CLIN. CHEM. 39/9, 1857-1860(1993) CLINICALCHEMISTRY,Vol. 39, No. 9, 1993 1857 Substantial Urinary Concentrations of Material Resembling 13-CoreFragment of Chorionic Gonadotropin /3-Subunit in Mid-Menstrual Cycle Patrick Neven,”4 Ray K. lies,’ Ian Howes,2 Kiran Sharma,’ John H. Shepherd,1 Ray Edwards,2 William P. Collins,3 and Tim Chard’ We measured the day-to-day variations in concentrations of p-core, luteinizing hormone (LH), and a-subunit in urine during the menstrual cycle. The a-subunit concentrations showed a pattern similar to that of the LH concentrations. p-Core-like material was increased during and up to 3 to 4 days after the surge in urine LH. The urine LH concen- tration was associated with the presence of a-core immu- noreactivity during the urine LH peak. Chromatography showed that, at the peak LH concentration and at 2 days after the LH peak, p-core immunoreactivity could be ac- counted for by the presence of a peptide of low molecular mass similar to the p-core molecule of I,CG, but probably originating from the degradation of LH. The prolonged excretion of gonadotropin metabolites in the midcycle must be considered when p-core is being assessed as a tumor marker. IndexIngTerms: luteinizinghormone tumor marker . varia- tion, source of The metabolism of circulating gonadotropins includes degradation in the kidneys and excretion in urine. The most abundant material relatedto human choriomc go- nadotropin (hCG) in urine from pregnant women is a fragment of the p-subunit known as p-core (1-3). A similar protein of small molecular mass, which reacts in assays for luteini.zing hormone (LH), is present in the urine of nonpregnant pre- and postmenopausal women (4). This cross-reactant probably results from degrada- tion of the p-subunit of LH and is likely to have consid- erable sequence homology with the p-core fragment of hCG (5). Consequently, this p-LH core reacts in assays directed against the p-core of hCG. Measurement of urinary metaholites of p-hCG, espe- daily of the core fragment, has been advocated as a marker in some nontrophoblastic gynecological malignan- cies (6-11). However, the presence of p-LH core iminuno- 1Wilhiamson Laboratory for Molecular Oncology, Joint Aca- demic Department of Obstetrics, Gynaecology and Reproductive Physiology, St. Bartholomew’s Hospital Medical College, West Smithfield, London EC1A 7BE, UK. 2North East Thames Regional Immunoassay Unit, St. Barthol- omew’s Hospital, West Smithfield, London EC1A 7BE, UK. ‘Diagnostics Research Unit, King’s College School of Medicine and Dentistry, Denmark Hill, London SE5 8RX, UK. 4Address for correspondence: Algemeen Ziekenhuis Sint Jan, Broekstraat 104, 1000 Brussel, Belgium. ‘Nonstandard abbreviations: LH, luteinizung hormone; hCG, human chorionic gonadotropun; IFMA, immunofluorometric assay; IRMA, immunoradiometric assay; NIH, National Institutes of Health; TSH, thyroid-stimulating hormone; and FSH, follicle- stimulating hormone. Received December 22, 1992; accepted March 9, 1993. reactivity in the urine of normal postmenopausal women must be considered when the concentration of /3-core-like material is being evaluated in cancer patients (5). There is little information on the amounts of urinary gonadotropin metabolites excreted during the normal menstrual cycle. Because gynecological malignancies may occur during the premenopausal period, the pres- ence of such metabolites in premenopausal women would be highly relevant to an assessment of the clinical value of urinary p-core as a cancer marker. We mea- sured p-core-like activity in early-morning urine speci- mens collected during the menstrual period, and present evidence that urinary gonadotropin metabolites are substantially increased during and after the midcycle LII surge. MaterIals and Methods Samples A total of 463 early-morning urine samples were col- lected from 22 healthy premenopausal women (ages 19-29 years) from day 2 of a menstrual cycle up to the following menses. The procedures followed were in ac- cordance with the ethical standards of our institution’s ethical committee. Sodium aside (1 g/L) was added to all samples prior to storage at -20 #{176}C. All samples were assayed for p-core-like material by RIA. The LH peak (day 0) was measured by immunofluorometric assay (IFMA) in each of the women by using samples from day -1 to day +5, the periovulatory period. Samples from the same period from nine randomly chosen cycles were also assayed for free a-subunit (RIA) and by another LH assay (immunoradiometric assay, IRMA). Assays p-Core immunoreactivity was measured by RIA with a polyclonal sheep antiserum raised against purified p-core from hCG (S504) and ‘251-labeled purified p-core (12). Standards were calibrated against material pro- vided by R. Wehmann and D. Blithe (NIH, Bethesda, MD). This assay shows partial cross-reaction with intact hCG (6.9%) and free p-hCG (18%), but negligible cross- reactivity with LH, thyroid-stimulating hormone (TSH), and follicle-stimulating hormone (FSH) (<0.7%). LH was measured by two methods: a time-resolved IFMA and an IRMA. The IFMA was performed in the Diagnostics Research Unit of King’s College School of Medicine and Dentistry. The assay is based on the direct sandwich technique in which a polyclonal antibody is used to capture the analyte and a europium-labeled monoclonal antibody against the a-subunit is used to