First trimester screening for pre-eclampsia and targeted aspirin prophylaxis: a cost-effectiveness cohort study.

OBJECTIVE: Investigate cost-effectiveness of first trimester pre-eclampsia screening using the Fetal Medicine Foundation (FMF) algorithm and targeted aspirin prophylaxis in comparison with standard care. DESIGN: Retrospective observational study. SETTING: London tertiary hospital. POPULATION: 5957 pregnancies screened for pre-eclampsia using the National Institute for Health and Care Excellence (NICE) method. METHODS: Differences in pregnancy outcomes between those who developed pre-eclampsia, term pre-eclampsia and preterm pre-eclampsia were compared by the Kruskal-Wallis and Chi-square tests. The FMF algorithm was applied retrospectively to the cohort. A decision analytic model was used to estimate costs and outcomes for pregnancies screened using NICE and those screened using the FMF algorithm. The decision point probabilities were calculated using the included cohort. MAIN OUTCOME MEASURES: Incremental healthcare costs and QALY gained per pregnancy screened. RESULTS: Of 5957 pregnancies, 12.8% and 15.9% were screen-positive for development of pre-eclampsia using the NICE and FMF methods, respectively. Of those who were screen-positive by NICE recommendations, aspirin was not prescribed in 25%. Across the three groups, namely, pregnancies without pre-eclampsia, term pre-eclampsia and preterm pre-eclampsia there was a statistically significant trend in rates of emergency caesarean (respectively 21%, 43% and 71.4%; P < 0.001), admission to neonatal intensive care unit (NICU) (5.9%, 9.4%, 41%; P < 0.001) and length of stay in NICU. The FMF algorithm was associated with seven fewer cases of preterm pre-eclampsia, cost saving of £9.06 and QALY gain of 0.00006/pregnancy screened. CONCLUSIONS: Using a conservative approach, application of the FMF algorithm achieved clinical benefit and an economic cost saving.

Current controversies in prenatal diagnosis: Noninvasive prenatal testing should replace other screening strategies for fetal trisomies 13, 18, 21.

This is a written summary of the oral debate presented at the International Society for Prenatal Diagnosis annual conference in Edinburgh in 2023. The topic under debate is whether noninvasive prenatal testing (NIPT) using cell-free fetal DNA should replace other screening strategies for the detection of fetal trisomies 13, 18, 21. There is no disagreement that NIPT is far more sensitive and has better positive predictive values for identifying trisomies 13, 18, and 21 than traditional screening approaches using biochemical markers and measurement of nuchal translucency. The major issue lies in the potential adverse consequences associated with abandoning traditional screening methods. The source of disagreement stems primarily from whether you consider the role of ultrasound in the context of screening to be strictly for nuchal translucency measurement or whether it should be combined with a fetal anatomy scan. The debate featured two experts who presented evidence in favor of each argument.

The evolving genetic etiology of conotruncal anomalies.

OBJECTIVE: To assess the diagnostic yield of genetic testing for antenatally detected conotruncal defects. METHOD: This was a retrospective analysis of all antenatally detected cases of conotruncal anomalies over a 4-year period. Patients were offered antenatal and postnatal genetic testing including QF-PCR, microarray and exome sequencing (ES) antenatally or genome sequencing (GS) postnatally on a case-by-case basis. RESULTS: There were 301 cases included. Overall, there were pathogenic genetic findings in 27.6% of the cases tested (53/192). The commonest finding was 22q11.21 deletion (20/192 cases, 10.4%), followed by trisomy 21 (6/192, 3.1%). There were 249 cases of isolated conotruncal anomalies, of which 59.8% (149/249) had genetic testing and 22.8% (34/149) had pathogenic findings. ES/GS was performed in five cases with no pathogenic findings. There were 52 cases of non-isolated contruncal anomalies, of which 82.7% (43/52) had genetic testing. ES/GS was performed in 11 cases in this group and increased the yield of clinically significant diagnoses from 32.6% (14/43) to 44.2% (19/43). CONCLUSION: Genetic abnormalities are present in over one quarter of cases of antenatally detected conotruncal anomalies. The commonest abnormality is 22q11.21 deletion. Exome sequencing or genome sequencing leads to a significant increase in genetic diagnosis in non-isolated cases.

'Something that helped the whole picture': Experiences of parents offered rapid prenatal exome sequencing in routine clinical care in the English National Health Service.

