Biology subjects

Monaco, D.

Publications and source records attributed to Monaco, D..

4 recordsLinked to original sources

bioRxiv2020

Utilizing Computational Machine Learning Tools to Understand Immunogenic Breadth in the Context of a CD8 T-Cell Mediated HIV Response

Predictive models are becoming more and more commonplace as tools for candidate antigen discovery to meet the challenges of enabling epitope mapping of cohorts with diverse HLA properties. Here we build on the concept of using two key parameters, diversity metric of the HLA profile of individuals within a population and consideration of sequence diversity in the context of an individuals CD8 T-cell immune repertoire to assess the HIV proteome for defined regions of immunogenicity. Using this approach, Analysis of HLA adaptation and functional immunogenicity data enabled the identification of regions within the proteome that offer significant conservation, HLA recognition within a population, low prevalence of HLA adaptation and demonstrated immunogenicity. We believe this unique and novel approach to vaccine design that, in combination with in vitro functional assays, offers a bespoke pipeline for expedited and rational CD8 T-cell vaccine design for HIV and potentially other pathogens with the potential for both global and local coverage.

immunology↗

bioRxiv2020

Highly multiplexed oligonucleotide probe-ligation testing enables efficient extraction-free SARS-CoV-2 detection and viral genotyping

The emergence of SARS-CoV-2 has caused the current COVID-19 pandemic with catastrophic societal impact. Because many individuals shed virus for days before symptom onset, and many show mild or no symptoms, an emergent and unprecedented need exists for development and deployment of sensitive and high throughput molecular diagnostic tests. RNA-mediated oligonucleotide Annealing Selection and Ligation with next generation DNA sequencing (RASL-seq) is a highly multiplexed technology for targeted analysis of polyadenylated mRNA, which incorporates sample barcoding for massively parallel analyses. Here we present a more generalized method, capture RASL-seq (“cRASL-seq”), which enables analysis of any targeted pathogen-(and/or host-) associated RNA molecules. cRASL-seq enables highly sensitive (down to ∼1-100 pfu/ml or cfu/ml) and highly multiplexed (up to ∼10,000 target sequences) detection of pathogens. Importantly, cRASL-seq analysis of COVID-19 patient nasopharyngeal (NP) swab specimens does not involve nucleic acid extraction or reverse transcription, steps that have caused testing bottlenecks associated with other assays. Our simplified workflow additionally enables the direct and efficient genotyping of selected, informative SARS-CoV-2 polymorphisms across the entire genome, which can be used for enhanced characterization of transmission chains at population scale and detection of viral clades with higher or lower virulence. Given its extremely low per-sample cost, simple and automatable protocol and analytics, probe panel modularity, and massive scalability, we propose that cRASL-seq testing is a powerful new surveillance technology with the potential to help mitigate the current pandemic and prevent similar public health crises.Competing Interest StatementJ.J.C. and H.B.L. are listed as inventors on a patent describing the cRASL-seq method. H.B.L. has founded a company to license and commercialize oligonucleotide probe ligation related technologies.View Full Text

pathology↗

bioRxiv2020

Protease Activity Profiling Via Programmable Phage Display

Endopeptidases catalyze the internal cleavage of proteins, playing pivotal roles in protein turnover, substrate maturation and the activation of signaling cascades. A broad range of biological functions in health and disease are controlled by proteases, yet assays to characterize their activities at proteomic scale do not yet exist. To address this unmet need, we have developed SEPARATE (Sensing EndoPeptidase Activity via Release and recapture using flAnking Tag Epitopes), which uses monovalent phage display of the entire human proteome at 90-aa peptide resolution. We demonstrate that SEPARATE is compatible with several human proteases from distinct catalytic classes, including Caspase-1, ADAM17, and Thrombin. Both well-characterized and newly identified substrates of these enzymes were detected in the assay. SEPARATE was used to discover a non-canonical Caspase-1 substrate, the E3 ubiquitin ligase HUWE1, a key mediator of apoptotic cell death. SEPARATE is a novel methodology to enable efficient, unbiased assessment of endopeptidase activity using a phage-displayed proteome.

molecular biology↗

bioRxiv2018

Deconvoluting Virome-Wide Antiviral Antibody Profiling Data

The ability to comprehensively characterize exposures and immune responses to viral infections will be critical to better understanding human health and disease. We previously described the VirScan system, a phage-display based technology for profiling antibody binding to a comprehensive library of peptides designed to represent the human virome. The previous VirScan analytical approach did not fully account for disproportionate representation of viruses in the library or for antibody cross-reactivity among sequences shared by related viruses. Here we present the AntiViral Antibody Response Deconvolution Algorithm ( AVARDA), a multi-module software package for analyzing VirScan datasets. AVARDA provides a probabilistic assessment of infection at species-level resolution by considering alignment of all library peptides to each other and to all human viruses. We employed AVARDA to analyze VirScan data from a cohort of encephalitis patients with either known viral infections or undiagnosed etiologies. By comparing acute and convalescent sera, AVARDA successfully confirmed or detected antibody responses to human herpesviruses 1, 3, 4, 5, and 6, thereby improving the rate of diagnosing viral encephalitis in this cohort by 62.5%. We further assessed AVARDAs utility in the setting of an epidemiological study, demonstrating its ability to determine infections acquired in a child followed prospectively from infancy. We consider ways in which AVARDAs conceptual framework may be further developed in the future and describe how its analyses may be extended beyond investigations of viral infection. AVARDA, in combination with VirScan and other pan-pathogen serological techniques, is likely to find broad utility in the epidemiology and diagnosis of infectious diseases.

bioinformatics↗