Racial Disparities in Pregnancy Complications Should we design diagnostic tests based on race? Alison Woodworth, PhD, DABCC, FACB AACC-SES Audio Conference Series 2/23/11 Infant Mortality • Infant mortality -death of an infant prior to 1 year • No significant change has been observed since 2000 (Significant reduction 1995 – 2000) • In the United States in 2006: Infant mortality 6.7 Infant Deaths / 1000 live births • Healthy people 2010 –Goal to reduce infant mortality rates to 4.5% and racial disparity • Research aimed at reduction of causes Infant mortality rates US, 2006 Southeast is well above national average and 4.5% goal!!! An infant death occurs within the first year of life. Source: National Center for Health Statistics, final mortality data, 1990-1994 and period linked birth/infant death data, 1995-present. Retrieved February 23, 2011, from www.marchofdimes.com/peristats. Infant Mortality Rates by Race US, 2004-2006 Average An infant death occurs within the first year of life. Source: National Center for Health Statistics, period linked birth/infant death data. Retrieved July 21, 2010, from www.marchofdimes.com/peristats. Infant Mortality by Cause US, 2006 SIDS is Sudden Infant Death Syndrome. RDS is Respiratory Distress Syndrome. "Maternal Preg. Comp." stands for "Maternal Complications of Pregnancy." Cause of death for 19961998 is based on the Ninth Revision, International Classification of Diseases (ICD-9); cause of death for after 1998 is based on the Tenth Revision, International Classification of Diseases (ICD-10). Source: National Center for Health Statistics, period linked birth/infant death data. Retrieved July 21, 2010, from www.marchofdimes.com/peristats. Racial Disparities in Pregnancy Outcomes Bryant et al. AJOG. 2010 Preeclampsia • Multisystem disorder due to poor placentation - 2nd half of pregnancy •Affects up to 8% of pregnancies •Leading cause of maternal death (50,000/yr) • A major cause of perinatal morbidity and mortality (900,000/yr) worldwide • Despite extensive research no methods of prediction or prevention Racial Disparities in PE • PE ~2X higher in African Americans than Caucasians Mortality Rate per 100 hospitalizations with delivery Healy, AJ et al. Obstet and gynecol 107(3): 625 – 31, 2006 and Tanaka, M et al. Am J Public Health. 97(2): 163 – 170, 2007. 4 3.5 3 2.5 2 1.5 1 0.5 0 3.4 2.9 2.1 1.7 White Black Hispanic 1999-2002 Other Adapted from Tanaka, M et al. Symptoms of Preeclampsia Maternal Syndrome • Hypertension, edema, proteinuria • Thrombocytopenia • Liver dysfunction HELLP syndrome • Seizures, stroke Eclampsia Fetal Syndrome • Intrauterine Fetal Growth Restriction (IUGR) • oligohydramnios • abnormal oxygenation Proposed Etiologies of Preeclampsia Increased in Maternal Serum Anti-angiogenic factors Placental Debris Reactive O2 species Proinflammatory cytokines C. W. Redman et al., Science 308, 1592 -1594 (2005) Decreased in Maternal Serum Angiogenic factors Predictors of Preeclampsia • Risk factors – Advanced maternal age, primiparity, obesity, insulin resistance, chronic hypertension, preexisting thrombophilias, and limited sperm exposure • History of preeclampsia • Multi-fetal gestation • Edema, Hypertension, Proteinuria • Doppler ultrasonography- Uterine