Markers for Senescence in Cells

Cells are complex machines that have many carefully regulated states. One of those states is senescence, in which the cell permanently exits the cell cycle, stops dividing, and begins to secrete a variety of molecules that, among other things, degrade surrounding extracellular matrix structures and encourage nearby cells to also become senescent or change their behavior in other ways. This senescent state seems to be a tool that originally evolved to help manage embryonic growth: senescent cells are found in embryos in places that suggest they are managing shape or tissue transitions during development.

Evolution promiscuously reuses everything that emerges in biology, and at some point cellular senescence became a reaction to damage likely to cause cancer. Toxins, stress, and the various forms of cellular and molecular damage of aging can provoke cells into more readily becoming senescent, either directly or through changes in the signaling environment in tissues. Cancers result from damage to nuclear DNA, producing cells capable of unfettered replication. Senescence is a form of defense that removes potentially cancerous cells from consideration, or at least to some degree. Cancers and cellular senescence are two aspects of our biology in the midst of a long-running evolutionary struggle: the story of the lengthening of human life span in comparison to other primates is one of a moving balance between death by cancer and death by failing tissue function. Many of the most important mechanisms in aging, cellular senescence included, can be viewed in that context. However, too much cellular senescence actually promotes some forms and aspects of cancer precisely because of the various molecules secreted by these cells. There is a tipping point between cancer suppression and cancer promotion determined by the changes in the cellular environment caused by the presence of senescent cells. It is a complex situation.

Senescent cells sometimes destroy themselves through programmed cell death mechanisms that serve to remove damaged cells from circulation before they cause issues, and are sometimes destroyed by the immune system. Those that linger stick around for the long term, however, accumulating in ever increasing numbers. To make things worse, as the immune system declines with aging it does an increasingly poor job of removing these cells, so even as the pace of creation accelerates due to age-damaged tissues, the policing mechanisms are in decline. A sizable percentage of adult skin cells are senescent by the time old age rolls around, for example.

In an ideal world, the perfect situation for adult cellular senescence would be if all these cells in fact destroyed themselves fairly promptly, say within a few days. That is a long way removed from the present situation, but it is something that near future medicine could achieve. The cancer research community is developing all sorts of ways to precisely target cells based on their particular biochemical differences, usually and most easily based on differences in surface chemistry, the molecules presented on the exterior of the cell membrane. Many cancers are distinctive enough to target in this way, and so an industry of research and development has for years been working on the use of altered viruses, designer nanoparticles, and the like, that can find and bind to particular combinations of cell surface chemistry, and once there deliver some form of traditional cancer therapy in the tiny dosage needed to destroy a single cell. Manufacturing tens of millions of such devices for each dose of a therapy promises a very effective next generation of treatments that nonetheless have few side-effects - a world removed from the present standards of chemotherapy and radiation therapy.

Turning these prototype cancer therapies into senescent cell clearance treatments is a very plausible path forward. The roadblock in the way is the need for a good, reliable marker to determine which cells are senescent and which are not. Some of the most interesting work on senescent cell clearance has focused on p16 as a marker for cellular senescence, but this is not as discerning as would be liked despite the successes achieved to date in mice. A couple of other lines of research have looked promising in recent years, such as using lysosomal hydrolases or TRF2 as markers. We are still waiting on a research group to pull this all together, but meaningful removal of senescent cells remains probably the closest item in the rejuvenation toolkit to actual realization.

Here is recent news in which the research team seem fairly confident they have a good marker for senesence in the form of a combination of proteins. They have tried a couple of different cell lines by the sound of it, but I'd want to see more diverse tissues tested; there is no necessary reason to expect cellular senescence to generate usefully similar surface chemistry in all of the most common cell types in the body, though it would be pleasant if that did turn out to be the case. That said, despite the lack of funding for senescent cell clearance with the explicit goal of treating aging it seems there is still some progress, as illustrated by the fact that these cancer researchers are aware enough of efforts in that direction to talk about it at all:

Study offers future hope for tackling signs of aging

"What we have found is a series of novel markers - a way to detect senescent cells. What is more, we have shown that they can be used to predict increased survival in certain types of cancer. Until now, good protocols to help spot these cells have been sadly lacking. Our research has described new markers located on the surface of the old cells. This makes these markers particularly useful to quickly identify these cells in laboratory and human samples using a range of techniques."

