Search

Purpose: The hyperactivation of epidermal growth factor receptor (EGFR) plays a crucial role in non-small cell lung cancer (NSCLC). Hedgehog (Hh) signaling has been implicated in the tumorigenesis and progression of various cancers, however, its function in NSCLC cells remains controversial. Herein, we present a novel finding that challenges the current understanding of Hh signaling in tumor growth., Methods: Expression of Hh ligands and receptor were assessed using TCGA datasets, immunoblotting and immunohistochemical. Biological function of Hh ligands and receptor in NSCLC were tested using colony formation, cell count kit-8 (CCK-8) and xenograft assays. Biochemical effect of Hh ligands and receptor on regulating EGFR stability and activity were checked via immunoblotting., Results: Expression of Hh ligands and receptor was suppressed in NSCLC tissues, and the lower expression levels of these genes were associated with poor prognosis. Ptch1 binds to EGFR and facilitates its poly-ubiquitylation and degradation independent of downstream transcriptional signaling. Moreover, Hh ligands cooperate with Ptch1 to regulate the protein stability and activity of EGFR. This unique mechanism leads to a suppressive effect on NSCLC tumor growth., Conclusion: Non-canonical Hh signaling pathway, involving cooperation between Hh ligands and their receptor Ptch1, facilitates the degradation of EGFR and attenuates its activity in NSCLC. These findings provide novel insights into the regulation of EGFR protein stability and activity, offer new diagnostic indicators for molecular typing of NSCLC and identify potential targets for targeted therapy of this challenging disease., (© 2024. Springer Nature Switzerland AG.)

In brain metastases, radiation necrosis (RN) is a complication that arises after single or multiple fractionated stereotactic radiosurgery (SRS/FSRS), which is challenging to distinguish from local recurrence (LR). Studies have shown increased RN incidence rates in non-small cell lung cancer (NSCLC) patients with oncogenic driver mutations (ODMs) or receiving tyrosine kinase inhibitors (TKIs). This study investigated enlarging brain lesions following SRS/FSRS, for which additional surgeries were performed to distinguish between RN and LR. We investigated seven NSCLC patients with ODMs undergoing SRS/FSRS for BM and undergoing surgery for suspicion of LR on MRI imaging. Descriptive statistics were performed. Among the seven patients, six were EGFR+, while one was ALK+. The median irradiation dose was 30 Gy (range, 20-35 Gy). The median time to develop RN after SRS/FSRS was 11.1 months (range: 6.3-31.2 months). Moreover, gradually enlarging lesions were found in all patients after 6 months post-SRS/FSR. Brain radiation necrosis was pathologically confirmed in all the patients. RN should be suspected in NSCLC patients when lesions keep enlarging after 6 months post-SRS/FSRS, especially for patients with ODMs and receiving TKIs. Further, this case series indicates that further dose reduction might be necessary to avoid RN for such patients., (© 2024 Scandinavian Societies for Pathology, Medical Microbiology and Immunology.)

Several generations of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors have been developed for the treatment of non-small cell lung cancer (NSCLC) in clinic. However, emerging drug resistance mediated by new EGFR mutations or activations by pass, leads to malignant progression of NSCLC. Proteolysis targeting chimeras (PROTACs) have been utilized to overcome the drug resistance acquired by mutant EGFR, newly potent and selective degraders are still need to be developed for clinical applications. Herein, we developed autophagosome-tethering compounds (ATTECs) in which EGFR can be anchored to microtubule-associated protein-1 light chain-3B (LC3B) on the autophagosome with the assistance of the LC3 ligand GW5074. A series of EGFR-ATTECs have been designed and synthesized. Biological evaluations showed that these compounds could degrade EGFR and exhibited moderate inhibitory effects on certain NSCLC cell lines. The ATTEC 12c potently induced the degradation of EGFR with a DC 50 value of 0.98 μM and a Dmax value of 81% in HCC827 cells. Mechanistic exploration revealed that the lysosomal pathway was mainly involved in this degradation. Compound 12c also exhibited promising inhibitory activity, as well as degradation efficiency in vivo. Our study highlights that EGFR-ATTECs could be developed as a new expandable EGFR degradation tool and also reveals a novel potential therapeutic strategy to prevent drug resistance acquired EGFR mutations., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Background: The effective therapeutic approach is still an unmet need for patients diagnosed with both lung cancer and interstitial lung disease (ILD). This is primarily due to the possible risk of ILD exacerbation caused by surgery or radiotherapy. The current study aimed to investigate the efficacy and safety of local ablative therapy (LAT) for this specific population., Methods: Consecutive patients with non-small cell lung cancer (NSCLC) and ILD who received LAT between January 2018 and August 2022 were enrolled, and propensity score matching (PSM) was utilized to match the non-ILD group. The primary endpoint was recurrence-free survival (RFS), and secondary endpoints included overall survival (OS), adverse events (AEs) and hospital length of stay (HLOS)., Results: The PSM algorithm yielded matched pairs in the ILD group (n = 25) and non-ILD group (n = 72) at a ratio of 1:3. There were no statistically significant differences in RFS (median 16.4 vs. 18 months; HR = 1.452, p = 0.259) and OS (median: not reached vs. 47.9 months; HR = 1.096, p = 0.884) between the two groups. Meanwhile, no acute exacerbation of ILD was observed in the ILD group. However, the incidence of pneumothorax, especially pneumothorax requiring chest tube drainage, was significantly higher (36.0% vs. 11.2%, p = 0.005) among patients with NSCLC and co-existing ILD, which resulted in longer HLOS (p = 0.045)., Conclusion: Although ILD was associated with a higher incidence of pneumothorax, the efficacy of LAT for NSCLC patients with ILD was comparable to those without ILD, suggesting that LAT might be a reliable and effective treatment option for this population, particularly in the early stage., (© 2024 The Authors. Thoracic Cancer published by John Wiley & Sons Australia, Ltd.)

