This is an administrative supplement proposal to the parent grant HD98389 (2019-24) titled ?Anti-inflammatory drug target to reduce adverse pregnancy outcomes?. The goal of the parent grant is to develop Anakinra (recombinant human IL1receptor antagonist) as a safe and effective therapeutic for intrauterine infection/inflammation mediated preterm labor and fetal inflammation. In the parent grant, pregnant Rhesus macaques will be given Anakinra in a model of intrauterine inflammation (IUI). We will now leverage the resources of the parent grant to expand the Aims of the parent grant. Fetal neuro-inflammation is associated with later development of cerebral palsy (CP) and autism spectrum disorders (ASD) with significant medical and public health implications. As part of the parent grant activities, we have demonstrated that inhibition of IL1 signaling using Anakinra significantly decreased fetal inflammation induced by intraamniotic injection of E. coli LPS. We have now developed capability of extracting intact nuclei from fetal frozen brain sample and to do single cell sequencing. Nuclear as compared to cytosolic RNA seq has the advantage of not needing protease digestion at 37oC thereby eliminating artifactual gene expression. Furthermore, nuclear RNA seq can be done in archived frozen brain sample thereby sparing precious animal use and reducing costs. In this administrative supplement grant, we will test the hypothesis that inhibition of IL1 signaling by Anakinra will decrease region specific fetal neuro inflammation induced by intraamniotic LPS. We propose doing single nuclear mRNA seq from 2 regions of the brain ? the peri-ventricular white matter, and the prefrontal cortex ? regions implicated in the pathogenesis of CP and ASD. We will compare region specific single nuclear mRNA seq in three groups of Rhesus macaque fetuses at about 80% gestation: 1) Controls ? intraamniotic injection (IA) of saline, 2) IA LPS, 3) IA LPS + Anakinra. All exposures are for 16h, a time point that we previously demonstrated to be optimum for the proposed studies. The animal resources funded by the parent grant are already available. Thus, single cell transcriptomic studies will be greatly facilitated and feasible within the year of funding. This fetal brain transcriptomic study will reveal mechanistic insights in the pathogenesis of neuroinflammation resulting from IUI, and identify leads for biomarker sets for CP and ASD. The study will also further assist in pre-clinical development of Anakinra as a therapeutic candidate for IUI.
Intrauterine inflammation is an important cause of preterm labor and injury to the brain of the unborn fetus increasing the risk for childhood neurologic problems like cerebral palsy and autism. This grant will discover genes responsible for brain injury using advanced transcriptomics and study whether Anakinra, an anti- inflammatory drug will decrease the injury. This knowledge will be important to develop new biomarkers of newborn brain injury, cerebral palsy, and autism.
