Estradiol, Sensitive, LC/MS

Estradiol is responsible for the regulation of the estrous and female menstrual reproductive cycles and for the development and maintenance of female secondary sex characteristics.^2,3 Estradiol plays a key role in germ cell maturation and numerous other, non−gender-specific processes, including growth, bone metabolism, nervous system maturation, and endothelial responsiveness. Estrogens are crucial for the normal development and maintenance of the breasts and the uterus.⁴ Excessive estrogen levels, however, can promote cell proliferation and may increase the risk of developing breast and uterine cancers as well as uterine endometriosis.⁴

The three major naturally occurring estrogens in women are estrone (E1), estradiol (E2), and estriol (E3). E2 is the predominant estrogen during reproductive years—both in terms of absolute serum levels as well as estrogenic activity.² During menopause, a dramatic drop in E2 production leaves estrone as the predominant circulating estrogen. Estriol is the main pregnancy estrogen, but it does not play a significant role in nonpregnant women or men.² The concentration of E2 in men is much lower than in women of reproductive age. All estrogens are synthesized from androgen precursors by the enzyme aromatase.^2,4 Aromatase converts the androgenic substrates androstenedione, testosterone, and 16-hydroxytestosterone to the corresponding estrogens: estrone, estradiol, and estriol.⁴ E2 is produced primarily in ovaries and testes by aromatization of testosterone.² A lesser amount of E2 is produced in the adrenal glands and some peripheral sites, most notably adipose tissue. Most of the circulating estrone is derived from peripheral aromatization of androstenedione (mainly in the adrenal glands). E2 and E1 can be converted into each other, and both are inactivated via hydroxylation and conjugation. E2 demonstrates two to five times the biological potency of E1.²

The importance of E2 testing and the need for reliable and accurate estradiol measurements throughout the analytic range are emphasized in several recent publications.^1,5,6 Measurement of serum E2 serves an integral role in the assessment of reproductive function in females and in the assessment of infertility, oligo-amenorrhea, and menopausal status. E2 is commonly measured for monitoring ovulation induction, as well as during preparation for in vitro fertilization. Because of the relatively high serum concentrations of E2 in these patients, readily available automated immunoassays methods with modest sensitivity meet the clinical requirements. E2 levels in children, postmenopausal women and men and are much lower than in women of reproductive age. The increased sensitivity and specificity that are achieved by LC/MS-MS are the more appropriate choice for these clinical situations.^7,8 LC/MS-MS offers superior analytical sensitivity, specificity and a larger dynamic range than immunoassays.⁷ The clinical applications benefiting from highly sensitive E2 measurement include the assessment of congenital defects in sex steroid metabolism and disorders of puberty. This sensitive assay also has application in the evaluation of estrogen deficiency in men menopausal women, fracture risk assessment in these populations, and increasingly, in therapeutic drug monitoring of low-dose female hormone replacement therapy or anti-estrogen treatment.

Adult Women. In premenopausal women, E2 levels, along with luteinizing hormone (LH) and follicle-stimulating hormone (FSH) levels, delineate the stage of the menstrual cycle.² E2 levels are lowest during the early follicular phase and rise gradually. Two to three days before ovulation, estradiol levels start to increase much more rapidly to a peak just before the ovulation. This dramatic increase in circulating E2 levels induces a surge in LH and FSH. E2 levels decline modestly during the ovulatory phase and then increase again gradually until the midpoint of the luteal phase and ultimately decline back to early follicular levels.

Assessment of E2 levels is useful for the evaluation of hypogonadism and oligo-amenorrhea in women. Decreased ovarian estrogen production is classified as hypergonadotropic or hypogonadotropic, depending on whether the disease is of gonadal or pituitary/hypothalamic origin.^9-11 Measurement of gonadotropins (LH and FSH) is fundamental in differentiating these two low estradiol states. The main causes of primary gonadal failure (hypergonadotropic) are genetic (Turner syndrome, familial premature ovarian failure), autoimmune (autoimmune ovarian failure, autoimmune polyglandular endocrine failure syndrome type II), and toxic (related to chemotherapy or radiation therapy for malignant disease).

Low E2 with low or inappropriately "normal," LH and/or FSH in young adult females is consistent with hypogonadotrophic hypogonadism.^11-13 This can be caused by hypothalamic or pituitary failure due to conditions, including multiple pituitary hormone deficiency and Kallmann syndrome. Diagnostic workup includes the measurement of E2 along with pituitary gonadotropins and prolactin—and possibly imaging. This endocrine presentation can be caused by starvation, over-exercise, severe physical or emotional stress, and drug/alcohol abuse. While early studies suggested that E2 levels could be used to predict ovarian reserve in women of reproductive age undergoing assisted reproduction procedures, more recent studies have found the marker less useful.¹⁴ Estradiol measurement is useful in assessing the status of ovulation induction in women with hypogonadotropic hypogonadism¹⁵ and for the prediction and prevention of ovarian hyperstimulation syndrome in patients undergoing assisted reproduction.¹⁶

Normal or high E2 with irregular or absent menstrual periods is suggestive of possible polycystic ovarian syndrome, androgen-producing tumors, or estrogen-producing tumors. In these cases, measurement of total and bioavailable testosterone, androstenedione, dehydroepiandrosterone (sulfate), and sex hormone-binding globulin can aid in differential diagnosis.

The main site of estrogen biosynthesis in the nonpregnant premenopausal woman is the ovarian granulosa cells, however, the adipose tissue becomes a major source of circulating estradiol in postmenopausal women.⁴ After menopause, androstenedione, secreted by the adrenal gland, is converted into estrone in the adipose tissue.⁴ The conversion of plasma androstenedione to estrone increases with excess body weight in both pre- and postmenopausal women.⁴ Estrone is then eventually converted to estradiol by 17 β-hydroxysteroid dehydrogenase enzymes present in peripheral tissues.⁴

Measurement of E2 level, together with FSH and/or anti-Müllerian hormone, can be useful in predicting the timing of the transition into menopause.^17,18 A large population study (Randolph) found that the mean E2 level started&