Estradiol Target Level in Treating Endometriosis

FERTILITY AND STERILITY Copyright © 1992 The American Fertility Society

Estradiol Target Level in Treating Endometriosis

To the Editor: In their editorial, Barbieri and Gordon (1) provide an excellent review of the abilities of different gonadotropin-releasing hormone (GnRH) regimes to suppress estradiol (E 2) to target levels of 30 or 15 pg/mL but fail to indicate where the original recommendation for suppression to these levels came from. In our paper published in 1984, (2) we showed that to obtain complete remission of extensive endometriosis (American Fertility Society's [AFS] original classification [3]), it was necessary to suppress E2 to <15 pg/mL for ::::.75 days. Likewise, complete remission of AFS class severe and moderate endometriosis (3) required suppression to <22 and <41 pg/mL, respectively. We believe that these levels were subsequently utilized to design protocols for GnRH analog studies. We also pointed out that "Serum E2 levels, particularly the low levels found in menopause, may vary with different RIA techniques. Clinicians should be aware of normal values for menopause in their own reference laboratory." Experience in our radioimmunoassay laboratory shows that this situation has not changed.

Richard P. Dickey, M.D., Ph.D. David N. Curole, M.D. Steven N. Taylor, M.D. The Fertility Institute and Tulane University School of Medicine New Orleans, Louisiana January 22, 1992 REFERENCES 1. Barbieri RL, Gordon AC. Hormonal therapy of endometriosis:

the estradiol target. Fertil Steril 1991;56:820-2. 2. Dickey RP, Taylor SN, Curole D N. Serum estradiol and danazol. I. Endometriosis response, side effects, administration interval, concurrent spironolactone and dexamethasone. Fertil SterilI984;42:709-16. 3. American Fertility Society. Classification of endometriosis. Fertil Steril 1979;32:633.

Reply of the Author: I would like to thank Drs. Dickey, Taylor, and Curole for their interest and kind comments conVol. 57, No.6, June 1992

Vol. 57, No.6, June 1992 Printed on acid-free paper in U.S.A.

cerning our recent Editor's Corner (1). In 1984, Drs. Dickey, Taylor, and Curole published convincing evidence that the efficacy of danazol in the treatment of endometriosis was dependent, in part, on the degree of hypoestrogenism achieved (2). This is a landmark publication and deserves careful review by clinicians involved in endometriosis and research. Because of space and reference limitations imposed on manuscripts submitted to the Editor's Corner, we could not review the publication of Dickey et al. (2) In addition, the pharmacology of danazol is very complex (3). Danazol may have direct effects on endometriosis lesions independent of the effects produced by changes in circulating estradiol (E 2). For example, danazol may cause regression in endometriosis lesions by a direct action on intracellular androgen and progestin receptors (3). Because the major actions of gonadotropin-releasing hormone analogues (GnRH-a) occur by binding to pituitary GnRH receptors, our review focused on these highly specific agents. Furthermore, in the study by Dickey et al., (2) many of the subjects (n = 40) were treated with a multidrug regimen that combined danazol therapy with spironolactone or dexamethasone. Dickey et al. (2) noted that when using danazol (often given in combination with spironolactone or danazol) to treat endometriosis, "complete remission" of extensive, severe, and moderate endometriosis required suppression of circulating E2 to concentrations of 15, 22, and 41 pg/mL, respectively. As a clinician, I find these results to be consistent with my observations in practice. However, Henzl and colleagues (4) reported results from a very well designed and executed clinical trial and observed somewhat different results. Nasal nafarelin, at doses of 400 and 800 JLg/d, produced circulating E2 concentrations of approximately 28 and 15 pg/mL, respectively. However, both agents were equally efficacious against stage II, III, and IV endometriosis as determined by post-treatment laparoscopy. This study by Henzl and colleagues (4) does not support the contention of Dickey and colleagues (2) that E2 must be <15 pg/mL to treat stage IV endometriosis. Further investigation will be needed to clarify this issue. As an aside, danazol decreases hepatic production of sex-hormone binding globulin, and this may result in elevated concentrations of free E2 (3). ConseLetters-to-the-editor

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quently, it may not be appropriate to compare the effects of danazol and GnRH -a by referring to the "total" E2 concentration. Because of an increase in free E 2, danazol treatment may be less efficacious than GnRH-a treatment at any given target concentration of total E 2. The main thrust of our review was that clinicians may want to "aim" their hormonal therapy of endometriosis at a specific "E2 target." Estradiol radioimmunoassays are commercially available that provide high sensitivity (10 pgjmL) and good reliability (interassay variation < 10%). Gynecologists should use a high-quality clinical laboratory to measure their E2 samples.

