Introduction
Historically, the concept of a drug that could extend lifespan and restore youthful vitality seemed more like fantasy than feasible science. However, recent advancements in single-cell technology and immunotherapy have brought us closer to making this dream a reality.
In this context, a recent study by Amor et al. (2024), published in the journal Nature Aging, has demonstrated the potential of a novel targeted therapy using engineered immune cells to alleviate the symptoms associated with physiological aging and offer promising insights into extending a healthy lifespan.
This study focuses on cellular senescence, a stress response where cells permanently stop dividing and produce inflammatory substances and enzymes that remodel tissue. In young individuals, this process helps heal wounds and suppress tumors by recruiting immune cells to clear out senescent cells and restore tissue health. However, as we age, increased tissue damage and a weakened immune system lead to the accumulation of these senescent cells, creating chronic inflammation and various age-related diseases. Current approaches to remove senescent cells, called senolytic therapies, mainly involve drugs that need frequent administration and are often not targeted. In contrast, CAR (chimeric antigen receptor) T cells—a therapy that uses engineered immune cells to target specific proteins on senescent cells—can persist and remain effective for years after a single dose. Previous research showed that CAR T cells targeting the uPAR (urokinase plasminogen activator receptor) protein could efficiently remove senescent cells and reverse liver fibrosis in young mice. This study explores whether these CAR T cells can safely and effectively eliminate senescent cells in aged mice and improve their health span.
Single-Cell RNA Sequencing Sheds Light on uPAR’s Role in Aging
uPAR is essential for remodeling the extracellular matrix in wound healing and tumor growth. Physiologically, uPAR is found in some immune cells and at low levels in the bronchial epithelium. Senescent cells are found to have elevated levels of uPAR. In addition, plasma levels of soluble uPAR correlate with aging in humans, and further analysis revealed that the gene encoding uPAR, Plaur, is part of a signature used to identify senescent cells in aged tissues, making it a promising target for CAR T cell therapy to eliminate these harmful cells. Studies using RNA sequencing data from aged mice showed increased Plaur expression in several organs. Immunohistochemistry confirmed this age-related rise in uPAR protein in the liver, adipose tissue, skeletal muscle, and pancreas. This increase was paralleled by a rise in senescence-associated markers.
Single-cell RNA sequencing provided detailed insights into the types of cells expressing uPAR in aged tissues, revealing that both immune and non-immune cells exhibit this marker. Major uPAR-positive cell types include endothelial and myeloid cells in the liver, preadipocytes and dendritic cells in adipose tissue, and endothelial cells, fibroblasts, and myeloid cells in the pancreas. These uPAR-positive cells were enriched in gene signatures linked to inflammation and other age-related pathways.
This study found that senescent cells in the liver are primarily endothelial,and myeloid dendritic,myeloid cells, and preadipocytes in adipose tissue; endothelial cells, fibroblasts, and myeloid cells in the pancreas. A significant proportion of these cells are uPAR-positive. Analysis of human pancreatic tissue confirmed that uPAR-expressing cells increase with age, suggesting that targeting uPAR could mitigate aging effects.
Rejuvenating Aged Mice: The Impact of uPAR CAR T Cells
To test the effectiveness and safety of uPAR-targeting CAR T cells in aged mice, researchers infused 18-20 month-old C57BL/6 mice with CAR T cells designed to target mouse uPAR. These CAR T cells were engineered to differentiate from within the T cells allowing them to be monitored over time. Control groups either received unmodified T cells or CAR T cells targeting human CD19, which does not affect mouse cells. The results were promising: mice treated with uPAR-targeting CAR T cells showed a significant reduction in senescent cells in the pancreas, liver, and adipose tissue. Additionally, mice treated with the uPAR-targeting CAR T cells exhibited lower levels of pro-inflammatory cytokines, linked to reduced “inflammaging,” and overall improved health. Notably, the treatment was well-tolerated. The mice did not show signs of morbidity, weight loss, or changes in blood chemistry; moreover, no tissue damage was observed during the microscopic evaluation.
Metabolic improvements were also notable. uPAR-targeting CAR T treated mice had lower fasting glucose levels and improved glucose tolerance relative to controls, which correlated to improved insulin sensitivity and enhanced pancreatic beta cell function. Exercise capacity also improved post-treatment. This metabolic improvement was accompanied by an expansion and activation of CAR T cells in the liver and spleen. Majority of these cells were cytotoxic CD8+ T cells in the liver and CD4+ T cells in the spleen, indicating that the CAR T cells were actively targeting and eliminating senescent cells, contributing to the observed health benefits. Overall,these findings demonstrate that uPAR CAR T cells can safely and effectively eliminate senescent cells in aged mice, improving metabolic and physical functions associated with aging.
