OBJECTIVE:As the body's largest organ, the skin plays a crucial role in defending against external stressors. Skin characteristics change with age, decreasing skin barrier integrity and compromising skin and body health. This study aimed to investigate the potential of a topical formulation containing OS-01 (a.k.a. Peptide 14), a senotherapeutic peptide, to counteract age-related skin changes and their systemic consequences. METHODS:A randomized, double-blinded clinical trial involving 60 female volunteers aged 60-90 was conducted over 12 weeks. Participants received either an OS-01 topical formulation or a commercially available moisturizer control formulation. Skin parameters, subjective perceptions, and circulating cytokine levels were assessed. Skin instrumental analysis included transepidermal water loss (TEWL), skin hydration, and pH measurements. RESULTS:Participants treated with the OS-01 topical formulation displayed significantly improved skin barrier function and hydration compared to the control group. Participant perceptions aligned with objective findings: after 12 weeks, 70% of participants in the OS-01 group noticed an improvement in general skin appearance versus 42% for the control group. The systemic levels of proinflammatory cytokines tended to normalize, with a significant decrease in IL-8 in the blood analysis of participants from the OS-01 group. On the other hand, the control group demonstrated an increase in a few circulating cytokines, particularly TNF-ɑ and IFN-γ. Moreover, GlycanAge analysis measuring participants' biological age suggested the slowing of systemic aging in the group treated with the OS-01 topical formulation. CONCLUSION:The study suggests that the OS-01 formulation can impact skin health by improving the skin barrier function, potentially influencing systemic inflammation and biological age. In conclusion, the study supports that targeting skin health may contribute to better longevity outcomes, underscoring the skin's pivotal role in systemic aging and supporting an integrated approach to health management.

BACKGROUND:The aging of the skin, particularly around the periorbital region, is a complex process characterized by the accumulation of senescent cells, decreased collagen production, and reduced skin elasticity, leading to visible signs such as fine lines, wrinkles, and sagging. OBJECTIVE:This study investigates the efficacy of a novel topical formulation, OS-01 EYE, containing the senomorphic peptide, OS-01, along with other active ingredients, in improving the skin around the eyes. METHODS:A 12-week clinical study was conducted with 22 participants who applied OS-01 EYE twice daily. Assessments included bioinstrumental measurements of skin hydration, barrier function, firmness, and elasticity, as well as expert photographs grading fine lines, wrinkles, under-eye puffiness, and dark circles. RESULTS:After 12 weeks of product use, transepidermal water loss (TEWL) decreased by 17.33%, hydration increased by 32.49%, skin firmness improved by 10.19%, and elasticity increased by 25.58%, compared to baseline, which was all statistically significant. Expert grading revealed a decrease in fine lines and wrinkles, under-eye puffiness, and dark circles over the study period. Furthermore, subjective assessments showed that 95.46% of participants reported improvements in overall appearance. CONCLUSION:The OS-01 EYE formulation was found to be safe and effective in mitigating the visible signs of aging in the periorbital region, making it a potent treatment option for enhancing periorbital skin health, function, and appearance.

Cellular senescence is known to play a role in age-related skin function deterioration which potentially influences longevity. Here, a two-step phenotypic screening was performed to identify senotherapeutic peptides, leading to the identification of Peptide (Pep) 14. Pep 14 effectively decreased human dermal fibroblast senescence burden induced by Hutchinson-Gilford Progeria Syndrome (HGPS), chronological aging, ultraviolet-B radiation (UVB), and etoposide treatment, without inducing significant toxicity. Pep 14 functions via modulation of PP2A, an understudied holoenzyme that promotes genomic stability and is involved in DNA repair and senescence pathways. At the single-cell level, Pep 14 modulates genes that prevent senescence progression by arresting the cell cycle and enhancing DNA repair, which consequently reduce the number of cells progressing to late senescence. When applied on aged ex vivo skin, Pep 14 promoted a healthy skin phenotype with structural and molecular resemblance to young ex vivo skin, decreased the expression of senescence markers, including SASP, and reduced the DNA methylation age. In summary, this work shows the safe reduction of the biological age of ex vivo human skins by a senomorphic peptide.

