Today’s open access paper reports on the use of a prodrug senolytic strategy to reverse aspects of aging in mice via the selective destruction of senescent cells. A prodrug is a small molecule, usually innocuous, that can be converted into an active drug molecule by the action of specific proteins in the body. For example a drug can be made into a prodrug by the addition of further chemical structure that (a) renders it inert, and (b) is cleaved away by an enzyme inside cells. Ideally, the inactive prodrug is designed such that this conversion to an active drug molecule only takes place where and when the drug is needed.
Senescent cell accumulation with age is an important cause of age-related degeneration and disease. Senescent cells are characterized by high levels of β-Galactosidase, known as senescence-associated β-Galactosidase (SA-β-Gal). Since β-Galactosidase is an enzyme that cleaves glycosidic bonds, it is possible to turn many types of drug into prodrugs that only activate to meaningful levels inside senescent cells by attaching structures that will be removed by β-Galactosidase. Researchers have recently demonstrated that this can be done with the chemotherapeutic drug navitoclax. Navitoclax is the worst of the effective first generation senolytics: it certainly kills senescent cells, and is somewhat specific, but it also kills far too many other cells for comfort. It has significant and unpleasant side-effects, but when it is made into a prodrug, these problems go away.
One doesn’t have to use senolytic drugs as a basis for the prodrug. The results below were obtained using a fairly generic cytotoxic chemotherapeutic drug. More or less any cell-killing drug will do, so long as (a) it can be made inert with a structure that will be cleaved away by β-Galactosidase, and (b) the difference in amount of β-Galactosidase between normal cells and senescent cells is enough to make the difference between too few drug molecules to produce any measurable effect and sufficient drug molecules to kill the cell.
Frailty is connected to cellular aging, which in turn is connected to cellular senescence. Senescent cells are permanently withdrawn from the cell cycle and generally develop a persistent pro-inflammatory phenotype called the senescence-associated secretory phenotype (SASP) which is comprised of proinflammatory cytokines and chemokines. Selective killing of senescent cells with therapeutics (i.e., senolytics) have gained attention as a new therapeutic approach for age-related diseases. Targeting of pro-survival Senescent Cell Anti-apoptotic Pathways (SCAPs) has emerged as the primary strategy for senescent cell killing.
The translational value of many senolytic drugs in vivo is limited due to their chronic toxicity. The identification of agents that selectively kill senescent cells while sparing other cell populations represents a scientific challenge. Current senolytic drugs target molecular pathways shared between senescent and proliferating cells, thus achieving cell killing but not specificity. As a matter of fact, many known senolytic agents were initially developed as cytotoxic anti-cancer agents and subsequently repurposed for ‘selective’ removal of senescent cell populations.
Senescent cells are characterized by a notable change in biological properties such as an increase in the levels of mitochondria, reactive oxygen species, lysosomal content, and upregulation of many lysosomal proteins, including the lysosomal enzyme senescence-associated β-galactosidase (SA-βGal). Recently, a promising strategy has been proposed based on galactose-derivative prodrugs. These prodrugs are selectively activated in senescent cells upon conversion into the parent active drug by the hydrolase activity of SA-βGal. In particular, specific senotoxic compounds such as duocarmycin, gemcitabine, and navitoclax have been modified into galacto-derivative prodrugs showing increased selectivity in targeting senescence cells and efficacy in treating cancer and aged mouse models.
Here, we report a novel prodrug design to target senescent cells, allowing systemic removal of senescent cells in geriatric mice without noticeable side effects. We took advantage of the senescence-specific activity of SA-βGal in the design of a non-toxic senolytic prodrug derivative of the compound 5-Fluorouridine, a metabolic precursor of the clinically approved anti-cancer medication 5-Fluorouracil. We first tested the specificity of this prodrug on senescent cells in vitro. We then confirmed safety and efficacy of the prodrug in young (5 month-old), aged (22 month old) and in geriatric (30 month old) mice. Importantly, we showed that geriatric mice that received the prodrug treatment for four weeks altogether improved significantly: 1) frailty profile; 2) skeletal muscle function; 3) muscle stem cell function; 4) cognitive function; and 5) survival.