DNA-PK is activated by SIRT2 deacetylation to promote DNA double-strand break repair by non-homologous end joining.

DNA-dependent protein kinase (DNA-PK) plays a critical role in non-homologous end joining (NHEJ), the predominant pathway that repairs DNA double-strand breaks (DSB) in response to ionizing radiation (IR) to govern genome integrity. The interaction of the catalytic subunit of DNA-PK (DNA-PKcs) with the Ku70/Ku80 heterodimer on DSBs leads to DNA-PK activation; however, it is not known if upstream signaling events govern this activation. Here, we reveal a regulatory step governing DNA-PK activation by SIRT2 deacetylation, which facilitates DNA-PKcs localization to DSBs and interaction with Ku, thereby promoting DSB repair by NHEJ. SIRT2 deacetylase activity governs cellular resistance to DSB-inducing agents and promotes NHEJ. SIRT2 furthermore interacts with and deacetylates DNA-PKcs in response to IR. SIRT2 deacetylase activity facilitates DNA-PKcs interaction with Ku and localization to DSBs and promotes DNA-PK activation and phosphorylation of downstream NHEJ substrates. Moreover, targeting SIRT2 with AGK2, a SIRT2-specific inhibitor, augments the efficacy of IR in cancer cells and tumors. Our findings define a regulatory step for DNA-PK activation by SIRT2-mediated deacetylation, elucidating a critical upstream signaling event initiating the repair of DSBs by NHEJ. Furthermore, our data suggest that SIRT2 inhibition may be a promising rationale-driven therapeutic strategy for increasing the effectiveness of radiation therapy.

Resilient Consensus Control for Multi-Agent Systems: A Comparative Survey.

Due to the openness of communication network and the complexity of system structures, multi-agent systems are vulnerable to malicious network attacks, which can cause intense instability to these systems. This article provides a survey of state-of-the-art results of network attacks on multi-agent systems. Recent advances on three types of attacks, i.e., those on DoS attacks, spoofing attacks and Byzantine attacks, the three main network attacks, are reviewed. Their attack mechanisms are introduced, and the attack model and the resilient consensus control structure are discussed, respectively, in detail, in terms of the theoretical innovation, the critical limitations and the change of the application. Moreover, some of the existing results along this line are given in a tutorial-like fashion. In the end, some challenges and open issues are indicated to guide future development directions of the resilient consensus of multi-agent system under network attacks.

Bcl2-induced DNA replication stress promotes lung carcinogenesis in response to space radiation.

Space radiation is characterized by high-linear energy transfer (LET) ionizing radiation. The relationships between the early biological effects of space radiation and the probability of cancer in humans are poorly understood. Bcl2 not only functions as a potent antiapoptotic molecule but also as an oncogenic protein that induces DNA replication stress. To test the role and mechanism of Bcl2 in high-LET space radiation-induced lung carcinogenesis, we created lung-targeting Bcl2 transgenic C57BL/6 mice using the CC10 promoter to drive Bcl2 expression selectively in lung tissues. Intriguingly, lung-targeting transgenic Bcl2 inhibits ribonucleotide reductase activity, reduces dNTP pool size and retards DNA replication fork progression in mouse bronchial epithelial cells. After exposure of mice to space radiation derived from 56iron, 28silicon or protons, the incidence of lung cancer was significantly higher in lung-targeting Bcl2 transgenic mice than in wild-type mice, indicating that Bcl2-induced DNA replication stress promotes lung carcinogenesis in response to space radiation. The findings provide some evidence for the relative effectiveness of space radiation and Bcl-2 at inducing lung cancer in mice.

Correction to: Small Molecule KRAS Agonist for Mutant KRAS Cancer Therapy.

An amendment to this paper has been published and can be accessed via the original article.

Small Molecule KRAS Agonist for Mutant KRAS Cancer Therapy.

