Inhibition Of Senescence Research Articles

Regulatory mechanisms associating innate leaves senescence of incongruent species

<p class="abstract"><strong>Background:</strong> Senescence is the final developmental phase of a leaf which starts with nutrient salvage and ends with cell death. The first visible event during senescence is leaf yellowing, which typically starts at the leaf margins and progresses to the interior of the leaf blade. Though, regulators of senescence adopt a range of physiological and developmental mechanisms which undergo senescence of plant.</p><p class="abstract"><strong>Methods:</strong> Leaves of different species were collected from the green house, and then rinsed several times with sterilized distilled water. For discs of leaves, two same sized leaves were collected and made the same sized discs. The samples were infiltrated with specific senescence inhibitor. The discs then kept in distilled water and placed under condition at 25<sup>0 </sup>C. Observed the phenotypes at two days interval, molecular based analysis was perfumed at 8<sup>th</sup> day of infiltration. </p><p class="abstract"><strong>Results:</strong> In this study, innate senescence approach comparison to inhibitor based senescence has been performed in order to check its consequences on leaves of different crops such as; cauliflower, apple, tobacco, rose and Arabidopsis. Arabidopsis and apple have resulted in a narrative phenotype with high level of ion leakage. While in case of rose and cauliflower, the phenotype was characterized with yellow fading of leaves. Interestingly, in the tobacco<em> </em>plants, intense yellowing of leaves developed along bottom. Further, in order to confirm the efficiency and pattern of senescence, we had also assessed the changes occurred during leaf senescence via ion leakage and chlorophyll content, expression of SAG12 (a senescence associated gene) and (PSA) photosynthetic associated genes expression as markers.</p><p><strong>Conclusions:</strong> It has been noted that progression of leaf senescence is a very critical and important factors affecting plant growth and development. It can be stated that initiation of leaves senescence can be controlled by using specific inhibitor.</p>

Over Expression of Long Non-Coding RNA PANDA Promotes Hepatocellular Carcinoma by Inhibiting Senescence Associated Inflammatory Factor IL8

It has been reported that long non-coding RNA PANDA was disregulated in varieties types of tumor, but its expression level and biological role in hepatocellular carcinoma (HCC) remains contradictory. We detected PANDA expression in two independent cohorts (48 HCC patients following liver transplantation and 84 HCC patients following liver resection), and found that PANDA was down-regulated in HCC. Thereafter we explored its function in cancer biology by inversing its low expression. Surprisingly, overexpression of PANDA promoted HCC proliferation and carcinogenesis in vitro and in vivo. Mechanistically, PANDA repressed transcriptional activity of senescence associated inflammatory factor IL8, which leaded to inhibition of cellular senescence. Therefore, our research help to better understand the complex role of PANDA in HCC, and suggest more thoughtful strategies should be applied before it can be treated as a potential therapeutic target.

TRIB3 Promotes APL Progression through Stabilization of the Oncoprotein PML-RARα and Inhibition of p53-Mediated Senescence

Acute promyelocytic leukemia (APL) is driven by the oncoprotein PML-RARα, which antagonizes myeloid differentiation and promotes APL-initiating cell self-renewal. Combined all-trans retinoic acid (ATRA) with arsenic trioxide (As2O3) or chemotherapy dramatically improves the prognosis of APL patients. Here we report that expression of pseudokinase Tribble 3 (TRIB3) associates positively with APL progression and therapeutic resistance. The elevated TRIB3 expression promotes APL by interacting with PML-RARα and suppressing its sumoylation, ubiquitylation, and degradation. This represses PML nuclear body assembly, p53-mediated senescence, and cell differentiation, and supports cellular self-renewal. Genetically inhibiting TRIB3 expression or combination of a peptide disturbing TRIB3/PML-RARα interaction with ATRA/As2O3 eradicates APL by accelerating PML-RARα degradation. Our study provides insight into APL pathogenesis and a potential therapeutic option against APL.