INTRODUCTION:Senescent cells contribute to age-related tissue deterioration, including the skin, which plays important roles in overall health and social interactions. This study aimed to assess the effects of the senotherapeutic peptide, OS-01 (a.k.a. Pep 14), on skin.METHODS:A 12-week split-face, double-blinded, vehicle-controlled study involving 22 participants was conducted. The OS-01-containing formulation was applied to one side of the face, while the other side received an identical control formulation lacking the peptide. Skin characteristics were assessed using instrumental measurements, expert clinical grading, and subjective questionnaires.RESULTS:Results showed that the OS-01 formulation significantly improved one aspect of skin barrier function, as evidenced by reduced trans-epidermal water loss compared to both baseline and vehicle control. Expert grading and Antera 3D image analysis revealed a reduction in wrinkle appearance and indentation in the periorbital area, and improved skin texture and radiance on both sides of the face, with the OS-01-containing formulation demonstrating superior results. Participants also perceived improvements in skin hydration, smoothness, radiance, and overall appearance.CONCLUSION:The findings suggest that the OS-01 formulation promotes skin health by strengthening the skin barrier, protecting against dehydration, reducing the appearance of wrinkles, and improving skin texture and radiance. These effects are likely attributed to the senotherapeutic properties of OS-01 in reducing cellular senescence and its associated detrimental effects.

mp3 file (6.8 MB). In the inaugural edition of the Cancer Discovery podcast, Executive Editor Mark Landis talks with Matthew Meyerson about his paper, which describes the identification of the DDR2 kinase as a therapeutic target in squamous cell lung cancer.

Supplementary Table 1 from Mutations in the DDR2 Kinase Gene Identify a Novel Therapeutic Target in Squamous Cell Lung Cancer

Supplementary Figure Legends 1-7 from Mutations in the DDR2 Kinase Gene Identify a Novel Therapeutic Target in Squamous Cell Lung Cancer

Radiation therapy damages and depletes total bone marrow (BM) cellularity, compromising safety and limiting effective dosing. Aging also strains total BM and BM hematopoietic stem and progenitor cell (HSPC) renewal and function, resulting in multi-system defects. Interventions that preserve BM and BM HSPC homeostasis thus have potential clinical significance. Here, we report that 50% calorie restriction (CR) for 7-days or fasting for 3-days prior to irradiation improved mouse BM regrowth in the days and weeks post irradiation. Specifically, one week of 50% CR ameliorated loss of total BM cellularity post irradiation compared to ad libitum-fed controls. CR-mediated BM protection was abrogated by dietary sulfur amino acid (i.e., cysteine, methionine) supplementation or pharmacological inhibition of sulfur amino acid metabolizing and hydrogen sulfide (H2S) producing enzymes. Up to 2-fold increased proliferative capacity of ex vivo-irradiated BM isolated from food restricted mice relative to control mice indicates cell autonomy of the protective effect. Pretreatment with H2S in vitro was sufficient to preserve proliferative capacity by over 50% compared to non-treated cells in ex vivo-irradiated BM and BM HSPCs. The exogenous addition of H2S inhibited Ten eleven translocation 2 (TET2) activity in vitro, thus providing a potential mechanism of action. Short-term CR or fasting therefore offers BM radioprotection and promotes regrowth in part via altered sulfur amino acid metabolism and H2S generation, with translational implications for radiation treatment and aging.

Senotherapeutic molecules decrease cellular senescence burden, constituting promising approaches to combat the accumulation of senescent cells observed in chronological aging and age-related diseases. Numerous molecules have displayed senotherapeutic potential, but toxicity has been frequently observed. Recently, a new senotherapeutic compound, Peptide 14, was developed to modulate cellular senescence in the skin. In order to assess the potential toxic and genotoxic effects of the peptide, we observed the viability of human primary dermal fibroblasts and epidermal keratinocytes with Peptide 14 treatment, and show that it is mostly non-toxic in concentrations up to 100 μM. Cancer lines were also used to investigate its potential of modulating proliferation. Different concentrations of the peptide promoted a discrete reduction in the proliferation of cancerous cells of the MeWo and HeLa lineages. In full-thickness human skin equivalents, topically formulated Peptide 14 also failed to exert any significant irritation, nor cellular toxicity when added to the culture media. Genotoxic assays including the Ames, micronucleus, and karyotyping tests also indicate the safety of the peptide. Finally, the irritative potential of the peptide was assessed in human subjects in a repeated insult patch test executed using 1 mM peptide. No visible skin reactions were observed in any of the 54 participants. Taken together, the present data support that Peptide 14 is a senotherapeutic molecule with a positive safety profile as tested with cruelty-free models, justifying further studies involving the peptide.