Purpose: To investigate the correlation of air bronchogram sign with clinicopathological characteristics and prognosis in patients with clinical stage (c-stage) I non-small cell lung cancer (NSCLC) with radiological pure-solid appearance., Method: We retrospectively evaluated 276 patients with pure-solid c-stage I NSCLC and assessed the correlation between the air bronchogram and clinicopathological characteristics. A Cox proportional hazards model was performed to identify the effect of air bronchogram and clinicopathological variables on oncological outcomes. Recurrence-free survival (RFS) and overall survival (OS) were calculated by Kaplan-Meier curves and were compared using log-rank tests., Results: Presence of air bronchogram was associated with a well differentiated degree (P =.026), higher incidence of EGFR mutation (P <.001) and lower recurrence(P =.021). Kaplan-Meier survival curves showed that air bronchogram group was associated with favorable RFS(67.0% vs. 50.2%; P =.015). A multivariable analysis revealed that air bronchogram and EGFR mutation were independent significant prognostic factors associated with RFS (hazard ratio [HR] = 0.495, 95% confidence interval [CI]: 0.322-0.761, P =.001; HR = 1.625, 95% CI: 1.074-2.457, P =.021; respectively), but not with OS. Additionally, we found that pathological lymph node metastasis was identified as an independent prognostic factor associated with poor RFS and OS(HR = 2.808, 95% CI: 1.913-4.123, P <.001 for RFS; HR = 1.983, 95% CI: 1.185-3.318, P =.009 for OS)., Conclusions: Presence of air bronchogram was associated with well differentiated degree, higher incidence of EGFR mutation and had additional positive prognostic value for RFS in c-stage I NSCLC with a radiological pure-solid appearance., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

This study aims to investigate the predictive value of preoperative whole-tumor histogram analysis of multi-parametric MRI for histological subtypes in patients with lung cancer brain metastases (BMs) and explore the correlation between histogram parameters and Ki-67 proliferation index. The preoperative MRI data of 95 lung cancer BM lesions obtained from 73 patients (42 men and 31 women) were retrospectively analyzed. Multi-parametric MRI histogram was used to distinguish small-cell lung cancer (SCLC) from non-small cell lung cancer (NSCLC), and adenocarcinoma (AC) from squamous cell carcinoma (SCC), respectively. The T1-weighted contrast-enhanced (T1C) and apparent diffusion coefficient (ADC) histogram parameters of the volumes of interest (VOIs) in all BMs lesions were extracted using FireVoxel software. The following histogram parameters were obtained: maximum, minimum, mean, standard deviation (SD), variance, coefficient of variation (CV), skewness, kurtosis, entropy, and 1st-99th percentiles. Then investigated their relationship with the Ki-67 proliferation index. The skewness-T1C, kurtosis-T1C, minimum-ADC, mean-ADC, CV-ADC and 1st - 90th ADC percentiles were significantly different between the SCLC and NSCLC groups (all p < 0.05). When the 10th-ADC percentile was 668, the sensitivity, specificity, and accuracy (90.80%, 76.70% and 86.32%, respectively) for distinguishing SCLC from NSCLC reached their maximum values, with an AUC of 0.895 (0.824 - 0.966). Mean-T1C, CV-T1C, skewness-T1C, 1st - 50th T1C percentiles, maximum-ADC, SD-ADC, variance-ADC and 75th - 99th ADC percentiles were significantly different between the AC and SCC groups (all p < 0.05). When the CV-T1C percentiles was 3.13, the sensitivity, specificity and accuracy (75.00%, 75.60% and 75.38%, respectively) for distinguishing AC and SCC reached their maximum values, with an AUC of 0.829 (0.728-0.929). The 5th-ADC and 10th-ADC percentiles were strongly correlated with the Ki-67 proliferation index in BMs. Multi-parametric MRI histogram parameters can be used to identify the histological subtypes of lung cancer BMs and predict the Ki-67 proliferation index., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: Lung cancer (LC) is one of the most frequent cancers worldwide, as well as the leading cause of cancer-related death. Non-small cell lung cancer (NSCLC, which accounts for 85% of occurrences) is the main type of LC. MiRNAs appear to play a role in the occurrence and progression of many malignancies, according to mounting data. The underlying mechanism of miRNAs in regulating NSCLC cell biological activity and progression, on the other hand, is still being investigated., Methods: QRT-PCR were used to detect miR-185-5p expression and YWHAZ mRNA in NSCLC. The CCK-8 assay was used to determine the tumor cells' ability to proliferate. Transwall assay was used to test the migratory and invasive properties of cells. Cell apoptosis was detected using flow cytometry. Tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein zeta (YWHAZ), E-Cadherin, N-Cadherin and cleaved-caspase3 protein expression were assessed using Western Blot. The bioinformatics analysis software StarBase2.0 predicted miR-185-5p downstream targets. To confirm the target association between miR-185-5p and YWHAZ, a luciferase experiment was used. In addition, an NCl-H1299 xenograft model was created to assess the anti-tumor impact of miR-185-5p in vivo. The expression level of YWHAZ in tumor tissues of small xenograft tumor model was detected by immunohistochemistry assay., Results: Decreased miR-185-5p expression levels were observed in NSCLC. In vitro, over-expressed miR-185-5p decreased cell viability, proliferation, invasion/migration, and induced cell apoptosis, while inhibiting tumor growth in vivo. Dual-luciferase gene experiments confirmed that YWHAZ binds to miR-185-5p. Overexpression of YWHAZ partially restored the inhibitory effects of miR-185-5p on cell behaviors., Conclusion: MiR-185-5p was down-regulated in NSCLC, and that overexpressed miR-185-5p inhibited malignant behaviors of cells and tumor growth by negatively regulating YWHAZ., (© 2023. The Author(s).)

Anaplastic lymphoma kinase (ALK) rearrangements occur in ∼3%-6% of patients with advanced non-small-cell lung cancer (NSCLC). Small molecular drugs that effectively inhibit ALK gene have revolutionized the therapeutic paradigm for patients with ALK rearrangements, resulting in significant improvements in objective response rate, progression-free survival, and overall survival compared with classical platinum-based chemotherapy. Several ALK tyrosine kinase inhibitors (ALK-TKIs), including crizotinib, alectinib, ceritinib, brigatinib, ensartinib, and lorlatinib, have been recommended as standard first-line treatment for advanced NSCLC patients with ALK rearrangements. Patients with ALK rearrangements typically exhibit long-term durable responses to ALK-TKIs; therefore, the management of adverse drug reactions (ADRs) with ALK-TKIs is crucial in clinical practice to maximize clinical benefits, prevent an adverse impact on quality of life, and improve patient compliance. In general, ALK-TKIs are well tolerated. There are, however, a number of serious toxicities that may necessitate dose modification or even discontinuation of treatment and the management of ADRs with ALK-TKIs has grown in importance. The therapeutic use of this class of medications still carries some risk because there are currently no pertinent guidelines or consensus recommendations for managing ADRs caused by ALK-TKIs in China. In order to improve the clinical management of ADRs with ALK-TKIs, the Chinese Society of Clinical Oncology (CSCO) Non-small Cell Lung Cancer Professional Committee led the discussion and summary of the incidence, diagnosis and grading standards, and prevention and treatment of ADRs caused by ALK-TKIs., Competing Interests: Disclosure The authors have declared no conflicts of interest., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Immune checkpoint inhibitors (ICI)-based combination strategies have improved the survival outcomes in advanced non-small cell lung cancers; however, data regarding their efficacy remains limited for uncommon histological types, including large-cell carcinoma (LCC) and large-cell neuroendocrine carcinoma (LCNEC)., Methods: We retrospectively analyzed a total of 60 patients with advanced LCC and LCNEC - 37 treatment-naïve and 23 pre-treated - who received pembrolizumab with or without chemotherapy. Treatment and survival outcomes were analyzed., Results: Of the 37 treatment-naïve patients who received first-line pembrolizumab combined with chemotherapy, the 27 patients with LCC had an overall response rate (ORR) of 44.4% (12/27) and a disease control rate (DCR) of 88.9% (24/27); whereas 10 patients with LCNEC had an ORR of 70% (7/10) and DCR of 90% (9/10). The median progression-free survival (mPFS) was 7.0 months (95% confidence intervals [CI]: 2.2-11.8) and median overall survival (mOS) was 24.0 months (95%CI: 0.0-50.1) for first-line pembrolizumab plus chemotherapy of LCC (n = 27), whereas mPFS was 5.5 months (95%CI: 2.3-8.7) and mOS was 13.0 months (95%CI: 11.0-15.0) for first-line pembrolizumab plus chemotherapy of LCNEC (n = 10). Of the 23 pre-treated patients who received subsequent-line pembrolizumab with or without chemotherapy, mPFS was 2.0 months (95% CI: 0.6-3.4) and mOS was 4.5 months (95% CI: 0.0-9.0) for LCC and mPFS was 3.8 months (95% CI: 0.0-7.6) and mOS was not reached for LCNEC., Conclusion: Our study provides real-world clinical evidence of the anti-tumor activity of pembrolizumab plus chemotherapy in advanced LCC and LCNEC, indicating that this regimen could serve as a treatment option, particularly as first-line therapy, for improving the survival outcomes of patients with these rare histological subtypes of lung cancer., Trial Registration: NCT05023837(ESPORTA, 27/08/2021)., (© 2023. The Author(s).)

