TPGS-b-PBAE Copolymer-Based Polyplex Nanoparticles for Gene Delivery and Transfection In Vivo and In Vitro.

Poly (ß-amino ester) (PBAE) is an exceptional non-viral vector that is widely used in gene delivery, owing to its exceptional biocompatibility, easy synthesis, and cost-effectiveness. However, it carries a high surface positive charge that may cause cytotoxicity. Therefore, hydrophilic d-α-tocopherol polyethylene glycol succinate (TPGS) was copolymerised with PBAE to increase the biocompatibility and to decrease the potential cytotoxicity of the cationic polymer-DNA plasmid polyplex nanoparticles (NPs) formed through electrostatic forces between the polymer and DNA. TPGS-b-PBAE (TBP) copolymers with varying feeding molar ratios were synthesised to obtain products of different molecular weights. Their gene transfection efficiency was subsequently evaluated in HEK 293T cells using green fluorescent protein plasmid (GFP) as the model because free GFP is unable to easily pass through the cell membrane and then express as a protein. The particle size, ζ-potential, and morphology of the TBP2-GFP polyplex NPs were characterised, and plasmid incorporation was confirmed through gel retardation assays. The TBP2-GFP polyplex NPs effectively transfected multiple cells with low cytotoxicity, including HEK 293T, HeLa, Me180, SiHa, SCC-7 and C666-1 cells. We constructed a MUC2 (Mucin2)-targeting CRISPR/cas9 gene editing system in HEK 293T cells, with gene disruption supported by oligodeoxynucleotide (ODN) insertion in vitro. Additionally, we developed an LMP1 (latent membrane protein 1)-targeting CRISPR/cas9 gene editing system in LMP1-overexpressing SCC7 cells, which was designed to cleave fragments expressing the LMP1 protein (related to Epstein-Barr virus infection) and thus to inhibit the growth of the cells in vivo. As evidenced by in vitro and in vivo experiments, this system has great potential for gene therapy applications.

Survival benefit of thoracic radiotherapy plus EGFR-TKIs in patients with non-oligometastatic advanced non-small-cell lung cancer: a single-center retrospective study.

Objectives: The study aims to evaluate the efficacy and safety of thoracic radiotherapy in epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI)-treated patients with stage IV non-small-cell lung cancer (NSCLC). Methods: Patients with non-oligometastatic NSCLC harboring EGFR mutations were recruited. All patients received the first-generation TKI treatment with or without radiotherapy. The irradiated sites included primary and/or metastatic lesions. Of all the patients who underwent thoracic radiotherapy, some received radiotherapy before EGFR-TKI resistance, others received radiotherapy after progressive disease. Results: No statistically significant difference was observed in progression-free survival (PFS) (median 14.7 versus 11.2 months, p = 0.075) or overall survival (OS) (median 29.6 versus 40.6 months, p = 0.116) between patients treated with EGFR-TKIs alone and those with additional radiotherapy to any sites. However, EGFR inhibitors with thoracic radiation significantly improved OS (median 47.0 versus 31.0 months, p < 0.001) but not PFS (median 13.9 versus 11.9 months, p = 0.124). Moreover, longer PFS (median 18.3 versus 8.5 months, p < 0.001) was achieved in the preemptive thoracic radiation cohort than in the delayed thoracic radiation cohort. However, OS was similar between the two cohorts (median 40.6 versus 52.6 months, p = 0.124). The lower incidence rate of grade 1-2 pneumonitis occurred in preemptive radiation cohort (29.8% versus 75.8%, p < 0.001). Conclusion: Non-oligometastatic NSCLC patients with EGFR mutations benefited from thoracic radiotherapy while using EGFR inhibitors. Preemptive thoracic radiotherapy could be a competitive first-line therapeutic option due to superior PFS and favorable safety.

Airway Fusobacterium is Associated with Poor Response to Immunotherapy in Lung Cancer.