BACKGROUND: Lung cancer patients with KRAS mutation(s) have a poor prognosis due in part to the development of resistance to currently available therapeutic interventions. Development of a new class of anticancer agents that directly targets KRAS may provide a more attractive option for the treatment of KRAS-mutant lung cancer. RESULTS: Here we identified a small molecule KRAS agonist, KRA-533, that binds the GTP/GDP-binding pocket of KRAS. In vitro GDP/GTP exchange assay reveals that KRA-533 activates KRAS by preventing the cleavage of GTP into GDP, leading to the accumulation of GTP-KRAS, an active form of KRAS. Treatment of human lung cancer cells with KRA-533 resulted in increased KRAS activity and suppression of cell growth. Lung cancer cell lines with KRAS mutation were relatively more sensitive to KRA-533 than cell lines without KRAS mutation. Mutating one of the hydrogen-bonds among the KRA-533 binding amino acids in KRAS (mutant K117A) resulted in failure of KRAS to bind KRA-533. KRA-533 had no effect on the activity of K117A mutant KRAS, suggesting that KRA-533 binding to K117 is required for KRA-533 to enhance KRAS activity. Intriguingly, KRA-533-mediated KRAS activation not only promoted apoptosis but also autophagic cell death. In mutant KRAS lung cancer xenografts and genetically engineered mutant KRAS-driven lung cancer models, KRA-533 suppressed malignant growth without significant toxicity to normal tissues. CONCLUSIONS: The development of this KRAS agonist as a new class of anticancer drug offers a potentially effective strategy for the treatment of lung cancer with KRAS mutation and/or mutant KRAS-driven lung cancer.

Prognostic significance of periostin in colorectal cancer.

Accumulating evidence suggests that periostin is frequently upregulated in tissue injury, inflammation, fibrosis and tumor progression. Periostin expression in cancer cells can promote metastatic potential of colorectal cancer (CRC) via activating PI3K/Akt signaling pathway. Moreover, periostin is observed mainly in tumor stroma and cytoplasm of cancer cells, which may facilitate aggressiveness of CRC. In this review, we summarize information regarding periostin to emphasize its role as a prognostic marker of CRC.

Targeting miR-21 with Sophocarpine Inhibits Tumor Progression and Reverses Epithelial-Mesenchymal Transition in Head and Neck Cancer.

A major challenge for cancer chemotherapy is the development of safe and clinically effective chemotherapeutic agents. With its low toxicity profile, sophocarpine (SC), a naturally occurring tetracyclic quinolizidine alkaloid derived from Sophora alopecuroides L, has shown promising therapeutic properties, including anti-inflammatory, anti-nociceptive, and antivirus activities. However, the antitumor efficacy of SC and its underlying mechanisms have not been completely delineated. In the present study, the inhibitory effect of SC on head and neck squamous cell carcinoma (HNSCC) progression and possible mechanisms for this effect involving microRNA-21 (miR-21) regulation were investigated. By cell viability, Transwell, and wound healing assays, we show that SC effectively inhibited proliferation, invasion, and migration of HNSCC cells. Moreover, SC exerted its growth-inhibitory effect via the downregulation of miR-21 expression by blocking Dicer-mediated miR-21 maturation. Furthermore, SC treatment led to the increased expression of PTEN and p38MAPK phosphorylation as well as the reversal of epithelial-mesenchymal transition (EMT), which was rescued by ectopic expression of miR-21 in cells. Notably, SC dramatically repressed tumor growth without observable tissue cytotoxicity in a mouse xenograft model of HNSCC. Our findings offer a preclinical proof of concept for SC as a leading natural agent for HNSCC cancer therapy.

Tumor pyruvate kinase M2: A promising molecular target of gastrointestinal cancer.

Gastrointestinal (GI) cancer is one of the most common causes of cancer-related deaths worldwide. Tumor markers are valuable in detecting post-surgical recurrence or in monitoring response to chemotherapy. Pyruvate kinase isoform M2 (PKM2), a glycolytic enzyme catalyzing conversion of phosphoenolpyruvate (PEP) to pyruvate, confers a growth advantage to the tumor cells and enables them to adapt to the tumor microenvironment. In this review, we have summarized current research on the expression and regulation of PKM2 in tumor cells, and its potential role in GI carcinogenesis and progression. Furthermore, we have also discussed the potential of PKM2 as a diagnostic and screening marker, and a therapeutic target in GI cancer.

Bcl2 inhibits recruitment of Mre11 complex to DNA double-strand breaks in response to high-linear energy transfer radiation.