artery blood flow • Circulating biochemical markers: Angiogenic (VEGF,PlGF) and antiangiogenic (sFlt1, sEng) sFlt-1 and PlGF • Antiangiogenic factor sFlt-1sequesters proangiogenic PlGF and VEGF • Ratio of sFLT-1/PlGF provides the best Predictive value • Prospective, multicenter trial • N= 409 controls (High and low risk) 48 cases • Method = Beckman Access IA Ideal Cut-off for Preterm PE =137 Sensitivity = 96%, Specificity = 97% Sunderji et al AJOG 2010 IL-6 and Preeclampsia • Increase in systemic inflammation in PEelevated CRP in PE • IL-6 elevation is maternal serum during PE • Secreted by decidual cells in response to inflammation Lockwood, CJ et al. AJP. 172(6): 1571-1577, 2008. • A role in the pathobiology of PE - IL-6 infused pregnant rats developed proteinuria and increased blood pressure Orshal and Khalil. Hypertension. 43: 434-444, 2004. Other Markers for Prediction of Preeclampsia Analyte/ Sample Cutoff GA (wks) Predictive Reference Collection Values SEng > 7.2 ng/mL 21 - 32 OR = 9.4 Levine et al. NEJM 2006 PP-13 < 0.39 MoM 8 - 13 Sens 100% Spec 80% Romero R et al. Am J Obs Gyn 2008 24 - 33 Sig in PE Zhong et al. Clin Chem 2005 Fetal DNA N/A sLSelectin < 1414 ng/mL 20 Sens = 84 Spec = 90 Chavarria et al. Eur J Obs Gyn Rep Biol. 2007 NO Inh. (ADMA) N/A 23 – 25 in PE, IUGR, abn doppler Savvidou et al. Lancet 2003 Prospects for Preeclampsia Predictive Markers • Limited knowledge of pathogenesis and genetic basis of preeclampsia limits identification of good predictive preeclampsia markers • Uniform definition of PE since 2000 has allowed for more accurate studies • Heterogeneity in results of different studies • Long-term prospective studies are needed with well defined patient populations and rigorous laboratory and study design • WHO Global Program to Conquer Preeclampsia mid-2006 Preterm Birth (PTB) • Delivery before 37 weeks gestation • Two types – Spontaneous and Induced • Occurred in 12.7% of births in 2007 (500,000 births/yr) • 20% increase since 1990 • Accounts for >75% of fetal & neonatal deaths in babies w/o anomalies • The majority of surviving preterm infants suffer serious morbidity Racial Disparities in Preterm Birth National Prevalence (%) Change -3% 20% 12% 35% Adapted from Heron et al. Pediatrics. 125:4-15, 2010. Etiologies of Preterm Birth Uterine Distension Hematologic Cervical Disease Hormonal/ STRESS Inflammation Infection Unknown GENETICS ? (Roberto Romero, MD) Predictors of Preterm Birth • Symptoms of labor • Genetics • History of preterm delivery • Multi-fetal gestation • Cervical Evaluation by sonography or digital exam • Biochemical markers: fFN, Salivary Estriol, Cytokines Fetal Fibronectin (fFN) • ECM Glycoprotein with unique “oncofetal domain” (FDC-6) • Located at choriodecidual interface – Key for implantation • In cervicovaginal fluid (CVF) prior to term labor –glycosylation • PTL - mechanical or local inflammation Courtesy of Hologic • Absence of fFN in CVF 24 – 36 weeks gestation good predictor of those who Won’t deliver Preterm or w/in 14 days PPV < 20%, NPV > 95%, Cytokines in Preterm Delivery • ~40-50% of preterm labor associated w/ infection • Chorioamnionitis (CAM) -Infection involving chorion, amnion, amniotic fluid, placental villi, and