A senolytic immunotoxin eliminates p16INK4a-positive T cells and ameliorates age-associated phenotypes of CD4+ T cells in a surface marker knock-in mouse

Abstract

Senescent cells were recently shown to play a role in aging-related malfunctions and pathologies. This consensus has been facilitated by evidence from senolytic model mice capable of eliminating senescent cells in tissues using well-characterized senescent markers, such as p16INK4a (hereafter p16). However, since the incomplete or artificial gene expression regulatory regions of manipulated marker genes affect their cognate expression, it currently remains unclear whether these models accurately reflect physiological senescence. We herein describe a novel approach to eliminate p16-expressing cells from mice at any given point in time, generating a new type of knock-in model, p16hCD2 mice and a toxin-conjugated anti-human CD2 antibody (hCD2-SAP) as an inducer. p16hCD2 mice possess an intact Cdkn2a locus that includes a p16 coding region and human CD2 (hCD2) expression unit. We confirmed cognate p16-associated hCD2 expression in mouse embryonic fibroblasts (MEFs) and in several tissues, such as the spleen, liver, and skin. We detected chronological increases in the hCD2-positive population in T lymphocytes that occurred in a p16-dependent manner, which reflected physiological aging. We then confirmed the high sensitivity of hCD2-SAP to hCD2 and validated its efficacy to remove p16-positive cells, particularly in T lymphocytes. The multiple administration of hCD2-SAP for a prolonged p16-positive cell deficiency partially restored aging-related phenotypes in T lymphocytes, such as the contraction of the CD4+ naïve population and expansion of senescence-associated T cells. Our novel approach of targeting p16-positive senescent cells will provide novel insights into the mechanisms underlying physiological aging in vivo.

Keywords

• Aging
• p16
• Immunotoxin
• Immunosenescence
• naïve T
• SA-T