Lebo, K. | Search bioRxiv

bioRxiv2024

A Ketogenic Diet Sensitizes Pancreatic Cancer to Inhibition of Glutamine Metabolism

Pancreatic cancer is the third leading cause of cancer death in the United States, and while conventional chemotherapy remains the standard treatment, responses are poor. Safe and alternative therapeutic strategies are urgently needed1. A ketogenic diet has been shown to have anti-tumor effects across diverse cancer types but will unlikely have a significant effect alone. However, the diet shifts metabolism in tumors to create new vulnerabilities that can be targeted (1). Modulators of glutamine metabolism have shown promise in pre-clinical models but have failed to have a marked impact against cancer in the clinic. We show that a ketogenic diet increases TCA and glutamine-associated metabolites in murine pancreatic cancer models and under metabolic conditions that simulate a ketogenic diet in vitro. The metabolic shift leads to increased reliance on glutamine-mediated anaplerosis to compensate for low glucose abundance associated with a ketogenic diet. As a result, glutamine metabolism inhibitors, such as DON and CB839 in combination with a ketogenic diet had robust anti-cancer effects. These findings provide rationale to study the use of a ketogenic diet with glutamine targeted therapies in a clinical context. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=186 SRC="FIGDIR/small/604377v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@1ebeefforg.highwire.dtl.DTLVardef@97e2ceorg.highwire.dtl.DTLVardef@1ab45c3org.highwire.dtl.DTLVardef@1713540_HPS_FORMAT_FIGEXP M_FIG C_FIG Graphical Abstract Description: Mechanistic rationale for combining a ketogenic diet and glutamine metabolism inhibitors. The combination of low glucose from a ketogenic diet and pharmacologic glutamine inhibition impairs nutrient input to mitochondria, reducing cancer growth.

cancer biology↗

bioRxiv2022

Human anelloviruses produced by recombinant expression of synthetic genomes.

Human anelloviruses are acquired universally in infancy, highly prevalent, abundant in blood, and extremely diverse. Their apparent lack of pathogenicity indicates that they are a major component of the commensal human virome. Despite their being extensively intertwined with human biology, these viruses are poorly understood. A major impediment in studying anelloviruses is the lack of an in vitro system for their production and/ or propagation. Here we show that the T cell-derived human cell line MOLT-4 can be transfected with plasmids comprising tandem anellovirus genomes to produce viral particles visualized by electron microscopy. We found that a previously described human anellovirus of the Betatorquevirus genus (LY2), as well as a second Betatorquevirus detected by sequencing DNA extracted from a human retinal pigmental epithelium (nrVL4619), can be synthesized and produced by these means, enabling further molecular virology studies. Southern blot was used to demonstrate replication, and site-directed mutagenesis of the viral genome was performed to show that the production of anellovirus in this cell line is dependent on the expression of certain viral proteins. Finally, experiments performed in mice using purified nrVL4619 particles produced in MOLT-4 cells demonstrated infectivity in vivo in the tissue of origin. These results indicate that anelloviruses can be produced in vitro and manipulated to improve our understanding of this viral family which is ubiquitous in humans and many other mammals. Applications of this work to gene therapy and other therapeutic modalities are currently under investigation. IMPORTANCEAnelloviruses are a major component of the human virome. However, their biology is not well understood mainly due to the lack of an in vitro system for anellovirus production and/or propagation. In this study, we used multiple orthogonal measures to show that two different anelloviruses belonging to the Betatorquevirus genus can be produced in a T-cell-derived human cell line, MOLT-4, via recombinant expression of synthetic genomes. Additionally, we show that anellovirus particles generated in this in vitro system demonstrate infectivity in vivo. Our findings enable new molecular virology studies of this highly prevalent, non-pathogenic, and weakly immunogenic family of viruses, potentially leading to therapeutic applications.

microbiology↗