ADAM12: a novel first-trimester maternal serum marker for Down syndrome

OBJECTIVES: The concentration of bioavailable insulin-like growth factor (IGF) I and II is important to foetal growth. It is regulated by insulin-like growth factor binding proteins (IGFBP) 1 through 6. Proteolytic cleavage of IGFBP-3 takes place in human pregnancy serum; accordingly, IGFBP-3 serum levels decrease markedly during pregnancy. ADAM12 (A disintegrin and metalloprotease) is an IGFBP-3 and IGFBP-5 protease and is present in human pregnancy serum. The goal of this study was to determine whether ADAM12 concentration in maternal serum is a useful indicator of foetal health. METHODS: We developed an enzyme-linked immunosorbent assay (ELISA) for the quantification of ADAM12 in serum. The assay range was 42 to 667 micro g/L. Recombinant ADAM12 was used as the standard for calibration. RESULTS: We found that ADAM12 was highly stable in serum. Serum concentration increased from 180 micro g/L at week 8 of pregnancy to 670 micro g/L at 16 weeks, and reached 12 000 micro g/L at term. In 18 first-trimester Down syndrome pregnancies, the concentration of ADAM12 was decreased, thus the median multiple of mean (MoM) value was 0.14 (0.01-0.76). A detection rate for foetal Down syndrome of 82% for a screen-positive rate of 3.2% and a 1:400 risk cut-off was found by Monte Carlo estimation using ADAM12 and maternal age as screening markers. CONCLUSION: ADAM12 is a promising marker for Down syndrome.     

ADAM 12 as a first-trimester maternal serum marker in screening for Down syndrome

BACKGROUND: A Disintegrin And Metalloprotease 12 (ADAM 12) is a glycoprotein synthesised by placenta and it has been shown to be a potential first-trimester maternal serum marker for Down syndrome (DS) in two small series. Here we analyse further, the potential of ADAM 12 as a marker for DS in a large collection of first-trimester serum samples. MATERIALS AND METHODS: The concentration of ADAM 12 was determined in 10-14-week pregnancy sera from 218 DS pregnancies and 389 gestational age-matched control pregnancies, which had been collected as part of routine prospective first-trimester screening programs (DS = 105) or as part of previous research studies (DS = 113). ADAM 12 was measured using a semi-automated time resolved immunofluorometric assay and median values for normal pregnancies were established by polynomial regression. These medians were then used to determine population distribution parameters for DS and normal pregnancy groups. Correlation with previously established PAPP-A and free beta-hCG multiple of the medians (MoMs) and delta nuchal translucency (NT) were determined and used to model the performance of first-trimester screening with ADAM 12 in combination with other first-trimester markers at various time periods across the first trimester. The benefits of a contingent testing model incorporating early measurement of PAPP-A and ADAM 12 were also explored. RESULTS: The maternal serum concentration of ADAM 12 was significantly reduced (p = 0.0049) with an overall median MoM of 0.79 in the DS cases and a log(10) MoM SD of 0.3734 in the DS cases and 0.3353 in the controls. There was a significant correlation of ADAM 12 MoM in DS cases with gestational age (r = 0.375) and the median MoM increased from 0.50 at 10-11 weeks to 1.38 at 13 weeks. ADAM 12 was correlated with maternal weight (r(controls) = 0.283), PAPP-A (r(controls) = 0.324, r(DS) = 0.251) but less so with free beta-hCG (r(controls) = 0.062, r(DS) = 0.049) and delta NT (r(controls) = 0.110, r(DS) = 0.151). ADAM 12 was significantly (p = 0.026) lower in smokers (0.87 vs 1.00) and elevated in Afro-Caribbean women compared to Caucasian women (1.34 vs 1.00).Population modelling using parameters from this and an earlier study showed that a combination of ADAM 12 and PAPP-A measured at 8-9 weeks and combined with NT and free beta-hCG measured at 12 weeks could achieve a detection rate of 97% at a 5% false-positive rate or 89% at a 1% false-positive rate. PAPP-A and ADAM 12 alone at 8-9 weeks could identify 91% of cases at a 5% false-positive rate. Using this as part of a contingent-screening model to select an intermediate risk group of women for NT and free beta-hCG at 11-12 weeks would enable the detection of 92% of cases with a 1% false-positive rate at a cost of providing NT and free beta-hCG for 6% of women with 94% of women having completed screening by the 10th week of pregnancy. CONCLUSION: ADAM 12 in early first trimester is a very efficient marker of DS. In combination with existing markers, it offers enhanced screening efficiency in a two-stage sequential first-trimester screening program or in a contingent-screening model, which may have benefits in health economies where universal access to high quality ultrasound is difficult. More data on early first-trimester cases with DS are required to establish more secure population parameters by which to assess further the validity of these models.     