Glutathione Peroxidase 3 (GPx3) Delivered by hiPSC-MSCs Ameliorated Hepatic IR Injuries Via Inhibition of Hepatic Senescence

Glutathione Peroxidase 3 (GPx3) Delivered by hiPSC-MSCs Ameliorated Hepatic IR Injuries Via Inhibition of Hepatic Senescence

Nucleus accumbens-1/GADD45GIP1 axis mediates cisplatin resistance through cellular senescence in ovarian cancer.

Nucleus accumbens-1 (NAC1), a nuclear factor belonging to the bric-a-brac-tramtrack-broad complex/pox virus and zinc finger gene family, is known to serve important roles in the proliferation and growth of tumor cells, and in chemotherapy resistance. However, the underlying molecular mechanisms through which NAC1 contributes to drug resistance remain unclear. In the present study, the role of NAC1 in drug resistance in ovarian cancer was investigated. NAC1 expression was markedly negatively associated with growth arrest and DNA-damage-inducible 45γ-interacting protein 1 (GADD45GIP1) expression in ovarian cancer. Increased NAC1 expression or decreased GADD45GIP1 expression was significantly associated with decreased progression-free survival (P=0.0041). Multivariate analysis demonstrated that NAC1/GADD45GIP1 expression was an independent prognostic factor of progression-free survival (P=0.0405). It was investigated whether cellular senescence was involved in NAC1-mediated resistance to cisplatin, a commonly used chemotherapeutic drug in the treatment of ovarian cancer. Treatment with cisplatin activated cellular senescence in ovarian cancer cell lines (SKOV3 and TOV-21G cells). Furthermore, knockdown of NAC1 by RNA interference significantly increased GADD45GIP1 expression and inhibited cisplatin-induced cellular senescence, resulting in increased cisplatin cytotoxicity in SKOV3 cells, which express increased levels of NAC1. To investigate whether the sensitizing effect of NAC1 inhibition on cisplatin-induced cytotoxicity may be attributed to the suppression of cellular senescence, the effects of NAC1 overexpression were assessed in TOV-21G cells, which do not express endogenous NAC1. Transfection with NAC1 in TOV-21G cells reduced the sensitivity of TOV-21G cells to cisplatin, indicating that suppression of cellular senescence was induced by GADD45GP1 activation. The results of the present study suggest that NAC1 is a negative regulator of cellular senescence and that NAC1-dependent suppression of senescence, mediated through GADD45GIP1, serves an important role in promoting cisplatin resistance. Therefore, the NAC1/GADD45GIP1 axis may be a potential target for the treatment of ovarian cancer, particularly in platinum-resistant cancers.

Perfluorodecanoic acid (PFDA) promotes gastric cell proliferation via sPLA2-IIA.

The association of perfluorodecanoicacid (PFDA) with tumor promotion and associated effects is not clear. Given that PDFA is mostly consumed with food and drinking water, we evaluated the effects of PFDA on a gastric cell line. When added to cell cultures, PFDA significantly increased growth rate and colony forming ability compared with control treatment. We found that suppression of cell senescence, but not apoptosis or autophagy was associated with PFDA-induced promotion of cell amount. To determine the molecular mechanism that was involved, DNA microarray assays was used to analyze changes in gene expression in response to PFDA treatment. Data analysis demonstrated that the vascular endothelial growth factor signaling pathway had the lowest p-value, with sPLA2-IIA (pla2g2a) exhibits the most altered expression pattern within the pathway. Moreover, sPLA2-IIA and its transcription factor TCF4, known as a direct target and a binding partner of Wnt/β-catenin signaling in gastric cells respectively, were the third and second most varied genes globally. Cells transfected with expression plasmids pENTER-tcf4 and pENTER-pla2g2a show reduced cell proliferation by more than 60% and 30% respectively. Knockdown with sPLA2-IIA siRNA provided additional evidence that sPLA2-IIA was a mediator of PFDA-induced cell senescence suppression. The results suggest for the first time that PFDA induced suppression of cell senescence through inhibition of sPLA2-IIA protein expression and might increased the proliferative capacity of an existing tumor.

Mcl-1 regulates reactive oxygen species via NOX4 during chemotherapy-induced senescence.

