The SENS Research Foundation has published its annual reports for 2022, for those interested. SENS, the Strategies for Engineered Negligible Senescence, is both (a) a laundry list of forms of cell and tissue damage that cause aging, with supporting evidence from the past century of scientific research into aging, and (b) a laundry list methods of intervention that should produce rejuvenation. Aging is damage accumulation, and rejuvenation is repair of that damage.
Funding for SENS programs, and initiatives to produce therapies based on the SENS view of damage repair, remain as relevant as ever. In fact, even more relevant now than was the case in the early 2000s, given the extensive evidence gathered over the past decade to support the SENS view on the role of senescent cells in aging. The view that accumulation of senescent cells is an important aspect of aging, and a viable point of intervention for the first rejuvenation therapies worthy of the name, was first published by Aubrey de Grey and others in an academic paper in 2002, well in advance of the 2011 technology demonstration of senescent cell clearance that convinced enough of the research community for further exploration to be prioritized.
Today, twenty years after the first call to action, and ten years after the first compelling demonstration, many biotech companies are working on the development of therapies to selectively destroy or modulate the behavior of senescent cells, scores of animal studies show reversal of measures of age-related disease following partial clearance of senescent cells in old mice, human clinical trials are underway, and countless research groups are investigating the biology of senescent cells, in search of new approaches to achieve these goals.
Senescent cells are just one of the seven categories of cell and tissue damage outlined in the SENS proposals for a rejuvenation biotechnology industry. The SENS Research Foundation and its allied researchers and spin-out companies remain necessary: the success achieved in turning senescent cell clearance from a compelling idea to (almost) a clinical reality must be repeated, and repeated many times, if we are to achieve the goal of cures for age-related disease, prevention of frailty, sickness, and death in the old, elimination of the largest cause of suffering and mortality in the human condition.
Like so much in our modern world, curing the diseases of aging is a collaborative effort. In 2021, SENS Research Foundation (SRF) found itself at the center of a brand new way to fundraise. The ingenuity and generosity of Richard Heart, and the willingness to envision a life free from age-related disease from a forward-thinking global community, provided SRF with unprecedented resources. We gained not only in dollars, but also in number of supporters. Our vision struck a chord that reverberated across a broader group of people than ever before. Our mission inspired so many to put their trust and resources behind us, and we could not be more grateful or more determined to honor their support through the acceleration and expansion of our vital research. At the same time, we had a changing of the guard at SRF. Undergoing internal investigations in the public eye, under intense scrutiny. Saying goodbye to our visionary founder, to a full half of our Board of Directors, and to our long-time Director of Education.
Within the last year, SRF has seen more upheaval, more incredible support, and more intense criticism, than in the entirety of the previous decade. And yet we remain, passionately in pursuit of the mission that drove our founding. Our dedication to making the ‘Strategies for Engineered Negligible Senescence’ a life-saving reality is rock-solid, as we hope this Annual Report will make clear. Our mission is vital; one hundred thousand people die every day of age-related disease. Millions more suffer due to age- related decline and disability. Our mission cannot be side-tracked, cannot be delayed, and must take precedence over all other concerns. Last year was difficult, but also empowering. Our leadership may change, but our founding vision is powerful and keeps us focused on the path ahead. Our mission is our defining priority. Together, we will build this new world, one brick at a time.
Catalytic Antibodies Targeting Intracellular Tau Oligomers
Therapeutic interventions with anti-tau immunotherapies have shown promise, but the efficacy seems to vary greatly. The tau LysoSENS group at SENS Research Foundation is investigating the therapeutic potential of catabodies (catalytic antibodies) targeted to the intracellular compartment to degrade tau aggregates and prevent or reverse tau-associated neurodegeneration. Unlike conventional binding antibodies, catabodies bind transiently to their targets and hydrolyze them into very small hydrolytic end-products, leaving the catabody free to attack the next target molecule.
Rejuvenating Immune Surveillance of Senescent Cells
Natural Killer (NK) cells are a known key immune cell type responsible for the immune-mediated senolysis of senescent cells. Moreover, NK cells are increasingly emerging as an important defense against several age-related diseases, the best-understood example of which is cancer. However, NK cell function declines as part of immunosenescence, and this likely includes immune surveillance of senescent cells, leaving the host increasingly vulnerable to diseases of aging, as recently reviewed in a paper published by the Sharma lab at SENS Research Foundation.
