Biology subjects

Böhnert, V.

Publications and source records attributed to Böhnert, V..

4 recordsLinked to original sources

bioRxiv2021

A cGAMP-containing hydrogel for prolonged SARS-CoV-2 RBD subunit vaccine exposure induces a broad and potent humoral response

The receptor binding domain (RBD) of the SARS-CoV-2 virus spike protein has emerged as a promising target for generation of neutralizing antibodies. Although the RBD subunit is more stable than its encoding mRNA, RBD-based subunit vaccines have been hampered by RBDs poor immunogenicity. We hypothesize that this limitation can be overcome by sustained co-administration with a more potent and optimized adjuvant than standard adjuvants. The endogenous innate immune second messenger, cGAMP, holds promise as potent activator of the anti-viral STING pathway. Unfortunately, delivery of cGAMP as a therapeutic exhibits poor performance due to poor pharmacokinetics and pharmacodynamics from rapid excretion and degradation by its hydrolase ENPP1. To overcome these limitations, we sought to create an artificial immunological niche enabling slow release of cGAMP and RBD to mimic natural infections in which immune activating molecules are co-localized with antigen. Specifically, we co-encapsulated cGAMP and RBD in an injectable polymer-nanoparticle (PNP) hydrogel. This cGAMP-adjuvanted hydrogel vaccine elicited more potent, durable, and broad antibody responses with improved neutralization as compared to dose-matched bolus controls and hydrogel-based vaccines lacking cGAMP. The cGAMP-adjuvanted hydrogel platform developed is suitable for delivery of other antigens and may provide enhanced immunity against a broad range of pathogens.

bioengineering↗

bioRxiv2021

Probing pathophysiology of extracellular cGAMP with substrate-selective ENPP1

The biology of the immune second messenger cGAMP depends on its cellular localization. cGAMP, which is synthesized in response to cytosolic double-stranded DNA, also exists in the extracellular space as a paracrine immunotransmitter that enhances the anticancer immune response. However, the role of extracellular cGAMP is unexplored outside of cancer due to a lack of tools to systemically manipulate it. The extracellular enzyme ENPP1, the only known hydrolase of cGAMP, is a promising target. However, because ENPP1 also degrades extracellular ATP, using genetic knockouts of ENPP1 to study extracellular cGAMP leads to confounding effects. Here we report the H362A point mutation in ENPP1, the dominant cGAMP hydrolase, which selectively abolishes ENPP1s ability to degrade cGAMP, while retaining activity toward other substrates. H362 is not necessary for binding cGAMP or the catalytically-essential zinc atoms but instead supports the in-line reaction geometry. H362 is evolutionarily conserved down to bacteria, suggesting an ancient origin for extracellular cGAMP biology. Enpp1H362A mice do not display the systemic calcification seen in Enpp1-/- mice, highlighting the substrate-specific phenotype of ENPP1. Remarkably, Enpp1H362A mice were resistant to HSV-1 infection, demonstrating the antiviral role of endogenous extracellular cGAMP. The ENPP1H362A mutation is the first genetic tool to enable exploration of extracellular cGAMP biology in a wide range of tissues and diseases.

immunology↗

bioRxiv2020

Murine M1 macrophages are among the direct responders to tumor-derived extracellular cGAMP and their human counterparts use SLC46A2 to import cGAMP

Administration of exogenous CDNs to activate the cGAMP-STING pathway is a promising therapeutic strategy to unleash the full potential of cancer immunotherapy. This strategy mirrors the role of endogenous extracellular cGAMP, an immunotransmitter that is transferred from cancer cells to cGAMP-sensing cells in the host, promoting immunity. However, the CDN import mechanisms used by host cells within tumors remain unknown. Here we identified the protein SLC46A2 as the dominant cGAMP importer in primary human monocytes. Furthermore, we discovered that monocytes and M1-polarized macrophages directly sense tumor-derived extracellular cGAMP in murine tumors. Finally, we demonstrated that SLC46A2 is the dominant cGAMP importer in monocyte-derived macrophages. Together, we provide the first cellular and molecular mechanisms of cGAMP as an immunotransmitter, paving the way for effective STING pathway therapeutics.

cancer biology↗

bioRxiv2020

The LRRC8A:C Heteromeric Channel Is a cGAMP Transporter and the Dominant cGAMP Importer in Human Vasculature Cells

Extracellular 23-cyclic-GMP-AMP (cGAMP) is an immunotransmitter secreted by cancer cells and taken up by host cells to activate the anti-cancer STING pathway. No cGAMP exporter has been identified, and SLC19A1, a recently identified cGAMP importer, does not account for the import activity in most cell types. Here, we identify the LRRC8A:C heteromeric channel, a volume-regulated anion channel (VRAC), as a cGAMP transporter. This channel mediates cGAMP import or export depending on the cGAMP chemical gradient, and channel activation or inhibition modulates cGAMP transport. Other 23-cyclic dinucleotides are also transported by LRRC8A:C channels, including the investigational cancer therapeutic ADU-S100. Furthermore, we demonstrate that the LRRC8A-containing channel is the dominant cGAMP importer in primary human vasculature cells. Given tumor vasculatures regulation of immune infiltration and its disruption in response to STING agonists, we have uncovered a leading molecular mechanism for extracellular cGAMP signaling in this important anti-cancer target.

biochemistry↗