Researchers Report on More New Senolytic Drug Candidates

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Now that clearing senescent cells as a therapy for aging finally has meaningful support in the research community, there is far more funding available to turn the wheels of the standard drug discovery and evaluation process. Researchers are in search of senolytic drugs, those that can kill senescent cells without harming normal cells. The process starts at first with an evaluation of the performance of each molecule in the standard compound libraries in cell cultures, in search of molecules that preferentially kill senescent cells. This can be automated to a fair degree, especially when the desired result is as black and white as destroying one distinctive class of cell. It is very similar to the sort of cancer drug screening that the research community has a great deal of experience in carrying out. At scale, this might cost a few dollars per molecule screened these days. In fact, the existing candidates for senolytic drugs have largely emerged from the cancer drug candidate databases, and were tested for their effects on cancer for some years without noticing their strong effects on senescent cells – it wasn’t in the list of items to evaluate at the time.

Given the starting point of a few promising compounds, preferably already tried in animals and humans, and thus with decent pharmacology data, researchers then branch out to examine other chemically similar compounds. It is usually the case that a better version with greater primary effects and lesser side-effects can be established one way or another. The level of work to achieve that end varies greatly, however, ranging from finding another well-characterized small molecule drug candidate in the archives to the researchers having to carry out all of the work to model and synthesize a novel molecule and prove it to be effective. It is usually the case that researchers and developers are far more willing to push ahead with a suboptimal compound that is already fairly well tested than to work with a less well explored but potentially better compound. Nonetheless, in theory the competition in the system weeds out worse drugs in favor of better drugs, though that process may never seem as efficient in practice or as fast as we’d like it to be. Personally, I’d like to see more funding going towards the sort of programmable gene therapy pioneered by Oisin Biotechnologies, a better approach than mining cancer chemotherapy drugs in search of those with side-effects that are minimal enough for patients to accept.

Then it is on to animal studies, starting companies, and human trials, the standard process for moving forward with the development of new medicine. Many candidates will turn out to be not so useful in human medicine, others will pass all the way through. Insofar as senolytic drug discovery goes, all of the groundwork has already taken place for a number of cancer drug candidates that can promote apoptosis in senescent cells, such as navitoclax and dasatinib, some of which are being carried forward into clinical trials by UNITY Biotechnology. In animal studies, these appear to remove on the order of 10-50% of senescent cells in a single course of treatment, varying widely by tissue type. The research community isn’t resting on its laurels, however, and is turning up new candidates on a fairly regular basis at the moment, such as piperlongumine. The paper below offers another few candidates for consideration, though I would say that they are less interesting in and of themselves at this stage, but rather as an indication that we should expect the list of potential drugs to expand quite rapidly in the next few years, and hopefully the quality of the best candidates along with it.

New agents that target senescent cells: the flavone, fisetin, and the BCL-XL inhibitors, A1331852 and A1155463

Senescent cells accumulate in numerous tissues with aging and at sites of pathogenesis of multiple chronic diseases. Small numbers of senescent cells can cause extensive local and systemic dysfunction due to their pro-inflammatory senescence-associated secretory phenotype (SASP). For example, transplanting only 200,000 senescent ear chondroblasts or preadipocytes around knee joints induces osteoarthritis in mice, while injecting similar numbers of non-senescent cells does not. Clearing senescent cells by activating a drug-inducible “suicide” gene in progeroid or naturally-aged mice alleviates a range of age- and disease-related phenotypes, including sarcopenia, frailty, cataracts, adipose tissue dysfunction, insulin resistance, and vascular hyporeactivity.