Skin health and longevity are a reflection of the skin's functionality and metabolism, which are controlled by a complex connection between genetic background and environmental conditions, reflecting the tissue's epigenomic, transcriptomic, proteomic, lipidomic, glycomic, and metabolomic profiles. It is increasingly clear that the manipulation of at least some components of this complex web may be effective in promoting the functional and/or molecular rejuvenation of a biological tissue. Epigenetic markers have been consolidated as parameters to assess skin biological age, health, and function. More recently, epigenetic alterations have also been recognized as actionable targets to manipulate gene expression and therefore skin phenotype. In the Epigenetics and Cosmetics book chapter, epigenetics basics are revisited, as well as the methods to assess epigenetic changes in biological samples. The epigenetic particularities of the skin and their modifications throughout aging are presented. Biological clocks are discussed, as well as the current efforts to calculate skin biological age and to harness such information to foster cosmetic product development. Finally, commercially available cosmetic ingredients that act through epigenetic modulation are presented, as well as perspectives in the theme.

Cockayne syndrome (CS) is a rare, autosomal genetic disorder characterized by premature aging-like features, such as cachectic dwarfism, retinal atrophy, and progressive neurodegeneration. The molecular defect in CS lies in genes associated with the transcription-coupled branch of the nucleotide excision DNA repair (NER) pathway, though it is not yet clear how DNA repair deficiency leads to the multiorgan dysfunction symptoms of CS. In this work, we used a mouse model of severe CS with complete loss of NER (Csa(-/-)/Xpa(-/-)), which recapitulates several CS-related phenotypes, resulting in premature death of these mice at approximately 20 weeks of age. Although this CS model exhibits a severe progeroid phenotype, we found no evidence of in vitro endothelial cell dysfunction, as assessed by measuring population doubling time, migration capacity, and ICAM-1 expression. Furthermore, aortas from CX mice did not exhibit early senescence nor reduced angiogenesis capacity. Despite these observations, CX mice presented blood brain barrier disruption and increased senescence of brain endothelial cells. This was accompanied by an upregulation of inflammatory markers in the brains of CX mice, such as ICAM-1, TNF alpha, p-p65, and glial cell activation. Inhibition of neovascularization did not exacerbate neither astro-nor microgliosis, suggesting that the pro-inflammatory phenotype is independent of the neurovascular dysfunction present in CX mice. These findings have implications for the etiology of this disease and could contribute to the study of novel therapeutic targets for treating Cockayne syndrome patients.

Skin aging has been primarily related to aesthetics and beauty. Therefore, interventions have focused on reestablishing skin appearance, but not necessarily skin health, function, and resilience. Recently, cellular senescence was shown to play a role in age-related skin function deterioration and influence organismal health and, potentially, longevity. In the present study, a two-step screening was performed to identify peptides capable of reducing cellular senescence in human dermal fibroblasts (HDF) from Hutchinson-Gilford Progeria (HGPS) patients. From the top four peptides of the first round of screening, we built a 764-peptide library using amino acid scanning, of which the second screen led to the identification of peptide 14. Peptide 14 effectively decreased HDF senescence induced by HGPS, chronological aging, ultraviolet-B radiation, and etoposide treatment, without inducing significant cell death, and likely by modulating longevity and senescence pathways. We further validated the effectiveness of peptide 14 using human skin equivalents and skin biopsies, where peptide 14 promoted skin health and reduced senescent cell markers, as well as the biological age of samples, according to the Skin-Specific DNA methylation clock, MolClock. Topical application of peptide 14 outperformed Retinol treatment, the current gold-standard in “anti-aging” skin care. Finally, we determined that peptide 14 is safe for long-term applications and also significantly extends both the lifespan and healthspan of C. elegans worms tested in two independent testings. This highlights the potential for geroprotective applications of the senotherapeutic compounds identified using our screening platform beyond the skin.

Background DNA methylation (DNAm) age constitutes a powerful tool to assess the molecular age and overall health status of biological samples. Recently, it has been shown that tissue-specific DNAm age predictors may present superior performance compared to the pan- or multi-tissue counterparts. The skin is the largest organ in the body and bears important roles, such as body temperature control, barrier function, and protection from external insults. As a consequence of the constant and intimate interaction between the skin and the environment, current DNAm estimators, routinely trained using internal tissues which are influenced by other stimuli, are mostly inadequate to accurately predict skin DNAm age. Results In the present study, we developed a highly accurate skin-specific DNAm age predictor, using DNAm data obtained from 508 human skin samples. Based on the analysis of 2,266 CpG sites, we accurately calculated the DNAm age of cultured skin cells and human skin biopsies. Age estimation was sensitive to the biological age of the donor, cell passage, skin disease status, as well as treatment with senotherapeutic drugs. Conclusions This highly accurate skin-specific DNAm age predictor constitutes a holistic tool that will be of great use in the analysis of human skin health status/molecular aging, as well as in the analysis of the potential of established and novel compounds to alter DNAm age.

