Failure of cervical fibronectin to predict premature delivery in a population of monofetal pregnancies with idiopathic preterm labor

European Journal of Obstetrics & Gynecology and Reproductive Biology 97 (2001) 35±39

Failure of cervical ®bronectin to predict premature delivery in a population of monofetal pregnancies with idiopathic preterm labor Jean-Luc VolumeÂniea, Jean Guibourdencheb, Virginie Doridota, Olivier Sibonya, Jean-FrancËois Ourya, Philippe Blota, Dominique Lutona,* a

Department of Obstetrics and Gynecology, HoÃpital Universitaire Robert DebreÂ, 48 boulevard SeÂrurier, 75019 Paris, France b Departement of Biochemistry, HoÃpital Universitaire Robert DebreÂ, 48 boulevard SeÂrurier, 75019 Paris, France Received 23 June 2000; accepted 18 September 2000

Abstract Objective: The purpose of the study was to evaluate the correlation between the presence of cervical ®bronectin in a high-risk population of women with symptoms of preterm labor and the occurrence of preterm delivery or the need for aggressive tocolysis. Study design: One hundred and thirty women presenting with symptoms of threatened preterm labor were included. Cervical sampling for detection of ®bronectin was performed on admission and every day until discharge or delivery. Time to delivery, length of hospital stay, use of indomethacin, delivery before 37 weeks of GA, mean term of delivery and failure of tocolysis to prevent delivery were compared to ®bronectin test results. Data were analyzed using Student's t-test for continuous variables and the w2 test or Fisher exact test for discrete variables. Results: No correlation could be found between the results of ®bronectin cervical sampling on admission and any of the outcome parameters studied. Test performances were low (sensitivity 28%, speci®city 57%, positive predictive value 19%, negative predictive value 69%). Results were not modi®ed when the ®ndings of repeated tests were taken into account. Conclusion: Cervical ®bronectin failed to discriminate a subgroup of symptomatic women delivering prematurely. The prognostic value of ®bronectin testing was not better than clinical data in our series. This observation is in disagreement with previous studies on the diagnostic value of vaginal or cervical ®bronectin in preterm labor. # 2001 Elsevier Science Ireland Ltd. All rights reserved. Keywords: Cervical ®bronectin; Preterm labor; Preterm delivery

1. Introduction Preterm delivery is the leading cause of neonatal morbidity and mortality. The de®nition of a group at risk relies mainly on clinical data such as history of preterm birth, assessment of cervical dilatation, monitoring of uterine activity and risk factor scoring. All these factors have been shown to be of limited value for prediction of premature delivery due to their low sensitivity and speci®city [1±5]. Thus, more accurate alternative markers have been evaluated such as ultrasonographic measurement of cervical length [6], or detection of vaginal or cervical fetal ®bronectin [4,5,7±16]. The fetal ®bronectin test is an enzymelinked immunosorbent assay which uses the monoclonal antibody FDC-6 [17]. This antibody is speci®c for an epitope

* Corresponding author. Tel.: ‡33-01-40-03-47-15; fax: ‡33-01-40-03-24-80. E-mail address: [email protected] (D. Luton).

on ®bronectin from fetal, placental and malignant cells and tissues. Fetal ®bronectin can be found in vaginal secretions up to 20 GA [12] but is seldom detected (3 to 4% [12]) after 22 GA until the end of pregnancy. It has been demonstrated that the ®bronectin test accurately predicts term delivery [18]. Fibronectin concentration is elevated in amniotic ¯uid and thus, may be released in cases of premature rupture of membranes [12]. Various studies have demonstrated an association between elevated vaginal ®bronectin and preterm delivery in patients with intact membranes [4,5,7± 15,19±21] in either low- or high-risk populations. In all studies, vaginal ®bronectin displayed a high negative predictive value (at least 80% but very often superior to 90%). Thus, a negative test is associated with a good prognosis [19] and seems to be reassuring in populations with clinical symptoms of preterm labor. Populations with symptoms suggestive of preterm labor were studied either in research conditions [7,10±13,15,21,23,24] or in clinical practice [20] in order to establish whether vaginal or cervical ®bronectin was able to distinguish between true and false threatened

0301-2115/01/$ ± see front matter # 2001 Elsevier Science Ireland Ltd. All rights reserved. PII: S 0 3 0 1 - 2 1 1 5 ( 0 0 ) 0 0 5 0 4 - 2

