Olink Proteomics-Based Exploration of Immuno-Oncology-Related Biomarkers Leading to Lung Adenocarcinoma Progression.

INTRODUCTION: It is crucial to investigate the distinct proteins that contribute to the advancement of lung cancer. MATERIAL AND METHODS: We analyzed the expression levels of 92 immuno-oncology-related proteins in 96 pairs of lung adenocarcinoma tissue samples using Olink proteomics. The differentially expressed proteins (DEPs) were successively screened in tumor and paraneoplastic groups, early and intermediate-late groups by a nonparametric rank sum test, and the distribution and expression levels of DEPs were determined by volcano and heat maps, etc., and the area under the curve was calculated. RESULTS: A total of 24 DEPs were identified in comparisons between tumor and paracancerous tissues. Among them, interleukin-8 (IL8) and chemokine (C-C motif) ligand 20 (CCL20) as potential markers for distinguishing tumor tissues. Through further screening, it was found that interleukin-6 (IL6) and vascular endothelial growth factor A (VEGFA) may be able to lead to tumor progression through the JaK-STAT signaling pathway, Toll-like receptor signaling pathway and PI3K/AKT signaling pathway. Interestingly, our study revealed a down-regulation of IL6 and VEGFA in tumor tissues compared to paracancerous tissues. CONCLUSIONS: IL8 + CCL20 (AUC: 0.7056) have the potential to differentiate tumor tissue from paracancerous tissue; IL6 + VEGFA (AUC: 0.7531) are important protein markers potentially responsible for tumor progression.

MDM2 provides TOP2 poison resistance by promoting proteolysis of TOP2ßcc in a p53-independent manner.

DNA topoisomerase II (TOP2) is an enzyme that performs a critical function in manipulating DNA topology during replication, transcription, and chromosomal compaction by forming a vital intermediate known as the TOP2-DNA cleavage complex (TOP2cc). Although the TOP2cc is often transient, stabilization can be achieved by TOP2 poisons, a family of anti-cancer chemotherapeutic agents targeting TOP2, such as etoposide (VP-16), and then induce double-strand breaks (DSBs) in cellular DNA. TOP2cc first needs to be proteolyzed before it can be processed by TDP2 for the removal of these protein adducts and to produce clean DNA ends necessary for proper repair. However, the mechanism by which TOP2ßcc is proteolyzed has not been thoroughly studied. In this study, we report that after exposure to VP-16, MDM2, a RING-type E3 ubiquitin ligase, attaches to TOP2ß and initiates polyubiquitination and proteasomal degradation. Mechanistically, during exposure to VP-16, TOP2ß binds to DNA to form TOP2ßcc, which promotes MDM2 binding and subsequent TOP2ß ubiquitination and degradation, and results in a decrease in TOP2ßcc levels. Biologically, MDM2 inactivation abrogates TOP2ß degradation, stabilizes TOP2ßcc, and subsequently increases the number of TOP2ß-concealed DSBs, resulting in the rapid death of cancer cells via the apoptotic process. Furthermore, we demonstrate the combination activity of VP-16 and RG7112, an MDM2 inhibitor, in the xenograft tumor model and in situ lung cancer mouse model. Taken together, the results of our research reveal an underlying mechanism by which MDM2 promotes cancer cell survival in the presence of TOP2 poisons by activating proteolysis of TOP2ßcc in a p53-independent manner, and provides a rationale for the combination of MDM2 inhibitors with TOP2 poisons for cancer therapy.

Construction of RNA Methylation Modification-immune-related lncRNA Molecular Subtypes and Prognostic Scoring System in Lung Adenocarcinoma.