PURPOSE: There is a major limitation in the immunotherapy for solid cancer is that it only benefited a minority of cancer patients. This study aims to investigate whether the differential composition of the lung microbiome could affect the sustained clinical responses in lung cancers treated with immunotherapy. METHODS: Twenty-seven non-responders and 19 responders treated with anti-PD-1 therapy were included in the discovery set. Bacterial load in bronchoalveolar lavage from lung cancer patients was examined by quantitative PCR of 16S rRNA copies. Bacterial 16S rDNA was sequenced using the Illumina HiSeq on the 16S rDNA V3-V4 variable region. Operational taxonomic unit (OTU) analysis was performed using VSEARCH v2. The α-diversity and ß-diversity were calculated using QIIME software. RESULTS: The mean copy number of bacterial 16S DNA levels significantly decreased after anti-PD-1 treatment (after: 1.8 ± 0.6×104 copies per milliliter vs prior to treatment: 3.3 ± 1.1x104, p = 0.0036). In addition, longitudinal analysis revealed that microbial diversity was reduced taxonomically after treatment compared to those prior to the treatment (Shannon values: before: 3.291 ± 0.067 vs after: 2.668 ± 0.168, p < 0.01). Further, we observed a reduction of Fusobacterium nucleatum, including phylum Fusobacteria (p < 0.01), class Fusobacteria (p < 0.01), order Fusobacteria (p < 0.01), family Fusobacteria (p < 0.01), genus Fusobacteria (p = 0.025) in the responders post anti-PD-1 treatment. However, there was no significant difference of Fusobacterium in non-responders. An independent cohort was used to validate the levels of Fusobacterium, demonstrating that patients with higher abundance of Fusobacterium prior to treatment were significantly more likely to have poor response to anti-PD-1 therapy (p < 0.001). CONCLUSION: Airway enriched Fusobacterium prior to anti-PD-1 therapy is associated with poor response in lung cancer, which indicated that potential resistance to immunotherapy can be attributed to lung microbiome.

A self-assembled nanoplatform based on Ag2S quantum dots and tellurium nanorods for combined chemo-photothermal therapy guided by H2O2-activated near-infrared-II fluorescence imaging.

A nanoplatform based on Ag2S quantum dots (QDs) and tellurium nanorods (TeNRs) was developed for combined chemo-photothermal therapy guided by H2O2-activated near-infrared (NIR)-II fluorescence imaging. Polypeptide PC10AGRD-modified TeNRs and Ag2S QDs were co-encapsulated in 4T1 cell membrane to prepare a nanoplatform (CCM@AT). Ag2S QDs and TeNRs in the CCM@AT were used as a fluorescence probe and photosensitizer, and a chemotherapeutic prodrug and quenching agent to quench the fluorescence of Ag2S QDs, respectively. After the CCM@AT was specifically targeted to the tumor site, the TeNRs were dissolved by the high concentration of H2O2 at the tumor site to light up the fluorescence of Ag2S QDs for NIR-II fluorescence imaging. In addition, the generated toxic TeO66- molecules decreased ATP production by selective cancer chemotherapy, which is beneficial for photothermal therapy. The elevated temperature due to photothermal therapy in turn promoted the chemical reaction in chemotherapy. In vitro and in vivo toxicity results showed that the CCM@AT possesses high biocompatibility. Compared to single photothermal therapy and chemotherapy, the synergistic chemo-photothermal therapy can effectively suppress the growth of 4T1 tumor. This all-in-one nanoplatform provides a boulevard for the combination therapy of tumors guided by NIR-II fluorescence imaging. STATEMENT OF SIGNIFICANCE: NIR-II fluorescence imaging shows the characteristics of low tissue absorption, reflection, and scattering, which can greatly reduce the influence of autofluorescence in vivo. However, the non-negligible effect of autofluorescence is still observed in fluorescence imaging in vivo. Therefore, there is an urgent need to develop a strategy of controlled release of fluorescence for accurate imaging and tumor therapy. Here, Ag2S quantum dots (QDs) with NIR-II fluorescence emission and good photothermal conversion efficiency are used as a fluorescence probe and photosensitizer, and tellurium nanorods (TeNRs) are used as a chemotherapeutic prodrug and quenching agent to quench the fluorescence of Ag2S QDs. This multiple nanoplatform provides an inspiration for the combination therapy of tumor guided by NIR-II fluorescence imaging.