Mcl-1, a Bcl-2 family member, is highly expressed in a variety of human cancers and is believed to enhance tumorigenic potential and chemotherapy resistance through the inhibition of apoptosis and senescence. We previously reported that Mcl-1′s regulation of chemotherapy-induced senescence (CIS) is dependent on its ability to prevent reactive oxygen species (ROS) generation. In this report, we demonstrate that Mcl-1-regulated CIS requires not only ROS, but specifically mitochondrial ROS, and that these events are upstream of activation of the DNA damage response, another necessary step toward senescence. Mcl-1′s anti-senescence activity also involves the unique ability to inhibit ROS formation by preventing the upregulation of pro-oxidants. Specifically, we found that NADPH oxidases (NOXs) are regulated by Mcl-1 and that NOX4 expression in particular is a required step for CIS induction that is blocked by Mcl-1. Lastly, we illustrate that by preventing expression of NOX4, Mcl-1 limits its availability in the mitochondria, thereby lowering the production of mitochondrial ROS during CIS. Our studies not only define the essential role of Mcl-1 in chemoresistance, but also for the first time link a key pro-survival Bcl-2 family member with the NOX protein family, both of which have significant ramifications in cancer progression.

Effect of lifelong football training on the expression of muscle molecular markers involved in healthy longevity

We investigated whether lifelong football training affects the expression of healthy longevity-related muscle molecular markers. Biopsies were collected from the vastus lateralis muscle of 10 lifelong football-trained men (68.2 ± 3.0years) and of 10 active untrained healthy men (66.7 ± 1.3years). Gene and protein expression was measured by RTqPCR on RNA and by western blotting on protein extracts from muscle biopsies, respectively. The expression of AMPKα1/α2, NAMPT, TFAM and PGC1α, which are markers of oxidative metabolism, and MyHC β isoform expression was higher in the muscle of football-trained men vs untrained men. Also citrate synthase activity was higher in trained than in untrained men (109.3 ± 9.2 vs 75.1 ± 9.2mU/mg). These findings were associated with a healthier body composition in trained than in untrained men [body weight: 78.2 ± 6.5 vs 91.2 ± 11.2kg; body mass index BMI: 24.4 ± 1.6 vs 28.8 ± 4.0kgm-2; fat%: 22.6 ± 8.0 vs 31.4 ± 5.0%)] and with a higher maximal oxygen uptake (VO2max: 34.7 ± 3.8 vs 27.3 ± 4.0ml/min/kg). Also the expression of proteins involved in DNA repair and in senescence suppression (Erk1/2, Akt and FoxM1) was higher in trained than in untrained men. At BMI- and age-adjusted multiple linear regression analysis, fat percentage was independently associated with Akt protein expression, and VO2max was independently associated with TFAM mRNA and with Erk1/2 protein expression. Lifelong football training increases the expression of key markers involved in muscle oxidative metabolism, and in the DNA repair and senescence suppression pathways, thus providing the molecular basis for healthy longevity.

Notch activation enhances mesenchymal stem cell sheet osteogenic potential by inhibition of cellular senescence

Our previous studies have confirmed the therapeutic effects of mesenchymal stem cell (MSC) monolayer sheet transplantation on allograft repair. A limiting factor in their application is the loss of MSC multi-potency as a result of high density sheet culture-induced senescence. In the study reported in this article, we tested whether Notch activation could be used to prevent or delay sheet culture-induced cell aging. Our results showed that, during in vitro long-term (5-day) cell sheet culture, MSCs progressively lose their progenitor characteristics. In contrast, Notch activation by Jagged1 in MSC sheet culture showed reduced cellular senescence and cell cycle arrest compared with control MSCs without Notch activation. Importantly, knockdown of Notch target gene Hes1 totally blocked the inhibition effect of Jagged1 on cellular senescence. Finally, the in vivo allograft transplantation data showed a significant enhanced callus formation and biomechanical properties in Notch activation cultured long-term sheet groups when compared with long-term cultured sheet without Notch activation. Our results suggest that Notch activation by Jagged1 could be used to overcome the stem cell aging caused by high density sheet culture, thereby increasing the therapeutic potential of MSC sheets for tissue regeneration.