Lung cancer is the most common cause of cancer related deaths worldwide with the highest mortality rate. Non-small cell lung cancer (NSCLC) accounts for about 85 % of lung cancers. Mitochondrial methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) is a bifunctional enzyme and is the most differentially expressed metabolic enzyme in various tumors including lung cancer. However, little is known about how MTHFD2 functions in NSCLC. Integrin-linked kinase (ILK) signaling plays key a role in tumor progression including metastasis, proliferation and migration. Here, we show that MTHFD2 inhibition results in suppression of cell growth, migration, invasion and epithelial-mesenchymal transition (EMT) in NSCLC. Microarray analysis suggests that MTHFD2 is positively associated with ILK signaling based on western blotting results. In addition, the phosphorylation of AMPKα plays an essential role in MTHFD2 regulation of ILK signaling. Further, the small-molecule compound C18 inhibits MTHFD2 with great efficiency. C18 blocks MTHFD2/ILK signaling pathway and restrains cell growth, migration, invasion, and EMT of NSCLC and induces apoptosis. In brief, our study found that the positive impact of MTHFD2 is mediated via ILK signaling pathway in NSCLC. Thus, blocking MTHFD2 represents a promising therapeutic strategy against NSCLC clinically., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Background: Lung cancer is frequently accompanied by interstitial lung disease (ILD), and the overall survival (OS) of patients with these comorbidities is poor. Thus, we developed a nomogram for the prediction of the OS of patients with advanced non-small cell lung cancer (NSCLC) and ILD., Patients and Methods: Patients with wild-type gene advanced NSCLC with and without ILD who underwent chemotherapy between 2014 and 2019 were enrolled in the present study. The 0.5- and 1-year progression-free survival (PFS) and overall survival (OS) times of patients with and without ILD were determined using the Kaplan-Meier method. Cox regression was used to assess the prognostic value of clinical factors for patients with ILD. Based on the multivariate regression results, a nomogram for survival prediction was developed. The nomogram was validated using calibration curve., Results: Data from 155 patients with lung cancer and ILD and 118 matched patients with lung cancer alone who were receiving first-line chemotherapy were analyzed. The first-line chemotherapy regimens were paclitaxel + carboplatin, pemetrexed + carboplatin, gemcitabine + carboplatin, and other. The median PFS and OS were significantly shorter in patients with than in those without ILD (3.0 vs. 7.0 months [p < 0.001] and 7.0 vs. 15.0 months (p < 0.001), respectively). Multivariate analysis showed that the lymphocyte count (hazard ratio [HR] 2.38; 95% confidence interval [CI], 1.44-3.94; p = 0.01), partial pressure of oxygen (PaO 2 ; HR, 1.37; 95% CI, 1.03-1.82; p = 0.03), and chemotherapy regimen were independently associated with prognosis. The nomogram showed good discriminatory ability [C-index = 0.69 (95% CI, 0.49-0.82)]. Calibration curves showed that predicted and actual prognoses were consistent., Conclusion: This nomogram can aid the prediction of the OS of patients with advanced NSCLC and ILD., (© 2023 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Lung cancer is the most frequent type of cancer affecting both men and women globally, and it is associated with a high mortality rate. It is clinically treated with cisplatin, a platinum-based drug that works by generating DNA lesions, which activates DNA damage response and induces cell death. However, chemoresistance by cancer cells limits the clinical usefulness of cisplatin as an anticancer drug. Here, we uncovered a role of ubiquitin-specific protease 51 (USP51) in the chemosensitivity of lung cancer cells to cisplatin by regulating DNA damage response. USP51 was more upregulated in lung cancer tissues of chemotherapy-resistant patients than those of chemotherapy-sensitive patients with adjacent, nontumor tissues. USP51 overexpression in lung cancer cells in vitro reduced γ-H2AX formation and promoted checkpoint kinase 1 (CHK1) phosphorylation, whereas USP51 knockdown showed opposite effects, indicating that USP51 played an important role in promoting DNA damage repair. Finally, USP51 knockdown weakened cisplatin resistance in A549/DDP cells and significantly suppressed tumor growth in vivo, suggesting that a USP51 inhibitor combined with cisplatin may be considered as an effective treatment strategy to eliminate drug-resistant lung cancer cells., (© 2022 International Union of Biochemistry and Molecular Biology, Inc.)

Non-small cell lung cancer (NSCLC) is the predominant form of lung cancer and is one of the most fatal cancers worldwide. Recently, the International Association for the Study of Lung Cancer (IASLC) proposed a novel grading system based on the predominant and high-grade histological patterns for invasive pulmonary adenocarcinoma (IPA). To improve outcomes for NSCLC patients, we combined serum metabolomics and fecal microbiology to screen biomarkers in patients with early-stage NSCLC and identified characteristic microbial profiles in patients with different grades of IPA. 26 genera and 123 metabolites were significantly altered in the early-stage NSCLC patients. Agathobacter , Blautia , Clostridium , and Muribaculacea were more abundant in the early-stage NSCLC patients compared with healthy controls. For the different grades of IPA, the characteristic microorganisms are as follows: Blautia and Marinobacter in IPA grade type 1; Dorea in IPA grade type 2; and Agathobacter in IPA grade type 3. In the metabolome results, the early-stage NSCLC group mainly included higher levels of sphingolipids (D-erythro-sphingosine 1-phosphate, palmitoyl sphingomyelin), fatty acyl (Avocadyne 1-acetate, 12(S)-HETE, 20-Carboxy-Leukotriene B4, Thromboxane B3, 6-Keto-prostaglandin f1alpha, Sebacic acid, Tetradecanedioic acid) and glycerophospholipids (LPC 20:2, LPC 18:0, LPC 18:4, LPE 20:2, LPC 20:1, LPC 16:1, LPC 20:0, LPA 18:2, LPC 17:1, LPC 17:2, LPC 19:0). Dysregulation of pathways, such as sphingolipid metabolism and sphingolipid signaling pathway may become an emerging therapeutic strategy for early-NSCLC. Correlation analysis showed that gut microbiota and serum metabolic profiles were closely related, while Muribaculacea and Clostridium were the core genera. These findings provide new biomarkers for the diagnosis of early-stage NSCLC and the precise grading assessment of prognostic-related IPAs, which are of clinical importance and warrant further investigation of the underlying molecular mechanisms., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Ni, Kong, Yan, Fu, Zhou and Xu.)