Long-term Therapeutic Outcome and Prognostic Factors of Patients with Nasopharyngeal Carcinoma Receiving Intensity-modulated Radiotherapy: An Analysis of 608 Patients from Low-endemic Regions of China.

OBJECTIVE: To evaluate the long-term outcome and prognostic factors of patients with nasopharyngeal carcinoma (NPC) from low-endemic regions of China who received definitive intensity-modulated radiation therapy (IMRT). METHODS: The clinical data from 608 patients with newly-diagnosed non-metastatic NPC who have received initial treatment at our cancer center from January, 2008 to December, 2013 were retrospectively reviewed. All patients received definitive IMRT, and 87.7% received platinum-based chemotherapy. RESULTS: The median follow-up duration was 51 months (follow-up rate, 98.5%; range, 10-106 months) for the entire cohort. The 5-year overall survival rate was 79.7%. The 5-year local relapse-free survival rate, regional relapse-free survival rate, distant metastasis-free survival rate and progression-free survival rate were 92.4%, 93.3%, 79.2% and 74.3%, respectively. A total of 153 patients had experienced treatment failure, with distant metastasis as the primary cause in 77.1% (118/153). Patients with T4 or N3 diseases had a significantly poorer prognosis than other subcategories. Stage T4 and N3 were closely associated with distant metastasis, with the metastatic rate of 29.3% and 45.5%, respectively. CONCLUSION: IMRT provides patients with non-metastatic NPC with satisfactory long-term survival. Both T stage and N stage are important prognostic factors for NPC patients. Patients with T4 or N3 diseases have significantly increased distant metastatic rates and poor survival time.

[Corrigendum] A serum microRNA signature as a prognostic factor for patients with advanced NSCLC and its association with tissue microRNA expression profiles.

Subsequently to the publication of this paper, an interested reader drew to the authors' attention that, in Fig. 5B on p. 4651, the images selected to represent the miR­486/mimic and miR­221/inhibitor conditions for the migration experiments with the H358 cell line bore some striking similarities. After having examined their original data, the authors realized that they uploaded an incorrect image to represent the miR­486/mimic data panel during the process of compiling this figure. The corrected version of Fig. 5, showing all the correct data for Fig. 5B, is shown on the next page. Note that the replacement of the erroneous data does not affect either the results or the conclusions reported in this paper, and all the authors agree to this Corrigendum. The authors are grateful to the Editor of Molecular Medicine Reports for granting them this opportunity to publish a Corrigendum, and apologize to the readership for any inconvenience caused [the original article was published in Molecular Medicine Reports 13: 4643­4653, 2016; DOI: 10.3892/mmr.2016.5114].

Evidence that dysplasia related microRNAs in Barrett's esophagus target PD-L1 expression and contribute to the development of esophageal adenocarcinoma.

