First-trimester hyperglycosylated human chorionic gonadotropin and development of hypertension.

OBJECTIVE: This study aimed to determine whether urine levels of hyperglycosylated human chorionic gonadotropin (HhCG) in the first trimester are predictive of subsequent development of hypertension during pregnancy METHOD: This prospective cohort study consisted of women seeking care before 12 weeks gestation. A clean catch urine was obtained at the first prenatal visit and tested for HhCG and creatinine levels. The median HhCG levels and multiples of the median (MoM) by gestational age were compared between the groups that either developed hypertension or did not. RESULTS: Urine HhCG were determined for 204 women between 4 weeks 4 days to 11 weeks 6 days. The median HhCG of those who developed gestational hypertension (n = 7) or preeclampsia (n = 15) did not differ from the group that did not (median: 284 ng/mg creatinine vs 365 ng/mg; p = 0.55). If the MoM of HhCG for the no hypertension group was 1.00, the MoM of HhCG for the hypertension group was 0.93 (p = 0.93). A possible association was observed after 10 weeks between low HhCG levels and the development of late-onset hypertension (≥34 weeks). CONCLUSIONS: Prenatal screening for subsequent hypertension is unreliable with a single measurement of maternal urine HhCG at 10 weeks or less.

Hyperglycosylated hCG, a review.

Hyperglycosylated hCG (hCG-H) is a glycoprotein with the same polypeptide structure as hCG, and much larger N- and O-linked oligosaccharides. The oligosaccharides increase the molecular weight of hCG from 36,000 - 37,000 u to 40,000 - 41,000 u, depending on the extent of hyperglycosylation. hCG-H has triantennary N-linked oligosaccharides and double molecular size O-linked oligosaccharides (hexasaccharide compared with predominantly trisaccharide structures). hCG is produced by syncytiotrophoblast cells while hCG-H is made by extravillous cytotrophoblast cells. hCG-H promotes trophoblast invasion during choriocarcinoma, growth of cytotrophoblast cells and placental implantation in pregnancy. hCG-H is an independent molecule to hCG with totally separate biological functions. hCG has numerous functions during pregnancy, it promotes progesterone production, promotes angiogenesis in uterine vasculature, immuno-suppresses the invading placental tissue, promotes the growth of the uterus in line with the growth of the fetus during pregnancy, promotes the differentiation of growing cytotrophoblast cells, promotes the quiescence of contractions in the uterine myometrium during the course of pregnancy, and also has function in growth and development of fetal organs. Monoclonal antibody B152 uniquely binds hCG-H. Using this monoclonal antibody in immunometric assays permits detection of pregnancy. It also permits management of gestational trophoblastic diseases and detection of quiescent gestational trophoblastic disease. This same test can be used to differentiate of aggressive and minimally-aggressive gestational trophoblastic disease, and discrimination of patients that respond to chemotherapy and who are chemorefractory. The hCG-H test can be used to screen for Down syndrome pregnancies and predict patients likely to generate hypertensive disorder in pregnancy. It also can be used to differentiate pregnancies that will miscarry and pregnancies that will go to term.

Hyperglycosylated hCG.

Hyperglycosylated hCG (hCG-H) is a glycosylation variant of the hormone hCG. Here we review all that is known about this independently functioning molecule. As discussed, it is a very different molecule to the hormone hCG. First, hCG-H is produced by cytotrophoblast cells while regular hCG is made in syncytiotrophoblast cell. Second, it is an autocrine acting directly on the cells which produce it, while regular hCG is an endocrine acting on maternal corpus luteal cells. Third, hCG-H has minimal biological activity in promoting progesterone production compared to regular hCG. Fourth, hCG-H functions unlike regular hCG as an invasion promoter, whether invasion as in choriocarcinoma and testicular germ cell malignancies, or as in implantation of pregnancy. These functions seemingly occur through action on cytotrophoblast cell TGFbeta receptors. Fifth, hCG-H is an essential component for successful human implantation to prevent early pregnancy loss and spontaneous abortion. Sixth, hCG-H is critical for promoting the midtrimester hemochorial implantation, and for preventing preeclampsia. Seventh, measurements of hCG-H have advantages over measurements of regular hCG or total hCG, in detecting pregnancy, pregnancy outcome (failing or term pregnancy), predicting preeclampsia in pregnancy, or as a tumor marker for gestational trophoblastic diseases.

Detection of early pregnancy forms of human chorionic gonadotropin by home pregnancy test devices.