High-linear energy transfer ionizing radiation, derived from high charge (Z) and energy (E) (HZE) particles, induces clustered/complex DNA double-strand breaks (DSBs) that include small DNA fragments, which are not repaired by the non-homologous end-joining (NHEJ) pathway. The homologous recombination (HR) DNA repair pathway plays a major role in repairing DSBs induced by HZE particles. The Mre11 complex (Mre11/Rad50/NBS1)-mediated resection of DSB ends is a required step in preparing for DSB repair via the HR DNA repair pathway. Here we found that expression of Bcl2 results in decreased HR activity and retards the repair of DSBs induced by HZE particles (i.e. (56)iron and (28)silicon) by inhibiting Mre11 complex activity. Exposure of cells to (56)iron or (28)silicon promotes Bcl2 to interact with Mre11 via the BH1 and BH4 domains. Purified Bcl2 protein directly suppresses Mre11 complex-mediated DNA resection in vitro. Expression of Bcl2 reduces the ability of Mre11 to bind DNA following exposure of cells to HZE particles. Our findings suggest that, after cellular exposure to HZE particles, Bcl2 may inhibit Mre11 complex-mediated DNA resection leading to suppression of the HR-mediated DSB repair in surviving cells, which may potentially contribute to tumor development.

HOXC10 up-regulation promotes gastric cancer cell proliferation and metastasis through MAPK pathway.

OBJECTIVE: As an important regulator of embryonic morphogenesis, homeodomain-containing gene 10 (HOXC10) has been found to promote progression of human cancers and its expression indicates poor survival outcome. However, very few studies are available on the role of HOXC10 in gastric carcinoma. Therefore, the aim of this study was to determine the role of HOXC10 in gastric cancer and the potential mechanism underlying its function for cancer biology. METHODS: A primary gastric cancer mouse model was obtained via intra-gastric wall injection of gastric cancer cells and was used to evaluate the function of HOXC10 during gastric cancer progression in vivo. Immunohistochemistry was performed to visualize and measure HOXC10 protein expression in gastric cancer tissue. Cells were transfected with plasmids to increase the expression of HOXC10, and siRNA transfection was performed to suppress HOXC10 expression. Reverse transcription polymerase chain reaction (RT-PCR) and western blotting were utilized to measure mRNA and protein expression, respectively. Proliferation, migration, and invasion were investigated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, wound healing assay, and matrigel invasion assay in vitro, respectively. RESULTS: HOXC10 expression was significantly increased in gastric cancer tissues compared to matched normal tissues. HOXC10 up-regulation significantly increased tumor volumes in nude mice. Plasmid transfection significantly increased HOXC10 protein and mRNA expressions and effectively promoted cell proliferation. Moreover, HOXC10 up-regulation significantly promoted migration and invasion of gastric cancer cells. Mechanistic investigation showed that HOXC10 up-regulation significantly increased mRNA and protein expression of mitogen-activated protein kinase (MAPK) signaling related genes, including c-myc, c-jun and p53, while also modulating the phosphorylation of c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK) and P38 but not their total protein levels. CONCLUSIONS: This study demonstrated the tight link between HOXC10 and gastric cancer cell proliferation and metastasis via involvement of the MAPK pathway.

Patient-derived xenografts faithfully replicated clinical outcome in a phase II co-clinical trial of arsenic trioxide in relapsed small cell lung cancer.

BACKGROUND: SCLC has limited treatment options and inadequate preclinical models. Promising activity of arsenic trioxide (ASO) recorded in conventional preclinical models of SCLC supported the clinical evaluation of ASO in patients. We assessed the efficacy of ASO in relapsed SCLC patients and in corresponding patient-derived xenografts (PDX). METHODS: Single arm, Simon 2-stage, phase II trial to enroll patients with relapsed SCLC who have failed at least one line of therapy. ASO was administered as an intravenous infusion over 1-2 h daily for 4 days in week 1 and for 2 days in weeks 2-6 of an 8-week cycle. Treatment continued until disease progression. Pretreatment tumor biopsy was employed for PDX generation through direct implantation into subcutaneous pockets of SCID mice without in vitro manipulation and serially propagated for five generations. Ex vivo efficacy of cisplatin (3 mg/kg i.p. weekly) and ASO (3.75 mg/kg i.p. every other day) was tested in PDX representative of platinum sensitive and platinum refractory SCLC. RESULTS: The best response in 17 evaluable patients was stable disease in 2 (12 %), progressive disease in 15 (88 %) patients and median time-to-progression of seven (range 1-7) weeks. PDX was successfully grown in 5 of 9 (56 %) transplanted biopsy samples. Serially-propagated PDXs preserved characteristic small cell histology and genomic stability confirmed by immunohistochemistry, short tandem repeat (STR) profiling and targeted sequencing. ASO showed in vitro cytotoxicity but lacked in vivo efficacy against SCLC PDX tumor growth. CONCLUSIONS: Cisplatin inhibited growth of PDX derived from platinum-sensitive SCLC but was ineffective against PDX from platinum-refractory SCLC. Strong concordance between clinical and ex vivo effects of ASO and cisplatin in SCLC supports the use of PDX models to prescreen promising anticancer agents prior to clinical testing in SCLC patients. Trial Registration The study was registered at http://www.clinicaltrials.gov (NCT01470248).