OBJECTIVES: In October 2020, rapid prenatal exome sequencing (pES) was introduced into routine National Health Service (NHS) care in England. This study aimed to explore parent experiences and their information and support needs from the perspective of parents offered pES and of health professionals involved in its delivery. METHODS: In this qualitative study, semi-structured interviews were conducted with 42 women and 6 male partners and 63 fetal medicine and genetic health professionals. Interviews were transcribed verbatim and analysed using thematic analysis. RESULTS: Overall views about pES were positive and parents were grateful to be offered the test. Highlighted benefits of pES included the value of the additional information for pregnancy management and planning for future pregnancies. An anxious wait for results was common, often associated with the need to make decisions near to 24 weeks in pregnancy when there are legal restrictions for late termination. Descriptions of dealing with uncertainty were also common, even when results had been returned. Many parents described pES results as informing decision-making around whether or not to terminate pregnancy. Some professionals were concerned that a non-informative result could be overly reassuring and highlighted that careful counselling was needed to ensure parents have a good understanding of what the result means for their pregnancy. Emotional support from professionals was valued; however, some parents felt that post-test support was lacking. CONCLUSION: Parents and professionals welcomed the introduction of pES. Results inform parents' decision-making around the termination of pregnancy. When there are no diagnostic findings or uncertain findings from pES, personalised counselling that considers scans and other tests are crucial. Directing parents to reliable online sources of information and providing emotional support throughout could improve their experiences of care.

First-trimester 3D fetal neurosonography: five standardised views.

BACKGROUND: There are several international guidelines for foetal anomalies scanning at 11-14 weeks' gestation. The aim of this study is to present our first-trimester specialist neurosonography protocol with examples of pathology in order to develop a systematic approach to evaluating the first-trimester foetal brain. METHODS: Women undergoing a first-trimester foetal medicine ultrasound scan between 2010 and 2020 for multiple indications underwent neurosonography according to a set protocol. 3D transvaginal brain examination was performed in all cases (2000 pregnancies scanned). We retrospectively reviewed all imaging to develop this protocol. RESULTS: We propose that the following five axial-plane parallel views should be obtained when performing neurosonography in the first trimester, moving from cranial to caudal: 1. Lateral ventricles; 2. Third ventricle; 3. Thalamus and mesencephalon; 4. Cerebellum; 5. Fourth ventricle. Examples of these images and abnormalities that can be seen in each plane are given. CONCLUSIONS: We have presented a specialist protocol for systematically assessing the foetal brain in the first trimester and given examples of pathology which may be seen in each plane. Further work is needed to prospectively assess detection rates of major abnormalities using this protocol and assess the reproducibility and learning curve of this technique.

This article suggests a way in which specialists scanning babies at 11­14 weeks of pregnancy can check the brain in a structured way. This involves looking at the brain at five levels or planes to view the developing structures. The suggested scan protocol is similar to images produced of the brain and heart at the second trimester (20 week) scan. We hope that specialists will find it useful to check the brain in this way if there are concerns raised at the dating (12 week) scan, and that this will lead to earlier detection of brain abnormalities or differences.

Detection rates of a national fetal anomaly screening programme: A national cohort study.

OBJECTIVE: To measure condition-specific detection rates for 14 physical conditions screened for by the NHS fetal anomaly screening programme (FASP) fetal anomaly (FA) ultrasound scan. DESIGN: Retrospective audit of 12 694 diagnoses across a 3-year national cohort. SETTING: All English NHS and crown-dependency hospital trusts providing maternity services. POPULATION: Pregnancies booked for maternity services with an expected date of delivery between 1 April 2017 and 31 March 2020 and at least one diagnosis of a condition screened for by FASP. METHODS: Active multi-source ascertainment, linkage, audit and validation of clinical information to identify the subset of diagnoses meeting the condition-specific positivity threshold for the FA scan. MAIN OUTCOME MEASURE: The accuracy of the FA scan compared with diagnostic reference standards. RESULTS: FA scan detection rates were: anencephaly 96.3% (95% confidence interval [CI] 81.7-99.3%), atrioventricular septal defect: 69.2% (95% CI 65.8-72.4%), bilateral renal agenesis: 98.7% (95% CI 95.4-99.6%), cleft lip: 89.5% (95% CI 87.8-90.9%), congenital diaphragmatic hernia: 60.8% (95% CI 56.5-65%), Edwards syndrome: 73.8% (95% CI 67.5-79.3%), exomphalos: 59.4% (95% CI 49.4-68.7%), gastroschisis: 88.6% (95% CI 79-94.1%), hypoplastic left heart syndrome: 92.7% (95% CI 90-94.8%), lethal skeletal dysplasia: 93.2% (95% CI 88.6-96%), Patau syndrome: 82.3% (95% CI 72.4-89.1%), spina bifida: 93.8% (95% CI 91.8-95.3%), tetralogy of Fallot: 75.4% (95% CI 72.1-78.4%) and transposition of the great arteries: 84.9% (95% CI 81.7-87.5%). CONCLUSIONS: The performance of the FA scan is above the expectations set in 2010 for most conditions. For the remaining conditions, the majority of fetuses and babies affected are detected before the FA scan.

