CDKL1 potentiates the antitumor efficacy of radioimmunotherapy by binding to transcription factor YBX1 and blocking PD-L1 expression in lung cancer.

BACKGROUND: The evasion of the immune response by tumor cells through programmed death-ligand 1 (PD-L1) has been identified as a factor contributing to resistance to radioimmunotherapy in lung cancer patients. However, the precise molecular mechanisms underlying the regulation of PD-L1 remain incompletely understood. This study aimed to investigate the role of cyclin-dependent kinase-like 1 (CDKL1) in the modulation of PD-L1 expression and the response to radioimmunotherapy in lung cancer. METHODS: The tumorigenic roles of CDKL1 were assessed via cell growth, colony formation, and EdU assays and an in vivo nude mouse xenograft model. The in vitro radiosensitization effect of CDKL1 was evaluated using a neutral comet assay, γH2AX foci formation analysis, and a clonogenic cell survival assay. The protein‒protein interactions were confirmed via coimmunoprecipitation and GST pulldown assays. The regulation of PD-L1 by CDKL1 was evaluated via chromatin immunoprecipitation (ChIP), real-time quantitative PCR, and flow cytometry analysis. An in vitro conditioned culture model and an in vivo C57BL/6J mouse xenograft model were developed to detect the activation markers of CD8+ T cells and evaluate the efficacy of CDKL1 overexpression combined with radiotherapy (RT) and an anti-PD-L1 antibody in treating lung cancer. RESULTS: CDKL1 was downregulated and suppressed the growth and proliferation of lung cancer cells and increased radiosensitivity in vitro and in vivo. Mechanistically, CDKL1 interacted with the transcription factor YBX1 and decreased the binding affinity of YBX1 for the PD-L1 gene promoter, which consequently inhibits the expression of PD-L1, ultimately leading to the activation of CD8+ T cells and the inhibition of immune evasion in lung cancer. Moreover, the combination of CDKL1 overexpression, RT, and anti-PD-L1 antibody therapy exhibited the most potent antitumor efficacy against lung cancer. CONCLUSIONS: Our findings demonstrate that CDKL1 plays a crucial role in regulating PD-L1 expression, thereby enhancing the antitumor effects of radioimmunotherapy. These results suggest that CDKL1 may be a promising therapeutic target for the treatment of lung cancer.

Targeting FBXO22 enhances radiosensitivity in non-small cell lung cancer by inhibiting the FOXM1/Rad51 axis.

Radioresistance is a major constraint on the efficacy of lung cancer radiotherapy, but its mechanism has not been fully elucidated. Here, we found that FBXO22 was aberrantly highly expressed in lung cancer and that FBXO22 knockdown increased the radiosensitivity of lung cancer cells. Mechanistically, FBXO22 promoted Rad51 gene transcription by increasing the level of FOXM1 at the Rad51 promoter, thereby inducing the formation of lung cancer radioresistance. Furthermore, we found that deguelin, a potential inhibitor of FBXO22, enhanced radiosensitivity in an FBXO22/Rad51-dependent manner and was safely tolerated in vivo. Collectively, our results illustrate that FBXO22 induces lung cancer radioresistance by activating the FOXM1/Rad51 axis and provide preclinical evidence for the clinical translation of this critical target.

OTUB1 Targets CHK1 for Deubiquitination and Stabilization to Facilitate Lung Cancer Progression and Radioresistance.