BACKGROUND: Home pregnancy testing devices claim >99% diagnostic accuracy for pregnancy and utility on the first day of the missed menses or earlier. We investigated the forms of human chorionic gonadotropin (hCG) in early pregnancy urines, the diagnostic accuracy claim, and the abilities of 15 devices to detect the different forms of hCG. METHODS: We measured the concentrations of regular hCG and hyperglycosylated hCG (H-hCG, a large hCG variant) in 592 urines. Fifteen home devices were tested according to manufacturers' instructions with regular hCG and H-hCG diluted in urine. RESULTS: H-hCG was the principal hCG-related molecule in pregnancy urine in the 2 weeks following the missed menses (61% and 50% of total immunoreactivity in the 4th and 5th completed weeks of pregnancy, respectively). Of 15 home test devices, 2 had a detection limit of 6.3 IU/L for regular hCG, but poorer detection of H-hCG. Two devices detected 13 IU/L regular hCG, one with similar detection and one with poorer detection of H-hCG. Ten devices detected 25 IU/L regular hCG, 6 with poorer detection of H-hCG. One device detected 50 IU/L regular hCG, but better detected H-hCG. Overall, 9 of 15 devices did not detect H-hCG as well as regular hCG. CONCLUSIONS: H-hCG is the principal hCG immunoreactivity in early pregnancy urine. Home tests vary widely in detection limits for regular hCG (6.3-50 IU/L), and 9 of 15 devices (60%) had poorer detection limits for H-hCG than for hCG. The variation in analytical detection limits appears contradictory to the common claim for all devices of >99% detection of pregnancy on the first day of the missed menses or earlier. We suggest that manufacturers calibrate devices for both hCG and H-hCG and determine the detection rates for pregnancy rather than the proportion of positive results at arbitrary hCG concentrations.

High levels of hCG in choriocarcinoma can result in renal failure and a false-negative pregnancy test in men.

The diagnosis of choriocarcinoma was delayed because a urinary pregnancy test for hCG was falsely negative. This man's hCG rose to 25,000,000 iu/l. He developed renal impairment, probably due to precipitation of hCG in his renal rubules.

Utility of commonly used commercial human chorionic gonadotropin immunoassays in the diagnosis and management of trophoblastic diseases.

BACKGROUND: Patients with trophoblastic diseases produce ordinary and irregular forms of human chorionic gonadotropin (hCG; e.g., nicked hCG, hCG missing the beta-subunit C-terminal segment, hyperglycosylated hCG, and free beta subunit) that are recognized to differing extents by automated immunometric hCG (or hCG beta) assays. This has led to low or false-negative results and misdiagnosis of persistent disease. False-positive hCG immunoreactivity has also been detected, leading to needless therapy for trophoblastic diseases. Here we compare seven commonly used hCG assays. METHODS: Standards for five irregular forms hCG produced in trophoblastic diseases, serum samples from 59 patients with confirmed trophoblastic diseases, and serum samples from 12 women with previous false-positive hCG results (primarily in the Abbott AxSYM assay) were blindly tested by commercial laboratories in the Beckman Access hCG beta, the Abbott AxSYM hCG beta, the Chiron ACS:180 hCG beta, the Baxter Stratus hCG test, the DPC Immulite hCG test, the Serono MAIAclone hCG beta tests, and in the hCG beta RIA. RESULTS: Only the RIA and the DPC appropriately detected the five irregular hCG standards. Only the Beckman, DPC, and Abbott assays gave results similar to the RIA in the patients with confirmed trophoblastic diseases (values within 25% of RIA in 49, 49, and 54 of 59 patients, respectively). For samples that were previously found to produce false-positive hCG results, no false-positive results were detected with the DPC and Chiron tests (5 samples, median <2 IU/L), but up to one-third of samples were false positive (>10 IU/L) in the Beckman (1 of 5), Serono (2 of 9), and Baxter assays (1 of 5), and the hCG beta RIA (3 of 9; median for all assays, <5 IU/L). These samples, which produced false-positive results earlier in the Abbott AxSYM assay, continued to produce high values upon reassessment (median, 81 IU/L). CONCLUSIONS: Of six frequently used hCG immunometric assays, only the DPC detected the five irregular forms of beta hCG, agreed with the RIA, and avoided false-positive results in the samples tested. This assay, and similarly designed assays not tested here, seem appropriate for hCG testing in the diagnosis and management of trophoblastic diseases.

Mice devoid of fer protein-tyrosine kinase activity are viable and fertile but display reduced cortactin phosphorylation.

