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.

SAMHD1 deacetylation by SIRT1 promotes DNA end resection by facilitating DNA binding at double-strand breaks.

Sterile alpha motif and HD domain-containing protein 1 (SAMHD1) has a dNTPase-independent function in promoting DNA end resection to facilitate DNA double-strand break (DSB) repair by homologous recombination (HR); however, it is not known if upstream signaling events govern this activity. Here, we show that SAMHD1 is deacetylated by the SIRT1 sirtuin deacetylase, facilitating its binding with ssDNA at DSBs, to promote DNA end resection and HR. SIRT1 complexes with and deacetylates SAMHD1 at conserved lysine 354 (K354) specifically in response to DSBs. K354 deacetylation by SIRT1 promotes DNA end resection and HR but not SAMHD1 tetramerization or dNTPase activity. Mechanistically, K354 deacetylation by SIRT1 promotes SAMHD1 recruitment to DSBs and binding to ssDNA at DSBs, which in turn facilitates CtIP ssDNA binding, leading to promotion of genome integrity. These findings define a mechanism governing the dNTPase-independent resection function of SAMHD1 by SIRT1 deacetylation in promoting HR and genome stability.

Overcoming acquired resistance to third-generation EGFR inhibitors by targeting activation of intrinsic apoptotic pathway through Mcl-1 inhibition, Bax activation, or both.

Treatment of EGFR-mutant non-small cell lung cancer (NSCLC) with mutation-selective third-generation EGFR-tyrosine kinase inhibitors (EGFR-TKIs) such as osimertinib has achieved remarkable success in the clinic. However, the immediate challenge is the emergence of acquired resistance, limiting the long-term remission of patients. This study suggests a novel strategy to overcome acquired resistance to osimertinib and other third-generation EGFR-TKIs through directly targeting the intrinsic apoptotic pathway. We found that osimertinib, when combined with Mcl-1 inhibition or Bax activation, synergistically decreased the survival of different osimertinib-resistant cell lines, enhanced the induction of intrinsic apoptosis, and inhibited the growth of osimertinib-resistant tumor in vivo. Interestingly, the triple-combination of osimertinib with Mcl-1 inhibition and Bax activation exhibited the most potent activity in decreasing the survival and inducing apoptosis of osimertinib-resistant cells and in suppressing the growth of osimertinib-resistant tumors. These effects were associated with increased activation of the intrinsic apoptotic pathway evidenced by augmented mitochondrial cytochrome C and Smac release. Hence, this study convincingly demonstrates a novel strategy for overcoming acquired resistance to osimertinib and other 3rd generation EGFR-TKIs by targeting activation of the intrinsic apoptotic pathway through Mcl-1 inhibition, Bax activation or both, warranting further clinical validation of this strategy.

Discovery of Small Molecule Bak Activator for Lung Cancer Therapy.

Rationale: Bak is a major proapoptotic Bcl2 family member and a required molecule for apoptotic cell death. High levels of endogenous Bak were observed in both small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC) cell lines. Increased Bak expression was correlated with poor prognosis of NSCLC patients, suggesting that Bak protein is an attractive target for lung cancer therapy. The BH3 domain functions as death domain and is required for Bak to initiate apoptotic cell death. Thus, the BH3 domain is attractive target for discovery of Bak agonist. Methods: The BH3 death domain binding pocket (aa75-88) of Bak was chosen as a docking site for screening of small molecule Bak activators using the UCSF DOCK 6.1 program suite and the NCI chemical library (300,000 small molecules) database. The top 500 compounds determined to have the highest affinity for the BH3 domain were obtained from the NCI and tested for cytotoxicity for further screening. We identified a small molecule Bak activator BKA-073 as the lead compound. The binding affinity of BKA-073 with Bak protein was analyzed by isothermal titration calorimetry (ITC) assay. BKA-073-mediated Bak activation via oligomerization was analyzed by a cross-linking with Bis (maleimido) hexane (BMH). Sensitivity of BKA-073 to lung cancer cells in vitro was evaluated by dynamic BH3 profiling (DBP) and apoptotic cell death assay. The potency of BKA-073 alone or in combination with radiotherapy or Bcl2 inhibitor was evaluated in animal models. Results: We found that BKA-073 binds Bak at BH3 domain with high affinity and selectivity. BKA-073/Bak binding promotes Bak oligomerization and mitochondrial priming that activates its proapoptotic function. BKA-073 potently suppresses tumor growth without significant normal tissue toxicity in small cell lung cancer (SCLC) and NSCLC xenografts, patient-derived xenografts, and genetically engineered mouse models of mutant KRAS-driven cancer. Bak accumulates in radioresistant lung cancer cells and BKA-073 reverses radioresistance. Combination of BKA-073 with Bcl-2 inhibitor venetoclax exhibits strong synergy against lung cancer in vivo. Conclusions: Development of small molecule Bak activator may provide a new class of anticancer agents to treat lung cancer.

