ABT-737 is Another New Senolytic Drug Candidate, Working via BCL-W, BCL-XL, and Induced Apoptosis

Senolytic therapies are those that cause senescent cells to die while causing minimal side-effects. Developing methods to selectively destroy senescent cells has been on the SENS rejuvenation research agenda for going on fifteen years, based on strong evidence from many fields, but only recently have factions within the broader research community started to pick up on this approach to treating one of the causes of aging. Over the past two years a tipping point of sorts was reached and passed, and now a number of drug candidates are emerging from research groups, and the startup Oisin Biotechnologies has a more selective gene therapy approach to achieve the same aim. The open access paper I'll point out today describes one of these candidates, along with animal data that shows it destroying between a quarter and a half of senescent cells in a few tissues. This is on a par with the performance of some of the other candidates that have produced improved measures of health in mice.

Why destroy senescent cells? Because they help to make us old. Cells become senescent in response to reaching the Hayflick limit to the number of divisions, or when suffering damage, especially DNA damage likely to produce cancerous mutations, or a toxic local environment that seems likely to produce that sort of damage. Senescent cells cease to replicate and most self-destruct via apoptosis, or are targeted by immune cells for destruction. Some linger however, secreting a problematic mix of signals that induce inflammation and remodeling of tissue structures, while also encouraging neighboring cells to become senescent. As the years pass ever greater numbers of these cells cause ever greater disarray, contributing meaningfully to the development and progression of all common age-related diseases, and ultimately the tissue and organ failures that cause death. If all senescent cells were periodically removed, however, never permitted to assemble in large numbers, then this part of the aging process would be eliminated, the span of healthy life extended, and age-related disease pushed off that much further into the future. This has been demonstrated in a life span study in mice, in which continuous senescent cell removal via genetic engineering produced a 25% extension of median life span.

How to destroy senescent cells? In past years, I predicted that targeted therapies like those under development in the cancer research community would be used, combining a smart detector of cell chemistry that delivers a not-so-smart kill mechanism, but only to specific cells. At present immunotherapies are the best of these, but there are also selective viral therapies, and others involving nanoparticles. As it turned out, however, all of the more advanced techniques for destroying senescent cells are not targeted at all, and focus on inducing apoptosis, a path to destruction that these cells are already primed to take in comparison to a normal cell. A gentle nudge to the right cellular pathways, such as by increasing or reducing the levels of proteins relevant to apoptosis, will be ignored by near all cells other than those in a senescent state. There, however, it can be enough to tip them over the edge in large numbers. So a senolytic therapy can be delivered generally without targeting. Interestingly, inducing apoptosis is a long-standing line of work in the cancer research community, so there are already stables of drug candidates to explore for use in destruction of senescent cells. If recent deals are any indication, we'll probably see a lot of cross-pollination between these fields in the next few years. Aside from that, a lot of the recent work on senolytic drugs has focused on bcl-2 inhibitors capable of reducing levels of bcl-2, bcl-xl, and bcl-w, all of which act in various ways to suppress apoptosis. This latest published research is along these lines:

Directed elimination of senescent cells by inhibition of BCL-W and BCL-XL

While the mechanisms driving senescence are well studied, understanding of the mechanisms endowing these cells with increased survival capacity is limited. The BCL-2 protein family plays a central role in cell death regulation by diverse mechanisms, including apoptosis and autophagy. This family includes the anti-apoptotic proteins BCL-2, BCL-W, BCL-XL, MCL-1 and A1, and is intensively studied as a target for pharmacological intervention in cancer. We set out to evaluate the individual contributions of each of these BCL-2 family members and their combinations to the viability of senescent cells. We found that the increased presence of BCL-W and BCL-XL underlies senescent cell resistance to apoptosis, and that their combined inhibition leads to senescent cell death. We show that a small-molecule inhibitor targeting the BCL-2, BCL-W and BCL-XL proteins (ABT-737) causes preferential apoptosis of senescent cells, both in vitro and in vivo, and eliminates these cells from tissues, opening the door for targeted elimination of senescent cells.

In light of the consistent upregulation of BCL-W, BCL-XL and BCl-2 observed in all tested types of senescent cells, we examined the effects of their inhibition on cell viability using ABT-737, a potent small-molecule inhibitor of BCL-2, BCL-XL and BCL-W24. Human senescent cells of all three types were significantly more sensitive than control cells to treatment with ABT-737, showing up to 65% death at the highest concentration tested. The same effect was observed following ABT-737 treatment of senescent and control mouse embryonic fibroblasts (MEFs). These findings indicate that BCL-W, BCL-XL and BCL-2 confer resistance of senescent cells to apoptosis, and their inhibition by ABT-737 triggers cell death specifically in these cells.