PURPOSE: Radioresistance of lung cancer poses a significant challenge when it comes to the treatment of advanced, recurrent, and metastatic cases. Ovarian tumor domain ubiquitin aldehyde binding 1 (OTUB1) is a key member of the deubiquitinase OTU superfamily. This protein is involved in various cellular functions, including cell proliferation, iron death, lipid metabolism, and cytokine secretion as well as immune response processes. However, its specific role and molecular mechanism in lung cancer radioresistance remain to be clarified. METHODS AND MATERIALS: The expression levels of OTUB1 in paired lung cancer tissues were determined by immunohistochemistry. In vitro and in vivo experiments were conducted to investigate the impact of OTUB1 on the growth and proliferation of lung cancer. Coimmunoprecipitation and Western blotting techniques were performed to examine the interaction between OTUB1 and CHK1. The DNA damage response was measured by comet tailing and immunofluorescence staining. KEGG pathways and Gene Ontology terms were analyzed based on RNA sequencing. RESULTS: Our findings reveal a high frequency of OTUB1 overexpression, which is associated with an unfavorable prognosis in patients with lung cancer. Through comprehensive investigations, we demonstrate that OTUB1 depletion impairs the process of DNA damage repair and overcomes radioresistance. In terms of the underlying mechanism, our study uncovers that OTUB1 deubiquitinates and stabilizes CHK1, which enhances CHK1 stability, thereby regulating DNA damage and repair. Additionally, we identify CHK1 as the primary downstream effector responsible for mediating the functional effects exerted by OTUB1 specifically in lung cancer. Importantly, OTUB1 has the potential to be a valuable marker for improving the efficacy of radiation therapy for lung adenocarcinoma. CONCLUSIONS: These findings unveil a novel role for OTUB1 in enhancing radioresistance by deubiquitination and stabilization of the expression of CHK1 in lung cancer and indicate that targeting OTUB1 holds great potential as an effective therapeutic approach for enhancing the efficacy of radiation therapy in lung cancer.

FIBP interacts with transcription factor STAT3 to induce EME1 expression and drive radioresistance in lung adenocarcinoma.

Cancer cells inevitably develop radioresistance during lung adenocarcinoma radiotherapy. However, the mechanisms are incompletely clarified. In this study, we show that FIBP protein expression in lung adenocarcinoma tissues is upregulated and associated with worse overall survival. Functionally, we find that depletion of FIBP inhibits lung adenocarcinoma progression and radioresistance in vitro and in vivo. Moreover, we uncover that FIBP interacts with STAT3 to enhance its transcriptional activity, thereby inducing the expression of the downstream target gene EME1. Importantly, we demonstrate that the biological effects of FIBP are partially dependent on EME1 in lung adenocarcinoma. Our work reveals that FIBP modulates the STAT3/EME1 axis to drive lung cancer progression and radioresistance, indicating that targeting FIBP may be a novel intervention strategy for lung adenocarcinoma radiotherapy.

KDM4C silencing inhibits cell migration and enhances radiosensitivity by inducing CXCL2 transcription in hepatocellular carcinoma.

KDM4C, which is a histone lysine demethylase, has been proposed to participate in the malignant transformation and progression of several types of cancer. However, its roles in hepatocellular carcinoma (HCC) remain poorly understood. Here, we find that KDM4C protein expression is increased in HCC and promotes HCC cell growth, proliferation and migration. Furthermore, we provide evidence that depletion of KDM4C leads to a defective G2/M checkpoint, increases radiation-induced DNA damage, impairs DNA repair and enhances radiosensitivity in HCC cells. Using RNA sequencing, we identify that the chemokine CXCL2 is a downstream effector of KDM4C. KDM4C knockdown increases the binding of H3K36me3 to the promoter of CXCL2, thus upregulating CXCL2 expression and promoting CXCL2 secretion in HCC cells. Importantly, the observed effects of KDM4C depletion in HCC cells can be partially rescued by CXCL2 silencing. Thus, our findings reveal that KDM4C is involved in cell migration and radiosensitivity by modulating CXCL2 transcription, indicating that KDM4C may be a potential therapeutic target in HCC.

Deficiency of Carbamoyl Phosphate Synthetase 1 Engenders Radioresistance in Hepatocellular Carcinoma via Deubiquitinating c-Myc.

