Radiosensitivity Of Non-small Cell Lung Cancer Research Articles

Abstract B015: PLK4 inhibition as a strategy to enhance non-small cell lung cancer radiosensitivity

Abstract Lung cancer is the leading cause of cancer-related mortality worldwide. Non-small cell lung cancer (NSCLC) is the most common subtype of lung cancer and comprises 85% of cases. Despite treatment advances, local control after curative-intent chemoradiation for NSCLC remains suboptimal. Polo-like kinase 4 (PLK4) is a serine-threonine kinase that plays a critical role in the regulation of centrosome duplication and cell cycle progression and is overexpressed in NSCLC, thus, making it a potential therapeutic target. CFI-400945 is an orally available PLK4 inhibitor currently undergoing clinical trial evaluation. As radiation causes cell death primarily by mitotic catastrophe, a process enhanced by alterations in centrosome amplification, we hypothesized that disruption of the mitotic machinery by inhibition of PLK4 would enhance the effects of radiation in NSCLC. PLK4 inhibition by CFI-400945 resulted in radiosensitization of NSCLC cell lines. In contrast, CFI-400945 had no effect on the radiosensitivity of normal lung fibroblasts. PLK4 inhibition did not affect cell-cycle phase distribution prior to radiation, but rather the combination of CFI-400945 and radiation resulted in increased G2/M cell cycle arrest, increased centrosome amplification, and a concomitant increase in cell death through mitotic catastrophe. Lastly, CFI-400945 treatment enhanced the radiation-induced tumor growth delay of NSCLC tumor xenografts. These data indicate that targeting PLK4 is a novel approach to enhance the radiation sensitivity of NSCLC in vitro and in vivo through potentiation of centrosome amplification and cell death through mitotic catastrophe. Citation Format: Irma G. Dominguez-Vigil, Kishore Banik, Marta John Baro, Joseph N. Contessa, Thomas J. Hayman. PLK4 inhibition as a strategy to enhance non-small cell lung cancer radiosensitivity. [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Translating Targeted Therapies in Combination with Radiotherapy; 2025 Jan 26-29; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(2_Suppl):Abstract nr B015.

Abstract P017: Circulating tumor DNA kinetics identify prodynorphin signaling as a target to radiosensitize non-small cell lung cancer

Abstract Introduction: Definitive chemoradiation therapy (CRT) is essential for controlling locoregionally advanced non-small cell lung cancer (NSCLC); however, up to 30% of patients experience local recurrences within the radiation field. Analyzing favorable responders to treatment could enable identification of genetic alterations associated with radiosensitivity, but standard imaging cannot distinguish residual disease from inflammation and fibrosis during treatment to assess treatment response. We hypothesized that circulating tumor DNA (ctDNA) kinetics during CRT could identify genetic alterations associated with radiosensitivity that could be validated in preclinical models and harnessed to develop new combination therapies with radiotherapy (RT). Methods: We applied cancer personalized profiling by deep sequencing (CAPP-Seq) ctDNA analysis to pre-CRT and mid-treatment (mid-CRT) plasma samples from 61 patients with Stage II-III NSCLC. We identified ctDNA rapid responders as the 10 patients with the largest decrease in ctDNA concentration mid-CRT without local progression and determined the prevalence of genetic alterations in rapid responders versus slow responders using Fisher’s exact tests corrected for multiple hypothesis testing. To investigate the therapeutic potential of targeting PDYN during RT, we performed clonogenic assays on isogeneic PDYN CRISPR-Cas9 knockout and wildtype H1299 and SW1573 human NSCLC cell lines and H1299 cells treated with prodynorphin-neutralizing antibodies. Results: Mid-CRT log-fold change in ctDNA concentration was significantly associated with progression-free survival (P=0.02) in Stage II-III NSCLC. Mutations in PDYN, which encodes the opioid peptide precursor protein prodynorphin, were observed in 30% of ctDNA rapid responders and 0% of ctDNA slow responders (adjusted P=0.04). In NSCLC patients from TCGA treated with RT, the cumulative incidence of local failure was significantly lower in tumors with PDYN mutations (0% vs. 17% at 3 years, P<0.001). PDYN knockout (PDYNNULL) radiosensitized human NSCLC cancer cells by clonogenic assay, and extracellular administration of prodynorphin or its cleavage product dynophin A partially restored radioresistance in PDYNNULL cells. Prodynorphin cleavage products have been reported to signal through opioid receptors and to antagonize N-methyl-D-aspartate receptor (NMDAR) signaling. Although treatment of wildtype H1299 cells with opioid receptor antagonists did not affect radiosensitivity, the NMDAR antagonist MK-801 partially restored RT resistance in PDYNNULL cells. Finally, prodynorphin-neutralizing antibodies significantly reduced clonogenic survival of human NSCLC cells. Conclusions: ctDNA kinetics enable the identification of patients responding favorably to treatment and putative targets to enhance the efficacy of RT. Targeting prodynorphin may enhance the radiosensitivity of NSCLC by increasing NMDAR signaling. Citation Format: Ziwei Wang, Angela B. Hui, Ash A. Alizadeh, Maximilian Diehn, Everett J. Moding.Circulating tumor DNA kinetics identify prodynorphin signaling as a target to radiosensitize non-small cell lung cancer.[abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Translating Targeted Therapies in Combination with Radiotherapy; 2025 Jan 26-29; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(2_Suppl):Abstract nr P017

