Cell-free DNA as a versatile analyte for the monitoring of sepsis

Sepsis is a deadly condition caused by a dysregulated immune response to infection which affects 700,000 patients per year in the US alone. Early and accurate diagnosis of sepsis, and infection underlying sepsis is critical to inform treatment choices and to prevent serious organ injury and death, but diagnostic options remain limited. This proposal addresses this unmet medical need with the development of a minimally invasive blood test based on a multi-omics profiling of circulating cfDNA to simultaneously inform host responses, quantify organ injury and identify infection underlying sepsis Our recent pilot work provides significant support for the feasibility of this proposal. First, we have found that metagenomic sequencing of circulating cell-free DNA (cfDNA) can be used to detect a broad range of viral and bacterial pathogens in blood. To overcome the challenge of environmental contamination which limits the specificity of metagenomic cfDNA sequencing we have developed Coffee-seq, a metagenomic sequencing assay that is robust against environmental contamination. The core idea of Coffee-seq is to tag the DNA in the sample prior to sample preparation with a label that can be recorded by DNA sequencing. Any contaminating DNA that is introduced in the sample after tagging can then be bioinformatically identified and removed. Our proof-of-principle work demonstrates that Coffee-seq leads to a reduction of noise due to contamination by up to three orders of magnitude. Coffee-seq thereby dramatically improves the specificity of metagenomic sequencing assays. Second, we have developed cfDNA assays to inform injury to host tissues and organs. This test employs genome-wide profiling of methylation marks of cfDNA that are cell, tissue and organ-type specific to trace their tissues-of-origin, and to quantify tissue-specific injury. We have demonstrated the utility of this assay to identify host-tissue injury related to urinary tract infection, COVID-19, and Graft-Versus Host Disease, a frequent complication of stem cell transplantation. We have two aims: In Aim 1, we will test the utility of Coffee-seq to identify infection underlying sepsis with high sensitivity and specificity. In Aim 2, we will investigate the utility of a cfDNA metagenomic sequencing assay to inform sepsis related organ dysfunction. This is an exploratory study that tests the utility of cfDNA as a versatile analyte to monitor sepsis. We will test this highly translational concept with a retrospective study of 300 bio- banked samples (Cornell Biobank of Critical Illness), including 120 samples from septic patients with positive blood culture, and 180 samples from patients with non-infectious critical illness. Successful implementation of this study will lead to new avenues to identify infection underlying sepsis, to classify sepsis subtypes, to monitor sepsis-related organ damage, and to guide treatment decisions.