PURPOSE: Tumor radiation resistance is the main obstacle to effective radiation therapy for patients with hepatocellular carcinoma (HCC). We identified the role of urea cycle key enzyme carbamoyl phosphate synthetase 1 (CPS1) in radioresistance of HCC and explored its mechanism, aiming to provide a novel radiosensitization strategy for the CPS1-deficiency HCC subtype. METHODS AND MATERIALS: The expression of CPS1 was measured by western blot and immunohistochemistry. Cell growth assay, EdU assay, cell apoptosis assay, cell cycle assay, clone formation assay, and subcutaneous tumor assay were performed to explore the relationship between CPS1 and radioresistance of HCC cells. Lipid metabonomic analysis was used for investigating the effects of CPS1 on lipid synthesis of HCC cells. RNA sequencing and coimmunoprecipitation assay were carried out to reveal the mechanism of CPS1 participating in the regulation of HCC radiation therapy resistance. Furthermore, 10074-G5, the specific inhibitor of c-Myc, was administered to HCC cells to investigate the role of c-Myc in CPS1-deficiency HCC cells. RESULTS: We found that urea cycle key enzyme CPS1 was frequently lower in human HCC samples and positively associated with the patient's prognosis. Functionally, the present study proved that CPS1 depletion could accelerate the development of HCC and induce radiation resistance of HCC in vitro and in vivo, and deficiency of CPS1 promoted the synthesis of some lipid molecules. Regarding the mechanism, we uncovered that inhibition of CPS1 upregulated CyclinA2 and CyclinD1 by stabilizing oncoprotein c-Myc at the posttranscriptional level and generated radioresistance of HCC cells. Moreover, inactivation of c-Myc using 10074-G5, a specific c-Myc inhibitor, could partially attenuate the proliferation and radioresistance induced by depletion of CPS1. CONCLUSIONS: Our results recapitulated that silencing CPS1 could promote HCC progression and radioresistance via c-Myc stability mediated by the ubiquitin-proteasome system, suggesting that targeting c-Myc in CPS1-deficiency HCC subtype may be a valuable radiosensitization strategy in the treatment of HCC.

Prediction of response and adverse drug reaction of pemetrexed plus platinum-based chemotherapy in lung adenocarcinoma by serum metabolomic profiling.

BACKGROUND: Pemetrexed plus platinum doublet chemotherapy regimen remains to be the standard first-line treatment for lung adenocarcinoma patients. However, few biomarkers can be used to identify potential beneficiaries with maximal efficacy and minimal toxicity. This study aimed to explore potential biomarker models predictive of efficacy and toxicity after pemetrexed plus platinum chemotherapy based on metabolomics profiling. METHODS: A total of 144 patients who received at least two cycles of pemetrexed plus platinum chemotherapy were enroled in the study. Serum samples were collected before initial treatment to perform metabolomics profiling analysis. Logistic regression analysis was performed to establish prediction models. RESULTS: 157 metabolites were found to be differentially expressed between the response group and the nonresponse group. A panel of Phosphatidylserine 20:4/20:1, Sphingomyelin d18:1/18:0, and Phosphatidic Acid 18:1/20:0 could predict pemetrexed and platinum chemotherapy response with an Area Under the Receiver Operating Characteristic curve (AUROC) of 0.7968. 76 metabolites were associated with hematological toxicity of pemetrexed plus platinum chemotherapy. A panel incorporating triglyceride 14:0/22:3/22:5, 3-(3-Hydroxyphenyl) Propionate Acid, and Carnitine C18:0 was the best predictive ability of hematological toxicity with an AUROC of 0.7954. 54 differential expressed metabolites were found to be associated with hepatotoxicity of pemetrexed plus platinum chemotherapy. A model incorporating stearidonic acid, Thromboxane B3, l-Homocitrulline, and phosphoinositide 20:3/18:0 showed the best predictive ability of hepatotoxicity with an AUROC of 0.8186. CONCLUSIONS: This study established effective and convenient models that can predict the efficacy and toxicity of pemetrexed plus platinum chemotherapy in lung adenocarcinoma patients before treatment delivery.

Targeting KDM4C enhances CD8+ T cell mediated antitumor immunity by activating chemokine CXCL10 transcription in lung cancer.