decidua • If CAM present 4X increase in neonatal mortality • Only 12.5% of those women show clinical evidence • CAM triggers cytokine production and synthesis and release of prostaglandins cervical ripening and uterine contractions Interleukin-6 in Preterm Delivery • Interleukin-6 (IL-6) and other in response to infection or injury. • >50% of PTL associated w/ infection and/or inflammation • IL-6 stimulates Delivery (left) • IL-6 in CVF w/ microbial invasion of the intra-amniotic cavity •Absence of IL-6 in CVF = good predictor of No delivery in 2 weeks Gomez R et al. Am J Obstet Gynecol 1998 PPV < 40%, NPV > 94% IL-6 and fFN Predict Delivery <14 Days N = 725 Left over fFN specimens – Multiple sights Analyte (cut-off) PPV (%) NPV (%) LR + LR - Odds ratio 95% CI IL-6 (250 pg/mL) 15 98 4.83 0.52 9.3* 4.1 – 20.8 fFN (50 ng/mL) 14 99 4.48 0.41 11.1* 4.8 – 25.5 *p <0.0001 Woodworth et. al. Clin Chem 2007 Neither is a strong diagnostic test for prediction of PTB in < 14 days IL-6 and fFN to Predict Delivery Clinical Variable Gestational age @ delivery (wks) (Pos.-neg.) [95% CI] Interval-collection to birth (days) (Pos.-neg.) [95% CI] Preterm Birth (Del > 37 wks) (Odds Ratio) [95% CI] *P<0.001, +P<0.007 fFN IL-6 (Pos > 50 ng/mL) (Pos >250 pg/mL) -2.4 [-3.4, -1.4]* -1.7 [-2.7, -0.7]+ -20.1 -17.0 [-29.6, -10.6]* [-26.2, -7.8]+ 7.5 [2.6, 21.6]* 4.4 [1.6, 12.1]+ N= 127 Patients Racial Disparities and Genetics • Evidence of genetic predisposition to PTB: Twin studies, History (personal or family), and Racial Disparities • Differences in AF concentrations of cytokines between Cases and Controls – not universal Menon, R. Acta Obstet Gynecol Scand. 87: 6, 590 – 600. 2008 SNPs in IL-6 Correlate with Differential Protein Expression European American African American Menon, R et al. Am J Obs Gynecol. 198:77. e1-7. Racial/Genetic Differences in Etiologies of PTB Genetic Parameter PTB SNP Pathway(s) Maternal or Fetal Influence IL-6 SNP Expression AF-IL-6 Expression African American Caucasian Dermatologic, Inflammatory Disease Inflammatory, and Hematologic diseases Fetal Maternal Significant Difference in frequencies of 13/20 tested (Allelic and Genotypic) Elevated in Term & PTB Menon et al Reprod Biol Endocrinol 2009; Velez et al Ann Hum Gen 2009 Elevated in PTB Diagnostic Utility of IL-6 and fFN in AA and Caucasian Women • Retrospective cohort study • Cervicovaginal samples sent to BJH for fFN testing collected for 5 years (2001 – 2005) • Symptomatic and Asymptomatic women (all races) • 24 0/7 - 34 6/7 weeks of gestation • Singleton gestation • C-sections & labor induction <14 days of delivery excluded • Frozen -70 ºC ® ® • IL-6 performed using DPC Immulite Woodworth et al. Clin Chim Acta 2011 Patient Samples 1498 leftover cervicovaginal specimens 773 specimens excluded (No chart, no race data, induction, twins, cervical dilation or cerclage, vaginal bleeding, wrong GA, sexual intercourse, placenta previa, result N/A) 725 samples with fFN and IL-6 testing 343 AA and 324 Caucasian 26 delivered < 14 days (3.6 %) 17 AA (5%) 7 Cauc (2.2%) 2 Other (3.4%) 699 delivered > 14 days (96.4%) 326 AA (95%) 317 Cauc (97.8%) 56 Other (96.6%) Clinical Data Categorized By Race Clinical Variable Race p AA (% AA) Cauc (% Cauc) Total