A chemistry-driven approach to Senolytic Bispecific T-Cell Engagers

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Cellular senescence involves the stress-induced, durable cell-cycle arrest of previously replication-competent cells and has been associated with a plethora of age-related pathologies, such as liver and lung fibrosis, atherosclerosis, diabetes, and osteoarthritis.Since most small molecules with senolytic activity lack potency and produce substantial side effects, there is growing interest in the development of senolytic agents that selectively eliminate senescent over healthy cells. The present project will tackle this problem by establishing a new, chemistry-driven approach for the assembly of unique Bispecific T-Cell Engagers (BiTEs) with senolytic activity.This will be achieved by designing two orthogonal reactions that are specific to cysteine (one of them exclusively targeting N-terminal Cys and therefore allowing asymmetric ligation), for the assembly of a library of bispecific antibodies (bsAbs) from individual Cys-tagged monoclonal antibodies (mAbs) and avoiding the need to clone, express, purify and test each bsAbs one by one. The approach will then consist on expressing two sets of Cys-tagged antibodies, one targeting cell-surface antigens on hematopoietic and immune cells, and the other targeting senescent cells, which will be ligated to each other through designed linkers into bsAbs specific to senescent and T-cells antigens, and therefore capable to trigger an immune cytotoxic response towards senescent cells. To profile this library in vitro, the immunomodulatory properties of the senolytic BiTEs will be inferred from cell lysis and viability assays from cocultures of senescent cells and peripheral blood mononuclear cells. Following this, the immunomodulatory effects of selected bsAbs with senolytic activity will be elucidated using -omics and immunological tools. The technology developed will not be limited to bsAbs but will allow being extended to the ligation of any type of proteins, enabling future applications in protein-based drug discovery.