BACKGROUND: RNA methylation modification is not only intimately interrelated with cancer development and progression but also actively influences immune cell infiltration in the tumor microenvironment (TME). RNA methylation modification genes influence the therapeutic progression of lung adenocarcinoma (LUAD), and mining RNA methylation modification prognosis-related markers in LUAD is crucial for its precise prognosis. METHODS: RNA-Seq data and Gene sets were collected from online databases or published literature. Genomic variation analysis was conducted by the Maftools package. RNA methylation-immune-related lncRNAs were obtained by Pearson correlation analysis. Then, Consistent clustering analysis was performed to obtain RNA methylation modification- immune molecular subtypes (RMM-I Molecular subtypes) in LUAD based on selected lncRNAs. COX and random survival forest analysis were carried out to construct the RMM-I Score. The receiver operating characteristic (ROC) curve and Kaplan Meier survival analysis were used to assess survival differences. Tumor immune microenvironment was assessed through related gene signatures and CIBERSORT algorithm. In addition, drug sensitivity analysis was executed by the pRRophetic package. RESULTS: Four RNA methylation modified-immune molecular subtypes (RMM-I1, RMM- I2, RMM-I3, RMM-I4) were presented in LUAD. Patients in RMM-I4 exhibited excellent survival advantages and immune activity. HAVCR2, CD274, and CTLA-4 expression were activated in RMM-I4, which might be heat tumors and a potential beneficial group for immunotherapy. OGFRP1, LINC01116, DLGAP1-AS2, CRNDE, LINC01137, MIR210HG, and CYP1B1-AS1 comprised the RMM-I Score. The RMM-I Score exhibited excellent accuracy in the prognostic assessment of LUAD, as patients with a low RMM- I Score exhibited remarkable survival advantage. Patients with a low RMM-I score might be more sensitive to treatment with Docetaxel, Vinorelbine, Paclitaxel, Cisplatin, and immunotherapy. CONCLUSION: The RMM-I molecular subtype constituted the novel molecular characteristic subtype of LUAD, which complemented the existing pathological typing. More refined and accurate molecular subtypes provide help to reveal the mechanism of LUAD development. In addition, the RMM-I score offers a reliable tool for accurate prognosis of LUAD.

Identification of novel gene signature for lung adenocarcinoma by machine learning to predict immunotherapy and prognosis.

Background: Lung adenocarcinoma (LUAD) as a frequent type of lung cancer has a 5-year overall survival rate of lower than 20% among patients with advanced lung cancer. This study aims to construct a risk model to guide immunotherapy in LUAD patients effectively. Materials and methods: LUAD Bulk RNA-seq data for the construction of a model, single-cell RNA sequencing (scRNA-seq) data (GSE203360) for cell cluster analysis, and microarray data (GSE31210) for validation were collected from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) database. We used the Seurat R package to filter and process scRNA-seq data. Sample clustering was performed in the ConsensusClusterPlus R package. Differentially expressed genes (DEGs) between two groups were mined by the Limma R package. MCP-counter, CIBERSORT, ssGSEA, and ESTIMATE were employed to evaluate immune characteristics. Stepwise multivariate analysis, Univariate Cox analysis, and Lasso regression analysis were conducted to identify key prognostic genes and were used to construct the risk model. Key prognostic gene expressions were explored by RT-qPCR and Western blot assay. Results: A total of 27 immune cell marker genes associated with prognosis were identified for subtyping LUAD samples into clusters C3, C2, and C1. C1 had the longest overall survival and highest immune infiltration among them, followed by C2 and C3. Oncogenic pathways such as VEGF, EFGR, and MAPK were more activated in C3 compared to the other two clusters. Based on the DEGs among clusters, we confirmed seven key prognostic genes including CPA3, S100P, PTTG1, LOXL2, MELTF, PKP2, and TMPRSS11E. Two risk groups defined by the seven-gene risk model presented distinct responses to immunotherapy and chemotherapy, immune infiltration, and prognosis. The mRNA and protein level of CPA3 was decreased, while the remaining six gene levels were increased in clinical tumor tissues. Conclusion: Immune cell markers are effective in clustering LUAD samples into different subtypes, and they play important roles in regulating the immune microenvironment and cancer development. In addition, the seven-gene risk model may serve as a guide for assisting in personalized treatment in LUAD patients.

ATM inhibitor KU60019 synergistically sensitizes lung cancer cells to topoisomerase II poisons by multiple mechanisms.