BACKGROUND: Although immune checkpoint blockade (ICB) has been proven to achieve a persistent therapeutic response in various tumor types, only 20%-40% of patients benefit from this treatment. Radiotherapy (RT) can enhance tumor immunogenicity and improve the ICB response, but the outcome achieved by combining these two modalities remains clinically unsatisfactory. We previously uncovered that lysine-specific demethylase 4C (KDM4C) is a regulator of radiosensitivity in lung cancer. However, the role of KDM4C in antitumor immunity has not yet been investigated. METHODS: Infiltrating immune cells in our mouse tumor model were screened by flow cytometry. An in vivo subcutaneous transplanted tumor model and in vitro conditioned culture model were constructed to detect the quantitative and functional changes in CD8+ T cells. RNA sequencing and chromatin immunoprecipitation-PCR assays were used to explore the downstream regulatory mechanism of KDM4C in antitumor immunity. A C57BL/6 mouse tumor model was developed to evaluate the efficacy and safety of a triple therapy (the KDM4C-specific inhibitor SD70 plus RT and an anti-PD-L1 antibody) in lung cancer in vivo. RESULTS: Genetical or pharmacological inhibition of KDM4C specifically increased CD8+ T cell infiltration; promoted the proliferation, migration and activation of CD8+ T cells; and alleviated CD8+ T cell exhaustion in mouse tumor tissues. Mechanistically, KDM4C inhibition increased the binding of H3K36me3 to the CXCL10 promoter region, thus inducing CXCL10 transcription and enhancing the CD8+ T cell mediated antitumor immune response. More importantly, among the tested regimens, the triple therapy achieved the best therapeutic efficacy with tolerable toxicity in lung cancer. CONCLUSIONS: Our data reveal a crucial role for KDM4C in antitumor immunity in lung cancer and indicate that targeting KDM4C in combination with radioimmunotherapy might be a promising synergistic strategy in lung cancer.

Arginine methyltransferase PRMT5 methylates and destabilizes Mxi1 to confer radioresistance in non-small cell lung cancer.

Radioresistance is regarded as the main cause of local recurrence and distant metastasis in non-small cell lung cancer. However, the underlying mechanisms of radioresistance remains incompletely understood. In this study, we find that the arginine methyltransferase PRMT5 interacts with and methylates Mxi1, which promotes the binding of the ß-Trcp ligase to Mxi1, facilitating the ubiquitination and degradation of Mxi1 in lung cancer. Furthermore, genetic blockade of PRMT5 impairs DNA damage repair and enhances lung cancer radiosensitivity in vitro and in vivo, and these phenotypes are partially reversed by Mxi1 silencing. More importantly, pharmacological inhibition of PRMT5 with the specific inhibitor EPZ015666 leads to extraordinary radiosensitization in vitro and in vivo in lung cancer. Altogether, our data indicate that PRMT5 methylates and destabilizes Mxi1 to confer radioresistance, suggesting that PRMT5 may be a promising radiosensitization target in non-small cell lung cancer.

BRD4-IRF1 axis regulates chemoradiotherapy-induced PD-L1 expression and immune evasion in non-small cell lung cancer.

BACKGROUND: Chemoradiotherapy-induced PD-L1 upregulation leads to therapeutic resistance and treatment failure. The PD-1/PD-L1 blocking antibodies sensitize cancers to chemoradiotherapy by blocking extracellular PD-1 and PD-L1 binding without affecting the oncogenic function of intracellular PD-L1. Reversing the chemoradiation-induced PD-L1 expression could provide a new strategy to achieve a greater anti-tumour effect of chemoradiotherapy. Here, we aimed to identify candidate small molecular inhibitors that might boost the anti-tumour immunity of chemoradiotherapy by decreasing treatment-induced PD-L1 expression in non-small cell lung cancer (NSCLC). METHODS: A drug array was used to recognize compounds that can suppress the cisplatin-induced and radiation-induced PD-L1 expression in NSCLC via the flow cytometry-based assay. We examined whether and how targeting bromodomain containing 4 (BRD4) inhibits chemoradiation-induced PD-L1 expression and evaluated the effect of BRD4 inhibition and chemoradiation combination in vivo. RESULTS: BRD4 inhibitors JQ1 and ARV-771 were identified as the most promising drugs both in the cisplatin and radiation screening projects in two NSCLC cell lines. Targeting BRD4 was supposed to block chemoradiotherapy inducible PD-L1 expression by disrupting the recruitment of BRD4-IRF1 complex to PD-L1 promoter. A positive correlation between BRD4 and PD-L1 expression was observed in human NSCLC tissues. Moreover, BRD4 inhibition synergized with chemoradiotherapy and PD-1 blockade to show a robust anti-tumour immunity dependent on CD8+ T cell through limiting chemoradiation-induced tumour cell surface PD-L1 upregulation in vivo. Notably, the BRD4-targeted combinatory treatments did not show increased toxicities. CONCLUSION: The data showed that BRD4-targeted therapy synergized with chemoradiotherapy and anti-PD-1 antibody by boosting anti-tumour immunity in NSCLC.