Antenatal findings and early postnatal outcomes in pregnancies with trisomy 21: a 10 year retrospective review at a tertiary centre.

OBJECTIVE: To examine the antenatal imaging features, intrapartum findings and early postpartum course of pregnancies with trisomy 21 (T21) at a tertiary hospital in the United Kingdom. METHODS: Women with pregnancies diagnosed with T21 on antenatal or post-mortem/postnatal karyotyping, from February 2010-2020. Outcome measures included antenatal imaging findings, fetal growth restriction (FGR), birthweight, mode of delivery and early neonatal outcomes. RESULTS: 76 women were included. There were six intrauterine deaths and 70 livebirths. Thirty-eight (50%) had an antenatal diagnosis and twenty-five (33%) had a suspected diagnosis but declined further testing. The diagnosis was unanticipated in 13 (17%). Cardiac anomalies (35.5%) were the most common antenatal anomaly. Doppler abnormalities were apparent in 48/73 (68%). Eighteen (25.7%) had antenatal FGR. The majority were delivered by Caesarean section, and 21.4% of babies weighed

Implementing a rapid fetal exome sequencing service: What do parents and health professionals think?

OBJECTIVES: Prenatal exome sequencing (pES) for the diagnosis of fetal abnormalities is being introduced more widely in clinical practice. Here we explore parents' and professionals' views and experiences of pES, to identify perceived benefits, concerns, and support needs. METHODS: Semi-structured interviews were conducted with 11 parents and 20 health professionals (fetal medicine and clinical genetics) with experience of rapid pES prior to implementation in the English National Health Service. Interviews were transcribed verbatim and analysed thematically. RESULTS: Parents and professionals were largely positive about pES, emphasising clinical and psychosocial benefits of a timely, definitive diagnosis in pregnancy. Concerns included parental anxiety related to the timing of pES results or uncertain findings, a need for guidelines for case selection and reporting, and ensuring sufficient capacity for counselling, phenotyping and variant interpretation. Professionals were concerned non-genetics professionals may not be equipped to counsel parents on the complexities of pES. CONCLUSION: These findings highlight important issues for clinical implementation of pES. Expert counselling is required to enable parents to make informed decisions during a stressful time. To achieve this, professionals need further education and training, and fetal medicine and genetics services must work closely together to ensure parental understanding and appropriate support.

Fetal dural sinus thrombosis: A systematic review.

INTRODUCTION: Fetal dural sinus thrombosis (DST) is a rare condition. Although numerous case reports exist, the findings appear heterogenous and providing accurate patient counselling remains challenging. METHODS: A systematic literature review was conducted in accordance with PRISMA guidance. RESULTS: Thirty-one studies including 78 patients were included in this review. No association with maternal or neonatal coagulopathy, infection or trauma was found. The average gestational age at diagnosis was 25 weeks (range 17-34 weeks). Approximately half of foetuses affected were female (48.7%); one quarter were male (25.6%) and one quarter had no sex stated (25.6%). Termination of pregnancy was chosen in 25.6% of cases (20/78). In continuing pregnancies,10.3% (6/58) experienced a perinatal death. Antenatally, the majority of lesions either decreased in size (38.5%) or completely resolved (32.7%). The neonatal or childhood outcome was normal in 88.0% of survivors (44/50). The average age at follow up was 16.4 months, ranging from birth to 6 years. CONCLUSION: This review found that 10% of DST cases experience in-utero or neonatal death. In survivors, the majority of cases reduce in size or completely resolve in pregnancy and 85% are reported to have a good outcome. However, further evidence is needed regarding long-term neurocognitive sequelae.

Application of supercritical fluid chromatography for separation and quantitation of 15 co-formulated binary anti-hypertensive medications using a single elution protocol.