Bcl2 negatively regulates DNA double-strand-break repair through a nonhomologous end-joining pathway.

Bcl2 can enhance susceptibility to carcinogenesis, but the mechanism(s) remains fragmentary. Here we discovered that Bcl2 suppresses DNA double-strand-break (DSB) repair and V(D)J recombination by downregulating Ku DNA binding activity, which is associated with increased genetic instability. Exposure of cells to ionizing radiation enhances Bcl2 expression in the nucleus, which interacts with both Ku70 and Ku86 via its BH1 and BH4 domains. Removal of the BH1 or BH4 domain abrogates the inhibitory effect of Bcl2 on Ku DNA binding, DNA-PK, and DNA end-joining activities, which results in the failure of Bcl2 to block DSB repair as well as V(D)J recombination. Intriguingly, Bcl2 directly disrupts the Ku/DNA-PKcs complex in vivo and in vitro. Thus, Bcl2 suppression of the general DSB repair and V(D)J recombination may occur in a mechanism by inhibiting the nonhomologous end-joining pathway, which may lead to an accumulation of DNA damage and genetic instability.

Targeting KRAS-mutant non-small cell lung cancer: challenges and opportunities.

Oncogenic mutations in Kirsten rat sarcoma viral oncogene homolog (KRAS) occur in 15%-30% of non-small cell lung cancer (NSCLC). However, despite decades of intensive research, there is still no direct KRAS inhibitor with clinically proven efficacy. Considering its association with poor treatment response and prognosis of lung cancer, developing an effective inhibitory approach is urgently needed. Here, we review different strategies currently being explored to target KRAS-mutant NSCLC, discuss opportunities and challenges, and also propose some novel methods and concepts with the promise of clinical application.

Distinct roles of Ape1 protein, an enzyme involved in DNA repair, in high or low linear energy transfer ionizing radiation-induced cell killing.

High linear energy transfer (LET) radiation from space heavy charged particles or a heavier ion radiotherapy machine kills more cells than low LET radiation, mainly because high LET radiation-induced DNA damage is more difficult to repair. Relative biological effectiveness (RBE) is the ratio of the effects generated by high LET radiation to low LET radiation. Previously, our group and others demonstrated that the cell-killing RBE is involved in the interference of high LET radiation with non-homologous end joining but not homologous recombination repair. This effect is attributable, in part, to the small DNA fragments (≤40 bp) directly produced by high LET radiation, the size of which prevents Ku protein from efficiently binding to the two ends of one fragment at the same time, thereby reducing non-homologous end joining efficiency. Here we demonstrate that Ape1, an enzyme required for processing apurinic/apyrimidinic (known as abasic) sites, is also involved in the generation of small DNA fragments during the repair of high LET radiation-induced base damage, which contributes to the higher RBE of high LET radiation-induced cell killing. This discovery opens a new direction to develop approaches for either protecting astronauts from exposure to space radiation or benefiting cancer patients by sensitizing tumor cells to high LET radiotherapy.

Bcl2 inhibition of mitochondrial DNA repair.

BACKGROUND: Accumulation of mitochondrial DNA (mtDNA) damage could enhance the frequency of mitochondrial mutations and promote a variety of mitochondria-related diseases, including cancer. However, the mechanism(s) involved are not fully understood. METHODS: Quantitative extended length PCR was used to compare mtDNA and nDNA damage in human lung H1299 cells expressing WT Bcl2 or vector-only control. mtAPE1 endonuclease activity was analyzed by AP oligonucleotide assay. mtDNA mutation was measured by single molecule PCR. Subcellular localization of Bcl2 and APE1 was analyzed by subcellular fractionation. RESULTS: Bcl2, an anti-apoptotic molecule and oncoprotein, effectively inhibits the endonuclease activity of mitochondrial APE1 (mtAPE1), leading to significant retardation of mtDNA repair and enhanced frequency of mtDNA mutations following exposure of cells to hydrogen peroxide (H2O2) or nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK, a carcinogen in cigarette smoke). Inversely, depletion of endogenous Bcl2 by RNA interference increases mtAPE1 endonuclease activity leading to accelerated mtDNA repair and decreased mtDNA mutation. Higher levels of mtAPE1 were observed in human lung cancer cells than in normal human bronchial epithelial cells (i.e. BEAS-2B). Bcl2 partially co-localizes with APE1 in the mitochondria of human lung cancer cells. Bcl2 directly interacts with mtAPE1 via its BH domains. Removal of any of the BH domains from Bcl2 abolishes Bcl2's capacity to interact with mtAPE1 as well as its inhibitory effects on mtAPE1 activity and mtDNA repair. CONCLUSIONS: Based our findings, we propose that Bcl2 suppression of mtDNA repair occurs through direct interaction with mtAPE1 and inhibition of its endonuclease activity in mitochondria, which may contribute to enhanced mtDNA mutations and carcinogenesis.

