Stimulus-responsive assembly of nonviral nucleocapsids.

Controlled assembly of a protein shell around a viral genome is a key step in the life cycle of many viruses. Here we report a strategy for regulating the co-assembly of nonviral proteins and nucleic acids into highly ordered nucleocapsids in vitro. By fusing maltose binding protein to the subunits of NC-4, an engineered protein cage that encapsulates its own encoding mRNA, we successfully blocked spontaneous capsid assembly, allowing isolation of the individual monomers in soluble form. To initiate RNA-templated nucleocapsid formation, the steric block can be simply removed by selective proteolysis. Analyses by transmission and cryo-electron microscopy confirmed that the resulting assemblies are structurally identical to their RNA-containing counterparts produced in vivo. Enzymatically triggered cage formation broadens the range of RNA molecules that can be encapsulated by NC-4, provides unique opportunities to study the co-assembly of capsid and cargo, and could be useful for studying other nonviral and viral assemblies.

Medicinal Chemistry of Oligonucleotide Drugs - Milestones of the Past and Visions for the Future.

The oligonucleotide therapeutics field has blossomed in recent years, with thirteen approved drugs today and the promise of accelerated growth in coming years. Much of the progress in this field is due to advances in the medicinal chemistry of oligonucleotides,combined with a judicious choice of molecular targets and disease areas. In this perspective, we describe the growth of this new class of drugs highlighting selected milestones in oligonucleotide medicinal chemistry.

IL-1 mediates microbiome-induced inflammaging of hematopoietic stem cells in mice.

Aging is associated with impaired hematopoietic and immune function caused in part by decreased fitness in the hematopoietic stem cell (HSC) population and an increased myeloid differentiation bias. The reasons for this aging-associated HSC impairment are incompletely understood. Here we demonstrate that older specific pathogen free (SPF) wild-type (WT) mice in contrast to young SPF mice produce more interleukin-1a and interleukin-1b (IL-1a/b) in steady-state bone marrow (BM), with most of the IL-1a/b being derived from myeloid BM cells. Furthermore, blood from steady-state older SPF WT mice contains higher levels of microbe-associated molecular patterns, specifically TLR4 and TLR8 ligands. In addition, BM myeloid cells from older mice produce more IL-1b in vitro, and older mice show higher and more durable IL-1a/b responses upon stimulation with lipopolysaccharide in vivo. To test whether HSC aging is driven by IL-1a/b, we evaluated HSCs from IL-1 receptor 1 (IL-1R1) knockout (KO) mice. Indeed, older HSCs from IL-1R1KO mice show significantly mitigated aging-associated inflammatory signatures. Moreover, HSCs from older IL-1R1KO and from germ-free mice maintain unbiased lymphomyeloid hematopoietic differentiation upon transplantation, thus resembling this functionality of young HSCs. Importantly, in vivo antibiotic suppression of microbiota or pharmacologic blockade of IL-1 signaling in older WT mice was similarly sufficient to reverse myeloid-biased output of their HSC populations. Collectively, our data define the microbiome/IL-1/IL-1R1 axis as a key, self-sustaining and also therapeutically partially reversible driver of HSC inflammaging.

Microglia are effector cells of CD47-SIRPα antiphagocytic axis disruption against glioblastoma.

Glioblastoma multiforme (GBM) is a highly aggressive malignant brain tumor with fatal outcome. Tumor-associated macrophages and microglia (TAMs) have been found to be major tumor-promoting immune cells in the tumor microenvironment. Hence, modulation and reeducation of tumor-associated macrophages and microglia in GBM is considered a promising antitumor strategy. Resident microglia and invading macrophages have been shown to have distinct origin and function. Whereas yolk sac-derived microglia reside in the brain, blood-derived monocytes invade the central nervous system only under pathological conditions like tumor formation. We recently showed that disruption of the SIRPα-CD47 signaling axis is efficacious against various brain tumors including GBM primarily by inducing tumor phagocytosis. However, most effects are attributed to macrophages recruited from the periphery but the role of the brain resident microglia is unknown. Here, we sought to utilize a model to distinguish resident microglia and peripheral macrophages within the GBM-TAM pool, using orthotopically xenografted, immunodeficient, and syngeneic mouse models with genetically color-coded macrophages (Ccr2RFP) and microglia (Cx3cr1GFP). We show that even in the absence of phagocytizing macrophages (Ccr2RFP/RFP), microglia are effector cells of tumor cell phagocytosis in response to anti-CD47 blockade. Additionally, macrophages and microglia show distinct morphological and transcriptional changes. Importantly, the transcriptional profile of microglia shows less of an inflammatory response which makes them a promising target for clinical applications.