Cell-free fetal DNA screening in the USA: a cost analysis of screening strategies.

OBJECTIVES: To determine whether implementation of primary cell-free fetal DNA (cffDNA) screening would be cost-effective in the USA and to evaluate potential lower-cost alternatives. METHODS: Three strategies to screen for trisomy 21 were evaluated using decision tree analysis: 1) a primary strategy in which cffDNA screening was offered to all patients, 2) a contingent strategy in which cffDNA screening was offered only to patients who were high risk on traditional first-trimester screening and 3) a hybrid strategy in which cffDNA screening was offered to all patients ≥ 35 years of age and only to patients < 35 years who were high risk after first-trimester screening. Four traditional screening protocols were evaluated, each assessing nuchal translucency (NT) and pregnancy-associated plasma protein-A (PAPP-A) along with either free or total beta-human chorionic gonadotropin (ß-hCG), with or without nasal bone (NB) assessment. RESULTS: Utilizing a primary cffDNA screening strategy, the cost per patient was 1017 US$. With a traditional screening protocol using free ß-hCG, PAPP-A and NT assessment as part of a hybrid screening strategy, a contingent strategy with a 1/300 cut-off and a contingent strategy with a 1/1000 cut-off, the cost per patient was 474, 430 and 409 US$, respectively. Findings were similar using the other traditional screening protocols. Marginal cost per viable case detected for the primary screening strategy as compared to the other strategies was 3-16 times greater than the cost of care for a missed case. CONCLUSIONS: Primary cffDNA screening is not currently a cost-effective strategy. The contingent strategy was the lowest-cost alternative, especially with a risk cut-off of 1/1000. The hybrid strategy, although less costly than primary cffDNA screening, was more costly than the contingent strategy.

Impact of nuchal translucency credentialing by the FMF, the NTQR or both on screening distributions and performance.

OBJECTIVE: In the USA, both The Fetal Medicine Foundation (FMF) and the Nuchal Translucency Quality Review Program (NTQR) have operated education, review and credentialing for physicians and sonographers for the measurement of nuchal translucency (NT). We sought to assess differences in the distribution of NT measurements based upon the system from which the operator obtained their education, review and credentialing. METHODS: 398 311 NT measurements by 1541 sonographers who had performed ≥ 50 exams from July 2008 to June 2010 were grouped by organization. Differences between grouped measurements were assessed using analysis of variance of log(10) NT multiples of the median (MoM), with sonographer and organization as factors. RESULTS: MoM values were significantly lower (P ≤ 0.001) and SD was significantly higher (P < 0.001) for the NTQR group compared with the FMF group or those sonographers credentialed by both. The percentage of individuals with negative bias ≥ 10% was greater for the NTQR group (P < 0.001). The difference was less but still significant (P = 0.009) when bias was adjusted for by the overall median for the organization. CONCLUSIONS: Although NT MoM measurements were significantly lower and had a wider variance when obtained by the NTQR group, our data cannot distinguish between bias in training or the attributes of the participating sonographers in each program. With these large numbers, it is unlikely that patient characteristics could explain the discrepancy in distributions. Ongoing efforts to monitor sonographer performance with remediation for poor performers may reduce discrepancies between organizations.

Anxiety among women who have undergone fertility therapy and who are considering multifetal pregnancy reduction: trends and implications.

OBJECTIVE: Our primary objective was to evaluate the assumption that women carrying multiple fetuses and who have decided upon multifetal pregnancy reduction (MFPR) have a constant high level of anxiety. METHODS: A total of 66 multigestation women considering MFPR were asked to consider how anxious they were when they first started fertility therapy. Using that level of anxiety as a reference point, and using their self-assessments as a vehicle for probing the meaning they attached to their emotional state through time, they then assessed their anxiety level at different points in their pregnancy. RESULTS: Self-reported anxiety across time displayed considerable variation: there was a large drop in anxiety with pregnancy diagnosis. The women's anxiety rose to very high levels with the diagnosis of carrying multiples. Anxiety moderated again on average with consultation, rose sharply during the course of the procedure, and finally dropped to lower levels on average after the procedure was over. CONCLUSIONS: We conclude that women with multigestation experience considerable fluctuations in their level of anxiety from the time that they first start fertility therapy until they learn that they are carrying multiple embryos. Their expectations for the future of becoming pregnant seem at last fulfilled (becoming pregnant), become complicated (with multiples), appear salvageable (with consultation), but with a morally complicated resolution (MFPR) that seems at last to have put the pregnancy back on a normal track (post-MFPR). Those working with MFPR patients before, during and after the operation must understand the nature and variability of the anxiety that their patients are confronting, and how they are attempting to construct a safe passage through the moral dilemma associated with the multiple-gestation situation.