Red Yeast Rice Protects Circulating Bone Marrow-Derived Proangiogenic Cells against High-Glucose-Induced Senescence and Oxidative Stress: The Role of Heme Oxygenase-1.

The inflammation and oxidative stress of bone marrow-derived proangiogenic cells (PACs), also named endothelial progenitor cells, triggered by hyperglycemia contributes significantly to vascular dysfunction. There is supporting evidence that the consumption of red yeast rice (RYR; Monascus purpureus-fermented rice) reduces the vascular complications of diabetes; however, the underlying mechanism remains unclear. This study aimed to elucidate the effects of RYR extract in PACs, focusing particularly on the role of a potent antioxidative enzyme, heme oxygenase-1 (HO-1). We found that treatment with RYR extract induced nuclear factor erythroid-2-related factor nuclear translocation and HO-1 mRNA and protein levels in PACs. RYR extract inhibited high-glucose-induced (30 mM) PAC senescence and the development of reactive oxygen species (ROS) in a dose-dependent manner. The HO-1 inducer cobalt protoporphyrin IX also decreased high-glucose-induced cell senescence and oxidative stress, whereas the HO-1 enzyme inhibitor zinc protoporphyrin IX and HO-1 small interfering RNA significantly reversed RYR extract-caused inhibition of senescence and reduction of oxidative stress in high-glucose-treated PACs. These results suggest that RYR extract serves as alternative and complementary medicine in the treatment of these diseases, by inducing HO-1, thereby decreasing the vascular complications of diabetes.

Mitotic catastrophe and cell cycle arrest are alternative cell death pathways executed by bortezomib in rituximab resistant B-cell lymphoma cells.

The ubiqutin-proteasome system (UPS) plays a role in rituximab-chemotherapy resistance and bortezomib (BTZ) possesses caspase-dependent (i.e. Bak stabilization) and a less characterized caspase–independent mechanism-of-action(s). Here, we define BTZ-induced caspase-independent cell death pathways. A panel of rituximab-sensitive (RSCL), rituximab-resistant cell lines (RRCL) and primary tumor cells derived from lymphoma patients (N = 13) were exposed to BTZ. Changes in cell viability, cell-cycle, senescence, and mitotic index were quantified. In resting conditions, RRCL exhibits a low-proliferation rate, accumulation of cells in S-phase and senescence. Exposure of RRCL to BTZ reduces cell senescence, induced G2-M phase cell-cycle arrest, and is associated with mitotic catastrophe. BTZ stabilized p21, CDC2, and cyclin B in RRCL and in primary tumor cells. Transient p21 knockdown alleviates BTZ-induced senescence inhibition, G2-M cell cycle blockade, and mitotic catastrophe. Our data suggest that BTZ can induce apoptosis or mitotic catastrophe and that p21 has a pivotal role in BTZ activity against RRCL.

ZEB1-induced tumourigenesis requires senescence inhibition via activation of DKK1/mutant p53/Mdm2/CtBP and repression of macroH2A1

ObjectiveUnderstand the role of ZEB1 in the tumour initiation and progression beyond inducing an epithelial-to-mesenchymal transition.DesignExpression of the transcription factor ZEB1 associates with a worse prognosis in most cancers, including...

Inhibition of Both Activated p38 Mapkα and mTORC1 Potentiates the Effect of SR1 on Promoting Hematopoietic Reconstitution of Ex Vivo Expanded Human Umbilical Cord Blood Hematopoietic Stem Cells