A facile supercritical fluid chromatography method is proposed to analyse 15 co-formulated binary anti-hypertensive drug combinations using a customized elution procedure. The effect of mobile phase composition, column back pressure and temperature was suitably optimized for adequate retention, analyte response and resolution. The chromatographic separation of the different drug combinations was performed on a DCPak poly(4-vinylpyridine) column (250 × 4.6 mm, 5 µm) at 125-bar pressure and 40°C using a photodiode array detector. A linear gradient of CO2 and 0.1% formic acid in methanol provided the best elution conditions for all drug combinations. Baseline separation of the drugs was possible with resolution factor Rs ranging from 1.42 to 12.58. The method was validated for specificity, sensitivity, accuracy and precision, recovery and robustness. The limit of detection and limit of quantitation for aliskiren, amlodipine, atenolol, candesartan, hydrochlorothiazide, lisinopril, losartan, metoprolol, olmesartan, telmisartan and valsartan were in the range of 0.26-2.56 and 0.77-7.75 µg/mL, respectively. The thermodynamic study revealed that interactions of the drugs with the stationary phase were spontaneous as evident from the negative free energy values, and the separation process was enthalpy driven. The developed method was successfully employed to analyse these drugs in their co-formulated tablet formulations.

Fetal Hemivertebra at 11 to 14 Weeks' Gestation.

Hemivertebra appears as an angulation of the spine on a coronal section. We evaluated the prevalence of chromosomal defects and outcome of fetuses with hemivertebra detected in the first trimester over a 9-year period in a single tertiary referral unit. There were 10 cases; 9 had other anomalies. Seven couples opted for termination of pregnancy. One pregnancy ended in fetal demise at 16 weeks, and the 2 isolated cases continued the pregnancy with delivery at term. A karyotype analysis was performed in 8 fetuses: 5 found to be euploid and 3 having trisomy 18. Comprehensive ultrasound screening allows early prenatal detection and appropriate counseling.

Ultrasound Features and Management of Body Stalk Anomaly.

OBJECTIVES: This study describes the prenatal diagnosis of body stalk anomaly (BSA) in a large series of affected pregnancies and proposes an algorithm for the differential diagnosis of abdominal wall defects in the first trimester. METHODS: This was a retrospective study of all the cases of BSA diagnosed in our tertiary unit between 2009 and 2015. In all cases, we performed a detailed ultrasound examination with particular emphasis on the appearance of the amniotic membrane and content of the amniotic sac and the coelomic cavity. RESULTS: The study included 17 cases of BSA (15 singleton pregnancies, one dichorionic-diamniotic twin pregnancy discordant for BSA and one case of conjoined twins) at a median gestational age of 12+3 (range 11+0 to 16+6) weeks. In all cases, the liver and bowel were herniated into the coelomic cavity and the amniotic sac was intact containing the rest of the fetus with a normal amount of amniotic fluid; additionally, there was severe kyphoscoliosis and positional abnormalities of the lower limbs. The umbilical cord was absent, but umbilical vessels were seen with color Doppler running from the placenta to the fetus marginally to the eviscerated abdominal content. CONCLUSION: BSA can be easily diagnosed from 11 weeks' gestation. Examination of the amniotic membrane continuity, content of both the amniotic sac and coelomic cavity and a short umbilical cord help in differentiating this condition from other abdominal wall defects.

Why do women invest in pre-pregnancy health and care? A qualitative investigation with women attending maternity services.

BACKGROUND: Despite the importance attributed to good pre-pregnancy care and its potential to improve pregnancy and child health outcomes, relatively little is known about why women invest in pre-pregnancy health and care. We sought to gain insight into why women invested in pre-pregnancy health and care. METHODS: We carried out 20 qualitative in-depth interviews with pregnant or recently pregnant women who were drawn from a survey of antenatal clinic attendees in London, UK. Interviewees were purposively sampled to include high and low investors in pre-pregnancy health and care, with variation in age, partnership status, ethnicity and pre-existing medical conditions. Data analysis was conducted using the Framework method. RESULTS: We identified three groups in relation to pre-pregnancy health and care: 1) The "prepared" group, who had high levels of pregnancy planning and mostly positive attitudes to micronutrient supplementation outside of pregnancy, carried out pre-pregnancy activities such as taking folic acid and making changes to diet and lifestyle. 2) The "poor knowledge" group, who also had high levels of pregnancy planning, did not carry out pre-pregnancy activities and described themselves as having poor knowledge. Elsewhere in their interviews they expressed a strong dislike of micronutrient supplementation. 3) The "absent pre-pregnancy period" group, had the lowest levels of pregnancy planning and also expressed anti-supplement views. Even discussing the pre-pregnancy period with this group was difficult as responses to questions quickly shifted to focus on pregnancy itself. Knowledge of folic acid was poor in all groups. CONCLUSION: Different pre-pregnancy care approaches are likely to be needed for each of the groups. Among the "prepared" group, who were proactive and receptive to health messages, greater availability of information and better response from health professionals could improve the range of pre-pregnancy activities carried out. Among the "poor knowledge" group, better response from health professionals might yield greater uptake of pre-pregnancy information. A different, general health strategy might be more appropriate for the "absent pre-pregnancy period" group. The fact that general attitudes to micronutrient supplementation were closely related to whether or not women invested in pre-pregnancy health and care was an unanticipated finding and warrants further investigation.