To understand the potential of NK cell transplantation as an immunosenolytic therapy, the ApoptoSENS team is collaborating with the Campisi lab at the Buck Institute to investigate the effect of aging on NK cell cytotoxicity toward senescent cells. Studies will test the ability of young vs. old donor derived NK cells to remove senescent cells in a mouse model. Additionally, the Sharma lab is in the process of developing CAR (Chimeric Antigen Receptor)-NK cells with enhanced ability to target senescent cells for adoptive cell therapy, and the above studies will be repeated using CAR-NK cells.
Studying age-related changes in the immune cells has led the ApoptoSENS team to discover a sub-population of T-cells that declines with age. Analysis indicates that these “X cells” constitute only approximately 5% of total peripheral blood mononuclear cells, so in order to investigate their interaction with senescent cells, the team established a protocol for enrichment of X cells in culture. These experiments indicated that X cells rapidly kill senescent cells. Based on these promising initial results, Sharma and coworkers are now further assessing the therapeutic potential of this sub- population of T cells, as they appear to be highly selective in eliminating senescent cells in a substantially shorter time than has been reported by others or observed in their own prior work with NK cells.
Engineering New Mitochondrial Genes to Restore Mitochondrial Function
The MitoSENS lab at SENS Research Foundation, led by Dr. Amutha Boominathan, is working to develop rejuvenation biotechnologies to repair or obviate the accumulation of mitochondrial DNA (mtDNA) mutations with age. Their principal focus is allotopic expression (AE), in which copies of the protein-encoding mtDNA genes are placed in the nucleus, with suitable modifications to allow them to be expressed in the nucleus and translated in the cytosol, following which they must be imported into the mitochondria. There, these gene copies can incorporate into the relevant electron transport chain complexes and contribute to sustaining oxidative phosphorylation. This would allow mitochondria to continue producing ATP, irrespective of the accumulation of particular mtDNA mutations.
Boominathan and colleagues implemented that strategy in the past to synthesize 2 versions of the 13 mtDNA genes: a) the minimally recoded version that is absolutely required for productive protein translation in the cytosol and b) the codon-optimized version, synchronizing the codon usage in these genes to the mammalian nuclear code. They were able to successfully demonstrate robust transient protein production and mitochondrial association for all the 13 mtDNA genes using the codon-optimized gene expression constructs. Cytosolic protein expression under transient expression was substantially higher for the codon-optimized than for minimally-recoded genes, and similarly for steady-state mRNA levels under stable selection. Eight of the re-engineered genes retained expression and targeting to the organelle after stable selection. Building on these early observations, the team validated the utility of these codon-optimized mtDNA gene constructs for additional mitochondrial protein targets that did not work in the past.
The mitochondrial DNA deletions that accumulate in aging cells have a strong selection advantage, amplifying within post-mitotic cells to the point of homoplasmy. The MitoSENS team is exploring a strategy to address this issue by transferring exogenous, viable mitochondria modified for sustained retention and for therapeutic activity. While mitochondrial transplantation is already being investigated by several groups and companies as a therapeutic intervention strategy, Dr. Boominathan’s team is advancing an improvement on this strategy using mitochondria with genomes engineered for dominance over the native genotype.
Target Prioritization of Extracellular Matrix Aging
As we age, changes occur not only in the cells within the extracellular matrix (ECM), but importantly also in the composition and chemistry of the ECM. Many of the characteristic physical changes that we associate with ageing, such as the changes in skin appearance or the decrease in flexibility of joints, are specifically the result of changes in the structure and composition of the ECM. Two important components of the ECM are elastin and collagen; indeed, collagen is estimated to account for about 30% of the protein in the body. The Clarke lab is investigating age-related changes in the chemical structure of elastin and collagen and how these changes impact the mechanical behavior of the tissues.
Prior to Clark’s investigations, the general consensus in the literature was that tendon increases in stiffness with age and that this was due to an increase in crosslinking between collagen molecules. His group has instead found was that it is not possible to say that tendon gets stiffer with age, particularly when comparing mature to genuinely aged animal tissue. Instead, Clark and colleagues report an increase in the breaking strain, a decrease in the ability to absorb stress, and an increase in the fragility (chance of rupture) with age. This is clearly a more refined and complex description of the physical properties, and accords better with the orthopedic vulnerabilities of aging human tissues.