Esophageal adenocarcinoma (EAC) is the cancer arising from the esophagus, which frequently develop from Barrett's esophagus (BE). Extracellular vesicles (EVs), particularly exosomes, are nanosized vesicles of endosomal origin released from various types of cells that have been implicated in cancers. However, the significance of circulating exosomes during the progression of BE to EAC remains unknown. Sera exosmal microRNAs were profiled from 13 EAC and 12BE patients compared to 12 healthy controls. We found a substantial dysregulation of exosomal miRNA levels in BE compared to healthy control, and identified a unique signature of 24 up regulated and 14 down regulated miRNAs. Further validation showed exosomal miR-196a, -26b, -21, and -143 expression was significantly higher in BE and continued to have higher levels in EAC compared to healthy controls; while sera exosomal miR-378, -210, -205, and -200c-3p were significantly lower expressed in BE patients compared to compared to controls. Further, miR-378, -210, -205, and -200c-3p continue to have even lower levels in EAC patients compared to BE. Interestingly, sera expression levels of exosomal miR-15a, -16, and -193a-3p were significantly down regulated in BE PD-L1(+) patients; Sera exosomal miR-15a, -15b, -16, and -193a-3p expression levels in EAC PD-L1(+) patients were significantly lower (all p < 0.01) when compared to EAC PD-L1(-) patients. More importantly, the BE-EAC group had longitudinally decreased exosomal expression levels of miR-15a, -15b, -16, and -193a-3p from BE status to their EAC progression. In conclusion, distinct microRNA expression patterns were demonstrated in circulating exosomes from Barrett's esophagus and esophageal adenocarcinoma; Furthermore exosomal microRNAs potentially targeting PD-L1 mRNA were down regulated in PD-L1 (+) BE and EAC patients.

Injectable polypeptide-engineered hydrogel depot for amplifying the anti-tumor immune effect induced by chemo-photothermal therapy.

The immunosuppressive tumor microenvironment has caused great obstacles to tumor immunotherapy, especially where less tumor-associated antigens are released from tumor sites. Herein, a Ag2S QD/DOX/Bestatin@PC10ARGD genetically engineered polypeptide hydrogel PC10ARGD as a sustained-release material was developed for mammary carcinoma treatment. A near-infrared silver sulfide (Ag2S) QD as a photosensitizer was encapsulated into the hydrophobic cavity formed by the self-assembly of the polypeptide nanogel (PC10ARGD) for photothermal therapy. The water-soluble drug DOX and Bestatin were integrated into the PC10ARGD hydrogel. The photothermal effect could trigger the sustained release of the DOX, which could be applied to initiate in situ vaccination. Bestatin as an immune-adjuvant drug could amplify the body's immune function. The results of in vivo therapy tests exhibited that the Ag2S QD/DOX/Bestatin@PC10ARGD hydrogel with laser irradiation could activate anti-tumor immune effects that inhibit the growth of primary tumors and distal lung metastatic nodules. Meanwhile, a safer lower-temperature with multiple laser irradiation treatment strategy exhibited more effective tumor-killing performance (84.4% tumor inhibition rate) and promoted the penetration of immune cells into the tumor tissue. The CD8+ and CD4+ cytotoxic T cells ratio was increased by 5.3 and 10 times, respectively, thus exhibiting a good prognostic signal. The multifunctional polypeptide hydrogel as a green manufacturing and engineering material is promising to serve as a cancer vaccine for anticancer applications.

Circulating microRNAs predict the response to anti-PD-1 therapy in non-small cell lung cancer.

Finding reliable markers for predicting the efficacy of immunotherapy is urgently needed. We sought to investigate the association between serum microRNAs (miRNAs) and checkpoint inhibitor response in non-small cell lung cancer (NSCLC). Discovery assay with sera miRNA profiling was performed, demonstrating 27 sera miRNAs (relative fold >2, p < .05), 22 higher expressed and 5 lower expressed miRNAs, were differentially expressed in 19 responders compared to those in 27 non-responders. Further validation validated miR-93, -138-5p, -200, -27a, -424, -34a, -28, -106b, -193a-3p, and -181a were significantly higher expressed (p < .01) in an independent cohort of 17 responders vs. 17 non-responders. Longitudinally, responders had increased sera expression levels of miR-93, -138-5p, -200, -27a, -424, -34a, -28, -106b, -193a-3p, and -181a from pre-treatment to post-treatment (p < .01). More importantly, statistically significant improvement in PFS of patients was associated with the 10-high expressed miRNA pattern (median PFS of 6.25 versus 3.21 months, p < .001; hazard ratio, HR, 0.45; 95% CI, 0.25-0.76). Further OS improvement was also significantly associated with the 10-high expressed miRNA pattern in responders versus non-responders (median OS of 7.65 versus 3.2 months, p < .001, HR, 0.39; 95% CI, 0.15-0.68). In conclusion, these results demonstrated that alterations in circulating miRNAs are associated with the response and outcome in NSCLC patients treated with anti-PD1 drugs.