Type II topoisomerases (TOP2) poisons represent one class of the most successful and widely prescribed chemotherapeutics, which is frontline therapy for a myriad of systemic cancers and solid tumors, including lymphomas, leukemias, and lung cancer. Despite this, treatment with this class of drugs induces unwanted side effects (including cardiovascular morbidity and secondary malignancies). Additionally, the emergence of drug resistance also greatly compromises the clinical use of these drugs. To enhance therapeutic efficiency while lowering unwanted side effects, new insights into effective combination therapy are required. In this study we found that KU60019, a novel, and highly specific ATM kinase inhibitor interferes with the association of ATM with TOP2ß and stabilizes TOP2ß-DNA cleavage complex, thereby impairing the repair of TOP2 poison-induced DSBs and contributes to genome stability, leading to accelerated cell death. In H1299 as well as in A549 lung cancer cell lines, biologically, KU60019 combined with VP-16 (one of the TOP2 poisons) synergistically suppressed the growth of cells and survival and triggered a much higher apoptosis rate. In summary, we provide a proof-of-concept strategy that ATM inhibitors combined with TOP2 poison would synergistically suppresses lung cancer cell survival as well as reduce DNA damage responses, thus may lowering the possibility of cardiotoxicity and secondary malignancy linked to therapy.

Effect of Somatosensory Interaction Transcutaneous Electrical Acupoint Stimulation on Cancer-related Fatigue and Immunity: A Randomized Controlled Trial.

BACKGROUND: This study was intended to evaluate the clinical effect of somatosensory interaction transcutaneous electrical acupoint stimulation (SI-TEAS) on cancer-related fatigue (CRF) and its safety. METHODS: The study protocol had been registered in China Clinical Trial Registration Center with registration number: ChiCTR2100045655. CRF patients were equally divided into SI-TEAS Group, Acupressure Group and Sham Acupressure Group to receive SI-TEAS, acupressure and sham acupressure treatments 5 times a week. The fatigue levels of patients in the 3 groups were measured by the Piper Fatigue Scale during the baseline period and after 4 and 8 weeks (of treatment). The cell immunity of these patients was determined by detecting the T-lymphocyte subsets and NK cells. RESULT: Of the 300 participants, 279 have gone through the independent rehabilitation intervention study, including 94 in the SI-TEAS Group, 92 in the Acupressure Group, and 93 in the Sham Acupressure Group. Intergroup comparisons of fatigue degree and cell immunity, namely SI-TEAS Group versus Acupressure Group, Acupressure Group versus Sham Acupressure Group, and SI-TEAS Group versus Sham Acupressure Group, showed that group changes observed during the baseline period and different time points after Week 4 and 8 were statistically different (P<0.05). The SI-TEAS Group had the sharpest decreases in the behavioral, sensory, emotional and cognitive dimensions of fatigue, and the total score, followed by the Acupressure Group, while the Sham Acupressure Group did not show significant changes; the SI-TEAS Group experienced the sharpest increases in the absolute counts of CD3+ T cells, CD4+ T cells, CD8+ T cells, CD4+/CD8+ T cells, and NK cells, followed by the Acupressure Group, while the Sham Acupressure Group did not show significant changes. CONCLUSION: SI-TEAS could significantly relieve the fatigue of CRF patients and improve their cell immunity, which maybe a useful and effective option for reducing CRF in clinical practice.

Induction of p27 contributes to inhibitory effect of isorhapontigenin (ISO) on malignant transformation of human urothelial cells.