Targeting senescence-like fibroblasts radiosensitizes non-small cell lung cancer and reduces radiation-induced pulmonary fibrosis.

Cancer cell radioresistance is the primary cause of the decreased curability of non-small cell lung cancer (NSCLC) observed in patients receiving definitive radiotherapy (RT). Following RT, a set of microenvironmental stress responses is triggered, including cell senescence. However, cell senescence is often ignored in designing effective strategies to resolve cancer cell radioresistance. Herein, we identify the senescence-like characteristics of cancer-associated fibroblasts (CAFs) after RT and clarify the formidable ability of senescence-like CAFs in promoting NSCLC cell proliferation and radioresistance through the JAK/STAT pathway. Specific induction of senescence-like CAF apoptosis using FOXO4-DRI, a FOXO4-p53-interfering peptide, resulted in remarkable effects on radiosensitizing NSCLC cells in vitro and in vivo. In addition, in this study, we also uncovered an obvious therapeutic effect of FOXO4-DRI on alleviating radiation-induced pulmonary fibrosis (RIPF) by targeting senescence-like fibroblasts in vivo. In conclusion, by targeting senescence, we offer a strategy that simultaneously decreases radioresistance of NSCLC and the incidence of RIPF.

Precise pathological classification of non-small cell lung adenocarcinoma and squamous carcinoma based on an integrated platform of targeted metabolome and lipidome.

BACKGROUND: Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related death worldwide. Lung adenocarcinoma (LUAD) and squamous cell carcinoma (LUSC) are the most common subtypes of NSCLC. Despite genetic differences between LUAD and LUSC have been clarified in depth, the metabolic differences of these two subtypes are still unclear. METHODS: Totally, 128 plasma samples of NSCLC patients were collected before initial treatments, followed by determination of LC-ESI-Q TRAP-MS/MS. Differentially expressed metabolites were screened based on a strict standard. RESULTS: Based on the integrated platform of targeted metabolome and lipidome, a total of 1141 endogenous metabolites (including 809 lipids) were finally detected in the plasma of NSCLC patients, including 16 increased and 3 decreased endogenous compounds in LUAD group when compared with LUSC group. Thereafter, a logistic regression model integrating four differential metabolites [2-(Methylthio) ethanol, Cortisol, D-Glyceric Acid, and N-Acetylhistamine] was established and could accurately differentiate LUAD and LUSC with an area under the ROC curve of 0.946 (95% CI 0.886-1.000). The cut-off value showed a satisfactory efficacy with 92.0% sensitivity and 92.9% specificity. KEGG functional enrichment analysis showed these differentially expressed metabolites could be further enriched in riboflavin metabolism, steroid hormone biosynthesis, prostate cancer, etc. The endogenous metabolites identified in this study have the potential to be used as novel biomarkers to distinguish LUAD from LUSC. CONCLUSIONS: Our research might provide more evidence for exploring the pathogenesis and differentiation of NSCLC. This research could promote a deeper understanding and precise treatment of lung cancer.

Benefit from Adjuvant TKIs Versus TKIs Plus Chemotherapy in EGFR-Mutant Stage III-pN2 Lung Adenocarcinoma.