Role of exosomes in modulating non-small cell lung cancer radiosensitivity.

Non-small cell lung cancer (NSCLC) constitutes a significant proportion of lung cancer cases, and despite advancements in treatment modalities, radiotherapy resistance remains a substantial hurdle in effective cancer management. Exosomes, which are small vesicles secreted by cells, have emerged as pivotal players in intercellular communication and influence various biological processes, including cancer progression and the response to therapy. This review discusses the intricate role of exosomes in the modulation of NSCLC radiosensitivity. The paper focuses on NSCLC and highlights how tumor-derived exosomes contribute to radioresistance by enhancing DNA repair, modulating immune responses, and altering the tumor microenvironment. We further explore the potential of mesenchymal stem cell-derived exosomes to overcome radiotherapy resistance and their potential as biomarkers for predicting therapeutic outcomes. Understanding the mechanisms by which exosomes affect radiotherapy can provide new avenues for enhancing treatment efficacy and improving the survival rates of patients with NSCLC.

Enhanced radiotherapy susceptibility in NSCLC through palbociclib-mediated PP5 inhibition

Radiotherapy remains a cornerstone in the treatment of non-small cell lung cancer (NSCLC), yet radioresistance often limits its efficacy. Identifying molecular targets that enhance radiosensitivity is crucial to offering both curative and palliative benefits for patients with NSCLC. Utilizing bioinformatics analysis, our study revealed significantly higher expression of PP5 in NSCLC tissues compared to normal tissues. Kaplan-Meier survival analysis also showed that high PP5 expression correlates with poorer overall survival, particularly in patients undergoing radiotherapy, suggesting a role for PP5 in radioresistance. We further demonstrated that PP5 is a critical target of palbociclib, distinct from CDK4/6, influencing radiosensitivity in NSCLC. Palbociclib enhanced radiotherapy susceptibility by inducing sustained DNA damage and AMPK activation. The subsequent cellular event is apoptosis rather than autophagy. Furthermore, the enhanced efficacy of combination therapy was counteracted by an AMPK inhibitor and PP5 activator, underscoring the importance of these pathways in mediating the response. Our findings provide compelling evidence that targeting PP5 can significantly enhance the therapeutic outcomes of radiotherapy in NSCLC. This research offers valuable insights into new combination therapy strategies, highlighting the potential of PP5 as a novel therapeutic target to overcome radioresistance.

SPIN1 accelerates tumorigenesis and confers radioresistance in non-small cell lung cancer by orchestrating the FOXO3a/FOXM1 axis

Despite the importance of radiation therapy as a nonsurgical treatment for non-small cell lung cancer (NSCLC), radiation resistance has always been a concern because of poor patient response and outcomes. Therefore, it is crucial to identify novel targets to increase the effectiveness of radiotherapy and investigate the mechanisms underlying radioresistance. Previously, we demonstrated that Spindlin 1 (SPIN1) was related to tumour initiation and progression. In this study, we found that SPIN1 expression was higher in NSCLC tissues and cell lines than in the corresponding controls. SPIN1 overexpression in NSCLC patients was closely correlated with disease progression and poor prognosis. Functionally, SPIN1 depletion inhibited cell proliferation, decreased the percentage of cells in the G2/M phase and suppressed cell migration and invasion. Moreover, SPIN1 knockdown decreased the clonogenic capacity, impaired double-strand break (DSB) repair and increased NSCLC radiosensitivity. Mechanistically, forkhead box M1 (FOXM1) was identified as a key downstream effector of SPIN1 in NSCLC cells. Furthermore, SPIN1 was found to facilitate MDM2-mediated FOXO3a ubiquitination and degradation, leading to FOXM1 upregulation. Moreover, restoration of FOXM1 expression markedly abolished the inhibitory effects and increased radiosensitivity induced by SPIN1 depletion. These results indicate that the SPIN1-MDM2-FOXO3a/FOXM1 signalling axis is essential for NSCLC progression and radioresistance and could serve as a therapeutic target for increasing radiotherapy efficacy.