Our experiments above showed that ABT-737 treatment causes selective elimination of senescent cells in tissue culture, including of cells that were induced to senesce by direct induction of DNA damage. We therefore set out to test the effectiveness of ABT-737 treatment in elimination of DNA-damage-induced senescent cells in vivo. To this end, we induced lung damage and senescence in mice by ionizing radiation, which causes long-lasting accumulation of senescent cells, readily identified by persistent DNA damage, in the lungs. Seven days after irradiation the mice were treated with ABT-737 for 2 days, and 1 day later the lungs were excised and analysed for the expression of senescence markers. SA-β-Gal staining showed a significant decrease in the amount of senescent cells following ABT-737 treatment. This reduction was accompanied by a significant decrease in the numbers of γH2AX-positive lung cells and a decrease in the expression of the senescence markers p53 and p21. The molecular targets of ABT-737, BCL-XL and BCL-W, were expressed in the irradiated lungs and their levels were reduced as a result of the treatment. The reduction in the expression of senescence markers and ABT-737 target proteins was accompanied by increased caspase-3 cleavage, suggesting an increase in apoptosis in the lung following the treatment. These findings establish that ABT-737 treatment leads to the elimination of senescent cells in vivo.

We next tested whether BCL protein family inhibition by ABT-737 could eliminate senescent cells induced by direct activation of p53 in the skin. To this end, we used transgenic mice in which the human p14ARF gene is inducibly expressed in the basal layer of the skin epidermis. Induction of p14ARF in these mice activates p53 and generates senescent epidermal cells that are retained in the tissue for weeks. To generate senescent cells, we activated expression of p14ARF in 3-week-old mice for a period of 4 weeks, and then treated the mice with ABT-737 for 4 consecutive days. The number of senescent cells in the epidermis, determined by SA-β-Gal staining, was dramatically reduced in the ABT-737-treated mice relative to control mice. A similar degree of elimination was observed after ABT-737 treatment of these mice for 2 days. Concomitantly, the percentage of epidermal cells in which the transgenic p14ARF protein could be detected was reduced, indicating preferred elimination of transgene-expressing cells. Increased levels of apoptosis were detected in the epidermis after 2 days of ABT-737 treatment, consistent with increased apoptosis as the mechanism of senescent cell elimination. These findings indicate that the survival signal provided by BCL-family proteins is an essential component of the ability of senescent cells to be retained in the tissue, and in its absence they rapidly die.

The ability to pharmacologically eliminate senescent cells in vivo opens the door to study the roles of senescent cells in a wide range of physiological settings in which they are detected, and to dissect their beneficial and detrimental functions. Importantly, this is an early step towards potential clinical application of senolytic drugs, in such settings as aging-associated diseases. The chemotherapeutic elimination of senescent cells from premalignant lesions and tumours may also prove beneficial, in particular settings in which a pro-tumorigenic function of senescent cells in the tumour or stroma will be proven. Overall, our findings reveal a central molecular mechanism maintaining the viability and retention of senescent cells in tissues, and suggest that the elimination of senescent cells by inhibition of this mechanism represents a promising strategy for targeting senescent cells during tumourigenesis and age-related diseases.

Comments

Yes this compound is certainly on our list for the second round of combination testing at MMTP.

Posted by: Steve Hill at May 3rd, 2016 6:50 AM

I'm still rooting for Oisin Biotechnology. It just seems more likely that a genetically targeted therapy will have fewer chances of side effects than a small molecule targeting downstream chemistry in the cells.

Posted by: Jim at May 4th, 2016 5:57 AM

Same result Jim but yes I agree the Oisin approach is good. We should be testing this too in our next phase as we have discussed with Oisin. It is good for us to hit this on a broad front so we can push progress faster.

Posted by: Steve Hill at May 4th, 2016 11:47 AM

This approach sounds good, but we explored ectopic expression of many markers in AD and apoptosis was one way the terminally differentiated neurons give up the ship, a word of caution this approach h may me ill advised or a flawed concept when it comes to the brain and AD.9450309699939917079730
But otherwise may be wonderful way to cull old junk ladened cells and should be explored.

Posted by: Mark Obrenovich at May 8th, 2016 3:31 PM

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