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preterm labor and thus, to help the clinician to identify a population requiring tocolysis, long-term hospital stay or corticosteroids [24]. We decided to evaluate the association of fetal ®bronectin and premature delivery in a population of patients presenting with clinical symptoms of preterm labor. We also described the relation between ®bronectin and length of hospital stay, use of indomethacin and failure of tocolysis to prevent delivery. 2. Methods Our longitudinal prospective study was carried out in a single tertiary referral unit from January to December 1998. During this year 3034 deliveries took place in the department. All patients presenting with clinical symptoms of threatened preterm labor were included when hospitalization was indicated. The diagnosis of threatened preterm labor was based on the presence of uterine contractions accompanied by substantial changes in cervical length or dilatation since the previous examination. Gestational age ranged from 24 to 36 GA and was established by the date of the last menstrual period or set by ®rst-trimester ultrasonography in the case of a discrepancy between the two dates of greater than 10 days. Patients with multiple gestations, preterm rupture of membranes (diagnosed on frank visualization of ¯uid ¯ow through cervical OS and con®rmed in doubtful cases by detection of diamine oxidase and insulin growth-factor binding protein-1 in vaginal sampling), cervical dilatation superior to 3±4 cm, in situ cervical cerclage, placenta previa, hemorrhage, history of sexual intercourse <24 h, or any intercurrent pathology interfering with the decision to maintain the pregnancy were excluded. All patients gave informed consent for participation in the study, which had been approved by our local ethics committee. Samples for fetal ®bronectin assay were obtained during speculum examination on a sterile Dacron swab left in the cervix for 10 s. Samples were immediately dispatched to the laboratory where a qualitative immunoassay was used to check for rupture of membranes. The ROM-check membrane immunoassay is a solid-phase, immunogold assay (Adeza Biomedical, USA) where a speci®c epitope within the III-CS region of fetal ®bronectin is recognized. An antihuman ®bronectin gold conjugate forms a complex with ®bronectin which passes through a membrane containing a bound monoclonal antibody speci®c for fetal ®bronectin and binds to it, displaying a visible spot. Positive and negative controls were processed simultaneously. The threshold for detection of ®bronectin was 50 ng/ml. Cervical examination was performed after sampling. The results obtained were separated into positive, negative and doubtful. Patients with doubtful results on ®rst sampling were excluded from further analysis. The attending clinician was not aware of the ®bronectin test result, the study was thus, blinded. Tocolytic therapy was administered at the discretion of the attending clinician. Intravenous salbutamol was generally used but

oral calcium channel blockers (nicardipin) could also be prescribed. The ®rst sampling was performed upon admission and was repeated every day until the patient's discharge from hospital. A patient was graded as ``positive'' when the ®bronectin was positive upon admission. Iterative sampling were performed in order to see if patient status could change with time and if the value of ®bronectin was different when several results, and not solely the ®rst one, were taken in account for an individual patient. To summarize these data, we calculated a ``negativity ratio'' de®ned as the ratio of negative ®bronectin samples/total number of samples in an individual patient in order to determine whether patients with several positive ®bronectin results were more at risk of premature delivery than patients with one or a few positive results. Attending clinicians and patients were unaware of the ®bronectin test result. Main outcome measures were preterm delivery (i.e. delivery before 37 completed weeks of gestational age or 259 days), term of delivery, length of hospital stay, failure of tocolysis to stop premature labor, and need for anti-prostaglandin medication (indomethacin), which constitutes second-line tocolytic therapy in our setting. Statistical analysis was performed using Statview4 software (Macintosh package). Student's t-test was used for continuous variables normally distributed and a Mann± Whitney test for nonparametric data. The w2 test or Fisher exact test were applied for discrete variables. Linear regression was realized for correlation between our negativity ratio and continuous outcome measures. P < 0:05 was considered statistically signi®cant. Sample size calculation was performed, basing our estimations of the rate of premature delivery in ®bronectin positive and negative groups on the results of previous studies in high-risk patients [7,9,11,12,20]. Ninety subjects were necessary to detect a shift from 20% (®bronectin negative group) to 50% (®bronectin positive group) in the rate of premature delivery, with a power of 80% and a type I error of 5%. 3. Results In our department 178 women were examined and considered as presenting symptoms of threatening preterm labor necessitating hospitalization without clinical preterm premature rupture of membranes. They accounted for 4.3% of the patients delivered in the department during this period, 48 were excluded for the following reasons: multiple pregnancy (34), maternal intercurrent pathology (3), fetal malformation (3), placental abruption (2), leiomyoma necrobiosis (1), polyhydramnios (1), vaginal bleeding (1), abnormal fetal heart rhythm pattern (2), and incomplete data (1). No one presented with symptomatic infection. One hundred and thirty patients entered the study which can thus, be considered as patient with idiopathic preterm threatened labor, 70/130 (53.8%) presented with cervical length inferior to 2 cm, 60/130 (46.2%) had cervical dilatation

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Table 1 Demographic and clinical data according to initial fibronectin status

Multiparous Cervical length <2 cm Cervical dilatation >1 cm Lower uterine segment distended a

Fibronectin positive 47 patients (%)

Fibronectin negative 73 patients (%)

Significancea

23 25 22 28

45 32 33 42

N.S. N.S. N.S. N.S.