Bladder cancer (BC) is the most expensive cancer to manage on a per-patient basis, costing about $4 billion in total healthcare expenditure per annum in America alone. Therefore, identifying a natural compound for prevention of BC is of tremendous importance for managing this disease. Previous studies have identified isorhapontigenin (ISO) as having an 85% preventive effect against invasive BC formation induced by N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN). The results showed here that ISO treatment inhibited EGF-induced cell transformation of human urothelial cells through induction of tumor suppressor p27 transcription secondary to activation of an E2F1-dependentpathway.ISOtreatmentrenderedcellsresistanttoEGF-induced anchorage-independent growth concurrent with p27 protein induction in both UROtsa and SV-HUC-1 cells. ISO inhibition of EGF-induced cell transformation could be completely reversed by knockdown of p27, indicating that this protein was essential for the noted ISO inhibitory action. Mechanistic studies revealed that ISO treatment resulted in increased expression of E2F1, which in turn bound to its binding site in p27 promoter and initiated p27 transcription. The E2F1 induction was due to the elevation of its translation caused by ISO-induced miR-205 downregulation. Consistently, miR-205 was found to be overexpressed in human BCs, and ectopic expression of miR-205 mitigated ISO inhibitory effects against EGF-induced outcomes. Collectively, the results here demonstrate that ISO exhibits its preventive effect on EGF-induced human urothelial cell transformation by induction of p27 through a miR-205/E2F1 axis. This is distinct from what has been described for the therapeutic effects of ISO on human BC cells.

Territorial Resilience of Metropolitan Regions: A Conceptual Framework, Recognition Methodologies and Planning Response-A Case Study of Wuhan Metropolitan Region.

As the key link and spatial form of urbanization in China, metropolitan region development has become a strategic frontier issue in the field of regional planning and territorial resilience. This paper defines the essence of territorial resilience of metropolitan regions, analyses the capacity of the system and its elements, and builds a regional planning framework. An evaluation indicator system is constructed to evaluate the territorial resilience level and identify the limiting factors in the Wuhan metropolitan region by utilizing the grey correlation model and the obstacle degree model. The results show that the resilience of Wuhan metropolitan region forms an overall pattern of one core area and four sub-regions in the east, west, north and south. According to the different limiting factors of resilience, cities can be divided into three types: cities limited by both policy and spatial resource factors, cities with lagging socioeconomic factors, and cities with insufficient innovation factors. This paper proposes planning response strategies to enhance resilience from two spatial levels. At the regional level this can be done by building a gradually balanced urban system, establishing three areas based on the degree of resilience factor agglomeration, while at the urban level it can be accomplished by maintaining ecological security, promoting economic agglomeration development and constructing innovation networks.

Spatiotemporal characteristics and influencing factors of urban resilience efficiency in the Yangtze River Economic Belt, China.

Urban resilience efficiency is an important indicator to explore the relationship between resource consumption and urban resilience, shedding new light on the study of urban sustainable development. Based on the panel data of 2008, 2012, and 2017, this paper makes a spatiotemporal assessment on the urban resilience efficiency of 126 cities in the Yangtze River Economic Belt (YREB) in China by applying an entropy weight-TOPSIS method and a slack-based measure (SBM) model. Combined with the analysis of a geographically weighted regression model (GWR), the influencing factors on resilience efficiency are also investigated. The results show that both the resource consumption index (RC, inputs) and the urban resilience index (UR, outputs) presented a steady upward trend, and their spatial distribution characteristics were similar, showing a gradual decrease from the eastern coastal cities to the central and western inland cities. Derived from inputs and outputs, the mean values of resilience efficiency index (RE) in three periods were 0.3149, 0.2906, and 0.1625, respectively, revealing that there had been a noticeable decline. Spatially, its spatial distribution has evolved from a relatively balanced pattern to an unbalanced one, showing a gradual decrease from west to east. The results of the GWR model analysis indicate that the total electricity consumption and area of construction land had a considerable correlation with the overall urban resilience of the YREB. Furthermore, total quantity of water supply and science and technology (S&T) expenditure continued to be the main driving factors on urban resilience of the upstream cities. The midstream regions mainly depended on the scale of construction land, and the influencing factors are relatively single. The influencing factors in the downstream areas have changed from dominance of resources and capital factors to the single dominance of resource factors, and total electricity consumption had a strong explanatory power. Based on these findings, we had put forward the overall and local regional policy implications.

Observation on the Curative Effect of Massage Manipulation Combined with Core Strength Training in Patients with Chronic Nonspecific Low Back Pain.