Hematopoietic stem cells (HSCs) transplantation is a curative treatment for various hematological disorders. However, its use has been limited in part by the difficulty to obtain sufficient numbers of transplantable HSCs, especially from human umbilical cord bloods (hUCBs). Ex vivo expansion of cord blood HSCs can potentially mitigate the shortfall if the expanded HSCs can maintain their stemness for long-term hematopoietic reconstitution.Our previous studies have shown that some hUCB HSCs lose their "stemness" during ex vivo expansion in part due to induction of cellular senescence via p38MAPKα(p38) or mTORC1 activation and inhibition of p38 and mTOR C1 promotes HSCs expansion (Ann Hematol. 2012; 91:813-823; Transplantation. 2014; 97:20-29). Recently, it was shown that StemRegenin 1 (SR1), an antagonist of the aryl hydrocarbon receptor (AHR), could significantly promote ex vivo expansion of HSCs, primarily via inhibition of HSCs differentiation. Therefore, we investigated whether the combination of SR1 with LY2228820 (LY) and rapamycin (Rapa) to inhibit p38 and mTOR C1, respectively, can enhance ex vivo expansion of hUBC HSCs by inhibiting HSCs differentiation and senescence.Next, we confirmed that co-inhibition of activated p38 and mTORC1 promotes SR1 induced HSCs expansion ex vivo through inhibition of senescence.Our combined culture system in 7 days led to a 16.2-fold increase for CD34+CD90+ subpopulation and a 6.1-fold increase for CD34+CD45RA- subpopulation compared to input cells (Blood,126(23):381-381) .To determine the hematopoietic reconstitution ability of the progeny of hUCB CD34+ cells expanded with inhibitors of p38, mTORC1, and AHR, we transplanted 1000 to 30000 uncultured hUCB CD34+ cells or a fraction of the final culture equivalent to 1000 to 10000 starting hUCB CD34+ cells into sub-lethally irradiated 6 to 10-week-old NOD-Prkdcscid Il2rgnull (NOG) mice. Donor cell engraftment was analyzed at 8 and 13 weeks after the transplantation. The results showed that the progeny of 10,000 hUCB CD34+ cells cultured with LY, Rapa and SR1 produced significant greater donor cell engraftment in the recipients’ peripheral blood compared with the same number of un-cultured hUCB CD34+ cells (2.19-fold, p<0.01), the progeny of 10,000 hUCB CD34+ cells cultured with vehicle (2.76-fold, p<0.01), or the progeny of 10,000 hUCB CD34+ cells cultured with SR1 alone (1.72-fold, p<0.05). The similar results were also found in the recipients’ bone marrow and spleen. In addition, the progeny of 3,000 hUCB CD34+ cells cultured with LY, Rapa and SR1 generated the levels of peripheral blood donor cell engraftment that similar to these produced by 30,000 un-cultured hUCB CD34+ cells and the progeny of 10,000 hUCB CD34+ cells cultured with SR1 alone. More importantly, the hUCB CD34+ cells cultured with LY, Rapa and SR1 retained multi-lineage differentiation potential as they were able to produce human B cells, myeloid cells, red blood cell progenitors, megakaryocytes, and NK cells in the recipients’ bone marrow. Furthermore, hUCB CD34+ cells cultured with LY, Rapa and SR1 produced the highest levels of human HSCs (CD34+CD38-/CD34+CD90+/CD34+CD45RA- cells) engraftment compared to all other cells.Collectively, these findings suggest that the combined inhibition of p38, mTORC1 and AHR is more effective in promoting the expansion and hematopoietic reconstitution of hUBC HSCs than inhibition of p38, mTORC1 or ARH alone probably via inhibition of HSCs senescence and differentiation. This new strategy may alleviate the limitation of hUBC for transplantation in adult patients. DisclosuresNo relevant conflicts of interest to declare.

Age-Associated Sirtuin 1 Reduction in Vascular Smooth Muscle Links Vascular Senescence and Inflammation to Abdominal Aortic Aneurysm.