(49) (53.2) (46.8) (59.6)

(61) (43.8) (45.2) (57.5)

N.S.: not significant.

superior to 1 cm, 73/130 (56.1%) had lower uterine segment distension and 100/130 (84%) had at least one uterine contraction by 10 min period on tocography. All patients received tocolysis (intravenous salbutamol in 129/130 cases (99.2%) and nicardipin in 34/130 cases (26%)), some patients being treated with both drugs. The mean gestational age on inclusion in the study was 213.8 days (or 30.5 GA) (95% con®dence interval: (24.5±36.7), range: (21.3±35.7)). Mean gravidity and parity were respectively 2.45 and 1.71. Thirty-two patients delivered prematurely (before 37 weeks of gestational age), which represents 25% of the study population. Delivery before 34 weeks of GA accounted for 9% of the deliveries in our series. Mean gestational age at delivery was 261 days (37.3 GA, 95% con®dence interval: (32.4±42.1), range: (25.8±41.8)). Mean fetal birthweight was 2980 g (95% con®dence interval: (2868±3091), range: (800±4620)). First fetal ®bronectin sampling was positive upon admission in 47 patients (36% of the population), negative in 73 patients (56%) and doubtful in the remaining 10 patients (8%) which were excluded from further analysis. Gestational age on admission and parity were not different between positive and negative patients for ®bronectin (31 GA, 95% con®dence interval: (24.6±37.3), range: (22±35.6)) and (30.3 GA, 95% con®dence interval: (24.3±36.4), range: (21.3±35.7)), respectively for gestational age on admission; 1.64 and 1.8, respectively for parity, difference not signi®cant for either result. We con®rmed that patients with doubtful results did not differ from patients with positive or negative results for term on admission, cervical length and dilatation and lower uterine segment distension. Overall, 83 patients were positive at least once for vaginal ®bronectin (64%) whereas 45 (34%) were constantly negative (and 2 doubtful). No signi®cant difference was observed in the incidence of preterm delivery between positive and negative patients for fetal ®bronectin (9/47 in ®bronectin positive patients versus 22/74 in ®bronectin negative patients), (calculated OR ˆ 0:55, 95% con®dence interval ˆ …0:21 1:43†) for premature delivery in the presence of a positive ®bronectin test, P > 0:1). Mean birth term was concordant between the two groups (263.3 days, 95% con®dence interval ˆ …258 268† for the ®bronectin positive group versus 259 days, 95% con®dence interval ˆ …255 263† for the ®bronectin negative group). Consequently, in our population the test was poor in predicting preterm birth, with a sensitivity of 28%, a speci®city of 57%, a predictive positive value of 19%

and a negative predictive value of 69%. No correlation could be observed between initial ®bronectin test results and clinical parameters of severe preterm labor as summarized in Table 1. These clinical criteria were unable to predict preterm delivery as well, with a similar mean term of delivery in the different groups (Table 2). Beta mimetics were given to all patients but anti-prostaglandins were limited to subjects displaying marked uterine activity in spite of salbutamol infusion or nicardipin administration. Thus, administration of indomethacin can be regarded as a sign of severe threatened preterm labor. No difference was observed between ®bronectin positive (23% were given indomethacin) and ®bronectin negative (34% were given indomethacin) patients (P ˆ 0:2). Similarly, the mean length of hospital stay was not statistically different between ®bronectin positive and negative patients (18.77 days in positive versus 16.34 days in negative patients, P > 0:1). The percentage of patients delivering before term in spite of ongoing tocolysis did not differ between positive (12.8%) and negative (11%) patients. Fibronectin status could change on successive samplings and a large array of combinations of positive or negative results was found. This is why we calculated our ``negativity ratio'' and analyzed the correlation between this index and the outcomes previously studied. Results are summarized in Table 3. Linear regression was established between the negativity index and the number of days in hospital on the one hand, and between the negativity index and the length of pregnancy on the other hand. Results revealed no obvious correlation between the variables (data not shown). Table 2 Duration of pregnancy according to clinical parameters Clinical parameters

Duration of pregnancy (days)

Significance

Cervical length <2 cm >2 cm

260.9 261.3

N.S.

Cervical dilatation >1 cm <1 cm

260.1 262

N.S.

Lower uterine segment distension Yes 259.1 No 263.8

N.S.a

a P ˆ 0:11: trend to shorter duration of pregnancy in the subgroup with distended lower uterine segment.