In this prospective study, we used the random number table method to equally divide 141 patients with chronic nonspecific low back pain (CNLBP) who met the inclusion criteria into 3 groups. One group who received massage manipulation therapy was recorded as the manipulation group (n = 47), one other group who received core strength training therapy was recorded as the training group (n = 47), and the other group who received massage manipulation combined with core strength training was recorded as the combination group (n = 47). All the patients were treated for 1w as the course of treatment, with the treatment conducted for 5 d/w for 4 w and 12 w of follow-up. The efficacies of the 3 groups were evaluated after treatment, and the visual analog scale (VAS) scores, Oswestry disability index (ODI) scores, and waist dynamic and static muscle endurance before and after treatment of the 3 groups were compared, and the long-term efficacies of the 3 groups were evaluated combined with the VAS scores at 6 w and 12 w after treatment. Our result was that, after treatment, the total effective rate of the combination group (95.74%) is significantly better than that of the manipulation group (80.85%) and the training group (78.72%) (P < 0.05). After treatment, the VAS and ODI scores of the 3 groups significantly decreased, and the waist dynamic endurance and static muscle endurance significantly increased, and the condition of the combination group was significantly improved compared with that of the manipulation group and the training group (P < 0.05). At 6w and 12w after treatment, the VAS scores of the manipulation group and training group were increased compared with those at the end of treatment (P < 0.05), there was no significant change in the combination group (P < 0.05), and the VAS scores of the combination group at 6 w and 12 w after treatment were lower than those of the manipulation group and training group at the same time point (P < 0.05). This suggests that the synergistic effect of massage manipulation combined with core strength training in the treatment of CNLBP can effectively alleviate patients' pain and waist dysfunction, enhance dynamic and static muscle endurance, and have significant short-term and long-term effects, which are worthy of promotion.

Regulation of topoisomerase II stability and activity by ubiquitination and SUMOylation: clinical implications for cancer chemotherapy.

DNA topoisomerases II (TOP2) are peculiar enzymes (TOP2α and TOP2ß) that modulate the conformation of DNA by momentarily breaking double-stranded DNA to allow another strand to pass through, and then rejoins the DNA phosphodiester backbone. TOP2α and TOP2ß play vital roles in nearly all events involving DNA metabolism, including DNA transcription, replication, repair, and chromatin remodeling. Beyond these vital functions, TOP2 enzymes are therapeutic targets for various anticancer drugs, termed TOP2 poisons, such as teniposide, etoposide, and doxorubicin. These drugs exert their antitumor activity by inhibiting the activity of TOP2-DNA cleavage complexes (TOP2ccs) containing DNA double-strand breaks (DSBs), subsequently leading to the degradation of TOP2 by the 26S proteasome, thereby exposing the DSBs and eliciting a DNA damage response. Failure of the DSBs to be appropriately repaired leads to genomic instability. Due to this mechanism, patients treated with TOP2-based drugs have a high incidence of secondary malignancies and cardiotoxicity. While the cytotoxicity associated with TOP2 poisons appears to be TOP2α-dependent, the DNA sequence rearrangements and formation of DSBs appear to be mediated primarily through TOP2ß inhibition, likely due to the differential degradation patterns of TOP2α and TOP2ß. Research over the past few decades has shown that under various conditions, the ubiquitin-proteasome system (UPS) and the SUMOylation pathway are primarily responsible for regulating the stability and activity of TOP2 and are therefore critical regulators of the therapeutic effect of TOP2-targeting drugs. In this review, we summarize the current progress on the regulation of TOP2α and TOP2ß by ubiquitination and SUMOylation. By fully elucidating the basic biology of these essential and complex molecular mechanisms, better strategies may be developed to improve the therapeutic efficacy of TOP2 poisons and minimize the risks of therapy-related secondary malignancy.

DEPTOR inhibits lung tumorigenesis by inactivating the EGFR-mTOR signals.