Epigenetic modulation of FBW7/Mcl-1 pathway for lung cancer therapy.

Methylation induces epigenetic silencing of tumor suppressor genes in human lung cancer. Inhibition of DNA methyltransferases by decitabine (DAC) can demethylate and activate epigenetically silenced tumor suppressor genes. Epigenetic therapy using DAC should be an attractive strategy for lung cancer therapy. FBW7 is a tumor suppressor that functions as an Mcl-1 E3 ligase to degrade Mcl-1 by ubiquitination. Here we discovered that treatment of various human lung cancer cells with DAC resulted in activation of FBW7 expression, decreased levels of Mcl-1 protein, and growth inhibition. DAC-activated FBW7 expression promoted Mcl-1 ubiquitination and degradation leading to a significant reduction in the half-life of Mcl-1 protein. Mechanistically, treatment of lung cancer cells or lung cancer xenografts with DAC induced the conversion of the FBW7 gene from a methylated form to an unmethylated form, which was associated with the increased expression of FBW7 and decreased expression of Mcl-1 in vitro and in vivo. DAC suppressed lung cancer growth in a dose-dependent manner in vivo. Combined treatment with DAC and a Bcl2 inhibitor, venetoclax, exhibited strong synergistic potency against lung cancer without normal tissue toxicity. These findings uncover a novel mechanism by which DAC suppresses tumor growth by targeting the FBW7/Mcl-1 signaling pathway. Combination of DAC with Bcl2 inhibitor venetoclax provides more effective epigenetic therapy for lung cancer.

Metabolomic Comparison of Patients With Colorectal Cancer at Different Anticancer Treatment Stages.

BACKGROUND: The difficulties of early diagnosis of colorectal cancer (CRC) result in a high mortality rate. The ability to predict the response of a patient to surgical resection or chemotherapy may be of great value for clinicians when planning CRC treatments. Metabolomics is an emerging tool for biomarker discovery in cancer research. Previous reports have indicated that the metabolic profile of individuals can be significantly altered between CRC patients and healthy controls. However, metabolic changes in CRC patients at different treatment stages have not been explored. METHODS: To this end, we performed nuclear magnetic resonance (NMR)-based metabolomic analysis to determine metabolite aberrations in CRC patients before and after surgical resection or chemotherapy. In general, a total of 106 urine samples from four clinical groups, namely, healthy volunteers (n = 31), presurgery CRC patients (n = 25), postsurgery CRC patients (n = 25), and postchemotherapy CRC patients (n = 25), were collected and subjected to further analysis. RESULTS: In the present study, we identified five candidate metabolites, namely, N-phenylacetylglycine, succinate, 4-hydroxyphenylacetate, acetate, and arabinose, in CRC patients compared with healthy individuals, three of which were reported for the first time. Furthermore, approximately ten metabolites were uniquely identified at each stage of CRC treatment, serving as good candidates for biomarker panel selection. CONCLUSION: In summary, these potential metabolite candidates may provide promising early diagnostic and monitoring approaches for CRC patients at different anticancer treatment stages.

TGF-ß causes Docetaxel resistance in Prostate Cancer via the induction of Bcl-2 by acetylated KLF5 and Protein Stabilization.

Prostate cancer is the second leading cause of cancer-related death in the United States. As a first line treatment for hormone-refractory prostate cancer, docetaxel (DTX) treatment leads to suboptimal effect since almost all patients eventually develop DTX resistance. In this study, we investigated whether and how TGF-ß affects DTX resistance of prostate cancer. Methods: Cytotoxicity of DTX in DU 145 and PC-3 cells was measured by CCK-8 and Matrigel colony formation assays. Resistance to DTX in DU 145 cells was examined in a xenograft tumorigenesis model. A luciferase reporter system was used to determine transcriptional activities. Gene expression was analyzed by RT-qPCR and Western blotting. Results: We found that KLF5 is indispensable in TGF-ß-induced DTX resistance. Moreover, KLF5 acetylation at lysine 369 mediates DTX resistance in vitro and in vivo. We showed that the TGF-ß/acetylated KLF5 signaling axis activates Bcl-2 expression transcriptionally. Furthermore, DTX-induced Bcl-2 degradation depends on a proteasome pathway, and TGF-ß inhibits DTX-induced Bcl-2 ubiquitination. Conclusion: Our study demonstrated that the TGF-ß-acetylated KLF5-Bcl-2 signaling axis mediates DTX resistance in prostate cancer and blockade of this pathway could provide clinical insights into chemoresistance of prostate cancer.