Robert Barbieri, M.D. Department of Obstetrics and Gynecology Health Sciences Center State University of New York at Stony Brook Stony Brook, New York February 17,1992 REFERENCES 1. Barbieri RL, Gordon AMC. Hormonal therapy of endometriosis: the estradiol target. Fertil Steril 1991;56:820-2. 2. Dickey RP, Taylor SN, Curole D N. Serum estradiol and danazol. 1. Endometriosis response, side effects, administration interval, concurrent spironolactone and dexamethasone. Fertil Steril 1984;42:709-16. 3. Barbieri RL, Ryan KJ. Danazol: endocrine pharmacology and therapeutic applications. Am J Obstet GynecoI1981;141:45363. 4. Henzl MR, Corson SL, Moghissi K, Buttram VC, Berquist C, Jacobson J. Administration of nasal nafarelin as compared with oral danazol for endometriosis. N Engl J Med 1988;318: 485-9.

Editorial Comment

The easiest way to bring this discussion into sharp focus before everyone starts "targeting" these lowrange estradiol (E,J levels is to simply reiterate that E2 is a small molecule and it has been difficult to develop two-site "sandwich" assays using monoclonal antibodies for the measurement of small molecules. This is unfortunate because two-site systems offer a distinct advantage over competitive immunoassays in terms of greater precision and working range of analyte. Estradiol continues to be quantitated using polyclonal technology and isotopically labeled antigens. The technical constraints on the concentration of antibody in these competitive systems diminish the working range of the assay and lead to nonlinearity of the calibration curve at low concentrations of E 2. 1362

Letters-to-the-editor

There is a conspiratorial movement that is festering that seems to imply that we can confidently and consistently distinguish values of serum E2 at 5 to 10-pg intervals in the lower limits of the calibration plot for the assay. The thought of it gives me spells of double vision. It is true that many immunoassay systems, depending on the amount of nonspecific binding present, may have a relatively consistent lower limit of detection value. However, the lower limits of detection should not be confused with the sensitivity of an assay system. It is a widespread misconception that sensitivity and lower limits of detection are synonymous. The lower limit of detection is the lowest concentration of analyte detected by the method. This number is represented by the lowest value of an analyte (usually expressed in concentration) that can be statistically distinguished from a blank. It can be calculated by several methods including the precision of repeated measurements of a blank. Lower limits of detection is properly used to determine the concentration below which a value should be reported as "less than" rather than a finite value. Sensitivity, on the other hand, is the smallest concentration change that the method is capable of detecting. It is determined from the slope of the previously described plot and varies with the concentration range of the analyte. The differences between lower limits of detection and sensitivity are more than academic because the assay conditions or profile that are optimal for detecting small amounts of the analyte are not necessarily the same as the optimal conditions to maximize discrimination in the low range. (1) Sensitivity should reflect the ability of the assay to discriminate between two values, such as 15 or 20 pgjmL. The important consideration in these discussions is not the lower limits of detection but the ability of the assay to be truly discriminative in the low range, so that the difference between 15, 20, and 25 pg of E2 is a true difference and not subject to considerable analytical error. This is obviously important if you subscribe to the idea that lowering serum E2 to a specific target level will treat endometriosis while protecting bone density. This type of fine discernment or fingertip feel is difficult to consistently achieve with E2 assays because of differences in antibody specificity, antibody affinity, isotopic labels (gamma or beta emitters), and variations in technique among laboratories. Differences in study population and preparation for the assay have also contributed to wide discrepancies and poor discrimination among results. The external quality control that is designed to alert a laboratory to shortfalls in performance by providing objective information on the comparability of their E2 results with those of Fertility and Sterility