DEPTOR plays vital roles in the regulation of cell proliferation and survival by directly modulating the activity of mTORC1/2. However, the physiological role of DEPTOR in lung tumorigenesis, as well as its clinical significance, remains elusive. In this study, we revealed that decreased DEPTOR expression correlated with increased tumor size, poor differentiation, and worse survival in patients with lung cancer. DEPTOR depletion promoted cell proliferation, survival, migration, and invasion in human lung cancer cells. Mechanistically, DEPTOR bound to the kinase domain of EGFR via its PDZ domain to inactivate EGFR signal. Thus, DEPTOR depletion not only directly activated mTORC1/2, but also relieved the inhibition of EGFR to subsequently activate mTOR signals, leading to the induction of cell proliferation and survival. Additionally, activated EGFR-mTOR signals upregulated the expression of ZEB1 and SLUG to induce epithelial-mesenchymal transition, resulting in enhanced migration and invasion. Importantly, Deptor deletion accelerated KrasG12D;p53fl/fl-induced lung tumorigenesis and shortened mouse life span via the activation of EGFR-mTOR signals. Collectively, our study demonstrated that DEPTOR acts as a tumor suppressor in lung tumorigenesis, and its reduction may advance the progression of human lung cancer.

SCFß-TrCP-mediated degradation of TOP2ß promotes cancer cell survival in response to chemotherapeutic drugs targeting topoisomerase II.

Topoisomerase II (TOP2)-targeting anticancer chemotherapeutic drugs, termed TOP2 poisons, are widely used and effective in the clinic by stabilizing TOP2-DNA covalent complexes to induce DNA double-strand breaks (DSBs) and ultimately, cause cell death. The stabilized TOP2-DNA complex is known to be degraded by proteasome, whereas the underlying mechanism for instant TOP2ß degradation in response to TOP2 poisons and the subsequent biological consequence remain elusive. Here, we reported that TOP2 poison-induced TOP2ß degradation is mediated by SCFß-TrCP ubiquitin ligase. Specifically, DNA damage signal, triggered by teniposide (VM-26) treatment, activates ATM, cooperating with CK1 to phosphorylate TOP2ß on Ser1134 and Ser1130, respectively, in a canonical degron motif to facilitate ß-TrCP binding and subsequent degradation. Inactivation of ATM, CK1 or SCFß-TrCP by small molecular inhibitors or genetic knockdown/knockout abrogates TOP2ß degradation. Biologically, blockage of TOP2ß degradation in combination with VM-26 treatment impairs DNA damage response and repair, leading to an accelerated cell death via apoptosis. Thus, it appears that TOP2ß degradation is a cellular defensive mechanism to facilitate the exposure of DSBs to trigger DNA damage response and repair. Collectively, our findings reveal a new strategy to improve the efficacy of TOP2 poisons in combination with small-molecule inhibitors against TOP2ß degradation.

FBXW7 Confers Radiation Survival by Targeting p53 for Degradation.

The tumor suppressor p53 plays a critical role in integrating a wide variety of stress responses. Therefore, p53 levels are precisely regulated by multiple ubiquitin ligases. In this study, we report that FBXW7, a substrate recognition component of the SKP1-CUL1-F-box (SCF) E3 ligase, interacts with and targets p53 for polyubiquitination and proteasomal degradation after exposure to ionizing radiation or etoposide. Mechanistically, DNA damage activates ATM to phosphorylate p53 on Ser33 and Ser37, which facilitates the FBXW7 binding and subsequent p53 degradation by SCFFBXW7. Inactivation of ATM or SCFFBXW7 by small molecular inhibitors or genetic knockdown/knockout approaches extends the p53 protein half-life upon DNA damage in an MDM2-independent manner. Biologically, FBXW7 inactivation sensitizes cancer cells to radiation or etoposide by stabilizing p53 to induce cell-cycle arrest and apoptosis. Taken together, our study elucidates a mechanism by which FBXW7 confers cancer cell survival during radiotherapy or chemotherapy via p53 targeting.

DEPTOR is an in vivo tumor suppressor that inhibits prostate tumorigenesis via the inactivation of mTORC1/2 signals.

