Targeting senescent cells in translational medicine.

Organismal ageing is a complex process driving progressive impairment of functionality and regenerative potential of tissues. Cellular senescence is a state of stable cell cycle arrest occurring in response to damage and stress and is considered a hallmark of ageing. Senescent cells accumulate in multiple organs during ageing, contribute to tissue dysfunction and give rise to pathological manifestations. Senescence is therefore a defining feature of a variety of human age-related disorders, including cancer, and targeted elimination of these cells has recently emerged as a promising therapeutic approach to ameliorate tissue damage and promote repair and regeneration. In addition, in vivo identification of senescent cells has significant potential for early diagnosis of multiple pathologies. Here, we review existing senolytics, small molecules and drug delivery tools used in preclinical therapeutic strategies involving cellular senescence, as well as probes to trace senescent cells. We also review the clinical research landscape in senescence and discuss how identifying and targeting cellular senescence might positively affect pathological and ageing processes

A Two-Photon Probe Based on Naphthalimide-Styrene Fluorophore for the In Vivo Tracking of Cellular Senescence

Cellular senescence is a state of stable cell cycle arrest that can negatively affect the regenerative capacities of tissues and can contribute to inflammation and the progression of various aging-related diseases. Advances in the in vivo detection of cellular senescence are still crucial to monitor the action of senolytic drugs and to assess the early onset or accumulation of senescent cells. Here, we describe a naphthalimide-styrene-based probe (HeckGal) for the detection of cellular senescence both in vitro and in vivo. HeckGal is hydrolyzed by the increased lysosomal β-galactosidase activity of senescent cells, resulting in fluorescence emission. The probe was validated in vitro using normal human fibroblasts and various cancer cell lines undergoing senescence induced by different stress stimuli. Remarkably, HeckGal was also validated in vivo in an orthotopic breast cancer mouse model treated with senescence-inducing chemotherapy and in a renal fibrosis mouse model. In all cases, HeckGal allowed the unambiguous detection of senescence in vitro as well as in tissues and tumors in vivo. This work is expected to provide a potential technology for senescence detection in aged or damaged tissues

Galacto-conjugation of Navitoclax as an efficient strategy to increase senolytic specificity and reduce platelet toxicity.

Pharmacologically active compounds with preferential cytotoxic activity for senescent cells, known as senolytics, can ameliorate or even revert pathological manifestations of senescence in numerous preclinical mouse disease models, including cancer models. However, translation of senolytic therapies to human disease is hampered by their suboptimal specificity for senescent cells and important toxicities that narrow their therapeutic windows. We have previously shown that the high levels of senescence-associated lysosomal β-galactosidase (SA-β-gal) found within senescent cells can be exploited to specifically release tracers and cytotoxic cargoes from galactose-encapsulated nanoparticles within these cells. Here, we show that galacto-conjugation of the BCL-2 family inhibitor Navitoclax results in a potent senolytic prodrug (Nav-Gal), that can be preferentially activated by SA-β-gal activity in a wide range of cell types. Nav-Gal selectively induces senescent cell apoptosis and has a higher senolytic index than Navitoclax (through reduced activation in nonsenescent cells). Nav-Gal enhances the cytotoxicity of standard senescence-inducing chemotherapy (cisplatin) in human A549 lung cancer cells. Concomitant treatment with cisplatin and Nav-Gal in vivo results in the eradication of senescent lung cancer cells and significantly reduces tumour growth. Importantly, galacto-conjugation reduces Navitoclax-induced platelet apoptosis in human and murine blood samples treated ex vivo, and thrombocytopenia at therapeutically effective concentrations in murine lung cancer models. Taken together, we provide a potentially versatile strategy for generating effective senolytic prodrugs with reduced toxicities

Cellular Senescence in Non-Small Cell Lung Cancer: from mechanisms to therapeutic opportunities