Hollow gold nanoshells-incorporated injectable genetically engineered hydrogel for sustained chemo-photothermal therapy of tumor.

BACKGROUND: Combined therapy has demonstrated to be an effective strategy for cancer therapy. Herein, an injectable hydrogel based on the genetically engineered polypeptide and hollow gold nanoshells (HAuNS) has been developed for chemo-photothermal therapy of HepG2 tumor. METHODS: PC10A/DOX/HAuNS nanogel was prepared with layer-by-layer through the adsorption of DOX and PC10A successively. DOX with positive charge and PC10A with negative charge were coated step by step onto the surface of negatively charged HAuNS. The multifunctional hydrogel PC10A/DOX/HAuNS were prepared via dissolving hybrid PC10A/DOX/HAuNS nanogel in polypeptide PC10A. Chemotherapy drug DOX in the PC10A/DOX/HAuNS hydrogel was absorbed on the HAuNS and directly embedded in the PC10A hydrogel, which contributes to sequentially release of the drug. Specifically, DOX adsorbed on the HAuNS could be released slowly for sustainable chemotherapy. RESULTS: The PC10A/DOX/HAuNS hydrogel could pass 26-gauge needle without clogging, indicating that it is injectable. In addition, the PC10A/DOX/HAuNS hydrogel possessed outstanding photothermal effect and photothermal stability. In both in vitro cell and in vivo tumor-bearing mice experiments, a remarkably enhance tumor inhibition was observed by the combined therapy of chemo-photothermal therapy compared with photothermal therapy or chemotherapy alone. CONCLUSIONS: The combined chemotherapy and photothermal therapy of PC10A/DOX/HAuNS hydrogels could significantly improve the therapeutic effect. Therefore, the multifunctional hydrogel PC10A/DOX/HAuNS is promising to provide a new strategy for sustained chemo-photothermal therapy.

Smoking Induced Extracellular Vesicles Release and Their Distinct Properties in Non-Small Cell Lung Cancer.

Purpose: Smoking is a strong relative risk factor for lung cancer. Extracellular vesicles (EVs), particularly exosomes, have been implicated in cancers. In this study, we characterized smoking induced extracellular vesicles in smokers with non-small cell lung cancer (NSCLC). Methods: EVs were isolated from bronchoalveolar lavage (BAL) from smokers and NSCLC patients. EV microRNAs (miRNAs) were analyzed by using a TaqMan microRNA assays. Vesicle mRNAs and long non-coding RNAs (lncRNAs) were measured with quantitative RT-PCR. Tumor associated antigens were examined by Western Blot. Results: Higher levels of local site EVs are found in the lung of smokers and NSCLC patients. Further, over 90% of lung EVs are round vesicles of approximately 50-200 nm, ie., exosomes. There are 21 EV miRNAs up regulated, while 10 miRNAs under regulated, in smokers when compared to controls (relative fold > 2, p < 0.05). These miRNAs were further observed to be dysregulated in NSCLC patients when compared to smokers. Bioinformatic analysis demonstrated that Proteoglycans, Fatty acid biosynthesis, ErbB, Hippo, TGF-beta, Wnt, Rap1, AMPK and Ras pathways were the most prominent pathways enriched in NSCLC EV miRNA signatures. In addition, messenger RNA transcripts including EGFR, KRAS, ALK, MET, LKB1, BRAF, PIK3CA, RET, and ROS1 were significantly higher expressed in lung EVs in smokers and NSCLC patients compared to controls. Long non-coding RNAs, including MALAT1, HOTAIR, HOTTIP, AGAP2-AS1, ATB, TCF7, FOXD2-AS1, HOXA11-AS, PCAF1, and BCAR4, were over expressed in EVs from smokers and NSCLC patients. Furthermore, protein levels of tumor associated antigens including BAGE, PD-L1, MAGE-3, and AKAP4 were significantly dysregulated in EVs of smokers and NSCLC patients compared to healthy controls. Conclusions: In conclusion, these data demonstrated an intrinsic relationship of smoking dysregulated EVs and EVs contained RNA, proteins which may involve in the development of NSCLC.