Uncontrolled growth of abdominal aortic aneurysms (AAAs) is a life-threatening vascular disease without an effective pharmaceutical treatment. AAA incidence dramatically increases with advancing age in men. However, the molecular mechanisms by which aging predisposes individuals to AAAs remain unknown. In this study, we investigated the role of SIRT1 (Sirtuin 1), a class III histone deacetylase, in AAA formation and the underlying mechanisms linking vascular senescence and inflammation. The expression and activity of SIRT1 were significantly decreased in human AAA samples. SIRT1 in vascular smooth muscle cells was remarkably downregulated in the suprarenal aortas of aged mice, in which AAAs induced by angiotensin II infusion were significantly elevated. Moreover, vascular smooth muscle cell-specific knockout of SIRT1 accelerated angiotensin II-induced formation and rupture of AAAs and AAA-related pathological changes, whereas vascular smooth muscle cell-specific overexpression of SIRT1 suppressed angiotensin II-induced AAA formation and progression in Apoe-/- mice. Furthermore, the inhibitory effect of SIRT1 on AAA formation was also proved in a calcium chloride (CaCl2)-induced AAA model. Mechanistically, the reduction of SIRT1 was shown to increase vascular cell senescence and upregulate p21 expression, as well as enhance vascular inflammation. Notably, inhibition of p21-dependent vascular cell senescence by SIRT1 blocked angiotensin II-induced nuclear factor-κB binding on the promoter of monocyte chemoattractant protein-1 and inhibited its expression. These findings provide evidence that SIRT1 reduction links vascular senescence and inflammation to AAAs and that SIRT1 in vascular smooth muscle cells provides a therapeutic target for the prevention of AAA formation.

Artesunate enhances the therapeutic response of glioma cells to temozolomide by inhibition of homologous recombination and senescence.

Glioblastoma multiforme (GBM), a malignant brain tumor with a dismal prognosis, shows a high level of chemo- and radioresistance and, therefore, attempts to sensitize glioma cells are highly desired. Here, we addressed the question of whether artesunate (ART), a drug currently used in the treatment of malaria, enhances the killing response of glioblastoma cells to temozolomide (TMZ), which is the first-line therapeutic for GBM. We measured apoptosis, necrosis, autophagy and senescence, and the extent of DNA damage in glioblastoma cells. Further, we determined the tumor growth in nude mice. We show that ART enhances the killing effect of TMZ in glioblastoma cell lines and in glioblastoma stem-like cells. The DNA double-strand break level induced by TMZ was not clearly enhanced in the combined treatment regime. Also, we did not observe an attenuation of TMZ-induced autophagy, which is considered a survival mechanism. However, we observed a significant effect of ART on homologous recombination (HR) with downregulation of RAD51 protein expression and HR activity. Further, we found that ART is able to inhibit senescence induced by TMZ. Since HR and senescence are pro-survival mechanisms, its inhibition by ART appears to be a key node in enhancing the TMZ-induced killing response. Enhancement of the antitumor effect of TMZ by co-administration of ART was also observed in a mouse tumor model. In conclusion, the amelioration of TMZ-induced cell death upon ART co-treatment provides a rational basis for a combination regime of TMZ and ART in glioblastoma therapy.

Cytokinin Response Factor 6 Represses Cytokinin-Associated Genes during Oxidative Stress.

Cytokinin is a phytohormone that is well known for its roles in numerous plant growth and developmental processes, yet it has also been linked to abiotic stress response in a less defined manner. Arabidopsis (Arabidopsis thaliana) Cytokinin Response Factor 6 (CRF6) is a cytokinin-responsive AP2/ERF-family transcription factor that, through the cytokinin signaling pathway, plays a key role in the inhibition of dark-induced senescence. CRF6 expression is also induced by oxidative stress, and here we show a novel function for CRF6 in relation to oxidative stress and identify downstream transcriptional targets of CRF6 that are repressed in response to oxidative stress. Analysis of transcriptomic changes in wild-type and crf6 mutant plants treated with H2O2 identified CRF6-dependent differentially expressed transcripts, many of which were repressed rather than induced. Moreover, many repressed genes also show decreased expression in 35S:CRF6 overexpressing plants. Together, these findings suggest that CRF6 functions largely as a transcriptional repressor. Interestingly, among the H2O2 repressed CRF6-dependent transcripts was a set of five genes associated with cytokinin processes: (signaling) ARR6, ARR9, ARR11, (biosynthesis) LOG7, and (transport) ABCG14. We have examined mutants of these cytokinin-associated target genes to reveal novel connections to oxidative stress. Further examination of CRF6-DNA interactions indicated that CRF6 may regulate its targets both directly and indirectly. Together, this shows that CRF6 functions during oxidative stress as a negative regulator to control this cytokinin-associated module of CRF6-dependent genes and establishes a novel connection between cytokinin and oxidative stress response.

