US 20120021442A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2012/0021442 A1 Buhimschi et al. (43) Pub. Date: Jan. 26, 2012 (54) MARKERS FOR DETECTION OF (60) Provisional application No. 61/206,125, filed on Jan. COMPLICATIONS RESULTING FROM IN 28, 2009. UTERO ENCOUNTERS Publication Classification (75) Inventors: Catalin S. Buhimschi, New Haven, CT (US); Irina Buhimschi, New (51) Int. C. Haven, CT (US); Vineet Bhandari, GOIN 33/566 (2006.01) Cheshire, CT (US) (52) U.S. Cl. ........................................ 435/7.92; 436/501 (73) Assignee: Yale University, New Haven, CT (US) (57) ABSTRACT Described herein are biomarkers, such as protein biomarkers, (21) Appl. No.: 13/193,421 which are diagnostic of and predictive for complications that result from an in utero encounter, such as an infection by the (22) Filed: Jul. 28, 2011 fetus, that can lead to premature birth (PTB). The biomarkers can be used to identify fetuses and newborns at risk for Related U.S. Application Data complications of PTB, such as (Early Onset Neonatal Sepsis) (63) Continuation-in-part of application No. PCT/US2010/ EONS, intra-ventricular hemorrhage (IVH) and other poor 000259, filed on Jan. 28, 2010. OutCOmeS. Patent Application Publication Jan. 26, 2012 Sheet 1 of 24 US 2012/0021442 A1 Asia xead Patent Application Publication Jan. 26, 2012 Sheet 2 of 24 US 2012/0021442 A1 (g-u)dnolôIonuoo-(C=u)noubApngS- Tu?õd6>9-Ti•Taelºd06<9;T!“ ºsisdes(-).s.sdes?oo=(+)· •º.einunoein?no-IV(-)av(+): •?ºspelºvº?;ºoaperfiwoH Patent Application Publication Jan. 26, 2012 Sheet 3 of 24 US 2012/0021442 A1 uouebusseroeobolo?uo(=Hinwe 9eun61 sisebpondºnopogo? ),eseuduo?epljen*(3 Kneulou?oedssseulluºpuºl (gºi-u)s’esseounului?o?quesem Patent Application Publication Jan. 26, 2012 Sheet 4 of 24 US 2012/0021442 A1 [10,1-09/1026 7eun61 ?oliepeseg?leuelepingun |uputp-?efdy ?idºleuelesingun ujulg-lebdy Patent Application Publication Jan. 26, 2012 Sheet 5 of 24 US 2012/0021442 A1 t).SNOE sa| |s?n?nsas G?In6|- s?iuojuureoluogoleopolo?si?elemes| Patent Application Publication Jan. 26, 2012 Sheet 6 of 24 US 2012/0021442 A1 [119 |19-11y 9eunfil (0)0(9%)u ?ebuerolluepow sain?nopoo?qaapisod (guru?sileo)pav! (%)o?em1:1| | Patent Application Publication Jan. 26, 2012 Sheet 7 of 24 US 2012/0021442 A1 dankepulujejoigneuoneuro?uluisen?uepipºus?ºuocz Patent Application Publication Jan. 26, 2012 Sheet 8 of 24 US 2012/0021442 A1 9ºun61 Patent Application Publication Jan. 26, 2012 Sheet 9 of 24 US 2012/0021442 A1 resseounului 09 6?un 61 Patent Application Publication Jan. 26, 2012 Sheet 10 of 24 US 2012/0021442 A1 ( |";} 8(dH)NIEOT901dVH,|- 0\\§?º??N(O3s?kSIN(O3 (~~~~~,~~~ N?Ioù?d?IvT?u-Nigolºolavº | Patent Application Publication Jan. 26, 2012 Sheet 11 of 24 US 2012/0021442 A1 0 8(dH)NIEOTSOIdVH|(Mah).N?lohaq?lvian-NaoTºolavH LLaun61 Patent Application Publication Jan. 26, 2012 Sheet 12 of 24 US 2012/0021442 A1 (sah)nHlohadalvianNiaoToolavº Z?aun61 (dH)NaoTºoldv?i,|-? ?upayoudpaye?al-ulqo?boideH unaouxunuog?stinae+ Patent Application Publication Jan. 26, 2012 Sheet 13 of 24 US 2012/0021442 A1 ?&&d#8dH d?)sa??uep?euroe?oldgoBu??osisÁieueleuoisuºup!!!nu* Patent Application Publication Jan. 26, 2012 Sheet 14 of 24 US 2012/0021442 A1 (100'0=d)q?eagdoHA! TT. T. ter y took (uffr) dh's dh to 7|.?un61 (€10'0=d)SNO3?*~~ t e e es wer ce N: were (uffrt) adh 8 dh so Patent Application Publication Jan. 26, 2012 Sheet 15 of 24 US 2012/0021442 A1 ;----------- (!)A.)— Patent Application Publication Jan. 26, 2012 Sheet 16 of 24 US 2012/0021442 A1 5. zzdi ex E. dh Patent Application Publication Jan. 26, 2012 Sheet 17 of 24 US 2012/0021442 A1 ZLeln6|- Patent Application Publication Jan. 26, 2012 Sheet 18 of 24 US 2012/0021442 A1 Fuaipei6 zigfjaaXame tenoorow 8]©Jnfil Patent Application Publication Jan. 26, 2012 Sheet 19 of 24 US 2012/0021442 A1 6)aun61 Patent Application Publication Jan. 26, 2012 Sheet 20 of 24 US 2012/0021442 A1 Égi risi gri chose sco 0 Patent Application Publication Jan. 26, 2012 Sheet 21 of 24 US 2012/0021442 A1 /**** …;…;););)*(^&********************* ?zeun61 rº.*.*.*..*„……--~~~~~~~~~**************** s Patent Application Publication Jan. 26, 2012 Sheet 22 of 24 US 2012/0021442 A1 {}{}}, sociveti tastate. A ZZ?un61 |popupNo.?išs?šš?