Immune checkpoint inhibitors (ICIs), which represent the new standard of care for advanced nonsmall cell lung cancer (NCSLC), are not effective in many patients. Biomarkers are needed to guide treatment. Sequencing data of an ICI-treated cohort were analyzed to identify genomic signatures predicting ICI efficacy, followed by validation using multiple independent cohorts. Their predictive mechanism was explored by evaluating the tumor immune microenvironment and tumor mutational burden (TMB). In the discovery cohort, patients carrying FGFR4 alterations (FGFR4 altered ) had a better objective response rate (ORR) (50.0% vs 19.4%; P = .057) and improved median progression-free survival (mPFS) (13.17 vs 3.17 months; HR 0.37; 95% CI 0.14-1; P = .04) than wild-type patients (FGFR4 wt ). In the publicly available validation cohorts, FGFR4 alterations correlated with higher ORR (100% vs 31%; P = .028), longer median overall survival (mOS) (not reached [NR] vs 11 months; HR 0.28, 95% CI 0.09-0.89, P = .02), and mPFS (NR vs 6.07 months; HR 0.05, 95% CI 0-3.94, P = .039). FGFR4 alterations were confirmed as an independent predictor of superior PFS (P = .014) and OS (P = .005). FGFR4 altered patients also exhibited a significantly improved disease control rate (100% vs 60%, P = .045) and prolonged mPFS (9.70 vs 3.16 months; P = .095) compared to FGFR4 wt patients in our Shanghai Pulmonary Hospital cohort. FGFR4 alterations associated with a higher TMB levels, more CD8 + T cells in the tumor stroma, and a higher M1/M2 ratio for tumor-associated macrophages in the tumor center and stroma. Thus, FGFR4 alterations may serve as a potential independent predictor of ICI efficacy in NSCLC., (© 2022 UICC.)

Background: Anti-PD-1(L1) therapies are less efficacious in patients with EGFR -mutated non-small-cell lung cancer. However, the underlying mechanism is poorly understood., Methods: The characteristics of T cells in EGFR -mutated and wild-type tumors were analyzed based on The Cancer Genome Atlas database and clinical samples. Plasma levels of 8 T-cell-related cytokines were evaluated and its association with immunotherapy efficacy were explored. Association between EGFR signaling pathway and IL-10 was examined through tumor cell lines and clinical tumor samples. In vitro restimulation model of human CD8 + T cells isolated from peripheral blood was used to analyze the impact of IL-10 on T cells. Doxycycline-inducible transgenic EGFR L858R mouse models were used to investigate the efficacy of combining recombinant mouse IL-10 protein and PD-1 blockade and its underlying mechanism in vivo ., Results: EGFR -mutated tumors showed a lack of CD8 + T cell infiltration and impaired CD8 + T cell cytotoxic function. The incompetent CD8 + T cells in EGFR -mutated tumors were characterized as absence of CD39 expression, which defined hallmarks of cytotoxic and exhausted features and could not be reinvigorated by anti-PD-1(L1) treatment. Instead, CD39 expression defined functional states of CD8 + T cells and was associated with the therapeutic response of anti-PD-1(L1) therapies. Mechanically, IL-10 upregulated CD39 expression and was limited in EGFR -mutated tumors. IL-10 induced hallmarks of CD8 + T cells immunity in CD39-dependent manner. Using autochthonous EGFR L858R -driven lung cancer mouse models, combining recombinant mouse IL-10 protein and PD-1 blockade optimized antitumor effects in EGFR -mutated lung tumors., Conclusions: Our study suggested that owing to low level of IL-10 to induce the expression of CD39 on CD8 + T cells, fewer phenotypically cytotoxic and exhausted CD39 + CD8 + T cells in EGFR -mutated tumors could be potentially reinvigorated by anti-PD-1(L1) treatment. Hence, IL-10 could potentially serve as a cytokine-based strategy to enhance efficacy of anti-PD-1(L1) treatment in EGFR -mutated tumors., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Immune checkpoint blockade therapy (ICB) is ineffective against cold tumors and, although it is effective against some hot tumors, drug resistance can occur. We have developed a Plasmodium immunotherapy (PI) that can overcome these shortcomings. However, the specific killing effect of PI on tumor cells is relatively weak. Radiotherapy (RT) is known to have strong specific lethality to tumor cells. Therefore, we hypothesized that PI combined with RT could produce synergistic antitumor effects. We tested our hypothesis using orthotopic and subcutaneous models of mouse glioma (GL261, a cold tumor) and a subcutaneous model of mouse non-small cell lung cancer (NSCLC, LLC, a hot tumor). Our results showed that, compared with each monotherapy, the combination therapy more significantly inhibited tumor growth and extended the life span of tumor-bearing mice. More importantly, the combination therapy could cure approximately 70 percent of glioma. By analyzing the immune profile of the tumor tissues, we found that the combination therapy was more effective in upregulating the perforin-expressing effector CD8 + T cells and downregulating the myeloid-derived suppressor cells (MDSCs), and was thus more effective in the treatment of cancer. The clinical transformation of PI combined with RT in the treatment of solid tumors, especially glioma, is worthy of expectation.