Sonographic nuchal markers for Down syndrome are more common but less ominous in gestations with a male fetus.

OBJECTIVE: A change in the normal male-to-female ratio has been reported in some autosomal trisomies (i.e., trisomy 21 or trisomy 18). The objective of the present study was to evaluate the male-to-female ratio in pregnancies with sonographic nuchal markers for Down syndrome. METHODS: The results of amniocenteses performed for isolated nuchal markers for Down syndrome were grouped by fetal sex and by maternal age. The male-to-female ratio in normal and trisomic gestations was compared. RESULTS: 584 fetal karyotypes were available for analysis. A significantly higher male-to-female ratio was observed. More affected gestations were observed in association with a female fetus. These differences were mainly attributed to the group of patients younger than 35 years that represents more than 80% of our study population. No difference was observed in pregnancies of patients older than 35 years of age. CONCLUSIONS: In patients younger than 35 years, sonographic nuchal markers for Down syndrome are more frequent (but apparently less ominous) in gestations with a male fetus. If the gender is known, counseling can be modified to include such differential risks.

Fetal gender and aneuploidy detection using fetal cells in maternal blood: analysis of NIFTY I data. National Institute of Child Health and Development Fetal Cell Isolation Study.

OBJECTIVES: The National Institute of Child Health and Human Development Fetal Cell Isolation Study (NIFTY) is a prospective, multicenter clinical project to develop non-invasive methods of prenatal diagnosis. The initial objective was to assess the utility of fetal cells in the peripheral blood of pregnant women to diagnose or screen for fetal chromosome abnormalities. METHODS: Results of fluorescence in situ hybridization (FISH) analysis on interphase nuclei of fetal cells recovered from maternal blood were compared to metaphase karyotypes of fetal cells obtained by amniocentesis or chorionic villus sampling (CVS). After the first 5 years of the study we performed a planned analysis of the data. We report here the data from 2744 fully processed pre-procedural blood samples; 1292 samples were from women carrying singleton male fetuses. RESULTS: Target cell recovery and fetal cell detection were better using magnetic-based separation systems (MACS) than with flow-sorting (FACS). Blinded FISH assessment of samples from women carrying singleton male fetuses found at least one cell with an X and Y signal in 41.4% of cases (95% CI: 37.4%, 45.5%). The false-positive rate of gender detection was 11.1% (95% CI: 6.1,16.1%). This was higher than expected due to the use of indirectly labeled FISH probes in one center. The detection rate of finding at least one aneuploid cell in cases of fetal aneuploidy was 74.4% (95% CI: 76.0%, 99.0%), with a false-positive rate estimated to be between 0.6% and 4.1%. CONCLUSIONS: The sensitivity of aneuploidy detection using fetal cell analysis from maternal blood is comparable to single marker prenatal serum screening, but technological advances are needed before fetal cell analysis has clinical application as part of a multiple marker method for non-invasive prenatal screening. The limitations of the present study, i.e. multiple processing protocols, are being addressed in the ongoing study.

Prenatal diagnosis of a de novo ring chromosome 11.

Ring chromosomes are uncommon findings in prenatal diagnosis. Growth retardation is the most significant manifestation, in particular among patients with rings of larger chromosomes. A 30-year-old gravida 1, para 0 white woman was referred for genetic counseling because of maternal anxiety. Cytogenetic analysis of amniotic fluid cells at 16 weeks gestation revealed an abnormal mosaic female chromosome complement; 46,XX,r(11)(p15q25)[14]/45,XX,-11[7]. The ring 11 showed no detectable loss of chromosomal material at 450 band level. Both parents had a normal karyotype. Fluorescence in situ hybridization demonstrated intact subtelomeric regions in the ring chromosome. A targeted ultrasound evaluation at the time of consultation suggested no significant abnormalities. The parents were counseled and subsequently decided to terminate the pregnancy. The autopsy revealed an immature female fetus with abnormal craniofacial features including brachycephaly, low-set ears and hypertelorism, bicornuate uterus, and calcifications in the renal tubules. The abnormal phenotypes could be a consequence of the ring instability, submicroscopic deletion, and/or alteration of genetic material at the site of fusion.