Hypertriglyceridemia is an independent risk factor for cardiovascular disease. Dietary interventions based on protein restriction (PR) reduce circulating triglycerides (TGs), but underlying mechanisms and clinical relevance remain unclear. Here, we show that 1 week of a protein-free diet without enforced calorie restriction significantly lowered circulating TGs in both lean and diet-induced obese mice. Mechanistically, the TG-lowering effect of PR was due, in part, to changes in very low-density lipoprotein (VLDL) metabolism both in liver and peripheral tissues. In the periphery, PR stimulated VLDL-TG consumption by increasing VLDL-bound APOA5 expression and promoting VLDL-TG hydrolysis and clearance from circulation. The PR-mediated increase in Apoa5 expression was controlled by the transcription factor CREBH, which coordinately regulated hepatic expression of fatty acid oxidation-related genes, including Fgf21 and Ppara. The CREBH-APOA5 axis activation upon PR was intact in mice lacking the GCN2-dependent amino acid-sensing arm of the integrated stress response. However, constitutive hepatic activation of the amino acid-responsive kinase mTORC1 compromised CREBH activation, leading to blunted APOA5 expression and PR-recalcitrant hypertriglyceridemia. PR also contributed to hypotriglyceridemia by reducing the rate of VLDL-TG secretion, independently of activation of the CREBH-APOA5 axis. Finally, a randomized controlled clinical trial revealed that 4-6 weeks of reduced protein intake (7%-9% of calories) decreased VLDL particle number, increased VLDL-bound APOA5 expression, and lowered plasma TGs, consistent with mechanistic conservation of PR-mediated hypotriglyceridemia in humans with translational potential as a nutraceutical intervention for dyslipidemia.

Angiogenesis, the formation of new blood vessels by endothelial cells (ECs), is an adaptive response to oxygen/nutrient deprivation orchestrated by vascular endothelial growth factor (VEGF) upon ischemia or exercise. Hypoxia is the best-understood trigger of VEGF expression via the transcription factor HIF1 alpha. Nutrient deprivation is inseparable from hypoxia during ischemia, yet its role in angiogenesis is poorly characterized. Here, we identified sulfur amino acid restriction as a proangiogenic trigger, promoting increased VEGF expression, migration and sprouting in ECs in vitro, and increased capillary density in mouse skeletal muscle in vivo via the GCN2/ATF4 amino acid starvation response pathway independent of hypoxia or HIF1 alpha. We also identified a requirement for cystathionine-gamma-lyase in VEGF-dependent angiogenesis via increased hydrogen sulfide (H2S) production. H2S mediated its proangiogenic effects in part by inhibiting mitochondrial electron transport and oxidative phosphorylation, resulting in increased glucose uptake and glycolytic ATP production.

Decreased growth hormone (GH) and thyroid hormone (TH) signaling are associated with longevity and metabolic fitness. The mechanisms underlying these benefits are poorly understood, but may overlap with those of dietary restriction (DR), which imparts similar benefits. Recently we discovered that hydrogen sulfide (H2S) is increased upon DR and plays an essential role in mediating DR benefits across evolutionary boundaries. Here we found increased hepatic H2S production in long-lived mouse strains of reduced GH and/or TH action, and in a cell-autonomous manner upon serum withdrawal in vitro. Negative regulation of hepatic H2S production by GH and TH was additive and occurred via distinct mechanisms, namely direct transcriptional repression of the H2S-producing enzyme cystathionine γ-lyase (CGL) by TH, and substrate-level control of H2S production by GH. Mice lacking CGL failed to downregulate systemic T4 metabolism and circulating IGF-1, revealing an essential role for H2S in the regulation of key longevity-associated hormones.