Acquired drug resistance and epithelial-mesenchymal transition (EMT) mediated metastasis are two highly interacting determinants for non-small-cell lung cancer (NSCLC) prognosis. This study investigated the common mechanisms of drug resistance and EMT from the perspective of metabolic reprogramming, which may offer new ideas to improve anticancer therapy. Acquired resistant cells were found to grow faster and have a greater migratory and invasive capacity than their parent cells. Metabolomics analysis revealed that acquired resistant cells highly relied on glutamine utilization and mainly fluxed into oxidative phosphorylation energy production. Further mechanistic studies screened out glutamate dehydrogenase 1 (GLUD1) as the determinant of glutamine addiction in acquired resistant NSCLC cells, and provided evidence that GLUD1-mediated α-KG production and the accompanying reactive oxygen species (ROS) accumulation primarily triggered migration and invasion by inducing Snail. Pharmacological and genetic interference with GLUD1 in vitro significantly reversed drug resistance and decreased cell migration and invasion capability. Lastly, the successful application of R162, a selective GLUD1 inhibitor, to overcome both acquired resistance and EMT-induced metastasis in vivo, identified GLUD1 as a promising and druggable therapeutic target for malignant progression of NSCLC. Collectively, our study offers a potential strategy for NSCLC therapy, especially for drug-resistant patients with highly expressed GLUD1., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Rho GTPases play an essential role in many cellular processes, including cell cycle progress, cell motility, invasion, migration, and transformation. Several studies indicated that the dysregulation of Rho GTPase signaling is closely related to tumorigenesis. Rho GEFs considered being positive regulators of Rho GTPase, promoting the dissociation of Rho protein from GDP and binding to GTP, thus activating the downstream signaling pathway. Herein, we demonstrated that ARHGEF3, a member of the Rho GEFs family, played an important role in non-small cell lung cancer (NSCLC). We found that ARHGEF3 was highly expressed in non-small cell lung cancer and facilitated cancer cell proliferation of NSCLC cells in vitro and in vivo. Further studies demonstrated that ARHGEF3 enhanced the protein homeostasis of ATP-citrate lyase (ACLY) by reducing its acetylation on Lys17 and Lys86, leading to the dissociation between ACLY and its E3 ligase-NEDD4. Interestingly, this function of ARHGEF3 on the protein homeostasis of ACLY was independent of its GEF activity. Taken together, our findings uncover a novel function of ARHGEF3, suggesting that ARHGEF3 is a promising therapeutic target in non-small cell lung cancer., (© 2022. The Author(s).)

Background: Glycogen-Interacting Protein 1 (GNIP1), an E3 ligase, is a member of the tripartite motif (TRIM) family proteins. Current studies on GNIP1 mainly focus on glycogen metabolism. However, the function and molecular mechanisms of GNIP1 in regulating autophagy still remains unclear. This study aimed to investigate the regulatory mechanism of GNIP1 in regulating autophagy in non-small cell lung cancer (NSCLC)., Methods: Crystal violet staining assays were used to evaluate the ability of cell growth and proliferation. Transwell and scratch wound healing assays were used to evaluate the cell migration ability. The protein expressions were measured by western blot and immunohistochemistry. Co-immunoprecipitation assays determined the protein-protein interactions. The in vivo effect of GNIP1 on tumor growth was determined by xenograft assay., Results: We found that GNIP1 was overexpressed in tumor tissues and the expression level of GNIP1 was related to the poor prognosis and the survival time of NSCLC patients. In non-small cell lung cancer (NSCLC), GNIP1 increased proliferation and migration of cancer cells by promoting autophagy. Mechanistic studies indicated that GNIP1, as a scaffold protein, recruited BECN1 and LC3B to promote the formation of autophagosomes. Besides, GNIP1 mediated the degradation of 14-3-3ζ, the negative regulator of VPS34 complex, thus promoting autophagy. Overexpressing GNIP1 promoted tumorigenesis and enhanced autophagy in xenograft models., Conclusion: GNIP1 promotes proliferation and migration of NSCLC cells through mediating autophagy, which provides theoretical basis for targeting GNIP1 as anti-cancer drugs. Video Abstract., (© 2022. The Author(s).)

It has been reported that microRNA-203a-3p (miR-203a-3p) modulates cell proliferation, migration and invasion in a variety of cancer cell types. However, little is known about its role in lung cancer progression. The present study found that miR-203a-3p was downregulated in non-small cell lung cancer (NSCLC) cell lines and tissues. Overexpression of miR-203a-3p inhibits NSCLC cell proliferation, migration and invasion, and promotes cellular apoptosis in vitro. Restoration of miR-203a-3p expression in A549 and NCI-H520 cells enhances their chemosensitivity. Further experiments showed that DNA methyltransferase 3B (DNMT3B) was a direct target of miR-203a-3p. In addition, the present results revealed that promoter hypermethylation was the potential mechanism responsible for low miR-203a-3p expression in NSCLC. Notably, feedback regulation between miR-203a-3p and DNMT3B was observed in NSCLC. Moreover, Overexpression of miR-203a-3p reduces tumor growth in vivo. In summary, the present study has identified an miR-203a-3p-DNMT3B feedback loop that facilitates NSCLC progression., (© 2022. The Author(s) under exclusive licence to Japan Human Cell Society.)

Lung cancer continues to be the world's leading cause of cancer death and the treatment of non-small cell lung cancer (NSCLC) has attracted much attention. The tubers of Bletilla striata are regarded as "an excellent medicine for lung diseases" and as the first choice to treat several lung diseases. In this study, seventeen phenanthrene derivatives, including two new compounds ( 1 and 2 ), were isolated from the tubers of B. striata . Most compounds showed cytotoxicity against A549 cells. An EdU proliferation assay, a cell cycle assay, a wound healing assay, a transwell migration assay, a flow cytometry assay, and a western blot assay were performed to further investigate the effect of compound 1 on A549 cells. The results showed that compound 1 inhibited cell proliferation and migration and promoted cell apoptosis in A549 cells. The mechanisms might correlate with the regulation of the Akt, MEK/ERK, and Bcl-2/Bax signaling pathways. These results suggested that the phenanthrenes of B. striata might be important and effective substances in the treatment of NSCLC.

Background: Inevitably developed resistance of the third-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) limited its clinical benefit on non-small cell lung cancer (NSCLC). Upfront combination therapy is promising to prevent this resistance. Compelling clinical evidence indicated the failure of third-generation EGFR TKIs combined with either immunotherapy or antiangiogenic agents. In comparison, combined treatment of third-generation EGFR TKIs and chemotherapy might be a favorable choice. Herein, we systematically analyzed and compared the effects of pemetrexed and a novel third-generation EGFR TKI aumolertinib combined in different sequences, subsequently revealed the potential mechanisms and proved the optimal combination schedule with clinical retrospective study., Methods: Three combination schedules involving pemetrexed and aumolertinib in different sequences were developed. Their inhibition effects on cell proliferation and metastasis were firstly compared upon three human NSCLC cell lines in vitro, by cell counting kit-8, colony formation, wound healing and transwell assays respectively. Further evaluation in vivo was proceeded upon H1975 and HCC827 xenograft model. Gene and protein expression were detected by Q-PCR and western blot. Drug concentration was determined by LC-MS/MS. VEGF secretion was determined by ELISA. Tumor vessel was visualized by immunofluorescence. Lastly, a clinical retrospective study was raised with 65 patients' data., Results: The combination of pemetrexed and aumolertinib exhibited a sequence-dependent and EGFR mutant-dependent synergistic effect in vitro and in vivo. Only treatment with aumolertinib following pemetrexed (P-A) exhibited synergistic effect with stronger anti-tumor growth and anti-metastasis ability than monotherapy and also other combination sequences. This synergism could exclusively be observed in H1975 and HCC827 but not A549. Pathway analysis showed that P-A significantly enhanced the suppression of EGFR pathway. In addition, our results intriguingly found an obvious reduction of VEGF secretion and the accompanying normalization of the intratumor vessel, consequently increasing intratumoral accumulation of pemetrexed in P-A group. Finally, the clinical retrospective study verified the synergistic effect of P-A combination by significantly superior tumor response than aumolertinib monotherapy., Conclusion: Aumolertinib-pemetrexed combined therapy is promising for EGFR mutant NSCLC but only in right administration sequence. P-A could become an advantageous combination strategy in clinical with synergistic inhibition of tumor growth and metastasis., (© 2022. The Author(s).)