The ubiquitous Fer protein-tyrosine kinase has been proposed to regulate diverse processes such as cell growth, cell adhesion, and neurite outgrowth. To gain insight into the biological function of Fer, we have targeted the fer locus with a kinase-inactivating missense mutation (fer(D743R)). Mice homozygous for this mutation develop normally, have no overt phenotypic differences from wild-type mice, and are fertile. Since these mice lack both Fer and the testis-specific FerT kinase activities, these proteins are clearly not essential for development and survival. No differences were observed in overall cellularity of bone marrow, spleen, or thymus in the absence of Fer activity. While most platelet-derived growth factor (PDGF)-induced tyrosine phosphorylation was unchanged in fer(D743R) homozygous embryonic fibroblasts, cortactin phosphorylation was reduced. However, Fer kinase activity was not required for PDGF-induced Stat3, p120(ctn), or epidermal growth factor (EGF)-induced beta-catenin phosphorylation. Also, no defects were observed in changes to the actin cytoskeleton, adherens junctions, or focal adhesions in PDGF- or EGF-stimulated fer(D743R) homozygous embryonic fibroblasts. Therefore, Fer likely serves a redundant role in regulating cell growth, cell adhesion, retinal development, and spermatogenesis but is required for efficient phosphorylation of cortactin.

Urinary hyperglycosylated hCG in first trimester screening for chromosomal abnormalities.

Hyperglycosylated human chorionic gonadotrophin (H-hCG), also known as Invasive Trophoblast Antigen or ITA, is a unique metabolic variant of hCG with more complex oligosaccharide side chains. Concentrations are independent of regular hCG. Urine H-hCG has recently proved to be a highly sensitive marker for Down syndrome screening in the second trimester of pregnancy. We evaluated H-hCG as a potential marker in the first trimester of pregnancy. Maternal urine samples were collected from 10(+0) to 11(+6) weeks of gestation prior to genetic analysis and stored in frozen form. Samples from eight cases of Down syndrome, two cases of trisomy 13, one case of trisomy 18, and 55 control pregnancies were hand-carried frozen to the USA and tested blindly. Samples were tested in a specific H-hCG immunoassay and values were normalized to creatinine concentration. Values were plotted against gestational age, and multiples of control pregnancy median (MoM) calculated. The median level of the MoMs of the eight Down syndrome cases was 3.6 MoM. Five of the eight Down syndrome cases exceeded the 90th centile of the 55 unaffected cases. The MoMs of the trisomy 13 and 18 pregnancies were 0.2, 0.2 and 0.3. All three cases were under the 10th centile of unaffected pregnancies. The results of this study indicate that H-hCG testing may be useful in screening for Down syndrome in the first trimester of pregnancy. Further studies are needed to assess the potential screening values of urine H-hCG and the combination of this test with free beta-subunit, PAPP-A and other markers for Down syndrome in the first trimester of pregnancy.

Comparison of urinary hyperglycosylated human chorionic gonadotropin concentration with the serum triple screen for Down syndrome detection in high-risk pregnancies.

OBJECTIVE: Both modest screening performance and declining patient and physician acceptance have stimulated interest in alternative markers to the triple screen for the detection of Down syndrome. Our purpose was to compare the concentration of a single urinary analyte, hyperglycosylated human chorionic gonadotropin, with the serum triple screen (alpha-fetoprotein, human chorionic gonadotropin, and unconjugated estriol concentrations combined with age) for second-trimester Down syndrome detection. STUDY DESIGN: Urine and blood were obtained from pregnant women in the second trimester undergoing genetic amniocentesis. Urinary hyperglycosylated human chorionic gonadotropin concentration and serum triple-screen values were measured. Individuals undergoing amniocentesis because of abnormal triple-screen results were excluded. Individual Down syndrome risks on the basis of urinary hyperglycosylated human chorionic gonadotropin concentration plus maternal age and on the basis of the triple-screen results were calculated. For each algorithm the sensitivity and false-positive rate for Down syndrome detection at different risk thresholds were determined. From these values receiver operating characteristic curves were constructed, and the area under the curve was determined for each algorithm. Finally, the performance of a new combination in which urinary hyperglycosylated human chorionic gonadotropin concentration replaced serum human chorionic gonadotropin concentration in the triple screen was ascertained. RESULTS: We studied 24 pregnancies complicated by Down syndrome and 500 unaffected pregnancies between 14 and 22 weeks' gestation in a mostly white (93.5%) population undergoing amniocentesis primarily because of advanced maternal age. The sensitivity and false-positive rate for urinary hyperglycosylated human chorionic gonadotropin concentration were 75. 0% and 5.6%, respectively, whereas those for the triple screen were 75.0% and 33.2%, respectively. Urinary hyperglycosylated human chorionic gonadotropin concentration was superior to the triple screen (area under the curve, 0.9337 vs 0.7887; P =.02). The substitution of urinary hyperglycosylated human chorionic gonadotropin concentration for serum human chorionic gonadotropin concentration in the triple screen resulted in a 91.7% sensitivity at a 10.0% false-positive rate, versus a 54.2% sensitivity for the traditional triple screen at the same false-positive rate. CONCLUSION: The performance of urinary hyperglycosylated human chorionic gonadotropin concentration was statistically superior to that of the serum triple screen in a high-risk population. The use of urinary hyperglycosylated human chorionic gonadotropin concentration as an alternative test or substitution of this measurement for serum human chorionic gonadotropin concentration in the triple screen would improve diagnostic accuracy and address many current concerns related to the triple screen.