ADAM 12 as a second-trimester maternal serum marker in screening for Down syndrome

BACKGROUND: ADAM 12 is a placenta-derived glycoprotein that is involved in growth and differentiation. The maternal serum concentration of ADAM 12 is a potential first-trimester maternal serum marker of Down syndrome (DS). Here we examine the potential of ADAM 12 as a second-trimester maternal serum marker of DS. MATERIALS AND METHODS: The concentration of ADAM 12 was determined in gestational week 14-19 in 88 DS pregnancies and 341 matched control pregnancies. Medians of normal pregnancies were established by polynomial regression and the distribution of log(10) MoM ADAM 12 values in DS pregnancies and controls determined. Correlations with alpha-fetoprotein (AFP) and free beta-human chorionic gonadotrophin (free beta-hCG) were established and used to model the performance of maternal serum screening with ADAM 12 in combination with other second-trimester serum markers. RESULTS: The ADAM 12 maternal serum concentration was significantly increased with a median MoM of 1.85 and a mean log(10) MoM (SD) of 0.268 (0.2678) compared to a mean log(10) MoM (SD) of 0.013 (0.4318) in controls. ADAM 12 correlated with maternal weight and ethnicity (with the serum concentration increased in Afro-Caribbeans), but neither with maternal age nor gestational age, and only marginally with AFP (r(DS) = 0.078, r(controls) = 0.093) and free beta-hCG (r(DS) = 0.073, r(controls) = 0.144. The increase in detection rate-for a false positive rate of 5%--by adding ADAM 12 to the double test (AFP + free beta-hCG) was 4%, similar to that of adding uE3 to the double test. CONCLUSION: ADAM 12 is an efficient second-trimester marker for DS. Further studies should be conducted to determine whether it may be a useful additional or alternative marker to those currently used in the second-trimester.     

Human placental lactogen is a first-trimester maternal serum marker of Down syndrome

BACKGROUND: Human placental lactogen (hPL) is synthesised by the placenta and found in maternal serum. We analysed the potential of hPL as a first-trimester maternal serum-screening marker for fetal Down syndrome (DS). MATERIALS AND METHODS: hPL was quantified by ELISA in 47 DS pregnancies and 136 controls in gestational weeks 8-13. Distributions of log multiples of the median (MoMs) were established. The quantity of hPL in DS screening was estimated using Monte Carlo simulation methods. RESULTS: The mean log10 MoM hPL was - 0.1995 (SD: 0.1993) in affected and 0.0026 (SD: 0.2129) in control pregnancies. This corresponds to a MoM of 0.63 in DS pregnancies. hPL correlated significantly with log10 MoM values of hCGbeta (r = 0.320) and PAPP-A (r = 0.590) in controls, but not with hCGbeta (r = 0.228) or PAPP-A (r = 0.090) in DS pregnancies. The inclusion of hPL in the double test (PAPP-A + hCGbeta) increased the detection rate from 67 to 75% for a false-positive rate of 5%. CONCLUSION: hPL is a DS screening marker that is applicable at weeks 9-13 and could be included in multiple marker first-trimester screening for DS.     

Down syndrome maternal serum marker screening after 18 weeks' gestation

Women having access to prenatal care late in pregnancy may still wish to benefit from maternal serum screening for Down syndrome. Therefore, we established reference values for alpha-feto protein (AFP) and free beta-human chorionic gonadotrophin (beta-hCG), and assessed the diagnostic value of maternal serum marker screening at 18-35 weeks' gestation based upon a series of 4072 sera from unaffected pregnancies and 118 sera from pregnant women with fetuses affected by Down syndrome. Using a 1/250 risk cut-off, a detection rate of 72.9% (95% CI = 71.5-74.3%) was achieved with a false-positive rate of 7.51% (95% CI = 6.71-8.3%). This was not significantly different from the percentages observed in our 14-17 weeks routine screening (50 596 patients): 71.9% (95% CI = 71.5-72.3%) and 6.48% (95% CI = 6.28-6.68%), respectively. Detection and screen-positive rates were, respectively, 51.3% (95% CI = 35.6-67.0%) and 5.95% (95% CI = 5.12-6.68%) in women aunder 35 years of age, and 84.8% (95% CI = 76.9-92.7%) and 24% (95% CI = 20.7-27.3%) in women aged 35 years and over. In conclusion, maternal serum marker screening is feasible at 18 weeks' gestation and later, which may be of interest in selected cases.     