To decrease the burden of senescent cells in non-genetically-modified individuals, we used a hypothesis-driven approach to identify senolytic compounds, which preferentially induce apoptosis in senescent rather than normal cells. Our approach was based on the observation that senescent cells are resistant to apoptosis. This suggested that senescent cells either have reduced engagement of pro-apoptotic pathways that serve to protect them from their own pro-apoptotic SASP or they have up-regulated pro-survival pathways. We demonstrated the latter to be the case and identified senescence-associated pro-survival pathways based on expression profiling of senescent vs. non-senescent cells. We confirmed the requirement of these pathways for survival of senescent but not non-senescent cells by RNA interference. These pathways included pro-survival networks related to PI3K / AKT, p53 / p21 / serpins, dependence receptor / tyrosine kinases, and BCL-2 / BCL-XL, among others.

We tested drugs that target these pro-survival pathways. We initially reported that the dependence receptor/ tyrosine kinase inhibitor, dasatinib (D) and the flavonoid, quercetin (Q), are senolytic in vitro and in vivo. D and Q induced apoptosis in senescent primary human preadipocytes and HUVECs, respectively. Combining D+Q broadened the range of senescent cells targeted, and, in some instances, proved synergistic in some types of senescent cells. D+Q alleviated cardiovascular, frailty-related, osteoporotic, neurological, radiation-induced, and other phenotypes and disorders in chronologically aged, progeroid, and high fat-fed atherosclerosis-prone mice, consistent with our observations in mice from which senescent cells had been removed by inducing the suicide gene in transgenic INK-ATTAC mice. Expanding upon our findings with Q, we tested if the related flavonoid fisetin is senolytic. Fisetin is widely available as a nutritional supplement and has a highly favorable side-effect profile.

Based on our earlier hypothesis-driven identification of senolytic drugs and identification of the BCL-2 pro-survival pathway as one of the “Achilles’ heels” of senescent cells, we and others simultaneously reported that the BCL-2 / BCL-W / BCL-XL inhibitor, navitoclax (ABT263; N), is senolytic. Like D and Q, N is senescent cell type-specific, being effective in inducing apoptosis in HUVECs but not human preadipocytes. We also found that the related BCL-2 family inhibitor, TW-37, is not senolytic. TW-37, unlike N, does not target BCL-XL. Others confirmed that N targets senescent cells, but Bcl-2 family inhibitors that do not target BCL-XL are not senolytic. We therefore tested if the relatively specific BCL-XL inhibitors, A1331852 and A1155463, are senolytic. Unlike N, these agents do not target BCL-2. Consequently, A1331852 or A1155463 may cause less BCL-2-induced neutrophil toxicity, a serious side-effect of N.

We found that fisetin and the BCL-XL inhibitors, A1331852 and A1155463, are senolytic in vitro, inducing apoptosis in senescent, but not non-senescent HUVECs. This adds three new agents to the emerging repertoire of senolytics reported since early 2015, which currently includes D, Q, N, and piperlongumine. Fisetin has a plasma terminal half-life of just over 3 hours in mice. It alleviates dysfunction in animal models of chronic disease, including diabetic kidney disease and acute kidney injury, attributes consistent with those expected from a senolytic agent. Here we demonstrate that fisetin is indeed senolytic in senescent HUVECs, but not in senescent IMR-90 cells or human preadipocytes. A1331852 and A1155463 are senolytic in HUVECs and IMR-90 cells but not primary human preadipocytes. We noted that these drugs increased cellular ATP levels significantly in senescent human preadipocytes, but not HUVECs, through an as yet unknown mechanism.

We predict many more senolytic drugs will appear at an accelerating pace over the next few years. Initially, most are likely to be based on re-purposed drugs or natural products. Increasingly, new senolytics will likely be derived using medicinal chemical approaches based on optimizing properties of the repurposed agents. Consistent with this, it appears that small changes in the senolytic drugs already discovered can interfere with senolytic activity, such as in the case of D vs. imatinib, with the latter not being senolytic, or N vs. the closely-related agent, TW-37. Conversely, we speculate that small structural changes to repurposed senolytic drugs could enhance senolytic activity, with increases in the percent and range of types of senescent cells eliminated, as well as better stability, bioavailability, and side-effect profiles.

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