Accumulation of DNA damage is intricately linked to aging, aging-related diseases and progeroid syndromes such as Cockayne syndrome (CS). Free radicals from endogenous oxidative energy metabolism can damage DNA, however the potential of acute or chronic DNA damage to modulate cellular and/or organismal energy metabolism remains largely unexplored. We modeled chronic endogenous genotoxic stress using a DNA repair-deficient Csa-/-|Xpa-/- mouse model of CS. Exogenous genotoxic stress was modeled in mice in vivo and primary cells in vitro treated with different genotoxins giving rise to diverse spectrums of lesions, including ultraviolet radiation, intrastrand crosslinking agents and ionizing radiation. Both chronic endogenous and acute exogenous genotoxic stress increased mitochondrial fatty acid oxidation (FAO) on the organismal level, manifested by increased oxygen consumption, reduced respiratory exchange ratio, progressive adipose loss and increased FAO in tissues ex vivo. In multiple primary cell types, the metabolic response to different genotoxins manifested as a cell-autonomous increase in oxidative phosphorylation (OXPHOS) subsequent to a transient decline in steady-state NAD+ and ATP levels, and required the DNA damage sensor PARP-1 and energy-sensing kinase AMPK. We conclude that increased FAO/OXPHOS is a general, beneficial, adaptive response to DNA damage on cellular and organismal levels, illustrating a fundamental link between genotoxic stress and energy metabolism driven by the energetic cost of DNA damage. Our study points to therapeutic opportunities to mitigate detrimental effects of DNA damage on primary cells in the context of radio/chemotherapy or progeroid syndromes.

Objective: Angiogenesis is crucial to maintain tissue homeostasis under nutrient and oxygen deprivation (ischemia). Although considerable evidence supports that angiogenesis is regulated by hypoxia-HIF1α induction of vascular endothelial growth factor (VEGF), the role of nutrient deprivation in angiogenesis is poorly defined. Approach and Results: We report that nutrient deprivation in the form of dietary sulfur amino acid restriction (Methionine/cysteine Restriction; MR) promotes VEGF expression and functional growth of new capillaries in skeletal muscle of mice (Fig.1 A, B). This occurred independently of hypoxia or HIF1α, but instead required the amino acid-sensing eIF2α kinase GCN2 and the transcription factor ATF4 (Fig. 1C). In addition to increased VEGF, nutrient deprivation boosted production of the pro-angiogenic gas hydrogen sulfide (H 2 S) via increased GCN2/ATF4-dependent expression of the H 2 S-generating enzyme cystathionine-gamma-lyase (CGL). The genetic requirement for CGL in angiogenesis triggered by nutrient deprivation, exercise or local VEGF overexpression, as well as the ability of local CGL overexpression to promote angiogenesis in vivo, revealed the critical importance of CGL-derived H 2 S in angiogenesis (Fig. 1D). Finally, plasma H 2 S was reduced in patients with vascular disease (versus non-diseased age-matched controls), and correlated with 2-year survival following vascular surgery (Fig.1 E, F). Conclusions: These data reveal a nutrient-sensing pathway targetable by diet as a previously unrecognized central regulator of VEGF expression and angiogenesis independent of canonical hypoxic signaling. This discovery points to novel dietary interventions and GCN2/ATF4/CGL/H 2 S-based strategies to manipulate angiogenesis. Figure 1

Dietary restriction (DR) without malnutrition encompasses numerous regimens with overlapping benefits including longevity and stress resistance, but unifying nutritional and molecular mechanisms remain elusive. In a mouse model of DR-mediated stress resistance, we found that sulfur amino acid (SAA) restriction increased expression of the transsulfuration pathway (TSP) enzyme cystathionine gamma-lyase (CGL), resulting in increased hydrogen sulfide (H2S) production and protection from hepatic ischemia reperfusion injury. SAA supplementation, mTORC1 activation, or chemical/genetic CGL inhibition reduced H2S production and blocked DR-mediated stress resistance. In vitro, the mitochondrial protein SQR was required for H2S-mediated protection during nutrient/oxygen deprivation. Finally, TSP-dependent H2S production was observed in yeast, worm, fruit fly, and rodent models of DR-mediated longevity. Together, these data are consistent with evolutionary conservation of TSP-mediated H2S as a mediator of DR benefits with broad implications for clinical translation.

Mitochondrial dysfunction is a common feature in neurodegeneration and aging. We identify mitochondrial dysfunction in xeroderma pigmentosum group A (XPA), a nucleotide excision DNA repair disorder with severe neurodegeneration, in silico and in vivo. XPA-deficient cells show defective mitophagy with excessive cleavage of PINK1 and increased mitochondrial membrane potential. The mitochondrial abnormalities appear to be caused by decreased activation of the NAD(+)-SIRT1-PGC-1α axis triggered by hyperactivation of the DNA damage sensor PARP-1. This phenotype is rescued by PARP-1 inhibition or by supplementation with NAD(+) precursors that also rescue the lifespan defect in xpa-1 nematodes. Importantly, this pathogenesis appears common to ataxia-telangiectasia and Cockayne syndrome, two other DNA repair disorders with neurodegeneration, but absent in XPC, a DNA repair disorder without neurodegeneration. Our findings reveal a nuclear-mitochondrial crosstalk that is critical for the maintenance of mitochondrial health.