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.

EZH2 has a non-catalytic and PRC2-independent role in stabilizing DDB2 to promote nucleotide excision repair.

Small cell lung cancer (SCLC) is a highly aggressive malignancy with poor outcomes associated with resistance to cisplatin-based chemotherapy. Enhancer of zeste homolog 2 (EZH2) is the catalytic subunit of polycomb repressive complex 2 (PRC2), which silences transcription through trimethylation of histone H3 lysine 27 (H3K27me3) and has emerged as an important therapeutic target with inhibitors targeting its methyltransferase activity under clinical investigation. Here, we show that EZH2 has a non-catalytic and PRC2-independent role in stabilizing DDB2 to promote nucleotide excision repair (NER) and govern cisplatin resistance in SCLC. Using a synthetic lethality screen, we identified important regulators of cisplatin resistance in SCLC cells, including EZH2. EZH2 depletion causes cellular cisplatin and UV hypersensitivity in an epistatic manner with DDB1-DDB2. EZH2 complexes with DDB1-DDB2 and promotes DDB2 stability by impairing its ubiquitination independent of methyltransferase activity or PRC2, thereby facilitating DDB2 localization to cyclobutane pyrimidine dimer crosslinks to govern their repair. Furthermore, targeting EZH2 for depletion with DZNep strongly sensitizes SCLC cells and tumors to cisplatin. Our findings reveal a non-catalytic and PRC2-independent function for EZH2 in promoting NER through DDB2 stabilization, suggesting a rationale for targeting EZH2 beyond its catalytic activity for overcoming cisplatin resistance in SCLC.

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.

Mcl-1 Interacts with Akt to Promote Lung Cancer Progression.

Mcl-1 is a unique antiapoptotic Bcl2 family protein that functions as a gatekeeper in manipulating apoptosis and survival in cancer cells. Akt is an oncogenic kinase that regulates multiple cellular functions and its activity is significantly elevated in human cancers. Here we discovered a cross-talk between Mcl-1 and Akt in promoting lung cancer cell growth. Depletion of endogenous Mcl-1 from human lung cancer cells using CRISPR/Cas9 or Mcl-1 shRNA significantly decreased Akt activity, leading to suppression of lung cancer cell growth in vitro and in xenografts. Mechanistically, Mcl-1 directly interacted via its PEST domain with Akt at the pleckstrin homology (PH) domain. It is known that the interactions between the PH domain and kinase domain (KD) are important for maintaining Akt in an inactive state. The binding of Mcl-1/PH domain disrupted intramolecular PH/KD interactions to activate Akt. Intriguingly, Mcl-1 expression correlated with Akt activity in tumor tissues from patients with non-small cell lung cancer. Using the Mcl-1-binding PH domain of Akt as a docking site, we identified a novel small molecule, PH-687, that directly targets the PH domain and disrupts Mcl-1/Akt binding, leading to suppression of Akt activity and growth inhibition of lung cancer in vitro and in vivo. By targeting the Mcl-1/Akt interaction, this mechanism-driven agent provides a highly attractive strategy for the treatment of lung cancer. SIGNIFICANCE: These findings indicate that targeting Mcl-1/Akt interaction by employing small molecules such as PH-687 represents a potentially new and effective strategy for cancer treatment.

Acetylation regulates ribonucleotide reductase activity and cancer cell growth.

Ribonucleotide reductase (RNR) catalyzes the de novo synthesis of deoxyribonucleoside diphosphates (dNDPs) to provide dNTP precursors for DNA synthesis. Here, we report that acetylation and deacetylation of the RRM2 subunit of RNR acts as a molecular switch that impacts RNR activity, dNTP synthesis, and DNA replication fork progression. Acetylation of RRM2 at K95 abrogates RNR activity by disrupting its homodimer assembly. RRM2 is directly acetylated by KAT7, and deacetylated by Sirt2, respectively. Sirt2, which level peak in S phase, sustains RNR activity at or above a threshold level required for dNTPs synthesis. We also find that radiation or camptothecin-induced DNA damage promotes RRM2 deacetylation by enhancing Sirt2-RRM2 interaction. Acetylation of RRM2 at K95 results in the reduction of the dNTP pool, DNA replication fork stalling, and the suppression of tumor cell growth in vitro and in vivo. This study therefore identifies acetylation as a regulatory mechanism governing RNR activity.

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.

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.