Lung cancer is the leading cause of cancer-related deaths in our society due to the inefficiency of early detection strategies and the high rate of treatment failure. Therefore, a better understanding of the mechanisms underlying its origin and the response to current treatment paradigms are crucial to improve lung cancer survival. Cellular senescence is a powerful tumour-suppressive mechanism whereby cells stably enter a cell-cycle arrest in response to oncogenic stress. However, the accumulation of senescent cells can alter the tumour microenvironment through a strong paracrine secretion of factors that can lead to detrimental and tumour-promoting effects. Intriguingly, senescence has been reported to be a defining feature of early lesions in Non-Small Cell Lung Cancer (NSCLC), a subtype that accounts for over 80% cases of lung cancer. In addition, senescence has also been reported to occur in response to the standard of treatment for this disease. It is thus conceivable that senescence may play a role in the origin and progression of this disease, despite a causal connection remains to be deciphered. Pharmacologic therapeutics that preferentially target senescent cells, known as senolytics, have been successful in preventing and even reversing senescence-driven detrimental effects in multiple pathological processes. However, their suboptimal specificity and toxicities hamper their clinical translation. Therefore, the targeting of senescent cells through the development of second-generation senolytics that can overcome these obstacles has the potential to revolutionise cancer treatment. The aim of this work is to define the role of cellular senescence at the origin and progression of lung cancer and in response to chemotherapy, and to develop safer and more effective therapeutic approaches to eliminate senescent cells in the context of lung malignancies. In this thesis, we studied the accumulation of senescent cells during the development of lung adenocarcinoma using a KrasG12V-driven lung cancer mouse model. We demonstrate that senolytic treatment of early lesions results in a significant reduction in lung tumour burden and increased survival, providing evidence of the tumour-promoting effect of senescence in early stages of NSCLC. Our research also reveals that platinum-based chemotherapy of human and murine lung adenocarcinoma cells induces senescence, which in turn promotes malignant phenotypes on untreated cancer cells in a paracrine manner in vitro, in xenografts and in orthotopic models of lung adenocarcinoma. Through high throughput unbiased transcriptomic and proteomic approaches, we show that secreted TGF-β ligands activate the Akt/mTOR pathway in untreated cells resulting in enhanced tumour growth. We further demonstrate that senolytic treatment and pharmacologic inhibition of TGFβR1 in tumours can prevent increased proliferation and enhance survival of lung tumour-bearing mice. In order to develop a novel approach for improved senolytic treatment, we show that the galacto-conjugation of senolytic ABT-263 (navitoclax) in the form of an activatable pro-drug significantly enhances cytotoxicity in combination with cisplatin, resulting in reduced lung cancer tumour growth. Importantly, our approach demonstrates decreased navitoclax-associated toxicities, including platelet apoptosis in human and murine blood treated ex vivo and decreased thrombocytopenia in mouse lung cancer models. In summary, this PhD thesis provides evidence of the incidence and role that cellular senescence plays in promoting the progression of early and advanced NSCLC and demonstrates that cisplatin chemotherapy drives pro-tumorigenic phenotypes in a paracrine fashion, which can be prevented with senolytic and TGFβR1 inhibitory treatments. Lastly, it proposes a novel second-generation therapeutic approach to mitigate senolytic toxicities and enhance the efficiency of targeting senescence in the context of lung cancer

A guide to assessing cellular senescence in vitro and in vivo.

Cellular senescence is a physiological mechanism whereby a proliferating cell undergoes a stable cell cycle arrest upon damage or stress and elicits a secretory phenotype. This highly dynamic and regulated cellular state plays beneficial roles in physiology, such as during embryonic development and wound healing, but it can also result in antagonistic effects in age-related pathologies, degenerative disorders, ageing and cancer. In an effort to better identify this complex state, and given that a universal marker has yet to be identified, a general set of hallmarks describing senescence has been established. However, as the senescent programme becomes more defined, further complexities, including phenotype heterogeneity, have emerged. This significantly complicates the recognition and evaluation of cellular senescence, especially within complex tissues and living organisms. To address these challenges, substantial efforts are currently being made towards the discovery of novel and more specific biomarkers, optimized combinatorial strategies and the development of emerging detection techniques. Here, we compile such advances and present a multifactorial guide to identify and assess cellular senescence in cell cultures, tissues and living organisms. The reliable assessment and identification of senescence is not only crucial for better understanding its underlying biology, but also imperative for the development of diagnostic and therapeutic strategies aimed at targeting senescence in the clinic