BACKGROUND: Recent studies have demonstrated benefits from adjuvant tyrosine-kinase inhibitors (TKIs) compared with chemotherapy in non-small cell lung cancer. We launched a multi-center retrospective study to evaluate the efficacy and toxicity of adjuvant TKIs with or without chemotherapy in epidermal growth factor receptor (EGFR)-mutant stage III-pN2 lung adenocarcinoma. METHODS: Two hundred and seventy-four consecutive cases with stage III-pN2 lung adenocarcinoma and complete resection have been investigated. Clinic-pathologic characteristics, adjuvant treatments, long-term survivals, and toxicities were documented. Risk factors of distant metastasis-free survival (DMFS), disease-free survival (DFS), and overall survival (OS) were evaluated. RESULTS: There were 52 (19.0%) patients treated with adjuvant TKIs alone, 199 (72.6%) with adjuvant chemotherapy alone, and 23 (8.4%) with both. After a median follow-up time of 29 months, the two-year DMFS, DFS, and OS was 61.2%, 54.1%, and 91.2%, respectively. According to univariable analyses, the risk factors were lymphovascular invasion (p < 0.001), extranodal extension (p = 0.005), and adjuvant systemic therapy (p = 0.006) for DMFS, EGFR mutation type (p = 0.025), lymphovascular invasion (p = 0.013), extranodal extension (p = 0.004), and adjuvant systemic therapy (p < 0.001) for DFS, and EGFR mutation type (p < 0.001) for OS. Multivariable analyses indicated that the independent prognostic factors were adjuvant systemic therapy (TKIs vs. TKIs+chemotherapy, Harzard ratio (HR) = 0.40; p = 0.036; TKIs vs. chemotherapy, HR = 0.38; p = 0.004), lymphovascular invasion (yes vs. no, HR = 2.22; p = 0.001) for DMFS, and adjuvant systemic therapy (TKIs vs. TKIs+chemotherapy, HR = 0.42; p = 0.034; TKIs vs. chemotherapy, HR = 0.33; p < 0.001) for DFS. No significant difference was found in the incidence of Grade 3-4 toxicities between groups (p = 0.445). CONCLUSIONS: Adjuvant TKIs might be a beneficial choice compared with adjuvant chemotherapy or combination systemic treatments.

USP9X-mediated KDM4C deubiquitination promotes lung cancer radioresistance by epigenetically inducing TGF-ß2 transcription.

Radioresistance is regarded as the main barrier to effective radiotherapy in lung cancer. However, the underlying mechanisms of radioresistance remain elusive. Here, we show that lysine-specific demethylase 4C (KDM4C) is overexpressed and correlated with poor prognosis in lung cancer patients. We provide evidence that genetical or pharmacological inhibition of KDM4C impairs tumorigenesis and radioresistance in lung cancer in vitro and in vivo. Moreover, we uncover that KDM4C upregulates TGF-ß2 expression by directly reducing H3K9me3 level at the TGF-ß2 promoter and then activates Smad/ATM/Chk2 signaling to confer radioresistance in lung cancer. Using tandem affinity purification technology, we further identify deubiquitinase USP9X as a critical binding partner that deubiquitinates and stabilizes KDM4C. More importantly, depletion of USP9X impairs TGF-ß2/Smad signaling and radioresistance by destabilizing KDM4C in lung cancer cells. Thus, our findings demonstrate that USP9X-mediated KDM4C deubiquitination activates TGF-ß2/Smad signaling to promote radioresistance, suggesting that targeting KDM4C may be a promising radiosensitization strategy in the treatment of lung cancer.

ß-Trcp and CK1δ-mediated degradation of LZTS2 activates PI3K/AKT signaling to drive tumorigenesis and metastasis in hepatocellular carcinoma.

Distant metastasis is the leading cause of treatment failure in patients with hepatocellular carcinoma (HCC). However, the underlying mechanisms have not been fully elucidated. Here, we report that Leucine zipper tumor suppressor 2 (LZTS2) is downregulated and correlated with poor prognosis in HCC. Furthermore, we provide evidence that LZTS2 associates with p85 to inhibit the activation of PI3K/AKT signaling and impairs HCC tumorigenesis and metastasis in vitro and in vivo. Moreover, we identify LZTS2 as a bona fide substrate of the E3 ligase ß-Trcp and protein kinase CK1δ, which are responsible for the ubiquitination and degradation of LZTS2. Importantly, we show that the ß-Trcp and CK1δ-mediated degradation of LZTS2 promotes HCC progression and metastasis by activating PI3K/AKT signaling. Collectively, our study not only illustrates the roles of LZTS2 in regulating HCC tumorigenesis and metastasis but also reveals a novel posttranslational modification of LZTS2 by ß-Trcp and CK1δ, indicating that the ß-Trcp/CK1δ/LZTS2/PI3K axis may be a novel oncogenic driver involved in HCC progression and metastasis.