Samples (n=667, [% Total Samples]) 343 324 Average GA at Sampling (wk, mean + SD) 30.4 + 2.7 29.6 + 2.7 * Average GA at Delivery (wk, mean + SD) 37.7 + 0.14 37.8 + 0.13 NS Interval–Sampling to Delivery (wk, mean ± SD) 7.3 + 0.2 8.2 ± 0.2 * FFN Positive (> 50ng/mL) (n=104 [16%]) 68 (20%) 36 (11%) * IL-6 Positive (> 250 pg/mL) (n=83 [12%]) 51 (15%) 29 (9%) * Delivery <7 Days of Sampling (n=16 [2.2%]) 11(3.2%) 5(1.5%) NS Delivery <14 Days of Sampling (n=26 [3.6%]) 17(5%) 7(2.2%) NS Total Deliveries (n=580) 316 (52%) 264 (43%) Preterm Deliveries < 37 Weeks GA (n=101 [17%]) 62 (20%) 39 (15%) NS Deliveries < 32 Weeks GA (n =14 [2.4%] ) 9 (2.8%) 5 (1.9%) NS Bacterial Vaginosis or Chorioamnionitis (n=124 [21%]) 89 (28%) 35 (13%) * ROC Curves- PTB < 14 days 250 250 250 Area Under the Curve Total Population 0.760 Caucasian 0.749 African American 0.762 Racial Disparity in Diagnostic Markers- Delivery = 14 d LR + LR- Odds Ratio 95% CI P fFN 4.48 0.41 11.1 4.8 – 25.5 <0.0001 IL-6 4.83 0.52 9.3 4.1 – 20.8 <0.0001 fFN 5.66 0.48 11.9 2.5 – 55.5 0.004 IL-6 3.24 0.78 4.1 0.8 – 22.2 0.13 fFN 3.7 0.43 8.7 3.1 – 24.4 <0.0001 IL-6 5.15 0.40 12.7 4.5 – 36.3 <0.0001 Race Analyte All Cauc AA Woodworth et al. Clin Chim Acta 2011 Racial Disparity in Diagnostic Markers- Delivery = 7 d LR + LR- Odds Ratio 95% CI P fFN 5.8 0.2 23.5 6.8 – 80.9 <0.0001 IL-6 5.6 0.4 13.2 4.7 – 37.4 <0.0001 fFN 6.4 0.2 35.9 3.9 – 331 0.0006 IL-6 4.6 0.7 6.9 1.1 – 43.2 0.07 fFN 4.6 0.2 20.8 4.4 – 99 <0.0001 IL-6 5.5 0.3 17.5 4.5 – 68.3 <0.0001 Race Analyte All Cauc AA Woodworth et al. Clin Chim Acta 2011 Genetic Influence on Preterm Birth GENETICS Uterine Distension Cervical Disease Hormonal/ Stress Hematologic Inflammation Infection (Roberto Romero, MD) Unknown Racial Disparities in Diagnostic Markers for PTB - Conclusions • Racial disparities in PTB rates may be related to differences in pathobiological pathways associated with PTB in AA versus Caucasian women • Cervicovaginal IL-6 and fFN have similar diagnostic strengths in a racially diverse population (NPV >98%). • IL-6 in AAs and fFN in Caucasians are the only tests with sufficient diagnostic strength to rule-in delivery in 14 days. • Diagnostic testing algorithms based upon race/genetics and/or etiology may more accurately predict PTB. Conclusions • Racial disparities in incidences of preeclampsia and preterm birth may lead to disparities in infant mortality rates. • No PE biomarkers are currently recommended for routine use – sFlt1/PlGF ratio is the most promising – IL-6 may be involved in pathobiology • Cervicovaginal IL-6 and fFN have similar diagnostic strengths in a racially diverse population (NPV >98%). • IL-6 in AAs and fFN in Caucasians are the only tests with sufficient diagnostic strength to rule-in delivery within 14 days. • Diagnostic testing algorithms based upon race may provide a more accurate prediction of women who will deliver preterm and those at risk for preeclampsia Acknowledgments Vanderbilt University Elizabeth O’Keeffe Emily Bishop Courtney Moffett Terri Scott Beckman Coulter DPC Immulite Washington University Ann Gronowski Jennifer Moore Christopher G’Sell Bram Verdoes University of North Carolina David Grenache Jennifer Snyder Lee Morris Catherine Wares