RSPO3 regulates the radioresistance of Non-Small cell lung cancer cells via NLRP3 Inflammasome-Mediated pyroptosis

PurposeRadioresistance is a significant challenge in the radiotherapy of non-small cell lung cancer (NSCLC). This study aimed to investigate the role of R-spondin 3 (RSPO3) in regulating NSCLC radioresistance. Methods and MaterialsRNA sequencing was performed to analyze genes that are differentially expressed in radioresistant NSCLC cell lines. RSPO3 overexpression and knockdown experiments were conducted to assess its impact on radiosensitivity. The involvement of the β-catenin-NF-κB signaling pathway and the NLRP3 inflammasome in RSPO3-mediated radiosensitivity was also evaluated. In vivo experiments were conducted using a clinical-grade anti-RSPO3 antibody (OMP-131R10/rosmantuzumab) to assess its impact on radiation-induced pyroptosis and subsequent anti-tumor immunity. ResultsRSPO3 expression was downregulated in radioresistant NSCLC cells. Overexpression of RSPO3 increased NSCLC radiosensitivity through the induction of pyroptosis, which was mediated by the β-catenin-NF-κB signaling pathway and the NLRP3 inflammasome. The anti-RSPO3 antibody effectively blocked radiation-induced pyroptosis and anti-tumor immunity in vivo. Conversely, upregulation of RSPO3 enhanced NSCLC tumor radiosensitivity. ConclusionsThe findings demonstrated that RSPO3 plays a crucial role in regulating NSCLC radioresistance via NLRP3 mediated pyroptosis. Targeting the RSPO3-NLRP3 inflammasome axis may offer a potential therapeutic strategy to enhance the efficacy of radiotherapy for NSCLC patients.

Targeting Nrf2/PHKG2 axis to enhance radiosensitivity in NSCLC

While ferroptosis shows promise in anti-cancer strategy, the molecular mechanisms behind this process remain poorly understood. Our research aims to highlight the regulation of radiotherapy-induced ferroptosis in non-small cell lung cancer (NSCLC) via the NRF2/PHKG2 axis-mediated mechanism. To identify ferroptosis-associated genes associated with radioresistance in NSCLC, this study employed high-throughput transcriptome sequencing and Lasso risk regression analysis. Clinical samples were analyzed to confirm PHKG2 expression changes before and after radiotherapy. The study further examined ferritinophagy-related factors, intracellular iron levels, mitochondrial function, and ferroptosis in NSCLC cells undergoing radiation exposure to explore the effect of PHKG2 on radiosensitivity or radioresistance. The research also demonstrated the transcriptional inhibition of PHKG2 by NRF2 and created in situ transplantation tumor models of NSCLC to examine the role of NRF2/PHKG2 axis in NSCLC radiosensitivity and resistance in vivo. The Lasso risk regression model that incorporated ferroptosis-associated genes effectively predicted the prognosis of patients with NSCLC. Radiotherapy-sensitive tissues exhibited an increased expression of PHKG2. Overexpression of PHKG2 led to elevated intracellular iron levels by promoting ferritinophagy and increased mitochondrial stress-dependent ferroptosis induced by radiotherapy. PHKG2 transcription repression was achieved through NRF2. The FAGs-Lasso risk regression model can accurately predict the prognosis of NSCLC patients. Targeting Nrf2 upregulates the expression of PHKG2 and reverses radiotherapy resistance in NSCLC by promoting iron autophagy and inducing mitochondrial dysfunction, thereby increasing radiotherapy sensitivity.

Downregulation of MMP-9 by epicatechin can improve the radiosensitivity of non-small cell lung cancer.

