Dried blood spot proteomics analysis of newborn screening cards to identify prognostic markers of SIDS risk

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PROJECT SUMMARY Despite fewer deaths as a result of the Safe Sleep campaign, the Sudden Infant Death Syndrome (SIDS) remains the leading cause of post-neonatal infant mortality in the United States (0.33/1000 live births). Our research strongly implicates biological abnormalities within the brainstem that underlie an infant's vulnerability to SIDS. We have also identified SIDS-associated abnormalities within the serum and platelets taken from SIDS victims at autopsy. Together, these data suggest molecular abnormalities in the blood and brain of SIDS infants at death. What is unknown, however, is to what extent abnormalities observed at autopsy are also present at birth, and whether these or other, yet to be discovered, abnormalities can identify SIDS risk during the neonatal period, thereby providing the opportunity for intervention. Although there are no known biomarkers for SIDS risk, reports of apnea, cardiac rate abnormalities, and arousal deficits in infants who subsequently die of SIDS support the possibility that an underlying vulnerability may exist in the infant subclinically. Such subclinical abnormalities then manifest themselves when the infant is sufficiently stressed during a critical developmental period. Our laboratory has now acquired a unique dataset of dried blood spots (DBS) from neonatal screening cards from infants who later died of SIDS (n=109), died of known causes of death (demise controls, n=34), and age- and sex-matched control infants, identified as healthy at birth based on newborn screening for metabolic disorders (healthy controls (n=60)). In addition, we have corresponding brainstem tissue from autopsy in ~50% of these SIDS and demise control infants. Utilizing this dataset, we hypothesize that infants dying of SIDS display at birth abnormal protein profiles in DBSs compared to controls. Using liquid chromatography/mass spectrometry (LC/MS), we will test this hypothesis by performing an unbiased proteomic analysis of DBSs from SIDS and control infants (Specific Aim 1). Using the matching brainstem tissue we will use targeted methodologies, such as Western blotting, immunohistochemistry, and/or RNAscope to address the question of whether putative blood protein abnormalities at birth are represented in brain tissue collected at autopsy (Specific Aim 2). The use of neonatal DBSs for unbiased proteomics in SIDS is novel and the data generated from this unprecedented sample collection has potential to identify yet unknown proteins and/or pathways that play a role in SIDS pathogenesis. Furthermore, these data will allow us to assess identified abnormalities for their predictive potential for SIDS death and therefore their potential as neonatal biomarkers of SIDS risk. Given our overall goal, this application is fully aligned with NICHD's recent notice of special interest (NOT-HD-21-032) focused on identifying the root cause of SIDS with the aim to reduce SIDS-related deaths. The data to be generated from this R21 will provide novel prognostic information about SIDS infants at birth which will then be utilized as pilot data to expand upon in an R01 with new hypotheses, larger newborn datasets, and animal models.