Indocyanine Green-based Nanoprobe for In Vivo Detection of Cellular Senescence.

There is an urgent need to improve conventional cancer-treatments by preventing detrimental side effects, cancer recurrence and metastases. Recent studies have shown that presence of senescent cells in tissues treated with chemo- or radiotherapy can be used to predict the effectiveness of cancer treatment. However, although the accumulation of senescent cells is one of the hallmarks of cancer, surprisingly little progress has been made in development of strategies for their detection in vivo. To address a lack of detection tools, we developed a biocompatible, injectable organic nanoprobe (NanoJagg), which is selectively taken up by senescent cells and accumulates in the lysosomes. The NanoJagg probe is obtained by self-assembly of indocyanine green (ICG) dimers using a scalable manufacturing process and characterized by a unique spectral signature suitable for both photoacoustic tomography (PAT) and fluorescence imaging. In vitro, ex vivo and in vivo studies all indicate that NanoJaggs are a clinically translatable probe for detection of senescence and their PAT signal makes them suitable for longitudinal monitoring of the senescence burden in solid tumors after chemotherapy or radiotherapy

Indocyanine Green‐based Nanoprobe for In Vivo Detection of Cellular Senescence

Publication status: PublishedThere is an urgent need to improve conventional cancer‐treatments by preventing detrimental side effects, cancer recurrence and metastases. Recent studies have shown that presence of senescent cells in tissues treated with chemo‐ or radiotherapy can be used to predict the effectiveness of cancer treatment. However, although the accumulation of senescent cells is one of the hallmarks of cancer, surprisingly little progress has been made in development of strategies for their detection in vivo. To address a lack of detection tools, we developed a biocompatible, injectable organic nanoprobe (NanoJagg), which is selectively taken up by senescent cells and accumulates in the lysosomes. The NanoJagg probe is obtained by self‐assembly of indocyanine green (ICG) dimers using a scalable manufacturing process and characterized by a unique spectral signature suitable for both photoacoustic tomography (PAT) and fluorescence imaging. In vitro, ex vivo and in vivo studies all indicate that NanoJaggs are a clinically translatable probe for detection of senescence and their PAT signal makes them suitable for longitudinal monitoring of the senescence burden in solid tumors after chemotherapy or radiotherapy.</jats:p

An Indocyanine Green‐Based Nanoprobe for In Vivo Detection of Cellular Senescence

Publication status: PublishedThere is an urgent need to improve conventional cancer‐treatments by preventing detrimental side effects, cancer recurrence and metastases. Recent studies have shown that presence of senescent cells in tissues treated with chemo‐ or radiotherapy can be used to predict the effectiveness of cancer treatment. However, although the accumulation of senescent cells is one of the hallmarks of cancer, surprisingly little progress has been made in development of strategies for their detection in vivo. To address a lack of detection tools, we developed a biocompatible, injectable organic nanoprobe (NanoJagg), which is selectively taken up by senescent cells and accumulates in the lysosomes. The NanoJagg probe is obtained by self‐assembly of indocyanine green (ICG) dimers using a scalable manufacturing process and characterized by a unique spectral signature suitable for both photoacoustic tomography (PAT) and fluorescence imaging. In vitro, ex vivo and in vivo studies all indicate that NanoJaggs are a clinically translatable probe for detection of senescence and their PAT signal makes them suitable for longitudinal monitoring of the senescence burden in solid tumors after chemotherapy or radiotherapy