Bioterrorism threats: learning from inappropriate responses.

Between April 1997 and June 1999, some 200 mailed or telephoned bioterrorism threats were received at a variety of locations. Usually claiming that anthrax had been released, the threats all proved to be hoaxes. In many instances, local emergency responders treated the more than 13,000 potential victims inappropriately, in particular requiring victims to strip and undergo decontamination with bleach solutions. Narratives of several incidents indicated that many victims were distressed and embarrassed by their treatment. Their experiences underscore the need for improved local response actions and the formulation of a uniform response protocol for public health agencies.

False diagnosis and needless therapy of presumed malignant disease in women with false-positive human chorionic gonadotropin concentrations.

BACKGROUND: 12 women were diagnosed of having postgestational choriocarcinoma on the basis of persistently positive human chorionic gonadotropin (hCG) test results in the absence of pregnancy. Most of the women had extirpative surgery or chemotherapy, or both, without significant diminution in hCG titre. Our aim was to assess whether the hCG concentrations were false-positive test results. METHODS: Samples were tested for hCG, hCG free beta subunit, and hCG beta-core fragment. Assay kinetics were also assessed, and samples were tested independently by competitive RIA. False-positive hCG concentrations were identified by two criteria: detection of hCG in serum and lack of detection of hCG and its degradation products in urine; and wide variations in results for different hCG assays. We corroborated false-positive hCG values by the lack of parallel changes in hCG results when serum was diluted, by false detection of other antigens, and by failure to detect hCG with in-house assays. FINDINGS: All 12 women met both criteria for false-positive hCG, and all had corroborating findings. In all 12 cases, a false diagnosis had been made, and most of the women had been subjected to needless surgery or chemotherapy. Assay kinetics indicated that heterophilic antibodies were responsible for the false-positive results. As a result of our findings all further therapy was stopped. INTERPRETATION: Current protocols for the diagnosis and treatment of choriocarcinoma should be modified to include a compulsory test for hCG in urine.

Hyperglycosylated human chorionic gonadotropin (invasive trophoblast antigen) immunoassay: A new basis for gestational Down syndrome screening.

BACKGROUND: Serum human chorionic gonadotropin (hCG) and hCG free beta-subunit tests are used in combination with unconjugated estriol and alpha-fetoprotein in the triple screen test, and with the addition of inhibin-A in the quadruple marker test for detecting Down syndrome in the second trimester of pregnancy. These tests have a limited detection rate for Down syndrome: approximately 40% for hCG or free beta-subunit alone, approximately 60% for the triple screen test, and approximately 70% for the quadruple marker test, all at 5%, or a relatively high, false-positive rate. New tests are needed with higher detection and lower false rates. Hyperglycosylated hCG (also known as invasive trophoblast antigen or ITA) is a new test. It specifically detects a unique oligosaccharide variant of hCG associated with Down syndrome pregnancies. We evaluated this new Down syndrome-directed test in prenatal diagnosis. METHODS: Hyperglycosylated hCG was measured in urine samples from women undergoing amniocentesis for advanced maternal age concerns at 14-22 weeks of gestation, 1448 with normal karyotype and 39 with Down syndrome fetuses. RESULTS: The median hyperglycosylated hCG value was 9.5-fold higher in Down syndrome cases (9.5 multiples of the normal karyotype median). The single test detected 80% of Down syndrome cases at a 5% false-positive rate. Urine hyperglycosylated hCG was combined with urine beta-core fragment (urine breakdown product of serum hCG free beta-subunit), serum alpha-fetoprotein, and maternal age-related risk. This urine-serum combination detected 96% of Down syndrome cases at a 5% false-positive rate, 94% of cases at a 3% false-positive rate, and 71% of cases at a 1% false-positive rate. These detection rates exceed those of any previously reported combination of biochemical markers. CONCLUSIONS: Hyperglycosylated hCG is a new base marker for Down syndrome screening in the second trimester of pregnancy. The measurement of hyperglycosylated hCG can fundamentally improve the performance of Down syndrome screening protocols.