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Table 3 Mean negativity ratioa for different pregnancy outcomes Outcomes analyzed

Mean values of the index

Significance

Prematurity Yes No

0.605 0.574

N.S.

Indomethacin use Yes No

0.629 0.563

N.S.

Failure of tocolysisb Yes No

0.48 0.59

N.S.c

a

Defined as the ratio of negative/total fibronectin samples. Premature delivery in spite of ongoing tocolysis. c Mann±Whitney test; N.S.: not significant. b

4. Comments Cervical ®bronectin was not a predictor of preterm delivery in our series, neither was it able to distinguish patients with severe preterm labor necessitating long-term hospitalization or intensive therapy with anti-prostaglandin drugs. Clinical data (cervical length, cervical dilatation, lower uterine segment distension) did not yield better results in predicting premature delivery or severe preterm labor, as previously noted [5]. Most published studies, including a recent meta-analysis [25], evaluating the predictive value of ®bronectin for preterm delivery in a high-risk population, such as ours found a relationship between a positive ®bronectin test and preterm birth with a relative risk of premature delivery varying from 1.85 to 22.5 between negative and positive subjects for ®bronectin [7,11,13,15,20,21]. Sensitivity of fetal ®bronectin for preterm delivery ranged from 44 to 100% and negative predictive value, which is particularly interesting in women presenting with symptoms of threatened preterm labor, ranged from 71 to 100%. Fibronectin samples were positive in 20 to 50% of those symptomatic patients, and our value of 36% is therefore, within this range. In our series, cervical samples were taken every day from admission until delivery. This pattern of sampling may lower the speci®city, positive predictive value and likelihood ratio of the test [14,19]. Nevertheless, no correlation between the ®bronectin result and premature delivery was found when only the ®rst test on admission was considered. Thus, the multiplicity of samples could not explain our observations. We also decided to use a qualitative test because we intended to implement a test which could be performed at the bedside in a practical clinical setting. The same procedure was used by Parker [15] and yielded an excellent correlation between test result and interval to delivery. Other studies in symptomatic patients used quantitative laboratory techniques [7,11,13,20,21]. It is therefore, unlikely our results could be due to the use of a qualitative test. The manufacturer's instructions were strictly followed and, in particular, contamination by maternal blood was

avoided, digital examination never preceded sampling, and recent (<48 h) sexual intercourse was noted, as these features may induce false positive results [22,23]. No technical error can give a clear explanation of the discrepancy between our results and those found previously in symptomatic patients. No difference was noted in the clinical and demographic characteristics of the positive and negative patients on admission. Multiparous women were encountered with similar frequencies in negative and positive groups (parous women tended to display a higher prevalence of positive ®bronectin tests in another study [4]). All the patients hospitalized underwent tocolysis according to our usual protocol, and the drugs were similar in the two groups. As Goldenberg et al. [8] established a relation between the results of repeated tests, and between the percentage of successive positive tests and delivery, we sought to determine if sequential sampling could provide more information on pregnancy outcome, and calculated a ``negativity index''. No correlation was noted between this index and premature delivery, treatment with anti-prostaglandins or length of hospital stay. We also decided to exclude all patients with intercurrent pathology and to limit our study group to subjects with idiopathic preterm labor. We searched to eliminate patients whose pathology may have necessitated other therapy than tocolysis (i.e. antibiotics, nonsteroidal anti-in¯ammatory drugs, analgesics) and to focus on preterm labor with no obvious aetiology, as it represents a frequent and puzzling problem. Consequently, our results cannot be extrapolated to patients with polyhydramnios, twin pregnancies, infection or myomas, for instance. This study was designed as a preliminary to the implementation of cervical ®bronectin testing in the management of threatened preterm labor. We intended to determine later whether knowledge of a patient's ®bronectin status would lead to a reduction in tocolytic treatment, in the number and length of hospitalizations and to a more optimal management of patients really at risk of premature delivery (timing of corticosteroids administration, transfer to a referral unit), as recently reported [16,24]. As a consequence of our observations, we went on resorting to clinical parameters to deal with patients presenting symptoms of premature labor. In spite of a rigorous strategy of selection, sampling and detection of ®bronectin, our results are not in agreement with a large body of published data on the subject, including a recent meta-analysis [25]. This experience underscores the need to evaluate new techniques with caution in a particular setting before considering their widespread use. References [1] Beckmann C, Beckman C, Stanziano G, Marth C, Bergauer N. Accuracy of maternal perception of preterm uterine activity. Am J Obstet Gynecol 1996;174:672±5. [2] Copper RL, Goldenberg RL, Dubard MB, Hauth JC, Cutter GR. Cervical examination and tocodynamometry at 28 weeks' gestation:

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