Protein phosphatase 2A and DNA-dependent protein kinase are involved in mediating rapamycin-induced Akt phosphorylation.

BACKGROUND: The mechanisms underlying rapamycin-induced Akt phosphorylation have not been fully elucidated. RESULTS: Inhibition of PP2A or DNA-PK attenuates or abrogates rapamycin-induced Akt phosphorylation and co-inhibition of mTOR and DNA-PK enhances anticancer activity. CONCLUSION: PP2A-dependent and DNA-PK-mediated mechanism is involved in rapamycin-induced Akt phosphorylation. SIGNIFICANCE: A previously unknown mechanism underlying rapamycin-induced Akt phosphorylation and a novel strategy to enhance mTOR-targeted cancer therapy may be suggested. Inhibition of mammalian target of rapamycin complex 1 (mTORC1), for example with rapamycin, increases Akt phosphorylation while inhibiting mTORC1 signaling. However, the underlying mechanisms have not been fully elucidated. The current study has uncovered a previously unknown mechanism underlying rapamycin-induced Akt phosphorylation involving protein phosphatase 2A (PP2A)-dependent DNA protein kinase (DNA-PK) activation. In several cancer cell lines, inhibition of PP2A with okadaic acid, fostriecin, small T antigen, or PP2A knockdown abrogated rapamycin-induced Akt phosphorylation, and rapamycin increased PP2A activity. Chemical inhibition of DNA-PK, knockdown or deficiency of DNA-PK catalytic subunit (DNA-PKcs), or knock-out of the DNA-PK component Ku86 inhibited rapamycin-induced Akt phosphorylation. Exposure of cancer cells to rapamycin increased DNA-PK activity, and gene silencing-mediated PP2A inhibition attenuated rapamycin-induced DNA-PK activity. Collectively these results suggest that rapamycin induces PP2A-dependent and DNA-PK-mediated Akt phosphorylation. Accordingly, simultaneous inhibition of mTOR and DNA-PK did not stimulate Akt activity and synergistically inhibited the growth of cancer cells both in vitro and in vivo. Thus, our findings also suggest a novel strategy to enhance mTOR-targeted cancer therapy by co-targeting DNA-PK.

Exercise-induced lung cancer regression: mechanistic findings from a mouse model.

BACKGROUND: It has been demonstrated that regular exercise improves the quality of life in patients undergoing treatment for lung cancer and has been associated with reductions in cancer-specific mortality in patients with colon and breast cancer. The direct effects of cardiovascular exercise on lung cancer tumor biology, however, remain unknown. The authors evaluated the effects of cardiovascular exercise in a mouse model of lung adenocarcinoma. METHODS: Luciferase-tagged A549 lung adenocarcinoma cells were injected through the tail vein of nude male mice. Then, the mice underwent weekly bioluminescent imaging until lung tumors were clearly identified. After lung tumors were identified, the mice were randomized to daily wheel running versus no wheel running, and they were imaged weekly. After 4 weeks, all mice were killed, and the lung tumors were harvested. Western blot and immunohistochemical analyses were conducted on tumor tissues to identify potential differences in protein expression levels in exercising mice versus sedentary mice. RESULTS: Lung tumors in exercising mice grew significantly more slowly relative to sedentary mice. There was no change in the development of metastatic lesions between the 2 groups. Protein analysis by Western blot or immunohistochemical analysis demonstrated increased p53 protein levels in exercising mice relative to sedentary mice as well as increased mediators of apoptosis, including Bax and active caspase 3, in tumor tissues. In both groups of mice, no normal tissue toxicity was observed in other organs. CONCLUSIONS: Daily cardiovascular exercise appears to mitigate the growth of lung adenocarcinoma tumors, possibly by activation of the p53 tumor suppressor function and increased apoptosis.