New Approaches to Mining the Complete Actinomycete Genome to Discover Novel Beta-Lactam Antibiotics

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ABSTRACT There is a critical need to discover and develop safe and effective antibiotics for the treatment of multidrug- resistant bacterial infections. Each year approximately 1.5 million patients in the US get hospital acquired infections (HAIs), with a mortality rate of ~6%. Nearly 70% of the bacteria causing HAIs are resistant to at least one commonly used antibiotic. Natural products (NPs) and their derivatives are the basis of many pharmaceutical advances, representing ~50% of FDA approved small molecule drugs over the past 5 decades including many important antibiotics. Genome mining for novel natural products is quickly replacing traditional approaches to antibiotic discovery. However approximately 90% of all actinomycete gene clusters are cryptic or silent, i.e., expressed at levels too low to detect using traditional antibacterial screening assays, or not expressed at all. Warp Drive Bio (WDB) has sequenced over 135,000 actinomycete strain genomes from diverse sources worldwide, and our proprietary genomic database contains ~3.5 million secondary metabolite gene clusters. Importantly ~75% of cluster families in our database have yet to be reported in the literature. This provides an unprecedented opportunity to discover structurally and mechanistically novel NPs. Our objective is to discover and develop novel β-lactams (BLs) as broad-spectrum antibiotics with improved therapeutic profiles aimed at combating current and emerging multidrug-resistant Gram (-)/(+) pathogens. BLs represent a clinically validated class widely used in antimicrobial therapy with good, broad-spectrum activity and favorable safety profiles. Unfortunately, the emergence of multi-drug resistance severely limits the clinical efficacy of current BL antibiotics, as single agents or in combination with β-lactamase inhibitors. WDB is deploying our proprietary natural product genome-mining platform to discover novel BL antibiotics. This platform offers a new paradigm in which drug discovery is initiated at the genome level, allowing rapid identification of novel compounds that would be otherwise overlooked in a traditional activity-based discovery paradigm. We first deploy our bioinformatics search engines to rapidly identify gene clusters within WDB's genomic database that encode novel BL molecules. To access the potential of silent biosynthetic gene clusters, we have developed methods to systematically replace promoters, inducing compound expression. Clusters are transferred into engineered heterologous expression hosts to produce novel molecules that are isolated as extracts in a miniaturized high throughput plate format for bioassay, and we utilize mass spectrometry analytics to rapidly identify the masses of novel BLs of interest to facilitate rapid purification for further characterization. We will apply our genome-mining platform to engineer, express, purify, and characterize the products from 10 novel BL biosynthetic clusters in this Phase I proof-of-concept study.