Maternal serum placental growth factor in prospective screening for aneuploidies at 8-13 weeks' gestation.

OBJECTIVE: To investigate whether measurement of maternal serum placental growth factor (PLGF) can improve the performance of first-trimester combined screening for trisomy-21 by fetal nuchal translucency (NT) thickness and serum free ß-human chorionic gonadotropin (ß-hCG) and PAPP-A. METHODS: In singleton pregnancies attending for routine care, serum PLGF, free ß-hCG and PAPP-A were measured at 8(+0)-13(+6) weeks' gestation, and fetal NT was measured at 11(+0)-13(+6) weeks. The population included 12,154 normal and 44 trisomy-21 pregnancies. We examined the effect of adding PLGF on the performance of screening by the combined test. RESULTS: In the trisomy-21 pregnancies the median multiple of the normal median PLGF, adjusted for gestational age, maternal weight, racial origin, smoking status and method of conception, was significantly reduced (0.6070, 95% CI 0.5543-0.6648), and this did not change significantly with gestational age. Adding PLGF to combined testing with a risk cut-off of 1 in 100 reduced the false positive rate from 2.7% (95% CI 2.5-3.0) to 2.6% (95% CI 2.4-2.8) and increased the detection rate from 85% (95% CI 75-93) to 88% (95% CI 78-95). CONCLUSIONS: Inclusion of serum PLGF improves the performance of the first-trimester combined test in screening for trisomy-21.

Clinical significance of Maspin promoter methylation and loss of its protein expression in invasive ductal breast carcinoma: correlation with VEGF-A and MTA1 expression.

Maspin is a serine protease inhibitor with tumor-suppressor activity. Maspin can suppress tumor growth and metastasis in vivo and tumor cell motility and invasion in vitro. Previous studies indicate that the loss of Maspin expression is closely linked to aberrant methylation of the Maspin promoter. We examined the promoter methylation status of Maspin in tumor and corresponding serum of breast cancer patients. In addition, protein expression of this gene was also assessed to determine possible correlation between promoter hypermethylation and gene silencing. Further, we investigated the correlation of Maspin expression with vascular endothelial growth factor (VEGF-A) and MTA1 expression. Maspin methylation was analyzed by methylation-specific PCR in 100 invasive ductal breast carcinoma patients' tumors and circulating DNA in a prospective study. Promoter hypermethylation was correlated with expression of the encoded protein in tumors by immunohistochemistry. Significant correlation was observed between promoter hypermethylation of Maspin (r = +0.88; p ≤ 0.0001) in tumors and paired sera. Significant association was found between Maspin promoter hypermethylation and loss of its protein expression (p = 0.01, OR = 3.1, 95% CI = 1.3-7.4). The expression of VEGF-A and MTA1 was lower in tumors with high Maspin expression compared to tumors with loss of Maspin expression. Our results indicate that aberrant promoter methylation is associated with loss of Maspin immunoreactivity in breast cancer tissues. Further, loss of Maspin expression is significantly correlated with increased expression of VEGF-A and MTA1.

Simultaneous enantioseparation and simulation studies of atenolol, metoprolol and propranolol on Chiralpak® IG column using supercritical fluid chromatography.