As a first clinical application of these markers, the researchers observed that they were present in high numbers in samples from different types of cancer and that this correlated with a better prognosis of the disease. This was particularly evident in breast cancer. "These markers could be useful tools not only to study senescent cells in the lab but also they could be developed into diagnostics to help predict survival in cancer patients. Moreover, they could also be used in the future to define strategies to selectively eliminate the old cells from the tissues and thus reduce their effects on promoting ageing in healthy subjects."

Characterization of novel markers of senescence and their prognostic potential in cancer

Cellular senescence is a terminal differentiation state that has been proposed to have a role in both tumour suppression and ageing. This view is supported by the fact that accumulation of senescent cells can be observed in response to oncogenic stress as well as a result of normal organismal ageing. Thus, identifying senescent cells in in vivo and in vitro has an important diagnostic and therapeutic potential.

The molecular pathways involved in triggering and/or maintaining the senescent phenotype are not fully understood. As a consequence, the markers currently utilized to detect senescent cells are limited and lack specificity. In order to address this issue, we screened for plasma membrane-associated proteins that are preferentially expressed in senescent cells. We identified 107 proteins that could be potential markers of senescence and validated 10 of them (DEP1, NTAL, EBP50, STX4, VAMP3, ARMX3, B2MG, LANCL1, VPS26A and PLD3). We demonstrated that a combination of these proteins can be used to specifically recognize senescent cells in culture and in tissue samples and we developed a straightforward fluorescence-activated cell sorting-based detection approach using two of them (DEP1 and B2MG).

Of note, we found that expression of several of these markers correlated with increased survival in different tumours, especially in breast cancer. Thus, our results could facilitate the study of senescence, define potential new effectors and modulators of this cellular mechanism and provide potential diagnostic and prognostic tools to be used clinically.

Comments

"Some of the most interesting work on senescent cell clearance has focused on p16 as a marker for cellular senescence, but this is not as discerning as would be liked despite the successes achieved to date in mice."

Are there any links or explanations of why P16 is not as discerning as would be liked? Does the Mayo clinic's research into mice with a gene for half a drug inserted next to P16 provide evidence that P16 is discerning enough? I tried googling this question, but got lost in the thicket of papers and articles not answering the question.

Posted by: Jim at December 10th, 2014 10:15 PM

Hi Jim,

First, p16 expression is not a simple binary, and simply finding that a cell is expressing it does not necessarily mean that it's durably senescent. Conversely, not all senescence is even initiated by p16: some goes through the largely independent p53 pathway:
http://www.nature.com/nrm/journal/v8/n9/fig_tab/nrm2233_F4.html

Some studies also find that senescence initially induced to enter senescence by the p16 pathway can maintain a senescent phenotype despite p16 being silenced or ablated.

So while a targeting system based just on p16 is good enough for down-and-dirty work like van Deursen and Kirkwood's proof-of-concept clearance of senescent cells from aging tissues (especially in a severe progeroid model like their hypomorphic BubR1 mice), it would neither be sensitive nor specific enough for good therapies.

I think this new work is extremely promising -- and the fact that they've got what appears to be a sensitive and selective system that uses more than one cell-surface marker but not a large number is just the kind of sweet spot you'd optimistically expect for a targeting system: relying on just one marker seems likely to lead to some off-target cells that display it for reasons other than senescence, and to not be redundant enough, while the more targets you need to add the more complex it becomes to design a manageable targeting system.

Posted by: Michael at December 14th, 2014 2:34 PM

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