UBE2O targets Mxi1 for ubiquitination and degradation to promote lung cancer progression and radioresistance.

UBE2O, an E2/E3 hybrid ubiquitin-protein ligase, has been implicated in the regulation of adipogenesis, erythroid differentiation, and tumor proliferation. However, its role in cancer radioresistance remains completely unknown. Here, we uncover that UBE2O interacts and targets Mxi1 for ubiquitination and degradation at the K46 residue. Furthermore, we show that genetical or pharmacological blockade of UBE2O impairs tumor progression and radioresistance in lung cancer in vitro and in vivo, and these effects can be restored by Mxi1 inhibition. Moreover, we demonstrate that UBE2O is overexpressed and negatively correlated with Mxi1 protein levels in lung cancer tissues. Collectively, our work reveals that UBE2O facilitates tumorigenesis and radioresistance by promoting Mxi1 ubiquitination and degradation, suggesting that UBE2O is an attractive radiosensitization target for the treatment of lung cancer.

Cyclin K interacts with ß-catenin to induce Cyclin D1 expression and facilitates tumorigenesis and radioresistance in lung cancer.

Rationale: Radioresistance remains the major cause of local relapse and distant metastasis in lung cancer. However, the underlying molecular mechanisms remain poorly defined. This study aimed to investigate the role and regulatory mechanism of Cyclin K in lung cancer radioresistance. Methods: Expression levels of Cyclin K were measured by immunohistochemistry in human lung cancer tissues and adjacent normal lung tissues. Cell growth and proliferation, neutral comet and foci formation assays, G2/M checkpoint and a xenograft mouse model were used for functional analyses. Gene expression was examined by RNA sequencing and quantitative real-time PCR. Protein-protein interaction was assessed by immunoprecipitation and GST pull-down assays. Results: We report that Cyclin K is frequently overexpressed and correlates with poor prognosis in lung cancer patients. Functionally, we demonstrate that Cyclin K depletion results in reduced proliferation, defective G2/M checkpoint and enhanced radiosensitivity in lung cancer. Mechanistically, we reveal that Cyclin K interacts with and promotes the stabilization of ß-catenin protein, thereby upregulating the expression of Cyclin D1. More importantly, we show that Cyclin D1 is the major effector that mediates the biological functions of Cyclin K in lung cancer. Conclusions: These findings suggest that Cyclin K positively modulates the ß-catenin/Cyclin D1 axis to promote tumorigenesis and radioresistance in lung cancer, indicating that Cyclin K may represent a novel attractive biomarker for lung cancer radiotherapy.

Risk Prediction for Locoregional Recurrence in Epidermal Growth Factor Receptor-Mutant Stage III-pN2 Lung Adenocarcinoma after Complete Resection: A Multi-center Retrospective Study.