Clarke’s research has also revealed an increase in irreversible crosslinks in the tendon with age, which increase the force required to break a tendon. This increase occurs even as the tendon gradually becomes depleted of reversible crosslinks that allow the tendon to adapt to and absorb force. Understanding how each of these different crosslinks affect the mechanical properties of a tissue and how they change in number with age will enable more targeted strategies for rejuvenation biotechnologies. Based their findings, Clarke and colleagues predict that to rejuvenate youthful tendon function would entail decreasing the number of irreversible crosslinks while greatly increasing the reversible crosslinks. Conceptually, this could be achieved through various means, some of which might not involve directly targeting the crosslinks themselves, but instead cell therapy or other approaches that rejuvenate the behavior of the cells that turn over the ECM. In principle, an unbalanced approach based exclusively on breaking a subset of crosslinks might improve some aspects of tissue function but also cause structural problems.
Lipofuscin Degradation by Bacterial Hydrolases
According to the “garbage catastrophe theory of aging“, the accumulation of lipofuscin aggregates limits the remaining life span of the organism by disturbing lysosomal function and inducing cell death. While humans have no enzymes capable of breaking down lipofuscin, microorganisms possess a wide array of enzymes that allow the degradation of any conceivable molecule formed in nature. Thus, the LysoSENS strategy seeks to identify microbes that are able to degrade lipofuscin via specific hydrolases as lead candidates for potential longevity therapeutics. To pursue this goal, the Grune lab will use authentic lipofuscin derived from human cardiac tissue.
Dr. Grune and colleagues extracted microorganisms from different soil samples collected at a residential yard, a forest, a compost heap, and a riverbed. The team used these extracts to select lipofuscin-degrading bacteria by growing cultures on isolated lipofuscin as the only energy, carbon, and sulphur source. Following 20 sets of sub-culture passaging, bacterial mixtures growing on human tissue-derived lipofuscin were extracted, and 12 bacterial strains were isolated. These strains and their specific enzymes will be isolated and further investigated. It bears noting that it is not expected that these bacterial enzymes will prove to be proteases, but hydrolases able to degrade complex crosslinks between proteins. This fact will complicate the identification of lead candidates, but on the other hand, such structures will be unique to lipofuscin and able to function in mammalian cells (after suitable modification) without the danger of digesting functional proteins. A future task will be the targeting of the identified hydrolases towards the lysosomal compartment.
SenoStem: Combinatorial Rejuvenation Biotechnologies
Age-related disease and disability results from the complex interaction of multiple forms of cellular and molecular aging damage. Prominent examples of this damage are the loss of stem cells and the accumulation of senescent cells. Senescent cells propagate damage and impose systemic metabolic derangement through the secretion of a senescence-associated secretory profile (SASP). The SenoStem project at SENS Research Foundation is testing the hypothesis that combination therapy using senolytics and stem cell transplantation will have a synergistic beneficial effect on aging mice and might be able to further improve health and lifespan – literally a remove-and-replace strategy. This approach builds toward SRF’s larger long-term goal to develop synergistic combinatorial rejuvenation biotechnology approaches.
Microglia as a Vehicle for Brain Rejuvenation
With SRF funding, the Hébert team has developed a protocol for using microglia as a delivery system for biologics over wide areas of the adult brain. With this protocol, endogenous microglia are replaced with transplanted microglia after a single superficial cell injection. Microglia are migratorily more active compared to neuronal progenitors and more easily spread throughout the brain. In addition to therapeutic proteins, this system can be used to deliver new neurons to all areas of the brain to counteract neuronal loss with age. The transplanted microglia can be engineered to produce a secreted biologic, or engineered to be reprogrammed to new neurons. In both cases, normal microglia density is innately re-established, minimizing any effect of transient microglia depletion while providing novel therapeutic support to brain function.
Identification and Targeting of Noncanonical Death Resistant Cells
It is well established that senescent cells (SCs) can result from a number of stressors, including replicative stress, telomere erosion and damage, and oncogene expression. They are also induced as part of tissue remodeling in wound healing and development. More recently, it was discovered that SCs can spread the senescent phenotype to other cells in the body. Characterization of secondary SCs and differentiating their biology and vulnerabilities from those of primary SCs is thus critical to developing longevity therapeutics targeting the full spectrum of senescence in aging, and is the central focus of Dr. Admasu’s work at SENS Research Foundation. With his SRF colleagues, Dr. Adamasu developed a novel protocol to overcome one major roadblock in this endeavor, which has allowed him to make new insights into secondary senescence and identify a highly significant therapeutic target for senolytic drugs with broad senolytic activity against both primary and secondary SCs.