Radiation therapy is a crucial treatment for nonsmall cell lung cancer (NSCLC), but its effectiveness is limited by the resistance of tumor cells to radiation. This study aimed to evaluate the effect of epicatechin (EC) on radiosensitivity in NSCLC and to determine its relationships with matrix metalloproteinase (MMP)-9. MMP-9 expression was detected by Western blotting, and the expression of the DNA damage marker protein was detected by immunofluorescence. Cell viability was assessed using the CCK-8 assay, and cell proliferation was evaluated using the clonogenesis assay. Flow cytometry was used to determine the cell apoptosis, whereas cell migration and invasion were detected using the transwell assays. The cells were treated with ionizing radiation (IR) and EC to verify the sensitizing effect of EC on radiation therapy. MMP-9 expression was elevated in the NSCLC cells and tissues. DNA damage and cell apoptosis were increased, whereas cell vigor, proliferation, migration, and invasion were significantly decreased after IR. MMP-9 knockdown strengthened the impact of IR on the biological behaviors of the cells. EC + IR had the best effect on promoting DNA damage and the biological behaviors of the NSCLC cells; alternatively, the overexpression of MMP-9 weakened the role of EC. This study shows that EC can downregulate MMP-9 expression, promote DNA damage, reduce cell viability, proliferation, migration, and invasion, and facilitate cell apoptosis, thus, showing potential as a radiosensitizer for NSCLC.

LILRB2 inhibition enhances radiation sensitivity in non-small cell lung cancer by attenuating radiation-induced senescence

Radiotherapy (RT) in non-small cell lung cancer (NSCLC) triggers cellular senescence, complicating tumor microenvironments and affecting treatment outcomes. This study examines the role of lymphocyte immunoglobulin-like receptor B2 (LILRB2) in modulating RT-induced senescence and radiosensitivity in NSCLC. Through methodologies including irradiation, lentivirus transfection, and various molecular assays, we assessed LILRB2’s expression and its impact on cellular senescence levels and tumor cell behaviors. Our findings reveal that RT upregulates LILRB2, facilitating senescence and a senescence-associated secretory phenotype (SASP), which in turn enhances tumor proliferation and resistance to radiation. Importantly, LILRB2 silencing attenuates these effects by inhibiting the JAK2/STAT3 pathway, significantly increasing radiosensitivity in NSCLC models. Clinical data correlate high LILRB2 expression with reduced RT response and poorer prognosis, suggesting LILRB2’s pivotal role in RT-induced senescence and its potential as a therapeutic target to improve NSCLC radiosensitivity.

Mogrol-mediated enhancement of radiotherapy sensitivity in non-small cell lung cancer: a mechanistic study.

This study investigated mogrol's impact on non-small cell lung cancer (NSCLC) radiosensitivity and underlying mechanisms, using various methods including assays, bioinformatics, and xenograft models. CCK-8, clonogenic, flow cytometry, TUNEL, and Western blot assays evaluated mogrol and radiation effects on NSCLC viability and apoptosis. Ubiquitin-specific protease 22 (USP22) expression in NSCLC patient tissues was determined by RT-qPCR and Western blot. A xenograft model validated mogrol's effects on tumor growth. Bioinformatics identified four ubiquitin-specific proteases, including USP22, in NSCLC. Kaplan-Meier analysis confirmed USP22's value in lung cancer survival. Human Protein Atlas (HPA) database analysis indicated higher USP22 expression in lung cancer tissues. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis implicated ERK1/2 in NSCLC progression, and molecular docking showed stability between mogrol and ERK1/2. Further in vivo and in vitro experiments have demonstrated that mogrol enhances the inhibitory effect of radiation on NSCLC cell viability and clonogenic capacity. Cell viability and clonogenic capacity are reduced by >50%, and an increase in cellular apoptosis is observed, with apoptotic levels reaching 10%. USP22 expression was significantly elevated in NSCLC tissues, particularly in radiotherapy-resistant patients. Mogrol downregulated USP22 expression by inhibiting the ERK/CREB pathway, lowering COX2 expression. Mogrol also enhanced radiation's inhibition of tumor growth in mice. Mogrol enhances NSCLC radiosensitivity by downregulating USP22 via the ERK/CREB pathway, leading to reduced COX2 expression.NEW & NOTEWORTHY Mogrol enhances non-small cell lung cancer (NSCLC) cell sensitivity to radiotherapy by downregulating USP22 through the ERK/CREB pathway, reducing COX2 expression. These findings highlight mogrol's potential as an adjunct to improve NSCLC radiotherapy and open avenues for further research and clinical applications.