Principles of screening.

Over the past several decades, the principles by which screening tests are performed have slowly been developed and refined. The key distinction for the clinician to understand is the difference between diagnostic tests, which give a definitive answer, and screening tests, which identify who among the low-risk population is at high risk.

Screening for neural tube defects.

Neural tube defects are separated into two main categories: (1) abnormalities of the skull and brain (anencephaly, acrania, and encephalocele) and (2) malformations of the spine (meningomyelocele or spina bifida). The cause of neural tube defects is not always clear, and include chromosomal abnormalities, single gene mutations, maternal disease, or maternal exposure to teratogens. Mostly the disorder emerges as a multifactorial trait. Routine screening for neural tube defects was introduced in the United Kingdom in the mid-1970s and the United States in the mid-1980s. The use of screening has resulted in a marked decline in the frequency of neural tube defects diagnosed at birth.

Second-trimester biochemical screening.

The past years have seen considerable progress in the area of biochemical screening. Increasing data have now clearly shown the advantages of multiple markers, particularly beta-hCG over AFP alone. There continues to be considerable controversy over the best mathematic algorithm and which markers are best (e.g., beta-HCG, uE3, and so forth). There seems to be a plateau of detection frequencies at about 65% to 70% with current methodologies. Further work needs to be done, however, including some new approaches, if there is to be substantial improvement of screening sensitivity. The combination of biochemical with biophysical parameters as discussed elsewhere in this issue represents the next level of sophistication in the attempt to identify the highest proportion of abnormalities with the fewest false-positives.

Future directions.

With the continued explosion of genetic technology, the number of disorders amenable to screening is expanding geometrically. Historically, most genetic screening has been in the newborn period. Much more can be done for the fetus if genetic screening and diagnosis are accomplished early in the pregnancy rather than after birth. The principal requirements to make neonatal screening disorders possible in the first trimester center around those tests that can be one on a molecular basis, and the development of fetal cell isolation from the maternal circulation. Over the course of the next decade, it is likely that many of the tests currently performed in the newborn period will be accomplished in the early or mid-gestational period.

Estrogen modulates the expression of L-arginine:glycine amidinotransferase in chick liver.

Identification of estrogen-responsive genes is important to understand the molecular mechanisms of estrogen action. Suppression subtractive hybridization was employed to screen estrogen-responsive genes in chick liver. A single injection of estrogen into 6-week-old chick induced up-regulation of several known genes encoded for yolk proteins, such as Vitellogenin I and II and very low density lipoprotein II (apo-VLDL II). One novel sequence displayed a dramatic change (3-fold increase) in response to estrogen treatment. This cDNA fragment was extended and the resultant sequence was analyzed. Translated amino acid sequence was 90, 88, 83 and 87% identical to the L-arginine:glycine amidinotransferase of pig, rat, frog and human, respectively. The sequence has a conservative catalytic site of L-arginine: glycine amidinotransferase. The expression pattern of this gene in organs is consistent with previous reports of L-arginine:glycine amidinotransferase in chick. Thus, this clone represented the chicken L-arginine:glycine amidinotransferase. It appeared that estrogen-induced alteration of arginine:glycine amidinotransferase was not dependent on protein synthesis, because concurrent administration of cycloheximide did not affect the estrogen-mediated expression pattern. This is the first study demonstrating that L-arginine:glycine amidinotransferase is a target of the estrogen receptor.

Bonding and coping with loss: examining the construction of an intervention for multifetal pregnancy reduction procedures.

Multifetal pregnancy reduction is a medical procedure by which pregnancies usually with three or more embryos are reduced in number early in gestation to improve their medical outcomes. A socially constructed intervention used as part of these procedures at Wayne State University is described. The intervention has as its goals the reduction of anxiety in the woman undergoing such a procedure and the refocusing of attention on the surviving 'twins' or singleton. Case materials from 36 cases are examined in detail to document how the intervention is being accomplished in practice, and how it may break down.

Improvement in outcomes of multifetal pregnancy reduction with increased experience.