Medically assisted reproduction and second-trimester maternal serum marker screening for Down syndrome

OBJECTIVES: To evaluate the effect of in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) on total hCG, free ss-hCG, AFP and unconjugated estriol (uE3) used as markers for second-trimester Down syndrome maternal serum screening. METHODS: Second-trimester maternal sera from 1515 singleton pregnancies (970 by IVF, 545 by ICSI) were compared with control sera (21 014 cases). Free ss-hCG, total hCG, AFP and uE3 were compared between the control group and the medically assisted reproduction groups. The percentages of at-risk patients (>/=1/250) were also compared. RESULTS: No differences in values of the maternal serum markers were observed between the medically assisted and control groups. When maternal age was taken into account, the screen-positive rate for Down syndrome screening did not differ between the two groups. CONCLUSION: Patients undergoing assisted reproduction techniques can be counseled for maternal serum Down syndrome screening with the same efficacy as patients with naturally conceived pregnancies.     

First-trimester maternal serum biochemical indicators in Down syndrome.

A set of 21 early maternal serum samples (19 first-trimester and two at 14 weeks) from pregnancies resulting in a child with Down syndrome was matched for gestation and length of storage with 63 samples from unaffected pregnancies. The concentrations of alpha-fetoprotein (AFP), unconjugated oestriol (uE3), human chorionic gonadotrophin (hCG), pregnancy-specific beta 1-glycoprotein (SP1), and placental alkaline phosphatase (PALP) were measured. The ratios of the medians for Down syndrome pregnancies compared with the medians for controls were AFP 0.71, uE3 0.67, hCG 1.43, SP1 0.79, and PALP 0.92. Although the differences between the medians for affected and unaffected pregnancies were not significant, the trends for AFP, uE3, and hCG confirm earlier findings on first-trimester samples.     

Weight-correction formula for maternal serum screening for Down syndrome in Taiwan.

BACKGROUND AND PURPOSE: This study examined the relationship between maternal weight and serum marker concentrations (alpha-fetoprotein [AFP] and human chorionic gonadotropin [hCG]) to develop a weight-correction formula for second-trimester Down syndrome screening in Taiwan. METHODS: We conducted a prospective observational study based on data from 18, 016 pregnant Taiwanese women who participated in a second-trimester Down syndrome screening program at Taiwan Adventist Hospital. The relationship between maternal weight and the multiple of median (MoM) concentrations of serum markers was analyzed using two methods; one involved all the observational data (all point method), while the other involved only the median concentrations at various gestational ages (median regression method). In each method, several statistical approaches were used, including simple-linear, reciprocal, quadratic, and log-transformation linear regression. RESULTS: The correlation between maternal body weight and serum marker concentrations using the all point regression method was very poor, while the median regression method achieved a good fit. The reciprocal regression analysis had the best fit between AFP MoM concentration and maternal weight, while the best fit between hCG MoM concentration and maternal weight was achieved with quadratic regression analysis. CONCLUSION: The present study has developed a race-specific weight-correction formula for Taiwanese women. This formula is expected to be helpful in second-trimester Down syndrome screening programs in Taiwan.     

Second-trimester maternal serum immunoreactive inhibin as a marker for fetal Down's syndrome.

We measured immunoreactive inhibin in the maternal serum of 80 pregnancies with a chromosomally normal fetus and ten Down's syndrome pregnancies in the second trimester. The inhibin level in all Down's syndrome pregnancies was above the normal median; the multiple of the normal median (MoM) was 1.9. We found a statistically significant difference between the levels of inhibin in unaffected and affected pregnancies (Kolmogorov-Smirnov test: p < 0.002). Using an arbitrarily chosen cut-off of 2.4 MoM, 40 per cent of Down's syndrome and 5 per cent of the normal pregnancies were found. We conclude that immunoreactive inhibin may be useful as a marker for fetal Down's syndrome.     

Screening for fetal Down syndrome with maternal serum hCG and oestriol: a prospective study.

We report the preliminary results of a prospective study aimed at evaluating the effectiveness of Down syndrome (DS) screening using second-trimester measurement of maternal serum human chorionic gonadotrophin (hCG) and unconjugated oestriol (uE3) together with maternal age. Reference values for hCG, uE3, and the hCG/uE3 ratio in normal pregnancies were established from more than 3000 normal gestations and found to follow a log-normal statistical distribution. Risk evaluation was made using reference values for affected pregnancies from retrospective studies. Screening of 10,000 women under 38 years resulted in 412 amniocenteses and the prenatal diagnosis of six cases of DS, whereas four cases remained undetected until term. In a parallel study, diagnostic amniocentesis was performed in women over 38 years and in women with a previous affected child, and an evaluation of the risk of fetal DS based on serum hCG and uE3 levels was made in all cases. Fourteen cases of DS were detected. Median values for hCG and uE3 in the 24 affected pregnancies were close to the 90th and tenth centiles of the normal reference values, respectively, and thus are in good agreement with the values reported by others in retrospective studies.     