Exosome miR-101–3p derived from bone marrow mesenchymal stem cells promotes radiotherapy sensitivity in non-small cell lung cancer by regulating DNA damage repair and autophagy levels through EZH2

Background and objectiveThe morbidity rate of non-small cell lung cancer (NSCLC) increases with age, highlighting that NSCLC is a serious threat to human health. The aim of this study was mainly to describe the role of exosomal miR-101–3p derived from bone marrow mesenchymal stem cells (BMSCs) in NSCLC. MethodsA549 or NCI-H1703 cells (1×105/mouse) were injected into nude mice to establish an NSCLC animal model. RTqPCR, Western blotting and comet assays were used to assess the changes in gene expression, proteins and DNA damage repair. ResultsmiR-101–3p and RAI2 were found to be expressed at low levels in NSCLC, while EZH2 was highly expressed. In terms of function, miR-101–3p downregulated EZH2. In addition, exosomal miR-101–3p derived from BMSCs promoted the expression of RAI2, inhibited DNA damage repair, and inhibited the activation of the PI3K/AKT/mTOR signaling pathway by inhibiting EZH2, thereby promoting autophagy and decreasing cell viability and finally enhancing the sensitivity of NSCLC to radiotherapy and inhibiting the malignant biological behavior of NSCLC. ConclusionExosomal miR-101–3p derived from BMSCs can inhibit DNA damage repair, promote autophagy, enhance the radiosensitivity of NSCLC, and inhibit the progression of NSCLC by inhibiting EZH2.

Silencing PinX1 enhances radiosensitivity and antitumor-immunity of radiotherapy in non-small cell lung cancer

BackgroundWe aimed to investigate the effects of PinX1 on non-small cell lung cancer(NSCLC) radiosensitivity and radiotherapy-associated tumor immune microenvironment and its mechanisms.MethodsThe effect of PinX1 silencing on radiosensitivity in NSCLC was assessed by colony formation and CCK8 assay, immunofluorescence detection of γ- H2AX and micronucleus assay. Western blot was used to assess the effect of PinX1 silencing on DNA damage repair pathway and cGAS-STING pathway. The nude mouse and Lewis lung cancer mouse model were used to assess the combined efficacy of PinX1 silencing and radiotherapy in vivo. Changes in the tumor immune microenvironment were assessed by flow cytometry for different treatment modalities in the Lewis luuse model. The interaction protein RBM10 was screened by immunoprecipitation-mass spectrometry.ResultsSilencing PinX1 enhanced radiosensitivity and activation of the cGAS-STING pathway while attenuating the DNA damage repair pathway. Silencing PinX1 further increases radiotherapy-stimulated CD8+ T cell infiltration and activation, enhances tumor control and improves survival in vivo; Moreover, PinX1 downregulation improves the anti-tumor efficacy of radioimmunotherapy, increases radioimmune-stimulated CD8+ T cell infiltration, and reprograms M2-type macrophages into M1-type macrophages in tumor tissues. The interaction of PinX1 and RBM10 may promote telomere maintenance by assisting telomerase localization to telomeres, thereby inhibiting the immunostimulatory effects of IR.ConclusionsIn NSCLC, silencing PinX1 significantly contributed to the radiosensitivity and promoted the efficacy of radioimmunotherapy. Mechanistically, PinX1 may regulate the transport of telomerase to telomeres through interacting with RBM10, which promotes telomere maintenance and DNA stabilization. Our findings reveal that PinX1 is a potential target to enhance the efficacy of radioimmunotherapy in NSCLC patients.

Differences in Radiosensitivity According to EGFR Mutation Status in Non-Small Cell Lung Cancer: A Clinical and In Vitro Study.

Unlike drug selection, radiation parameters (field, dose) are not based on driver gene mutations in patients with metastatic non-small cell lung cancer (NSCLC). This study aimed to compare radiosensitivity in NSCLC with and without EGFR driver gene mutations using clinical and in vitro data. The clinical study included 42 patients who underwent whole-brain radiotherapy for brain metastases from NSCLC; of these, 13 patients had EGFR mutation-positive tumors. The Kaplan-Meier method was used to calculate the cranial control rate without intracranial recurrence. In the in vitro study, colony formation and double-strand DNA breaks were examined in two EGFR mutation-negative and three EGFR mutation-positive NSCLC-derived cell lines. Colony formation was assessed 14 days after irradiation with 0 (control), 2, 4, or 8 Gy. DNA double-strand breaks were evaluated 0.5 and 24 h after irradiation. EGFR mutation-positive patients had a significantly better cranial control rates than EGFR mutation-negative patients (p = 0.021). EGFR mutation-positive cells formed significantly fewer colonies after irradiation with 2 or 4 Gy than EGFR mutation-negative cells (p = 0.002, respectively) and had significantly more DNA double-strand breaks at 24 h after irradiation (p < 0.001). Both clinical and in vitro data suggest that EGFR mutation-positive NSCLC is radiosensitive.