Reactive Oxygen Species-Mediated Cezanne Inactivation by Oxidation of its Catalytic Cysteine Residue in Hepatocellular Carcinoma.

Cysteine oxidation occurs at the active site of deubiquitinases (DUBs) during many biologic signaling cascades. Here we report that hepatocellular carcinoma cells (HCCs) generated higher levels of endogenous reactive oxygen species (ROS). This elevated ROS production was inhibited by NADPH oxidase inhibitor diphenylene iodonium (DPI) and mitochondria electron chain inhibitor rotenone in HCC cells. Moreover, we found that H2O2 could activate NF-κB-dependent inflammatory effect through increased induction of matrix metalloproteinase 2 (MMP2), MMP9, and intercellular adhesion molecule 1 (ICAM1) expression levels. In addition, we found that H2O2 could prolong NF-κB activation by suppressing the negative regulatory functions of Cezanne in HCC cells. Ubiquitin-derived thiol-reactive probe (HA-UbVME) assay and biotin-tagged 1,3-cyclohexadione derivative (DCP-Bio1) assay showed that H2O2 has the capacity to inhibit the catalytic activity of Cezanne, and the reducing agent, DTT, could reactivate the Cezanne deubiquitinating enzyme activity. Taken all together, these findings demonstrated an important role for oxidation of Cezanne by ROS in regulation of the inflammatory effect of hepatocellular carcinoma.

Oxidative stress levels and dynamic changes in mitochondrial gene expression in a radiation-induced lung injury model.

The purpose of this study was to set up a beagle dog model, for radiation-induced lung injury, that would be able to supply fresh lung tissues in the different injury phases for research into oxidative stress levels and mitochondrial gene expression. Blood serum and tissues were collected via CT-guided core needle biopsies from dogs in the various phases of the radiation response over a 40-week period. Levels of reactive oxygen species (ROS) and manganese superoxide dismutase 2 (MnSOD) protein expression in radiation-induced lung injury were determined by in situ immunocytochemistry; malondialdehyde (MDA) content and reductase activity in the peripheral blood were also tested; in addition, the copy number of the mitochondrial DNA and the level of function of the respiratory chain in the lung tissues were assessed. ROS showed dynamic changes and peaked at 4 weeks; MnSOD was mainly expressed in the Type II alveolar epithelium at 8 weeks; the MDA content and reductase activity in the peripheral blood presented no changes; the copy numbers of most mitochondrial genes peaked at 8 weeks, similarly to the level of function of the corresponding respiratory chain complexes; the level of function of the respiratory chain complex III did not peak until 24 weeks, similarly to the level of function of the corresponding gene Cytb. Radiation-induced lung injury was found to be a dynamically changing process, mainly related to interactions between local ROS, and it was not associated with the levels of oxidative stress in the peripheral blood. Mitochondrial genes and their corresponding respiratory chain complexes were found to be involved in the overall process.

Right atrial epithelioid angiosarcoma with multiple pulmonary metastasis confirmed by multimodality imaging-guided pulmonary biopsy: A case report and literature review.