Background: This study aimed to develop a predictive model based on the risk of locoregional recurrence (LRR) in epidermal growth factor receptor (EGFR)-mutant stage III-pN2 lung adenocarcinoma after complete resection. Methods: A total of 11,020 patients with lung surgery were screened to determine completely resected EGFR-mutant stage III-pN2 lung adenocarcinoma. Patients were excluded if they received preoperative therapy or postoperative radiation therapy (PORT). The time from surgery to LRR was recorded. Clinicopathological variables with statistical significance predicting LRR in the multivariate Cox regression were incorporated into the competing risk nomogram. Patients were then sub-grouped based on different recurrence risk as a result of the nomogram. Results: Two hundred and eighty-eight patients were enrolled, including 191 (66.3%) with unforeseen N2 (IIIA1-2), 75 (26.0%) with minimal/single station N2 (IIIA3), and 22 (7.6%) with bulky and/or multilevel N2 (IIIA4). The 2-year overall cumulative incidence of LRR was 27.2% (confidence interval [CI], 16.3%-38.0%). IIIA4 disease (hazard ratio, 2.65; CI, 1.15-6.07; P=0.022) and extranodal extension (hazard ratio, 3.33; CI, 1.76-6.30; P<0.001) were independent risk factors for LRR and were incorporated into the nomogram. Based on the nomogram, patients who did not have any risk factor (low-risk) had a significantly lower predicted 2-year incidence of LRR than those with any of the risk factors (high-risk; 4.6% vs 21.9%, P<0.001). Conclusions: Pre-treatment bulky/multilevel N2 and pathological extranodal extension are risk factors for locoregional recurrence in EGFR-mutant stage III-pN2 lung adenocarcinoma. Intensive adjuvant therapies and active follow-up should be considered in patients with any of the risk factors.

Targeting NEK2 impairs oncogenesis and radioresistance via inhibiting the Wnt1/ß-catenin signaling pathway in cervical cancer.

BACKGROUND: NEK2, a serine/threonine kinase involved in mitosis, has been found to function in chromosome instability, tumor progression and metastasis, but its role in cervical cancer radioresistance remains unknown. METHODS: We detected the protein levels of NEK2 in cervical carcinoma tissues and paired paracarcinoma tissues by immunohistochemistry. The roles of NEK2 in oncogenesis were examined using cell growth and colony formation assays, EdU assay, apoptosis assay as well as in vivo mouse model. γ-H2AX and Rad51 foci formation, neutral comet assay and clonogenic cell survival assay were applied to determine the radiosensitivity of cervical cancer cells. RNA-seq was performed to identify the downstream effector of NEK2. The gene expression levels were measured by Real-time PCR. RESULTS: We report that NEK2 protein level is overexpressed and correlated with the tumor stage and lymph node metastasis in cervical cancer tissues. Furthermore, we provided evidence that depletion of NEK2 impairs oncogenesis and enhances radiosensitivity in cervical cancer. Using RNA sequencing, we identify Wnt1 as a key downstream effector of NEK2. Knockdown of NEK2 downregulates the mRNA and protein levels of Wnt1, thereby inhibiting the activation of the Wnt/ß-catenin signaling pathway. More importantly, the observed consequences induced by NEK2 depletion in cervical cancer cells can be partially rescued by Wnt1 overexpression. CONCLUSIONS: Our results demonstrate that NEK2 activates the Wnt/ß-catenin signaling pathway via Wnt1 to drive oncogenesis and radioresistance in cervical cancer, indicating that NEK2 may be a promising target for the radiosensitization of cervical cancer.

IQGAP3 interacts with Rad17 to recruit the Mre11-Rad50-Nbs1 complex and contributes to radioresistance in lung cancer.

IQ motif containing GTPase-activating protein 3 (IQGAP3) has been implicated in diverse cellular processes, including neuronal morphogenesis, cell proliferation and motility, and epithelial-mesenchymal transition. However, its role in cancer radioresistance is completely unknown. Here, we report that IQGAP3 is overproduced in lung cancer patients and correlates with poor clinical outcomes. Functionally, we demonstrate that depletion of IQGAP3 impairs oncogenesis and overcomes radioresistance in lung cancer in vitro and in vivo. Mechanistically, we uncover that IQGAP3 interacts with Rad17 and controls its expression to activate the ATM/Chk2 and ATR/Chk1 signaling pathways by recruiting the Mre11-Rad50-Nbs1 (MRN) complex in response to DNA damage. Moreover, Rad17 is identified as the major downstream effector that mediates the functions of IQGAP3 in lung cancer. Clinically, IQGAP3 overexpression positively correlates with Rad17 upregulation in human lung cancer tissues. Collectively, these data support key role for IQGAP3 in promoting lung cancer radioresistance by interacting with Rad17 and suggest that targeting IQGAP3 may be an attractive strategy for lung cancer radiotherapy.