Circular RNAs (circRNAs) are closely linked with human cancer development such as non-small-cell lung cancer (NSCLC). However, the characteristics and specific functions of most circRNAs in NSCLC remained unknown. Previous studies have suggested that circRNA SOD2 (CircSOD2) expression was upregulated in a number of cancers. This study aimed to explore the functions of circSOD2 in NSCLC advancement with epithelial-mesenchymal transition (EMT). Expression profile analysis of circSOD2, miR-2355-5p, and calmodulin-regulated spectrin-associated protein 2 (CAMSAP2) was detected by real-time quantitative PCR (RT-qPCR). Transwell assay, cell migration assay, CCK8, ELISA, RIP assay, RNA pull-down assay, and Western blot analysis were performed to evaluate the functions of circSOD2, miR-2355-5p, and CAMSAP2. We found elevated expression of circSOD2 and CAMSAP2 while reduced expression of miR-2355-5p in NSCLC tumor tissues. Silencing or overexpression of CircSOD2 resulted in increased or decreased expression of miR-2355-5p, respectively. Mechanically, we showed that silencing of CircSOD2 and overexpression of miR-2355-5p resulted in the reduced rate of NSCLC cell proliferation. Inhibition of miR-2355-5p reversed the changes induced via silencing of CircSOD2. MiR-2355-5p binds to the CircSOD2 promoter and triggered its stimulation, which further activated circSOD2 expression. CircSOD2 suppression impaired lung cancer cell growth, cell migration, prohibited cell cycle progression, and in vivo tumor growth by targeting miR-2355-5p expression in NSCLC tissues. Meanwhile, increased expression of CAMSAP2 reversed the changes stimulated by the elevated level of miR-2355-5p in NSCLC progression. This innovative signaling axis CircSOD2/miR-2355-5p/CAMSAP2 illustrated the new horizon to investigate NSCLC tumorigenesis and provided new prognosis and treatment of NSCLC.

Background: Immune checkpoint inhibitors (ICIs) brought about a major paradigm shift in non-small cell lung cancer (NSCLC) treatment. However, the use of ICIs is related to an unforeseeable pattern of immune-related adverse events (irAEs). Hence, more precise biomarkers are needed to predict the incidence of irAEs to prevent overtreatment of ICIs and decrease occurrences of irAEs. This study was designed to identify capable clinical features and plasma inflammatory factors for predicting irAEs., Methods: A total of 67 patients who received ICI monotherapy or ICI-based combination therapy were retrospectively identified. Clinical characteristics and plasma inflammatory cytokines were collected and analyzed to screen potential biological markers associated with irAEs. The chi-square test, Fisher's test, and the Mann-Whitney U test were performed for the primary analysis. The optimal cutoff value was determined by a receiver operating characteristic (ROC) curve. Univariate and multivariate logistic regression models were used to identify risk factors of irAEs. Univariate and multivariate Cox proportional hazards were also performed., Results: Out of 67 patients, 40 (59.7%) experienced irAEs, and 7 (10.4%) experienced severe adverse events (grade ≥ 3). Among these analyzed immune profile biomarkers, only interleukin-10 (IL-10) was related to the risk of irAEs. A high baseline IL-10 plasma level (odds ratio (OR) = 5.318, 95% CI 1.174-24.081, p = 0.030) was found to be a tremendous and independent risk factor for the development of irAEs. Also, for the dynamic analysis, upregulation of IL-10 after one cycle of ICI treatment was positively related to the occurrence of irAEs (OR = 5.712, 95% CI 1.088-29.993, p = 0.039). When pneumonitis, the most common irAEs, was analyzed, only baseline high-expression IL-10 was accompanied with the incidence of pneumonitis (OR = 9.969, 95% CI 1.144-86.843, p = 0.037)., Conclusion: Baseline and dynamic IL-10 plasma levels are tremendously and independently related to higher risk in the development of irAEs and could be utilized for medical practice to monitor adverse events in patients with ICI treatment., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Wang, Zhou, Zhao, Cheng, Zhou, Qiao, Li and Chen.)

Background: LINC00511 has been reported as a biomarker related to the prognosis of non-small cell lung cancer (NSCLC), but the molecular mechanism and exact functions of LINC00511 in chemoresistance of NSCLC remain to be elucidated., Methods: RT-qPCR was used to evaluate the mRNA expression of LINC00511, miR-625, and leucine rich repeat containing 8 volume-regulated anion channel subunit E (LRRC8E). Western blotting detected the protein levels of Ki-67, MMP-9, cleaved-caspase-3. The interaction between miR-625 and LINC00511 or LRRC8E was verified by luciferase reporter assays. CCK-8, TUNEL, and Transwell assays were used to evaluate IC 50 value, proliferation, migration, and invasion of NSCLC cells., Results: In our study, it was discovered that the levels of LINC00511 and LRRC8E were increased, while miR-625 expression was decreased in NSCLC tissues, DDP-resistant NSCLC cells, and non-resistant NSCLC cells. LINC00511 depletion significantly curbed cell growth, IC 50 value, and metastasis in DDP-resistant NSCLC cells. In addition, the influence of LINC00511 deficiency on the DDP resistance in NSCLC was overturned by suppressing miR-625. Furthermore, LRRC8E overexpression abolished the promotive effect of miR-625 abundance on the DDP sensitivity in DDP-resistant NSCLC cells., Conclusion: Our results demonstrated that LINC00511 increased DDP resistance in NSCLC by suppressing miR-625 to upregulate LRRC8E.