Optimal risk cut-offs for Down syndrome contingent maternal serum screening

Objectives: The objective of this study is to identify the optimal cut-off points of contingent serum screening excluding nuchal translucency (NT) measurement, to categorize the risk level in the first trimester.

Methods: A prospective database of women undergoing contingent serum screening, without NT measurement, was reviewed. In conventional categorization, the results of first-trimester screening were categorized into high risk (>1:30) (invasive diagnosis was offered); intermediate risk (1:30–1:1500) (second-trimester screening was needed); and low risk (<1:1500) (no further test). We recategorized the risk levels using various upper and lower cut-offs and compared detection rates, false-positive rates, and rates of intermediate risk.

Results: Among 24,874 women, the prevalence of Down syndrome was 1:691. The previously agreed cut-offs had a detection rate of 88.9% and a false-positive rate of 8.5% with high rate of intermediate risk (38.2%). Re-categorization provided the optimal lower and upper cut-offs 1/900 and 1/50, respectively, giving a detection rate of 86.1%, a false-positive rate of 8.1%, and a rate of intermediate risk of 24.8%.

Conclusions: This is the first study on contingent serum screening without NT measurement which shows a high detection rate with an acceptable false-positive rate. The optimal cut-offs to categorize the risk levels of the upper and the lower cut-off was 1:30–1:50 and 1:900, respectively.     

Plasma levels of eosinophil granule major basic protein in pregnant cynomolgus monkeys.

The sera of all pregnant women contain increased amounts of a protein biochemically and immunologically similar to the eosinophil granule major basic protein (MBP). Immunofluorescence shows that the pregnancy-associated MBP is localized to placental trophoblastic cells. This information raises important questions about the function of pregnancy-associated MBP because of the potential biological functions attributed to its eosinophil counterpart (namely, its potent toxic and cytostimulatory activities). Previous studies demonstrated the presence of an immunologically cross-reactive protein in the placentae and plasma of pregnant non-human primates. Here, plasma MBP levels were measured throughout gestation in cynomolgus monkeys. In early pregnancy, only modest increases in MBP were found in contrast to the sharp rise observed in the first 20 weeks of human pregnancy. During the final one-third of gestation, striking increases in plasma MBP occurred in the monkeys. This parallels the late rise in MBP seen in humans in the third trimester. Thus, the cynomolgus monkey may serve as a model to clarify the role of the MBP in the biochemical events that culminate in parturition.     

Second-trimester maternal serum inhibin-A screening for fetal Down syndrome in Asian women.

Case control studies in the Caucasian population showed that maternal serum inhibin-A is elevated in Down syndrome pregnancies and may be a useful second-trimester marker in addition to human chorionic gonadotrophin (hCG) and alpha-fetoprotein (AFP). Data in the Asian population are lacking. We measured inhibin-A levels in the stored maternal sera of 49 Down syndrome pregnancies and 341 controls with a commercially available assay and expressed them as the multiples of the median of the gestational week. The log means and standard deviations for case and control inhibin-A MOMs were 0.209, 0.226, and 0.002 and 0.177, respectively. Median inhibin-A MOM in Down syndrome cases was elevated to 1.62 (95 per cent confidence interval, 1.29-1.82). 36 per cent of Down syndrome cases were expected to be detected at a 5 per cent false-positive rate. However, inhibin-A MOMs were strongly correlated with hCG MOMs in the cases (r=0.73, p<0.001) and the controls (r=0.56, p<0.001). This will diminish the value of adding inhibin-A to the existing hCG and AFP screening protocol.     

Chinese weight-correction model for maternal serum markers in Down syndrome prenatal screening.

OBJECTIVE: To determine the best weight-correction model by means of analyzing the relationship between maternal weight and maternal serum markers when screening for Down syndrome in China. METHODS: Serum levels of alpha-fetoprotein (AFP) and free beta-human chorionic gonadotropin (hCG) were measured in 35,917 Chinese women during the second semester of a normal singleton pregnancy and converted to multiple of median (MoM) values. Using 2 methods of statistical analysis, the all-point method and the median regression method, 4 weight-correction models were then tried, the simple linear, reciprocal, quadratic, and log-linear regression models. RESULTS: The median regression method performed better than the all-point method, and the quadratic regression model showed the best fit for both AFP and hCG in the median regression method, with adjusted R(2)s of 0.987 and 0.988, respectively. CONCLUSION: The quadratic regression model was found to be the most suitable for Chinese pregnant women.     

