Engineered CAR T cells to the rescue: fighting the senescence

Biological aging, an intrinsic feature of living organisms, is the result of the combination of environmental factors, time and genetic. The most important factors that occurs during aging are inflammation, immune aging and senescence [1]. Inflammation and immune aging are relatively easy to understand, but... how about senescence? Senescence is defined as an irreversible form of long-term cell-cycle arrest (which means that cells are no longer capable to proliferate or exert its functions) caused by an excessive stress or damage. In a physiological context, senescence limits the proliferation of damaged cells to eliminate accumulated harmful factors and acts as a tumour-suppressive mechanism with a beneficial role in wound healing responses [2, 3]. The dark side of senescence arises in a pathological context, when the accumulation of senescent cells generates an inflammatory environment that leads to chronic tissue damage. There are several pathologies, such as lung fibrosis, atherosclerosis, diabetes and osteoarthritis [4] in which pathological senescence processes are implicated. Targeting and eliminating senescent cells from damaged tissues can ameliorate the symptoms of these pathologies, and the elimination of senescent cells from healthy organisms can even promote longevity. 

A recent paper published in Nature by Corina Amor et al proposed the use of chimeric antigen receptor (CAR) T cells targeting senescent cells as a therapeutic senolytic agents [5]. The development of senolytic agents that specifically eliminates senescent cells has been of great interest in recent years. Authors hypothesize that CAR T cells directed against senescence specific surface antigens may constitute a robust approach to eliminate senescent cells while keeping the integrity of the health tissue. CAR T cells are engineered T cells which express a modified T cell receptor (TCR) that allows the specific recognition of a given protein. CAR receptors combine many facets of a normal TCR into a single protein. They link an extracellular antigen recognition domain to an intracellular signaling domain which activates the T cell when an antigen (the protein of interest) is bound. CARs are composed of four regions: an antigen recognition domain, and extracellular hinge region, a transmembrane domain and an intracellular T-cell signaling domain.  

Researchers first needed to identify a cell surface protein broadly and specifically upregulated in senescent cells, so they compared the transcriptome of three different models of senescence: 1) therapy-induced senescence cells in mouse lung adenocarcinoma; 2) oncogene-induced senescence cells in mouse hepatocytes; and 3) culture-induced senescence in mouse hepatic stellate cells. Transcript candidates should encode molecules located in the plasma membrane, upregulated in all data sets and highly expresses on target cells but not in vital tissues. They found PLAUR, a gene encoding the urokinase-type plasminogen activator receptor (uPAR) as the best candidate because it was highly upregulated in senescent cells but absent in many vital organs. uPAR promotes the degradation of the extracellular matrix during fibrinolysis, wound healing or tumorigenesis, functioning as a signaling receptor that promotes motility, invasion and survival of tumour cells. 

Next step was to develop a specific CAR comprising an anti-mouse uPAR single-chain variable fragment linked to human CD28 costimulatory and CD3ζ signaling domains. This CAR was transduced in human T cells and the cells were validated in cytotoxicity assays using target cells that expressed a mouse uPAR cDNA. Once the senolytic effect of the uPAR CAR T cells was confirmed in vitro, the authors evaluated the capacity of these cells in immunocompetent settings in vivo transducing T cells from C57BL/6 mice with a fully mouse CAR. Their results confirmed that uPAR-directed CAR T cells can deplete senescent cells both in vitro and in vivo without inducting severe cytokine-release (commonly known as ‘cytokine storm’) symptoms. Furthermore, they successfully applied uPAR CAR T cells against fibrosis induced by non-alcoholic steatohepatitis, efficiently eliminating senescent cells, reducing fibrosis and improving liver function.

Fig 2. Chimeric antigen receptor: How it works. 
Obtained from Leukemia & Lymphopenia society. 

More work is needed to test whether an appropriately dosed of senolytic CAR T cells have the required safety profile to be developed clinically, if they can infiltrate the areas of senescence and if they can efficiently target senescent cells producing therapeutic benefits without toxicity. Senescent cells are not as easy to reach as tumour cells because they do not divide or create an immunosuppressive microenvironment. For that reason, senescent cells are harder to reach and to be identified, targeted and eliminated by CAR T cells within a given tissue. Nevertheless, as senescence has been linked to disorders of chronic tissue damage associated with aging, the senolytic CAR T cells may have broad therapeutic potential. 

In the present there are several CAR T cells trials generating impressive results in patients with blood cancers such as acute lymphoblastic leukemia, chronic lymphocytic leukemia and multiple myeloma. Nevertheless, and due to some side effects of this therapy (such as cytokine release syndrome due to the clonal proliferation of CAR T cells, and an occasional neurotoxicity that causes swelling, confusion, seizures or severe headaches), larger study samples and the application of these molecules in different pathologies will help researchers to further understand how to reduce its potential toxicity and improve the management of adverse side effects. 

References

1. Dodig, S., I. Cepelak, and I. Pavic, Hallmarks of senescence and aging. Biochem Med (Zagreb), 2019. 29(3): p. 030501.

2. Demaria, M., et al., An essential role for senescent cells in optimal wound healing through secretion of PDGF-AA. Dev Cell, 2014. 31(6): p. 722-33.

3. Kang, T.W., et al., Senescence surveillance of pre-malignant hepatocytes limits liver cancer development. Nature, 2011. 479(7374): p. 547-51.

4. He, S. and N.E. Sharpless, Senescence in Health and Disease. Cell, 2017. 169(6): p. 1000-1011.

5. Amor, C., et al., Senolytic CAR T cells reverse senescence-associated pathologies. Nature, 2020. 583(7814): p. 127-132.