Inhibition of demethylase by IOX1 modulates chromatin accessibility to enhance NSCLC radiation sensitivity through attenuated PIF1

Chromatin accessibility is a critical determinant of gene transcriptional expression and regulated by histones modification. However, the potential for manipulating chromatin accessibility to regulate radiation sensitivity remains unclear. Our findings demonstrated that the histone demethylase inhibitor, 5-carboxy-8-hydroxyquinoline (IOX1), could enhance the radiosensitivity of non-small cell lung cancer (NSCLC) in vitro and in vivo. Mechanistically, IOX1 treatment reduced chromatin accessibility in the promoter region of DNA damage repair genes, leading to decreased DNA repair efficiency and elevated DNA damage induced by γ irradiation. Notably, IOX1 treatment significantly reduced both chromatin accessibility and the transcription of phytochrome interacting factor 1 (PIF1), a key player in telomere maintenance. Inhibition of PIF1 delayed radiation-induced DNA and telomeric DNA damage repair, as well as increased radiosensitivity of NSCLC in vitro and in vivo. Further study indicated that the above process was regulated by a reduction of transcription factor myc-associated zinc finger protein (MAZ) binding to the distal intergenic region of the PIF1. Taken together, IOX1-mediated demethylase inactivation reduced chromatin accessibility, leading to elevated telomere damage which is partly due to PIF1 inhibition, thereby enhancing NSCLC radiosensitivity.

LINC00921 reduces lung cancer radiosensitivity by destabilizing NUDT21 and driving aberrant MED23 alternative polyadenylation

Alternative polyadenylation (APA) plays a major role in controlling transcriptome diversity and therapeutic resistance of cancers. However, long non-coding RNAs (lncRNAs) involved in pathological APA remain poorly defined. Here, we functionally characterize LINC00921, a MED13L/P300-induced oncogenic lncRNA, and show that it is required for global regulation of APA in non-small cell lung cancer (NSCLC). LINC00921 shows significant potential for reducing NSCLC radiosensitivity, and high LINC00921 levels are associated with a poor prognosis for patients with NSCLC treated with radiotherapy. LINC00921 controls NUDT21 stability by facilitating binding of NUDT21 with the E3 ligase TRIP12. LINC00921-induced destabilization of NUDT21 promotes 3' UTR shortening of MED23 mRNA via APA, which, in turn, leads to elevated MED23 protein levels in cancer cells and nuclear translocation of β-catenin and thereby activates expression of multiple β-catenin/T cellfactor (TCF)/lymphoid enhancer-binding factor (LEF)-regulated core oncogenes (c-Myc, CCND1, and BMP4). These findings highlight the importance of functionally annotating lncRNAs controlling APA and suggest the clinical potential of therapeutics for advanced NSCLC.

LINC00921 Diminishes Lung Cancer Radiosensitivity by Bestabilizing NUDT21 and Driving Aberrant MED23 Alternative Polyadenylation