OBJECTIVE: This study was undertaken to evaluate a decade of data on multifetal pregnancy reductions at centers with extensive experiences. STUDY DESIGN: A total of 3513 completed cases from 11 centers in 5 countries were analyzed according to year (before 1990, 1991-1994, and 1995-1998), starting and finishing numbers of embryos or fetuses, and outcomes. RESULTS: With increasing experience there has been a considerable improvement in outcomes, with decreases in rates of both pregnancy loss and prematurity. Overall loss rates in the last few years were correlated strongly with starting and finishing numbers (starting number > or =6, 15.4%; starting number 5, 11.4%; starting number 4, 7.3%; starting number 3, 4.5%; starting number 2, 6.2%: finishing number 3, 18.4%; finishing number 2, 6.0%; finishing number 1, 6.7%). Birth weight discordance between surviving twins was increased with greater starting number. The proportion of cases with starting number > or =5 diminished from 23.4% to 15.9% to 12.2%. The proportion of patients >40 years old increased in the last 6 years to 9.3%. Gestational age at delivery did not vary with increasing maternal age but was inversely correlated with starting number. CONCLUSION: Multifetal pregnancy reduction outcomes at our centers for both losses and early prematurity have improved considerably with experience. Reductions from triplets to twins and now from quadruplets to twins carry outcomes as good as those of unreduced twin gestations. Patient demographic characteristics continues to change as more older women use assisted reproductive technologies. In terms of losses, prematurity, and growth, higher starting numbers carry worse outcomes.

First-trimester embryo size discordance: a predictor of premature birth following multifetal pregnancy reduction.

OBJECTIVE: We have previously reported a correlation between the starting number of embryos for multifetal pregnancy reduction (MFPR) and discordance in size during the first trimester. Here we evaluated the correlation between the degree of discordance and length of gestation in the remaining fetuses. DESIGN: Observational clinical series. SETTING: Academic medical center with a single physician who performs a large number of MFPRs. PATIENT(S): Analysis of 252 consecutive MFPRs from a 2.5-year period (1996-1998). INTERVENTION(S): MFPR for patients with multifetal pregnancies. MAIN OUTCOME MEASURE(S): We evaluated the correlation between the degree of discordance in embryo size, as measured by the greatest difference in crown-rump length (CRL) (delta max), and the length of gestation. RESULT(S): Embryo size discordance was related to length of gestation of the remaining fetuses after MFPR. Of 72 patients with a delta max >5 mm, the rate of severe premature birth (delivery at <28 weeks' gestation) was 9.7%, compared with 1.7% for patients with a delta max <5 mm (P<.01). Of patients with severe premature birth, 70% had delta max >5 mm, compared with less than 30% in patients who delivered after 28 weeks (P<.05). CONCLUSION(S): Variations in embryo growth patterns in multifetal pregnancies may be observed even in the first trimester, which may be predictive of late pregnancy outcomes. With a delta max > or =5 mm, there is a significant increase in the risk of severe premature birth (delivery at <28 weeks).

Fetal obstructive uropathy in trisomy syndromes.

Fetal obstructive uropathy has seldom been described in trisomy syndromes, and its relationship to these syndromes remains unclear. Five trisomic male fetuses, four with trisomy 18 and one with trisomy 21, were identified out of 110 fetuses evaluated for fetal obstructive uropathy. We performed detailed examination on the urinary tracts of four of these fetuses, three with trisomy 18 and one with trisomy 21, following termination in the second trimester. All four had a markedly distended urinary bladder (megacystis), abdominal wall distension, and a small, poorly developed urethra thoughout its full length. All four also had poor development of the prostate with virtual absence of glandular development, as compared to age-matched controls. Posterior urethral valves were not identified in any case. Three of the fetuses (two with trisomy 18 and one with trisomy 21) had unilateral or bilateral hydroureters, and resulting renal tubulocystic or glomerulocystic change. Review of this database reveals an unexpectedly high frequency of trisomies, particularly trisomy 18, suggesting that the relationship may not be coincidental. Abnormal prostate development may be causally related to fetal obstructive uropathies and may be an under-recognized trait in trisomy syndromes. Karyotypic analysis of all fetuses with obstructive uropathy is important since in utero surgical intervention may be contraindicated in cases of fetal aneuploidy.