š?5O?????????????????????????????s?osgosysoü?e?p Patent Application Publication Jan. 26, 2012 Sheet 23 of 24 US 2012/0021442 A1 Figure 23 criterior; 3,37}rgical sersity': Six see:cay. 80% s as 3: 2.x 23 3 x 3 3s so go go 23 4. Sc. s. 33 ha&HRP stati?ficeactivity by west-ity atok ico-specificity Acided it coastazar girl at:tice, attrary aersity units a ---.283 Patent Application Publication Jan. 26, 2012 Sheet 24 of 24 US 2012/0021442 A1 Figure 24 Cord blood itp & Hip-RP switch parter scarf biao-is pesinai ECNS to a it star 2 paoisability likely exposed likely rion-exposed is ster 2 probabišity Y50%; {Caster probability >50% US 2012/0021442 A1 Jan. 26, 2012 MARKERS FOR DETECTION OF gestation also increased the risk of cerebral palsy.'" The COMPLICATIONS RESULTING FROM IN underlying mechanisms remain unclear, but this data high UTERO ENCOUNTERS lights the need for a paradigm shift in prematurity research from the unilateral goal of extending the duration of gestation RELATED APPLICATION to concurrently improving neonatal outcomes. Concerned by these issues and the alarming increase in the rate of PTB, the 0001. This application is a continuation-in-part of PCT/ March of Dimes Scientific Advisory Committee on Prematu US2010/000259 filed Jan. 28, 2010, which claims the benefit rity and the most recent report issued by the Institute of of the filing date of U.S. Provisional application 61/206,125, Medicine in 2008: Preterm Birth: Causes, Consequences and entitled “Novel biomarkers for detection of early onset neo Prevention suggested that studies to identify biomarkers that natal sepsis (EONS) and other complications of prematurity” may predict adverse outcomes for infants born preterm to and filed Jan. 28, 2009. The entire teachings of the referenced allow for early intervention should become a priority. applications are incorporated herein by reference. Clearly, additional approaches to assessing neonatal and early postnatal risk independent of gestational age (GA), and GOVERNMENT FUNDING particularly approaches that can provide an early diagnosis, 0002 This invention was made with government support are needed. under Grant Number R01 HD047321 and R01 HD062007-01, awarded by the National Institutes of Health. The government SUMMARY OF THE INVENTION has certain rights in the invention. 0004. Described herein are biomarkers, such as protein biomarkers, which are diagnostic of and predictive for com BACKGROUND OF THE INVENTION plications that result from an in utero encounter, such as an 0003 Premature birth (PTB) is a significant public health infection (e.g., bacterial, viral, parasitic or fungal), by the problem. The technological advances in newborn intensive fetus. The biomarkers described herein are useful in methods care of the past decades have increased Survival of preterm of predicting fetal and neonatal outcome and can be assessed, infants and the awareness of the need to improve outcomes. for example, in cord blood (CB), in blood or in other body Infection-induced PTB represents a unique environment, fluids obtained from the neonate/newborn. In some embodi given that attempts to prolong pregnancy raise the risk for ments, the biomarkers can be assessed in fetal blood from the early onset neonatal sepsis (EONS). In such cases, there is an umbilical cord, for example, by cordocentesis a procedure increased risk of poor neonatal outcomes, including intra sometimes called Percutaneous Umbilical Cord Blood Sam Ventricular hemorrhage (IVH), which is a significant cause of pling (PUBS). brain injury, cerebral palsy and developmental disability. A 0005. A specific embodiment described herein relates to key problem is that most hemorrhages occur in the first 24 protein biomarkers, originally identified in cord blood, that hours and therapies aimed to prevent IVH must address the are diagnostic of and predictive for Early Onset Neonatal complexity of this condition. In 2001 the World Health Orga Sepsis (EONS). The risk of EONS increases when a pregnant nization established the external Child Health Epidemiology woman is treated in an attempt to prolong pregnancy (to avoid Reference Group (CHERG) to develop epidemiological esti PTB), such as in cases in which intrauterine infection occurs. mates for the various etiologies of death in young children." As described herein, levels of biomarkers in cord blood In 2003, building on the work of CHERG, it was established samples from newborns who subsequently developed EONS that prematurity accounts for 75% of infant mortality and were increased or decreased by 3-fold or more (at least three 10% of the 10.6 million yearly deaths in children younger fold), compared to controls. Newborns matched for gesta than five years.' The latest U.S. vital statistics (2007) report a tional age (GA), low cord blood (CB) CB IL-6 (about 9 12.7% rate of PTB. Compared to 1990, the percentage of pg/mL) and delivered in the setting of idiopathic PTB or cord infants delivered <37 completed weeks of gestation has blood from a group of term babies with uncomplicated preg climbed 20%, resulting in ~550,000 premature infants born nancy and delivery may serve as negative controls. These annually: 60,000 of them have a birthweight <1500 grams. groups may serve to establish the methodological baseline or As support in the NBSCU has improved, more low- and reference point. Alternatively, a control or reference can be a very-low-birth weight (VLBW) infants survive. It has thus premature neonate who progressed to EONS and/or at least become clear that improving neonatal outcomes associated one related complication of prematurity (preterm birth).