Alternative polyadenylation (APA) plays a major role in controlling transcriptome diversity and therapeutic resistance of cancers. However, long-noncoding RNAs (lncRNAs) involved in pathological APA remain poorly defined. Here, we functionally identified a MED13L/P300-induced oncogenic lncRNA, LINC00921, diminished lung cancer radiosensitivity by destabilizing NUDT21 and driving aberrant MED23 alternative polyadenylation. ChIP-seq screening, RNA-seq and real-time PCR were used to identified LINC00921 in NSCLC. We performed RNA pulldown, RIP-qPCR, western blotting and Co-immunoprecipitation to investigate the function of LINC00921, which induced destabilization of NUDT21 and promoted 3' UTR shortening of MED23 via APA. Through H3K27ac ChIP-seq screening, we functionally characterize LINC00921, a MED13L/P300-induced oncogenic lncRNA, required for global regulation of APA in non-small-cell lung cancer (NSCLC). LINC00921 shows significant potential for reducing radiosensitivity of NSCLC and high LINC00921 levels were associated with poor prognosis for NSCLC patients treated with radiotherapy. Mechanistically, LINC00921 directly interacts withNUDT21 via binding to its RNA-binding motif-2. LINC00921 controls NUDT21 stability via facilitating binding of NUDT21 with its newly identified E3 ligase TRIP12. Intriguingly, 3'UTR APA profiles reveal that LINC00921-induced destabilization of NUDT21 decreases the percentage of distal polyadenylation sites (PAS) usage index, resulting in the 3' UTR shortening of MED23 mRNA, which, in turn, leads to elevated MED23 protein levels in cancer cells. MED23 further increases nuclear translocation of β-catenin, and, thereby, activates expression of multiple β-Catenin/TCF/LEF-regulated core oncogenes (c-Myc, CCND1, and BMP4). Taken together, our data revealed a novel model that integrates a lncRNA into regulation of malignant APA, radiotherapy resistance and NSCLC progression. These findings highlight the importance of functionally annotating lncRNAs controlling APA and unlock the clinical potential of novel therapeutics for advanced NSCLC.

GALNT2, an O-glycosylating enzyme, is a critical regulator of radioresistance of non-small cell lung cancer: evidence from an integrated multi-omics analysis.

Radioresistance is the primary reason for radiotherapy failure in non-small cell lung cancer (NSCLC) patients. Glycosylation-related alterations are critically involved in tumor radioresistance. However, the relationship between glycosylation and NSCLC radioresistance is unclear. Here, we generated radioresistant NSCLC cell models by using fractionated irradiation. The aberrant glycosylation involved in NSCLC-related radioresistance was elucidated by transcriptomic, proteomic, and glycomic analyses. We conducted in vitro and in vivo investigations for determining the biological functions of glycosylation. Additionally, its downstream pathways and upstream regulators were inferred and verified. We demonstrated that mucin-type O-glycosylation and the O-glycosylating enzyme GALNT2 were highly expressed in radioresistant NSCLC cells. GALNT2 was found to be elevated in NSCLC tissues; this elevated level showed a remarkable association with response to radiotherapy treatment as well as overall survival. Functional experiments showed that GALNT2 knockdown improved NSCLC radiosensitivity via inducing apoptosis. By using a lectin pull-down system, we revealed that mucin-type O-glycans on IGF1R were modified by GALNT2 and that IGF1R could affect the expression of apoptosis-related genes. Moreover, GALNT2 knockdown-mediated in vitro radiosensitization was enhanced by IGF1R inhibition. According to a miRNA array analysis and a luciferase reporter assay, miR-30a-5p negatively modulated GALNT2. In summary, our findings established GALNT2 as a key contributor to the radioresistance of NSCLC. Therefore, targeting GALNT2 may be a promising therapeutic strategy for NSCLC.

Ce6/PTX2-NP/G@NHs Confer Radiosensitivity in Non-Small Cell Lung Cancer via Promotion of Apoptotic Body-Mediated Neighboring Effects.

This study fabricates a nanoparticle delivery system of gold nanoparticles-dextran nanoparticles loaded with hypoxia-activated paclitaxel dimeric prodrug nanoparticles (PTX2-NP) and photosensitizer chlorin e6/paclitaxel-nanoparticle/gold@N-(2-hydroxypropyl) (Ce6/PTX2-NP/G@NHs) and analyzed the possible molecular mechanism for enhancing the radiosensitivity of non-small cell lung cancer (NSCLC). Ce6/PTX2-NP/G@NHs were prepared by a coupling reaction and dextran inclusion, followed by characterization using spectroscopy techniques. The cellular uptake and cytotoxicity of Ce6/PTX2-NP/G@NHs were analyzed. Radiosensitizing effects of the nanoparticles were evaluated by determining the malignant phenotypes and reactive oxygen species production of A549 cells and PI3K/AKT pathway-related proteins under 685 nm laser irradiation. A549 tumor-bearing nude mice were modeled to further confirm the radiosensitizing effect. Ce6/PTX2-NP/G@NHs were effectively internalized by A549 cells, producing cytotoxicity under laser irradiation. Ce6/PTX2-NP/G@NHs reduced cell viability, clonogenic potential, migration, and invasion along with reactive oxygen species (ROS) production while promoting apoptosis in A549 cells under laser irradiation. By inhibiting the PI3K/AKT pathway, Ce6/PTX2-NP/G@NHs increased the sensitivity of A549 cells to radiotherapy where apoptotic body (ApoBD)-mediated neighboring effects also played a key role. Ce6/PTX2-NP/G@NHs accumulated in tumor sites of nude mice and enhanced the radiosensitivity of NSCLC. Ce6/PTX2-NP/G@NHs showed no obvious toxicity or side effects in vivo. Collectively, the new Ce6/PTX2-NP/G@NHs nanoparticle delivery system can enhance the radiosensitivity of NSCLC via the promotion of ApoBD-mediated neighboring effects and inactivation of the PI3K/AKT pathway.