RATIONALE: Primary cardiac tumors are very rare, and angiosarcoma accounts for about 33% of all primary malignant cardiac tumors. Primary cardiac epithelioid angiosarcoma is a highly aggressive and difficult to diagnose tumor, with early systemic metastasis and poor prognosis. PATIENT CONCERNS: A 35-year-old Han male experienced sudden severe palpitation and moderate dyspnea. The patient received a whole body F-18 fluoro-deoxyglucose positron emission tomography (18F-FDG PET)/computed tomography (CT) scan, the scan showed a large mass in the right atrium (RA) and numerous pulmonary nodules in both lungs. DIAGNOSES: The patient was diagnosed as right atrial epithelioid angiosarcoma with multiple pulmonary metastasis by pulmonary biopsy through CT-guided percutaneous transthoracic fine needle aspiration. INTERVENTIONS: The patient received a cycle of chemotherapy with docetaxel and gemcitabine, followed by another cycle with epirubicin and ifosfamide. OUTCOMES: The chemotherapy was ineffective. After the two cycles, the bilateral pleural effusion steadily increased, the patient had severe dyspnea and palpitation, and died three weeks later, with an overall survival of 2.5 months. LESSONS: Primary angiosarcoma of heart is a very rare and aggressive disease, and its diagnosis and treatment are difficult. Most patients may have systemic metastasis at diagnosis, and have a very short survival without surgical resection. Hence, early diagnosis and surgical resection is extremely important to treat this disease.

S6K1 phosphorylation-dependent degradation of Mxi1 by ß-Trcp ubiquitin ligase promotes Myc activation and radioresistance in lung cancer.

Rationale: Mxi1 is regarded as a potential tumor suppressor protein that antagonizes the transcriptional activity of proto-oncogene Myc. However, the clinical significances and underlying mechanisms by which Mxi1 is regulated in lung cancer remain poorly understood. Methods: Mass spectrometry analysis and immunoprecipitation assay were utilized to detect the protein-protein interaction. The phosphorylation of Mxi1 was evaluated by in vitro kinase assays. Poly-ubiquitination of Mxi1 was examined by in vivo ubiquitination assay. Lung cancer cells stably expressing wild-type Mxi1 or Mxi1-S160A were used for functional analyses. The expression levels of Mxi1 and S6K1 were determined by immunohistochemistry in lung cancer tissues and adjacent normal lung tissues. Results: We found that Mxi1 is downregulated and correlated with poor prognosis in lung cancer. Using tandem affinity purification technology, we provided evidence that ß-Trcp E3 ubiquitin ligase interacts with and promotes the ubiquitination and degradation of Mxi1. Furthermore, we demonstrated that Mxi1 is phosphorylated at S160 site by the protein kinase S6K1 and subsequently degraded via the ubiquitin ligase ß-Trcp. Moreover, a phosphorylation mutant form of Mxi1 (Mxi1-S160A), which cannot be degraded by S6K1 and ß-Trcp, is much more stable and efficient in suppressing the transcriptional activity of Myc and radioresistance in lung cancer cells. More importantly, a strong inverse correlation between S6K1 and Mxi1 expression was observed in human lung cancer tissues. Conclusion: Our findings not only establish a crosstalk between the mTOR/S6K1 signaling pathway and Myc activation, but also suggest that targeting S6K1/Mxi1 pathway is a promising therapeutic strategy for the treatment of lung cancer.

LZTS2 inhibits PI3K/AKT activation and radioresistance in nasopharyngeal carcinoma by interacting with p85.