Prophylactic administration of uPAR CAR T cells in young mice also showed promising results in limiting metabolic decline as they aged. Treated mice also exhibited significantly lower fasting glucose levels, improved glucose tolerance, and enhanced pancreatic beta cell function compared to controls. These mice also exhibited higher exercise capacity at nine months of age, though this effect waned over time. These improvements correlated with a significant decrease in uPAR-positive cells in the pancreas, liver, and adipose tissue. These findings highlight the significant contribution of uPAR-positive cells to metabolic dysfunction in both aged and obese mice. Targeting these cells with CAR T cell therapy offers a promising approach to mitigating age-related and obesity-induced metabolic decline.
Extending Youth: The Persistent Effects of uPAR CAR T Cells
Unlike small molecules, CAR T cells can persist in the organism and exert their effects over time. In human cancer patients, CAR T cells have been noted to persist for up to 10 years after the initial infusion. This raises the question of whether administering uPAR CAR T cells to young animals could prevent or delay age-related conditions later in life. To explore this, researchers infused young mice (3 months old) with a single dose of uPAR-targeting CAR T cells and monitored them over their natural lifespan. Despite initially lower numbers of uPAR-positive cells compared to aged animals, uPAR CAR T cells were detectable in the spleens and livers of treated mice 12 months after infusion. These persisting cells, mostly cytotoxic CD8+ T cells with memory and effector phenotypes, suggest that uPAR CAR T cells can persist and expand over time due to increased antigen stimulation as the frequency of target cells rises with age.
uPAR CAR T Cells: A Novel Treatment for Metabolic Syndrome
Many features of metabolic syndrome in aged mice can be reproduced in young animals on a high-fat diet (HFD), which accelerates the aging process and accumulates senescent cells. To test the therapeutic potential of uPAR CAR T cells in this context, researchers modeled metabolic syndrome by feeding mice an HFD and then treating them with uPAR-targeting CAR T cells. Mice treated with uPAR CAR T cells showed significantly lower body weight, better-fasting blood glucose levels, and improved glucose and insulin tolerance compared to controls. These benefits persisted for 2.5 months after infusion and were accompanied by a decreased senescent cell burden in the pancreas, liver, and adipose tissue.
To test the preventive potential of uPAR CAR T cells, researchers administered them to young mice for 1.5 months before placing them on an HFD. Remarkably, this prophylactic treatment blunted the accumulation of senescent cells over time and was associated with decreased weight gain and glucose levels 3.5 months after infusion. The uPAR CAR T cells were detectable and enriched in the spleens and livers of treated mice, composed mainly of CD8+ T cells with an effector phenotype. This preventive effect on metabolic dysfunction was sustained for at least 5.5 months despite continuous exposure to an HFD. The persistence and effectiveness of uPAR CAR T cells in both therapeutic and prophylactic contexts underscore their potential as a long-lasting treatment strategy.
Conclusion
This study highlights the critical role of single-cell transcriptomics in advancing our understanding of cellular senescence and the potential of CAR T cell therapy. Using single-cell RNA sequencing, researchers identified uPAR as a critical marker of senescent cells across various tissues. This detailed mapping enabled the development of uPAR-targeting CAR T cells, which precisely eliminated senescent cells and improved metabolic and physical functions in aged mice. The findings underscore the transformative potential of single-cell transcriptomics in identifying therapeutic targets and developing innovative treatments for age-related diseases.
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Outsourcing Bioinformatics Analysis: How Bridge Informatics (BI) Can Help
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Shahrzad Ghazisaeidi, PhD, Data Scientist, Bridge Informatics
Shahrzad specializes in high-throughput sequencing, data pre-processing, and downstream analysis of transcriptomic and epigenetic landscapes. She is particularly passionate about developing innovative tools for drug repurposing.
Prior to joining Bridge Informatics, Shahrzad served as a Postdoctoral Associate at the Hospital for Sick Children in Toronto, Canada where she researched the epigenetics of peripheral nerve injury models.
Shahrzad earned her Ph.D. in Physiology and Neuroscience from the University of Toronto. Her studies focused on the sex-dependent and independent gene regulation of peripheral nerve injury. Currently based in Toronto, Shahrzad balances her professional pursuits with personal interests by making time for yoga, martial arts, and poetry.