Enantioseparation of three ß-blockers, i.e., atenolol, metoprolol and propranolol, was studied on amylose tris(3-chloro-5-methylphenylcarbamate) immobilized chiral stationary phase using supercritical fluid chromatography (SFC). The effect of organic modifiers (methanol, isopropanol and their mixture), column temperature and back pressure on chiral separation of ß-blockers was evaluated. Optimum chromatographic separation with respect to resolution, retention, and analysis time was achieved using a mixture of CO2 and 0.1% isopropyl amine in isopropanol: methanol (50:50, V/V), in 75:25 (V/V) ratio. Under the optimized conditions, the resolution factors (R s) and separation factors (α) were greater than 3.0 and 1.5, respectively. Further, with increase in temperature (25-45 °C) and pressure (100-150 bars) there was corresponding decrease in retention factors (k), α and R s. However, a reverse trend (α and R s) was observed for atenolol with increase in temperature. The thermodynamic data from van't Hoff plots revealed that the enantioseparation was enthalpy driven for metoprolol and propranolol while entropy driven for atenolol. To understand the mechanism of chiral recognition and the elution behavior of the enantiomers, molecular docking studies were performed. The binding energies obtained from simulation studies were in good agreement with the elution order found experimentally and also with the free energy values. The method was validated in the concentration range of 0.5-10 µg/mL for all the enantiomers. The limit of detection and limit of quantitation ranged from 0.126 to 0.137 µg/mL and 0.376-0.414 µg/mL, respectively. The method was used successfully to analyze these drugs in pharmaceutical preparations.

Super-resolution Reconstruction MRI Application in Fetal Neck Masses and Congenital High Airway Obstruction Syndrome.

OBJECTIVE: Reliable airway patency diagnosis in fetal tracheolaryngeal obstruction is crucial to select and plan ex utero intrapartum treatment (EXIT) surgery. We compared the clinical utility of magnetic resonance imaging (MRI) super-resolution reconstruction (SRR) of the trachea, which can mitigate unpredictable fetal motion effects, with standard 2-dimensional (2D) MRI for airway patency diagnosis and assessment of fetal neck mass anatomy. STUDY DESIGN: A single-center case series of 7 consecutive singleton pregnancies with complex upper airway obstruction (2013-2019). SETTING: A tertiary fetal medicine unit performing EXIT surgery. METHODS: MRI SRR of the trachea was performed involving rigid motion correction of acquired 2D MRI slices combined with robust outlier detection to reconstruct an isotropic high-resolution volume. SRR, 2D MRI, and paired data were blindly assessed by 3 radiologists in 3 experimental rounds. RESULTS: Airway patency was correctly diagnosed in 4 of 7 cases (57%) with 2D MRI as compared with 2 of 7 cases (29%) with SRR alone or paired 2D MRI and SRR. Radiologists were more confident (P = .026) in airway patency diagnosis when using 2D MRI than SRR. Anatomic clarity was higher with SRR (P = .027) or paired data (P = .041) in comparison with 2D MRI alone. Radiologists detected further anatomic details by using paired images versus 2D MRI alone (P < .001). Cognitive load, as assessed by the NASA Task Load Index, was increased with paired or SRR data in comparison with 2D MRI. CONCLUSION: The addition of SRR to 2D MRI does not increase fetal airway patency diagnostic accuracy but does provide improved anatomic information, which may benefit surgical planning of EXIT procedures.

Demethylating agent 5-aza-2-deoxycytidine enhances susceptibility of breast cancer cells to anticancer agents.

DNA methylation plays an important role in regulation of gene expression and is increasingly being recognized as a determinant of chemosensitivity of human cancers. With the aim of improving the chemotherapeutic efficacy of breast carcinoma, the effect of DNA methyltransferase inhibitor, 5-Aza-2'-deoxycytidine (5-aza-CdR), on the chemosensitivity of anticancer drugs was investigated. The cytotoxicity of paclitaxel (PTX), adriamycin (ADR), and 5-fluorouracil (5-FU) was analyzed against human breast cancer cell lines, MDA MB 231 and MCF 7 cell lines using the MTT assay, and the synergy of 5-aza-CdR and these agents was determined by Drewinko's fraction method. The effects of each single agent or the combined treatment on cell cycle arrest were analyzed by flow cytometric analysis. We also investigated the effect of each single agent or the combined treatment of anticancer drugs with 5-aza-CdR on the methylation status of the selected genes by methylation specific PCR. In MDA MB 231 cells, a synergistic antiproliferative effect was observed with a combination of 10 microM 5-aza-CdR and these three anticancer drugs, while in MCF 7 cells, a semiadditive effect was observed. Treatment with 5-aza-CdR and anticancer drug resulted in partial demethylation of a panel of genes including RARbeta2, Slit2, GSTP1, and MGMT. Based on these findings, we propose that 5-aza-CdR enhances the chemosensitivity of anticancer drugs in breast cancer cells and may be a promising approach for increasing the chemotherapeutic potential of these anticancer agents for more effective management of breast carcinomas.