as a female.

At times, intersex conditions occur in individuals who have a normal set of chromosomes. According to Estrada, "anything that will affect the genes that code for the protein and the hormones can have an effect [on gonadal development]. "

Teratogens can also affect the fetus. Teratogenic episodes like androgens and progestenes ingested in the first trimester of pregnancy can induce virilization in the unborn child if it is female. Upon birth, the baby may manifest with an enlarged clitoris that can be confused for a penis. Testosterone enanthate, testosterone propionate, methylandrostenediol, 6a methyltestosterone, ethisterone, norethindrone, and danazol have been shown to cause genital ambiguity.

Botched Chemistry

Androgen insensitivity and 5 reductase deficiency cause testicular feminization in males (they don't affect females) who may have completely feminine-looking external genitalia. But their testes still develop, though they fail to descend, and no ovary or uterus develops. In androgen insensitivity, fetal tissues supposed to develop into male external genitalia fail to do so because they do not respond to the hormone dihydrotestosterone (DHT).

Testosterone fails to convert to DHT when 5 reductase is deficient, inhibiting the development of male external genitalia. But unlike those with androgen deficiency, individuals with 5a reductase deficiency may still develop male external genitalia and manifest with body hair, large muscle mass, and deepening of the voice during puberty.

According to Estrada, the most common cause of ambiguous genitalia is CAH. CAH is a group of enzymatic disorders that depending on the type upset a child's sexual differentiation, promote early puberty, induce electrolyte imbalance, and cause death.

"21 hydroxylase deficiency is the most common of all the CAH subtypes, about 90 to 95 percent of all CAH fall under this category," says Estrada. "This is the CAH subtype picked up by newborn screening, and the physician is given a window period of 10 days to 14 days after birth to confirm the CAH and determine if the patient is the salt-losing type. If untreated [salt losers are given] 14 days to crash."

21 hydroxylase deficiency causes increased testosterone levels, resulting in virilization of females. Affected females though are not devoid of estrogens since testosterone and its precursor androstenedione can be converted into estradiol and estrone by the enzyme aromatase. Explains Estrada: "There is just too much androgens versus the estrogens. Besides you need aromatase to tick off that conversion but that would only take place at puberty. Plus, you also need puberty to make your estrogen receptors receptive. So what happens is you have excess testosterone that should be peripherally converted to estrogens."

Even if males with 21 hydroxylase deficiency are spared the ambiguous genitalia, the more pressing problem, Estrada warns, is the salt wasting caused by the nonproduction of cortisol and aldosterone seen in three out of four newborns affected. If untreated 10 to 14 days after birth, this will lead to dehydration, hypotension, shock, and death.

Another CAH subtype is 11 hydroxylase deficiency, which accounts for five to eight percent of cases. Infants with this condition are also prone to high testosterone levels, thus affected females experience virilization as in 21 hydroxylase deficiency. The hallmark of this condition is the severe hypertension brought about by accumulation of 11-deoxycorticosterone (DOC), one of the intermediate metabolites of aldosterone synthesis.
Less than five percent of CAH cases involve 3 hydroxysteroid dehydrogenase deficiency. Infants with this condition have decreased synthesis of cortisol, aldosterone, and androgens. Salt wasting occurs in the classic form of the disease, and ambiguity of the genitalia could result in both males and females. A milder nonclassic form exists where genital ambiguity and salt wasting do not occur but early pubarche, menstrual disorders, infertility, hirsutism, or polycystic ovaries develop in affected females.

In 17 hydroxylase deficiency, overproduction of DOC leads to hypertension, hypokalemia, suppression of renin and aldosterone, and inability to produce normal amounts of sex hormones. In male infants, lack of sex hormones leads to ambiguous or female external genitalia. Females, on the other hand, will not develop secondary sexual characteristics like breast enlargement and menstruation during puberty.

Rarer still is lipoid adrenal hyperpalasia that causes cholesterol and lipids to accumulate in the adrenals. The problem has been traced to failure of conversion of cholesterol to pregnenolone leading to non-production of aldosterone, cortisol, and sex hormones. The problem is traced to a defective gene in chromosome 15. Affected females do not present with genital ambiguity, but males present as females because of failure of testosterone production. These infants die early due to salt wasting.

Elusive Cure

CAH treatment entails the administration of glucocorticoids to inhibit the excessive production of androgens. "Hydrocortisone, which is your physiologic steroid, is ideally given. But because it is not available in the country, we rely on prednisone which is four times more potent and has side effects when given in excess," says Estrada.
Hydrocortisone is administered at 10-20mg/m2/24 hrs while prednisone is administered at 2.5-5mg/m2/24 hrs. Mineralocorticoids in the form of flourohydrocortisone are needed to normalize elevated plasma renin activity in both salt losers and non-salt losers. Flourohydrocortisone is not available in the Philippines.

Estrada remains hopeful: "Our plan at NIH is to make hydrocortisone and flourohydrocortisone available at cost for patients. The way these medicines are made available now is through networking. Like if some patients have excess, they sell it at cost to other patients who need it too. Hydrocortisone is more expensive than prednisone. If prednisone is PhP5 per tablet, hydrocotisone costs double or triple. Flourocortisone is PhP24 now."
Estrada adds that the enlarged clitoris in females with 21 hydroxylase deficiency may be surgically corrected at six to 12 months of age, but clitoral recession usually occurs over time and is preferred over surgical reduction.

For androgen insensitivity and 5 reducatase deficiency, gonadectomy and feminizing genitoplasty are recommended. Gonadectomy reduces the risk of developing gonadal tumors (specifically seminomas), which are associated with androgen insensitivity. Estrada notes that surgery to masculinize such patients is difficult and yields poor results.

Relates Estrada: "I had cases of 5 reducatase deficiency where the patients were reared as females but in the end [the parents] wanted [them] to become males. In one case, the patient started to have palpable inguinal masses at 13 years of age and started to look like a boy. I explained to the parents that we had to augment the penis and penile function may not be achieved. But they were willing to go through the whole process. The patient was sent for psychological therapy because psychiatrists and psychologists recommend that for a child beyond 10, it is simpler to continue with the gender he was accustomed to."

Estrada reiterates that genetic counseling is essential for parents who have children with intersex conditions or ambiguous genitalia. CAH, androgen insensitivity, and 5 reducatase deficiency are caused by mutations inherited as either autosomal recessive or X-linked genes. Klinefelter syndrome and MGD are casued by chromosomal aberrations

Estrada also stresses that 21 hydroxylase deficiency, the most common cause of ambiguous genitalia, can easily be detected through newborn screening for early management.