The human amniotic fluid stem cell secretome effectively counteracts doxorubicin-induced cardiotoxicity.

The anthracycline doxorubicin (Dox) is widely used in oncology, but it may cause a cardiomyopathy with bleak prognosis that cannot be effectively prevented. The secretome of human amniotic fluid-derived stem cells (hAFS) has previously been demonstrated to significantly reduce ischemic cardiac damage. Here it is shown that, following hypoxic preconditioning, hAFS conditioned medium (hAFS-CM) antagonizes senescence and apoptosis of cardiomyocytes and cardiac progenitor cells, two major features of Dox cardiotoxicity. Mechanistic studies with mouse neonatal ventricular cardiomyocytes (mNVCM) reveal that hAFS-CM inhibition of Dox-elicited senescence and apoptosis is associated with decreased DNA damage, nuclear translocation of NF-kB, and upregulation of the NF-kB controlled genes, Il6 and Cxcl1, promoting mNVCM survival. Furthermore, hAFS-CM induces expression of the efflux transporter, Abcb1b, and Dox extrusion from mNVCM. The PI3K/Akt signaling cascade, upstream of NF-kB, is potently activated by hAFS-CM and pre-treatment with a PI3K inhibitor abrogates NF-kB accumulation into the nucleus, modulation of Il6, Cxcl1 and Abcb1b, and prevention of Dox-initiated senescence and apoptosis in response to hAFS-CM. These results support the concept that hAFS are a valuable source of cardioprotective factors and lay the foundations for the development of a stem cell-based paracrine treatment of chemotherapy-related cardiotoxicity.

Abstract 1127: KIT/D816V induces SRC-mediated tyrosine phosphorylation of MITF and altered transcription program in melanoma

Abstract The oncogenic D816V mutation of the KIT receptor is well characterized in systemic mastocytosis and acute myeloid leukemia. Although KITD816V has been found in melanoma, its function and involvement in this malignancy is not understood. Here we show that KITD816V induces tyrosine phosphorylation of the microphthalmia-associated transcription factor (MITF) through a triple protein complex formation between KIT, MITF and SRC family kinases. In turn, phosphorylated MITF activates target genes that are involved in melanoma proliferation, cell cycle progression, suppression of senescence, survival and invasion. By blocking the triple protein complex formation, thus preventing MITF phosphorylation, the cells become hypersensitive to SRC inhibitors. We have therefore delineated the mechanism behind the oncogenic effects of KITD816V in melanoma and provide a rationale for testing SRC family kinase inhibitors to treat KITD816V-transformed melanomas. Citation Format: Bengt Phung, Julhash U. Kazi, Alicia Lundby, Kristin Bergsteinsdottir, Jianmin Sun, Colin R. Goding, Göran Jönsson, Jesper V. Olsen, Eiríkur Steingrímsson, Lars Rönnstrand. KIT/D816V induces SRC-mediated tyrosine phosphorylation of MITF and altered transcription program in melanoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1127.

Abstract 3528: Induction of cytoprotective autophagy and senescence in response to chemotherapy and ionizing-radiation as possible mechanisms of tumor dormancy