Aim and Objective: Lung cancer is the most commonly occurring cancer, which contributes to the majority of death caused by cancer, where non-small-cell lung cancer (NSCLC) accounts for approximately 85% of lung cancer. To treat NSCLC, STAT3 has been identified as a target with therapeutic potential. The neobavaisoflavone (NBIF) is one of the flavonoids of traditional Chinese medicine Psoralea corylifolial., Materials and Methods: Human NSCLC cell lines, PC-9, H460, and A549, were applied to determine NBIF's anti-proliferative effects through cell viability and colony formation detection. The effect of NBIF on cell apoptosis was determined through flow cytometry-based assay. Western blotting was used in this study to confirm the levels of P-STAT3, Bcl-2, and Bax, which are apoptotic proteins., Results: It was observed that NBIF could decrease the cell viability and its migration and induce apoptosis in human NSCLC cell lines dose-dependently. Levels of P-STAT3, as well as the downstream signals of the STAT3 pathway, were downregulated, suggesting that the tumorsuppression effects of NBIF might be related to the inhibition of STAT3 signaling. Furthermore, NBIF could contribute to the upregulation of BAX and downregulation of BCL2., Conclusion: NBIF might perform the anti-NSCLC efficacy as a result of the inhibition of the STAT3 pathway. Besides, our work suggests that NBIF could provide therapeutic alternatives for NSCLC., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Objectives: Immune checkpoint inhibitors (ICI) monotherapy was standard of care in second-line treatment of patients with advance non-small cell lung cancer (NSCLC). This study aims to investigate the efficacy of ICI plus chemotherapy in patients with previously treated advanced NSCLC., Patients and Methods: An investigator-initiated trial (IIT) aiming to evaluate the efficacy and safety of ICI in combination with chemotherapy as second line and beyond for patients with advanced NSCLC was undergone at Shanghai Pulmonary Hospital (ChiCTR1900026203). Patients who received ICI monotherapy as second or later line setting during the same period were also collected as a comparator., Results: From April 2018 to June 2019, 31 patients were included into this IIT study, simultaneously 51 patients treated with ICI monotherapy were selected as a comparator. ICI plus chemotherapy showed a significantly higher ORR (35.5% vs. 15.7%, p=0.039), prolonged PFS (median: 5.6 vs. 2.5 months, p = 0.013) and OS (median: NE vs. 12.6 months, p = 0.038) compared with ICI alone. In the subgroup of negative PD-L1 expression (9 patients in combination group and 12 patients in monotherapy group), ICI plus chemotherapy also had a favorable ORR (44.4% vs. 8.3%, p = 0.119), longer PFS (median: 6.5 vs 3.0 months, p < 0.05) and OS (median: NE vs. 8.2 months, p = 0.117). Meanwhile, the addition of chemotherapy did not increase immune-related adverse events., Conclusions: ICI plus chemotherapy showed superior ORR, PFS and OS than ICI alone patients with previous treated advanced NSCLC. These findings warrant further investigation., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

CD303 + plasmacytoid dendritic cells (pDCs) play an important role in the induction of immune tolerance and antitumor immunity. Here, we focused on the effect of NSCLC cells on the development of CD303 + pDC subsets expressing CD205 and/or CD103. The NSCLC cell line H1299 and primary NSCLC cells were incubated with DCs. The protein expression of costimulatory molecules on CD303 + pDCs, the production of pro-inflammatory and anti-inflammatory cytokines by CD303 + pDCs and the development of CD303 + pDC subsets were detected by using flow cytometry. Coculture with NSCLC cells modulates the protein expression of CD86 and HLA-DR on CD303 + pDCs. Moreover, NSCLC cells suppressed the production of IL-12 and IL-23 but facilitated the secretion of IL-27 and TGF-β by CD303 + pDCs. There were new CD303 + pDC subsets expressing CD205 and/or CD103 in healthy donors and NSCLC patients: CD303 + CD205 + CD103 + , CD303 + CD205 + CD103 - , CD303 + CD205 - CD103 + and CD303 + CD205 - CD103 - pDCs. NSCLC cells modulated the differentiation of CD303 + pDC subpopulations by regulating the protein expression of CD205 and/or CD103 on CD303 + pDCs. NSCLC cells may regulate the immune functions of CD303 + pDCs by modulating the expression of costimulatory molecules on DCs and the production of pro-inflammatory/anti-inflammatory cytokines by DCs. NSCLC cells also regulate the development of CD303 + pDC subsets expressing CD205 and/or CD103. These outcomes may reveal a new cellular mechanism leading to the NSCLC-induced immune-suppressive microenvironment., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Although immune checkpoint inhibitors (ICIs) that target programmed cell death protein-1/programmed cell death ligand-1 axis have significantly shifted the treatment paradigm in advanced NSCLC, clinical benefits of these agents are limited in patients with EGFR-mutated NSCLC. Several predictive biomarkers (e.g., programmed cell death ligand-1 expression, tumor mutation burden), which have been validated in EGFR-wild type NSCLC, however, are not efficacious in EGFR-mutated tumors, suggesting the unique characteristics of tumor microenvironment of EGFR-mutated NSCLC. Here, we first summarized the clinical evidence on the efficacy of ICIs in patients with EGFR-mutated NSCLC. Then, the cancer immunogram features of EGFR-mutated NSCLC was depicted to visualize the state of cancer-immune system interactions, including tumor foreignness, tumor sensitivity to immune effectors, metabolism, general immune status, immune cell infiltration, cytokines, and soluble molecules. We further discussed the potential subpopulations with EGFR mutations that could benefit from ICI treatment. Lastly, we put forward future strategies to adequately maximize the efficacy of ICI treatment in patients with EGFR-mutated NSCLC in the upcoming era of combination immunotherapies., (Copyright © 2021 International Association for the Study of Lung Cancer. Published by Elsevier Inc. All rights reserved.)

Lung cancer is a highly heterogeneous disease. Cancer cells and cells within the tumor microenvironment together determine disease progression, as well as response to or escape from treatment. To map the cell type-specific transcriptome landscape of cancer cells and their tumor microenvironment in advanced non-small cell lung cancer (NSCLC), we analyze 42 tissue biopsy samples from stage III/IV NSCLC patients by single cell RNA sequencing and present the large scale, single cell resolution profiles of advanced NSCLCs. In addition to cell types described in previous single cell studies of early stage lung cancer, we are able to identify rare cell types in tumors such as follicular dendritic cells and T helper 17 cells. Tumors from different patients display large heterogeneity in cellular composition, chromosomal structure, developmental trajectory, intercellular signaling network and phenotype dominance. Our study also reveals a correlation of tumor heterogeneity with tumor associated neutrophils, which might help to shed light on their function in NSCLC.

