Targeting immune cells in the aged brain reveals that engineered cytokine IL-10 enhances neurogenesis and improves cognition.

T cells in the aged brain exhibit an exhaustion signature distinct from those in the young brain.

• T cells with an exhaustion signature were detected in the old brain using single-cell transcriptomic approaches.
• These exhausted T cells were identified as a target population for immune-based intervention in the aging brain.
• The presence of exhausted T cells in the aged brain suggested a rationale for checkpoint inhibitor-based approaches.

Targeting exhausted T cells in the aged brain with an engineered checkpoint inhibitor (RIPR-PD1) led to T cell expansion and strong pro-inflammatory responses in multiple brain cell types, notably microglia.

• RIPR-PD1 is described as a 'potent engineered checkpoint inhibitor' delivered directly to the brain.
• T cell expansion was observed following RIPR-PD1 treatment in the old brain.
• Strong pro-inflammatory responses were detected in many brain cell types, with microglia being notably affected.
• This pro-inflammatory outcome indicated that checkpoint inhibition alone was not sufficient to restore beneficial immune balance in the aged brain.

Wild-type IL-10 boosted anti-inflammatory responses in old microglia but also triggered pro-inflammatory signaling, indicating a mixed response.

• IL-10 was used to target age-related inflammatory imbalances in microglia.
• Anti-inflammatory responses were upregulated in old microglia following IL-10 treatment.
• Unexpectedly, IL-10 also triggered pro-inflammatory signaling, representing an undesirable side effect in the aged brain context.
• This dual response motivated the development of an engineered IL-10 variant that could uncouple the two signaling outcomes.

An engineered IL-10 variant that uncouples pro- and anti-inflammatory responses positively impacted the transcriptome of multiple cell types in the aged brain.

• The engineered IL-10 variant was designed to selectively activate anti-inflammatory signaling while avoiding pro-inflammatory signaling.
• Transcriptomic analysis revealed positive impacts across multiple brain cell types following treatment with the engineered IL-10 variant.
• The variant was delivered directly to the brain, enabling targeted action on resident immune and other brain cell populations.
• Direct brain delivery was used as the strategy to administer engineered proteins to aged brain tissue.

The engineered IL-10 variant enhanced neurogenesis in aged mice.

• Neurogenesis, which declines with age, was boosted following treatment with the engineered IL-10 variant.
• This finding implicates microglial inflammatory balance as a regulator of adult neurogenesis in the aged brain.
• Enhanced neurogenesis was identified as one of the functional outcomes associated with the engineered IL-10 variant treatment.

The engineered IL-10 variant improved cognition in aged mice.

• Cognitive improvement was observed in aged mice treated with the engineered IL-10 variant.
• This represents a functional behavioral outcome in addition to transcriptomic and cellular changes.
• Cognitive improvement was achieved through direct brain delivery of the engineered cytokine, targeting the aged brain immune microenvironment.

A combined strategy of engineered proteins and direct brain delivery was used to target immune cell populations within the old brain.

• The study employed direct intracranial or brain-targeted delivery to bypass systemic distribution and focus effects on the aged brain.
• Both an engineered checkpoint inhibitor (RIPR-PD1) and an engineered cytokine (IL-10 variant) were tested using this delivery approach.
• This approach allowed interrogation of specific immune cell populations, particularly T cells and microglia, within the aged brain microenvironment.