Abstract Proliferative recovery in tumor cells subsequent to chemotherapy and/or radiation-induced prolonged growth arrest may prove to represent a state of autophagy and/or senescence. This may also be useful as an in vitro model of tumor dormancy. To explore this hypothesis, studies were performed in primary mouse mammary carcinoma cells (MMC) treated with Adriamycin (ADR) (0.25uM - 1uM) for 3 days (2h each) and ionizing radiation (IR) (6Gy). Promotion of autophagy was confirmed based on detection of autophagosomes based on acridine orange staining, the appearance of the autophagy marker, LC3.B, associated with autophagosome, degradation of p62/SQSTM1, a hallmark of autophagic flux (i.e. the completion of autophagy and degradation of autophagosome content). Autophagosome formation was observed 24h post ADR treatment and 72h post IR treatment. Furthermore, to understand the function of autophagy (cytoprotective, nonprotective or cytotoxic) autophagy was blocked either pharmacologically using chloroquine (CQ) or genetically by silencing of the key autophagy gene, ATG5. LC3.B appearance and p62/SQSTM1 degradation were more pronounced in wt (autophagy proficient) cells compared to ATG5 silenced cells, confirming that autophagy was inhibited by these pharmacological and genetic strategies. Furthermore, interference with autophagy sensitized the MMC cells to ADR and radiation. These observations indicated that ADR and IR induced autophagy was cytoprotective in the MMC cells. Unexpectedly, apoptosis (as measured by PI/ Annexin V assay) was not increased in response to either ADR alone or the combination of ADR and CQ, suggesting that sensitization was likely occurring through alternative pathways. An evaluation of select markers of senescence indicated enhanced expression of p21 as well as enhanced secretion of the autophagy/senescence associated marker, HMGB1, in response to Adriamycin and IR. Overall, these data suggest that both autophagy and senescence are induced in breast cancer cells by conventional therapies and could contribute to tumor dormancy. Inhibition of autophagy and/or senescence associated signaling pathways could lead to therapeutic strategies for interfering with proliferative recovery of tumor cells after standard modes of therapy. Studies are currently in progress to determine whether the senescence associated secretory phenotype (SASP) accompanies the induction of senescence. SASP has been associated with promotion of tumor growth and alternatively, with activation of an immune response to therapy. Citation Format: Theresa Thekkudan, Supriya Joshi, David Gewirtz. Induction of cytoprotective autophagy and senescence in response to chemotherapy and ionizing-radiation as possible mechanisms of tumor dormancy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3528.

Abstract 2846: Proliferative recovery after chemotherapy-induced senescence in non-small cell lung cancer (NSCLC) cells as a model of tumor dormancy and disease recurrence

Abstract Lung cancer is the leading cause of cancer-related death in both men and women in the United States. Most lung cancer cases are diagnosed in advanced, inoperable stages and are treated with chemoradiation; while chemoradiation is effective in suppressing tumor progression, recurrence following treatment is not infrequent. As there is no well-accepted preclinical model for tumor dormancy and disease recurrence, we hypothesize that the promotion of transient autophagy and senescence could be developed as a model of tumor growth and recovery following treatment. Studies were performed utilizing H460 NSCLC cells as well as a primary NSCLC cell line isolated and grown in our laboratory from a stage IV adenocarcinoma. After treatment with etoposide (1 uM), growth arrest was accompanied by the robust induction of autophagy and senescence. This growth arrest was transient and was followed by proliferative recovery in the course of several days. Genetic and pharmacological inhibition of autophagy failed to interfere with the ability of the cells to regrow, indicating a non-cytoprotective function of autophagy in response to etoposide (in contrast to the cytoprotective function of autophagy exhibited in response to radiation). Quantification of senescence over time based on C12FDG staining and flow cytometry demonstrated that the reversal of growth arrest coincided with a decline in the extent of senescence. To more precisely define the source of the recovered cells, senescent and non-senescent but growth arrested cells were separated by flow cytometry based on their relative β-galactosidase expression and replated. Both cell populations demonstrated the ability to re-emerge from the growth-arrested state and recover proliferative capacity. These observations suggest that senescence is ultimately a transient process in that at least a subpopulation of tumor cells can and will recover proliferative capacity. Furthermore, the reversibility of therapy-induced senescence (TIS) might prove to be a useful model both in cell culture and in vivo to study tumor dormancy and disease recurrence. Studies are currently in progress to determine the impact of senescence inhibition on tumor recovery as well as sensitivity to subsequent therapy given that recurrent disease tends to be relatively refractory to further treatment. Citation Format: Tareq Saleh, Theresa Thekkudan, Moureq R. Alotaibi, David A. Gewirtz. Proliferative recovery after chemotherapy-induced senescence in non-small cell lung cancer (NSCLC) cells as a model of tumor dormancy and disease recurrence. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2846.