Abstract 2811: CDK4/6 inhibition plus radiotherapy enhances anti-tumor immunity in non-small cell lung cancer

Abstract Lung cancer is the main cause of cancer-related mortality, of which non-small cell lung cancer (NSCLC) accounts for 85%. Abemaciclib, a selective CDK4/6 inhibitor, enhanced the radiosensitivity of NSCLC in vivo and in vitro and improved the prognosis of advanced NSCLC patients according to a Phase I clinical trial. This study aimed to explore the impact of CDK4/6 inhibition combined with irradiation and immunotherapy on NSCLC. The in vivo xenograft tumor mouse model was used to investigate the synergistic effects of CDK4/6 inhibitor abemaciclib and irradiation together with anti-PD-1 antibodies. The tumor infiltrating lymphocytes (TILs) in tumor microenvironment (TME) were analyzed by flow cytometry. The complete blood count and serum tests were used to determine the safety of this combination therapy. The human phospho-kinase array and dual-luciferase report assay were used to study the regulatory mechanisms of abemaciclib and irradiation combination on PD-L1 transcription in NSCLC cells in vitro. Our results indicated the significantly synergistic effects of abemaciclib, irradiation and anti-PD-1 antibodies on NSCLC growth in vivo, accompanied by increased CD8+ T cell infiltration and decreased regulatory T cells and myeloid-derived suppressor cells (MDSCs), suggesting a strong anti-tumor combination of CDK4/6 inhibition, radiotherapy and immunotherapy. This synergistic effect could be partially impaired by CD8 depletion, and alleviated the anemia and inflammation caused by tumor burden without hepatorenal toxicity. Abemaciclib induced PD-L1 transcription through JNK/c-Jun pathway in vitro, which might elevate PD-L1 expression synergistically with irradiation through TBK1/IRF3 pathway. Our studies suggested that CDK4/6 inhibition plus irradiation enhanced anti-tumor immune responses in NSCLC. Together with immunotherapy, the combination of CDK4/6 inhibitors and radiotherapy could lower the doses of individual treatment, thus alleviating the potential toxicity and side effects of CDK4/6 inhibition and irradiation on normal cells. Citation Format: Zhengrong Huang, Jiang Luo, Mengqin Wang, Hongxin Xie, Yuxin Zeng, Yu Yuan, Wan Xiang, Ziyu Jiang, Conghua Xie, Yan Gong. CDK4/6 inhibition plus radiotherapy enhances anti-tumor immunity in non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2811.

Radiosensitivity in Non-Small-Cell Lung Cancer by MMP10 through the DNA Damage Repair Pathway.

NSCLC (non-small-cell lung cancer) is an aggressive form of lung cancer and accompanies high morbidity and mortality. This study investigated the function and associated mechanism of MMP10 during radiotherapy of NSCLC. MMP10 expression in patients and their overall survival rate were assessed through GEPIA. Protein expression was tested by western blotting. Radioresistance was detected in vitro by apoptosis and clonogenic assay. The extent of DNA damage and repair was revealed by the comet test and γH2AX foci test. High MMP10 levels in specimens of lung adenocarcinoma were related to poor patient outcomes. Clonogenic and apoptosis assays revealed that MMP10 knockdown in A549 cells initiated radiosensitization. Furthermore, MMP10 siRNA increased damage to the DNA in NSCLC cells, while MMP10 was observed to participate in DNA damage repair post-ionizing radiation. Thus, after irradiation, MMP10 plays an essential role in NSCLC through the repair pathway of DNA damage; regulating MMP10 for NSCLC radiosensitivity might have potential treatment implications in radiotherapy of NSCLC.