Maternal urine hyperglycosylated hCG in pregnancies with Down syndrome.

Stored maternal urine samples were used to determine the distribution of hyperglycosylated human chorionic gonadotrophin (H-hCG) levels in pregnancies with Down syndrome. A total of 349 samples from singleton pregnancies, including 45 with Down syndrome, were tested at 10-19 weeks' gestation. Urinary concentration was allowed for by expressing H-hCG in ng per mmol creatinine. The median level in Down syndrome was 3.63 multiples of the gestation-specific median in unaffected pregnancies (p<0.0001, Wilcoxon rank-sum test, two-tail). However, creatinine levels were relatively low in cases and creatinine did not fully correct for concentration. When this bias was allowed for, the median level was 3.34 multiples of the normal median (MoM). The H-hCG elevation in affected pregnancies was more marked at 14 weeks' gestation or later: a median of 4.64 MoM and allowing for creatinine bias 4.46 MoM.

Urinary screening tests for fetal Down syndrome: I. Fresh beta-core fragment.

Variable results have been reported using urine beta-core fragment as a marker for fetal Down syndrome. Initial studies by Cuckle et al. (1994) and Canick et al. (1995) indicated that beta-core fragment was an outstanding marker, detecting >80 per cent of Down syndrome cases. Since these reports, widely varying results have been published, indicating between 20 per cent and 66 per cent detection of cases at 5 per cent false-positive rate. The wide variation in the reported data has led to a loss of enthusiasm for this marker as a useful test for Down syndrome screening. Here we report the results of a three-year prospective study in which urine samples were collected daily from women undergoing fetal karyotype analysis for advanced maternal age. Samples were tested within one week of collection and then frozen. We also investigated the likely causes of the variability observed in beta-core fragment data. We collected 1157 urine samples over 955 days. Beta-core fragment levels were measured. A regression line was calculated for the weekly medians of the 1134 control samples and multiples of the control median (MoM) were determined. The median MoM for the controls was 1.0 and the logarithmic standard deviation (log SD) was 0.41. The median MoM for the 23 Down syndrome cases was 5.44 and the log SD was 0.45. Over the study period, 65 per cent of Down syndrome cases exceeded the 95th centile of the control group. The median MoM of control samples and the proportion of Down syndrome cases detected by the test was relatively constant during the study period. The unaffected cases were divided into three equal divisions, corresponding to approximately the first, second and third year of sample collection. No trend was found in the median control MoM values in three sample collection periods (r2=0.04). A similar number of cases exceeded the 95th centile of control samples in the three sample collection periods, 63 per cent, 66 per cent and 66 per cent. Consistent results were indicated during the three years of sample testing. Levels of total oestriol were determined in urine samples and MoM statistics derived. The median oestriol level in Down syndrome cases was 0.59 MoM. Only 12 per cent of cases had MoM levels below the fifth centile. Gaussian models were prepared combining biochemical data and maternal age distribution. While beta-core fragment by itself detected 65 per cent of Down syndrome cases, beta-core fragment modelled with maternal age detected 66 per cent, and modelled with age and total oestriol levels detected 82 per cent of cases at 5 per cent false-positive rate. At the completion of the study, we thawed and reassayed 20 random urine samples (10 control and 10 Down syndrome) collected at different times during the study period. While the control samples (74-1700 ng/ml) had slightly increased values when reassayed (mean value 137 per cent of original prospective value), the Down syndrome samples (360-20,500 ng/ml) all had decreased values when reassayed (mean=53 per cent, t-test, controls versus cases, p = 0.0003). The Down syndrome samples were decreased to between 93 per cent and 12 per cent of the original value. A relationship was identified between the magnitude of the original beta-core fragment value and the change in immunoreactivity when reassayed (r2=0.998). The higher the initial beta-core fragment value the greater the loss of immunoreactivity. We considered the possibility that the beta-core fragment molecules aggregate upon storage in the freezer. We repeated the assay of the 20 samples after treatment with a high salt buffer. Down syndrome samples recovered half of the lost beta-core fragment immunoreactivity (mean increase in beta-core fragment levels 56 per cent, t-test, controls versus cases, p=0.004). We infer that aggregation of beta-core fragment upon storage interferes with beta-core fragment measurements. This may be the cause of the poor beta-core fragment screening performance reported using sto