The DEPTOR-mTORC1/2 axis has been shown to play an important, but a context dependent role in the regulation of proliferation and the survival of various cancer cells in cell culture settings. The in vivo role of DEPTOR in tumorigenesis remains elusive. Here we showed that the levels of both DEPTOR protein and mRNA were substantially decreased in human prostate cancer tissues, which positively correlated with disease progression. DEPTOR depletion accelerated proliferation and survival, migration, and invasion in human prostate cancer cells. Mechanistically, DEPTOR depletion not only activated both mTORC1 and mTORC2 signals to promote cell proliferation and survival, but also induced an AKT-dependent epithelial-mesenchymal transition (EMT) and ß-catenin nuclear translocation to promote cell migration and invasion. Abrogation of mTOR or AKT activation rescued the biological consequences of DEPTOR depletion. Importantly, in a Deptor-KO mouse model, Deptor knockout accelerated prostate tumorigenesis triggered by Pten loss via the activation of mTOR signaling. Collectively, our study demonstrates that DEPTOR is a tumor suppressor in the prostate, and its depletion promotes tumorigenesis via the activation of mTORC1 and mTORC2 signals. Thus, DEPTOR reactivation via a variety of means would have therapeutic potential for the treatment of prostate cancer.

The cross talk of two family members of ß-TrCP in the regulation of cell autophagy and growth.

ß-transducin repeat-containing protein (ß-TrCP), one of the best-characterized substrate recognition components of the SKP1-CUL1-F-box (SCF) E3 ligase, has two distinct paralogs, ß-TrCP1 and ß-TrCP2, expressed in mammals. Through governing the ubiquitination and degradation of numerous key regulators, ß-TrCP1/2 regulates various cellular physiological and pathological processes. However, whether and how these two proteins cross talk and whether they regulate cell autophagy and proliferation in different manners is completely unknown. Herein, we report that ß-TrCP1 and ß-TrCP2 are the physiological substrates of SCF E3 ligase and target each other for degradation that is dependent on their ß-TrCP degron sequences. Furthermore, glucose deprivation activates AMPK kinase to phosphorylate ß-TrCP1 and promotes the subsequent ubiquitination and degradation of ß-TrCP1 by ß-TrCP2, but does not promote ß-TrCP2 degradation by ß-TrCP1. Finally, we found that ß-TrCP2, not ß-TrCP1, preferentially degrades DEPTOR and REDD1, the inhibitors of mTORC1, to activate mTORC1, leading to autophagy inhibition and cell growth. Thus, our study demonstrates that ß-TrCP1 and ß-TrCP2 mutually target each other for degradation and that ß-TrCP2 acts as a dominant paralog in the regulation of cell autophagy and growth, which might be a promising anticancer target.

AKT inhibitor MK-2206 sensitizes breast cancer cells to MLN4924, a first-in-class NEDD8-activating enzyme (NAE) inhibitor.

Breast cancer is a common type of cancer among female cancer patients and the main cause of cancer-related deaths. During the last decades, targeted therapies for breast cancer have been rapidly developing. Among them, MLN4924, a first-in-class NEDD8-activating enzyme (NAE) inhibitor, has performed antitumor activity by inactivating the cullin-RING ligases and causing the accumulation of their substrates to induce apoptosis in a number of studies. In this study, we found that MLN4924 activates the AKT pathway in both HER2-positive and triple-negative breast cancer (TNBC) cell lines. Given that AKT signaling is responsible for tumor progression and drug resistance in some types of cancers, we hypothesized that the AKT inhibitor may synergistically enhance the tumor suppression capability in breast cancer by MLN4924. To demonstrate the sensitizing effect, MK-2206 was chosen as the adjuvant treatment, and cell growth, migration and apoptosis were detected. The results showed that MLN4924 treatment inhibited cell growth and migration and induced apoptosis in both SK-BR3 and MDA-MB231 breast cancer cell lines. More importantly, the combined treatment of MLN4924 and MK-2206 indeed caused stronger cytotoxicity and inhibition of migration and a much higher induction of apoptosis compared with MLN4924 treatment alone. Our study provides the proof-of-concept evidence for strategic drug combination of MLN4924 with an AKT inhibitor for maximal killing of breast cancer cells via the enhancement of apoptosis.