Prenatal diagnosis of trisomy 20 mosaicism by maternal serum screening for Down syndrome.

We report two cases of prenatally diagnosed trisomy 20 mosaicism associated with positive Down syndrome screening at 16 weeks' gestation. Both infants exhibited normal growth and mental development.These cases suggest that the multiple-marker screening test may play an important role in prenatal detection and diagnosis of chromosomal anomalies in addition to Down syndrome.     

Compromise ultrasound dating policy in maternal serum screening for Down syndrome

Routine ultrasound biometry is the method of choice for gestational dating when screening for Down syndrome. However, it is costly and an alternative policy is to restrict ultrasound to women most likely to have menstrual dating errors. This was evaluated by statistical modelling with parameters from 14,274 women screened between January 1997 and July 2001 using free beta-human chorionic gonadotrophin (free beta-hCG), alpha-fetoprotein (AFP) and unconjugated estriol (uE(3)). A total of 12,711 (89%) women had both ultrasound and menstrual gestations, but in 4101 (29%) women either the last menstrual period (LMP) was uncertain or a pill-withdrawal period, or there were irregular or abnormal length cycles. The LMP was not entered in the test request form for a further 1404 (9.8%) women. Routine ultrasound dating yielded a predicted detection rate higher than for menstrual dating by 3.9-7.1%, depending on the marker combination and cut-off. The false-positive rate was reduced by 0.2-1.1%. Selectively scanning the 39% with unreliable dates increased detection by 2.6-4.6%, and reduced the false-positive rate by 0.04-0.6%. Some centres only use the ultrasound estimate of gestation when it differs from the menstrual estimate by more than 7 days. Such a rule reduces the gain in detection rate to 2.5-4.6% for routine ultrasound and 1.7-3.1% with the compromise policy; the false-positive rate reductions are 0.06-0.6% and 0.0-0.3%, respectively. We conclude that if routine ultrasound is not financially and practically feasible, the compromise policy yields a clinically important improvement in screening performance compared to menstrual dating.     

Down syndrome screening in women under 35 with maternal serum hCG.

Human chorionic gonadotropin and maternal serum alpha-fetoprotein (MSAFP) concentrations were measured in frozen maternal serum samples from 16 pregnancies with Down syndrome and from 614 unaffected pregnancies in women younger than 35. By using only maternal age and MSAFP level to determine Down syndrome risk, four of 16 (25%) Down syndrome pregnancies were identified, using risk estimates as a screening variable. This detection rate required performing amniocentesis on 4.8% of pregnancies with a normal outcome. By adding hCG level as a third risk parameter, ten of 16 (62.5%) Down syndrome pregnancies were detected. To achieve this detection rate, 4.7% of women under 35 would be recommended for amniocentesis. These results indicate that the estimation of risk for Down syndrome based on the addition of maternal hCG level to MSAFP level and maternal age will substantially improve the detection rate for Down syndrome, with no change in the amniocentesis rate. These findings are in agreement with other studies that suggest that hCG is a valuable addition to screening programs for Down syndrome.     

Prenatal screening for Down syndrome with maternal serum human chorionic gonadotropin levels

Human chorionic gonadotropin levels in midtrimester pregnancies may be predictive of Down syndrome. A commercially available enzyme immunoassay kit was used to measure the beta-subunit of human chorionic gonadotropin in maternal sera from 38 Down syndrome pregnancies and 114 gestational age matched controls. The human chorionic gonadotropin levels were also assayed in 236 normal sera and plasma samples to determine normative values and appropriate individual corrections. Serum and plasma human chorionic gonadotropin levels are closely correlated and are stable at room temperature, during refrigeration, and throughout freeze-thaw cycles. There is no correlation between the human chorionic gonadotropin level and maternal age, weight, or race. However, the human chorionic gonadotropin level decreases with each week of gestation from 15 to 19 weeks. Medians for each week of gestation were established to account for this variable. Up to 63% of the Down syndrome pregnancies were detected with a cutoff of 2.0 multiples of the normal median. A computational combination of human chorionic gonadotropin and maternal serum alpha-fetoprotein testing will detect additional Down syndrome pregnancies and decrease the false-positive rate. The measurement of human chorionic gonadotropin appears to be a valuable addition to maternal serum alpha-fetoprotein screening programs that can significantly increase the proportion of Down syndrome cases diagnosed.