Phosphoinositide 3-kinase (PI3K) activity is aberrantly activated in nasopharyngeal carcinoma. However, the underlying mechanisms remain unclear. Here, we found that Leucine zipper tumor suppressor 2 (LZTS2) was downregulated and predicted poor prognosis in nasopharyngeal carcinoma patients. Furthermore, we identified the PI3K subunit p85 as a novel LZTS2-interacting protein using an unbiased proteomics approach. Moreover, we demonstrated that LZTS2 competes with p110 for p85 binding and inhibits activation of the PI3K/AKT signaling pathway. Functionally, we showed that LZTS2 suppresses tumorigenesis and radioresistance in nasopharyngeal carcinoma in a p85-dependent manner. Taken together, our results not only provide understanding of the molecular mechanisms by which PI3K/AKT signaling is activated but also suggest that targeting the LZTS2/PI3K/AKT signaling axis is a promising therapeutic strategy for radiosensitization of nasopharyngeal carcinoma.

ß-Trcp ubiquitin ligase and RSK2 kinase-mediated degradation of FOXN2 promotes tumorigenesis and radioresistance in lung cancer.

Aberrant expression of FOXN2, a member of the Forkhead box transcription factors, has been found in several types of cancer. However, the underlying mechanisms of FOXN2 deregulation in tumorigenesis remain largely unknown. Here, we find that FOXN2 binds to and is ubiquitinated by ß-Trcp ubiquitin ligase and RSK2 kinase for degradation. Furthermore, we demonstrate that the Ser365 and Ser369 sites in a conserved DSGYAS motif are critical for the degradation of FOXN2 by ß-Trcp and RSK2. Moreover, gain-of-function and loss-of-function studies show that FOXN2 impairs cell proliferation in vitro and in vivo and enhances the radiosensitivity of lung cancer. Importantly, ß-Trcp-mediated and RSK2-mediated degradation of FOXN2 promotes tumorigenesis and radioresistance in lung cancer cells. Collectively, our study reveals a novel post-translational modification of FOXN2 and suggests that FOXN2 may be a potential therapeutic and radiosensitization target for lung cancer.

CDP138 silencing inhibits TGF-ß/Smad signaling to impair radioresistance and metastasis via GDF15 in lung cancer.

CDP138, a CDK5 binding partner, regulates cell proliferation and migration. However, the mechanisms by which CDP138 functions in these processes remain unclear. In this study, we show that CDP138 is frequently overexpressed and that high levels of CDP138 are correlated with lymph node metastasis in lung cancer. Furthermore, we provide evidence that CDP138-depleted lung cancer cells exhibit enhanced radiosensitivity as well as reduced migration and invasion. Mechanistically, we identify GDF15, a member of the TGF-ß superfamily, as a key downstream effector of CDP138. CDP138 silencing attenuates TGF-ß/Smad signaling activation at least in part through the downregulation of GDF15. More importantly, the observed phenotypes caused by CDP138 knockdown are partially dependent on GDF15 inhibition. Together, our findings demonstrate that CDP138 positively modulates the TGF-ß/Smad signaling pathway via GDF15 to promote radioresistance and metastasis, suggesting CDP138 as a potential oncogenic biomarker and a promising therapeutic target in the treatment of lung cancer.

The BET bromodomain inhibitor JQ1 radiosensitizes non-small cell lung cancer cells by upregulating p21.

Radiotherapy is an important treatment modality in the management of locally advanced non-small cell lung cancer (NSCLC). However, radioresistance markedly impairs its efficacy in clinic. Bromodomain and extra-terminal (BET) bromodomain inhibitors have demonstrated dramatic antitumor activity in several preclinical human cancer models. In this study, we investigated for the first time the effect of JQ1, a novel BET bromodomain inhibitor, on tumor cell radiosensitivity of NSCLC in vitro and in vivo. Our results demonstrated that JQ1 significantly enhanced the effect of irradiation in NSCLC cell lines through a c-myc-independent mechanism. The notable findings in response to this combined treatment were prolonged delay in IR-induced DNA double-strand break (DSB) repair, induced robust G2/M checkpoint arrest and increased cell apoptosis. Additional investigations revealed that induction of p21 played an important role in its radiosensitizing effects. In conclusion, these results suggested that BET bromodomain inhibition might offer a potential strategy for enhancing the effects of radiotherapy and reducing radioresistance.