others using the same methodology is not in place at the present time. Circulation of spiked samples and reference values obtained by gas chromatographymass spectrometry (2) are a mandatory part of any assessment program for steroid assays. Until the time that such changes occur, a random doctor with a random E2 sample from a random patient sent to a random laboratory with a random method still has a random chance of obtaining a consistently accurate serum E2 value in these low analyte ranges. REFERENCES 1. Ezan E, TiberghIen C, Dray F. Practical method for opti-

mizing radioimmunoassay detection and precision limits. Clin Chern 1991;37:226-30. 2. Dikkeschei LD, deRuyter-Buitenhuis AW, Nagel GT, Schade JR, Wolthers BG, Kraan GPB, et al. GC-MS as a reference method in immunochemical steroid hormone analyses. J Clin Immunoassay 1991;14:37-43.

Paul G. McDonough, M.D., Editor, Letters

Biochemical Pregnancies or Heterophil Antibodies?

To the Editor: We were surprised and pleased to see the case report by Maier and Metzger (1) because we had prepared an almost identical case for publication. A 36-year-old white woman presented because of secondary infertility. She reported a history of right tubal pregnancy treated by right salpingectomy and of left ectopic pregnancy that permitted salvage of that tube. A hysterosalpingogram revealed a Tshaped uterus, characteristic of diethylstilbestral (DES) exposure in utero. Because she was sperm-antibody positive and demonstrated a persistently elevated total immunoglobulin (Ig)M and intermittently autoantibody abnormalities, especially ofthe IgG antiphospholipid phosphatidylserine, she was considered at risk for pregnancy loss (2). Because ovulation induction with clomiphene citrate had been unsuccessful elsewhere, she was treated with human menopausal gonadotropin and human chorionic gonadotropin (hCG). Progesterone (P) substitution by intramuscular injection was given in the luteal phase based on peripheral P levels. Vol. 57, No.6, June 1992

When antibody abnormalities were detected, the female was also treated with 20 mg of prednisone daily. Six such treatment cycles demonstrated positive hCG serum values in the luteal phase, when hCG values in nonpregnant subjects usually have reverted to zero (Table 1). In view of the patient's autoantibody findings, this was initially interpreted as early pregnancy wastage. When the pattern became repetitive, alternative explanations were explored. Ectopic hCG production from a tumor was ruled out through ovarian ultrasound scannings and tumor marker screening with a-fetoprotein, cancer antigen CA-125, and carcino embryonic antigen. At that point a "fake" cycle was instigated at the request of the husband (Table 1) entailing routine ovarian stimulation and ovulation induction with hCG and sexual abstinence by the couple. Human chorionic gonadotropin values were assessed throughout by the Stratus hCG Fluorometric Enzyme Immunoassay (Baxter Healthcare Corporation, Miami, FL). The assay has been standardized against the World Health Organization First International Reference Preparation (IRP no. 75/537). The level of sensitivity is 1.9 mID /mL. The interand intra-assay coefficients of variation are <10%. The hCG assay procedure consists of a sandwich immunoassay that uses two mouse monoclonal antibodies (IgG fraction) specific for the two subunits of hCG. As a result, this methodology permits the detection of the intact (covalently linked a and (3 subunits) hCG molecule. Damewood et al. (3) recently reported that with currently used highly sensitive and specific immunoassays, hCG levels were detectable up to 12 to 14 days past injection. A level exceeding 10% of the initial level on day 10 was, in their opinion, suggestive of pregnancy. The patient reported here and the one reported by Maier and Metzger (1) prove otherwise. Although the disappearance curve of hCG, reported by Damewood et al. (3), appears characteristic for a vast majority of patients, selected cases can clearly demonstrate a prolonged disappearance. As these cases also demonstrate, delayed disappearance of hCG can lead to the false diagnosis of ("chemical") pregnancies, especially if patients have a history that may predispose them to pregnancy loss. Therefore, one has to conclude that the diagnosis of pregnancy based on absolute (or relative) hCG levels after hCG administration can be misleading. Consequently, pregnancy should not be diagnosed before a significant rise in hCG levels can be observed. Letters-to-the-editor

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