Objective: CXCR1, a member of the seven-transmembrane chemokine receptor family, promotes cell proliferation and metastasis in many tumors. The present study was undertaken to explore the interrelation between CXCR1 expression and the prognosis of advanced non-small cell lung cancer (NSCLC) in addition to the efficacy of epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) in lung adenocarcinoma (LUAD)., Methods: The expression of CXCR1 in NSCLC tissues was assessed by immunohistochemistry. The relationships between CXCR1 expression and clinical-pathological factors were investigated. Concomitantly, the relationship between CXCR1 expression and EGFR-TKI treatment efficacy was investigated. Gene set enrichment analysis (GSEA) was employed for the exploration of pathway enrichment, tumor immune estimation resource (TIMER) and gene expression profiling interactive analysis (GEPIA) for the inspection of the interrelationship between infiltration immune cells and CXCR1. After gain-and loss-of-function of CXCR1 in NSCLC cells, qRT-PCR and Western blot were applied to measure the levels of proteins associated with the chemokine pathway (CCL3 and CXCL2) and the JAK/STAT pathway (IL9R, PIAS4 and STAT5A)., Results: CXCR1 significantly correlated with poor prognosis of NSCLC patients. Additionally, CXCR1 limited the clinical efficacy of EGFR-TKIs in advanced LUAD (P = 0.029). In the tumor microenvironment, CXCR1 was positively associated with infiltration levels of immune markers in lung squamous cell carcinoma (LUSC) and LUAD. High expression of CXCR1 was implicated in the NOD-like receptor (NLR), cytokine/cytokine receptor, JAK/STAT and chemokine signaling pathways in LUAD and LUSC. Overexpression of CXCR1 in NSCLC cell lines enhanced expressions of CCL3, CXCL2, IL9R, PIAS4 and STAT5A, while knockdown of CXCR1 repressed expressions of CCL3, CXCL2, IL9R, PIAS4 and STAT5A., Conclusion: CXCR1 is correlated with poor prognosis of NSCLC and affects the efficacy of EGFR-TKIs in LUAD., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Introduction: Previous clinical investigations have demonstrated that patients with non-small-cell lung cancer (NSCLC) harboring epidermal growth factor receptor (EGFR) mutation have moderate responses to programmed cell death-1 and it's ligand-1 (PD-1/PD-L1) inhibitors, while some patients who failed EGFR-tyrosine kinase inhibitor (TKI) therapy could benefit from immunotherapy. As a result, we have explored alterations in the tumor immune microenvironment (TIME) before and after EGFR-TKI treatment to detect the chances and proper timing of immunotherapy among patients., Methods: We identified 16 paired tissue samples pre- and post-EGFR-TKI treatment. Sections 4 μm thick were utilized to evaluate CD8, PD-L1, PD-1, LAG-3, and TIM-3 expressions by multiplexed fluorescent immunohistochemical staining. Five to ten representative original multispectral images of each sample were employed in the analysis., Results: Patients with positive CD8 + T-cell infiltration accounted for 37.5 % at baseline. Positive expression of PD-L1, PD1, LAG-3, and TIM-3 cells were observed in seven (43.8 %), four (25 %), one (6.25 %) and five (31.25 %) of the patients, respectively. PD-1 expression and infiltration of CD8 + PD-1 + -exhausted T cells increased significantly in patients with EGFR L858R mutation compared to patients with EGFR 19DEL. Patients who acquired T790 M after TKI treatment had less infiltrations of CD8 + PD-1 + T cells and CD8 + TIM-3 + T cells in the TIME at baseline. Positive expression of checkpoint proteins-including PD-1, TIM-3, and LAG-3-significantly correlated with shorter progression-free survival. LAG-3 was significantly upregulated after TKI treatment (p = 0.003), while other checkpoint proteins remained stationary., Conclusion: The present study is the first work to report LAG-3 upregulation after EGFR-TKI failure in advanced NSCLC, which proposed novel insights for rational use of LAG-3 inhibitors in advanced NSCLC patients with EGFR mutation., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Background: A series of studies have demonstrated that NPAS2 plays a critical role in the development and progression of several cancers. However, the association between genetic variants in the  NPAS2 gene and the clinical outcome of patients with non-small-cell lung cancer (NSCLC) has not been investigated. Methods: Six functional SNPs in NPAS2  were selected and genotyped using the Sequenom iPLEX genotyping system in a cohort of 484 Chinese NSCLC patients undergoing surgery. Multivariate Cox proportional hazards model were used for the prognosis analysis. Results: We found that SNP rs2305158 exhibited a significant association with overall survival of NSCLC patients in the dominant model (hazard ratio [HR]: 0.68; 95% CI: 0.49-0.95; p = 0.02). Lymph node metastasis was significantly associated with increased death risk (HR: 1.73; 95% CI: 1.24-2.40; p = 0.001) in patients with the homozygous wildtype (WW) genotype of rs2305158. However, no significant association was observed between them in patients carrying a heterozygous variant (WV) or homozygous variant (VV) genotype of rs2305158. Finally, in the joint and interaction analysis, the patients carrying homozygous wildtype (WW) genotype and lymph node metastasis from N1 to N3 conferred a significant increased effect on death (HR: 2.29; 95% CI: 1.40-3.76; p = 0.001). Conclusions: Our results suggest that NPAS2  polymorphisms may serve as an independent prognostic marker for NSCLC patients.

Background: Cyclin-dependent kinase 9 (CDK9) is a promising prognostic marker and therapeutic target in cancers. Bufalin is an effective anti-tumour agent; however, the clinical application of bufalin is limited due to its high toxicity. Acetyl-bufalin, the bufalin prodrug, was designed and synthesised with higher efficiency and lower toxicity., Methods: Three non-small-cell lung cancer (NSCLC) cell lines, a xenograft model and a patient-derived xenograft (PDX) model were used to examine the effects of acetyl-bufalin. CDK9/STAT3 involvement was investigated by knockdown with siRNA, proteome microarray assay, western blot analysis and co-immunoprecipitation experiments. Acute toxicity test and pharmacokinetics (PK) study were conducted to assess the safety and PK. The human NSCLC tissues were analysed to verify high CDK9 expression., Results: We showed that CDK9 induced NSCLC cell proliferation and that this effect was associated with STAT3 activation, specifically an increase in STAT3 phosphorylation and transcription factor activity. Acetyl-bufalin is an effective and safety inhibitor of the CDK9/STAT3 pathway, leading to the impediment of various oncogenic processes in NSCLC. Molecular docking and high-throughput proteomics platform analysis uncovered acetyl-bufalin directly binds to CDK9. Consequently, acetyl-bufalin impaired the complex formation of CDK9 and STAT3, decreased the expressions of P-STAT3, and transcribed target genes such as cyclin B1, CDC2, MCL-1, Survivin, VEGF, BCL2, and it upregulated the expression levels of BAX and caspase-3 activity. Acetyl-bufalin inhibited tumour growth in NSCLC xenograft and PDX models., Conclusions: Acetyl-bufalin is a novel blocker of the CDK9/STAT3 pathway thus may have potential in therapy of NSCLC and other cancers.

Great advances in immune checkpoint blockade have resulted in a paradigm shift in patients with lung cancer. Immune-checkpoint inhibitor (ICI) treatment, either as monotherapy or combination therapy, has been established as the standard of care for patients with locally advanced/metastatic non-small cell lung cancer without EGFR/ALK alterations or extensive-stage small cell lung cancer. An increasing number of clinical trials are also ongoing to further investigate the role of ICIs in patients with early-stage lung cancer as neoadjuvant or adjuvant therapy. Although PD-L1 expression and tumor mutational burden have been widely studied for patient selection, both of these biomarkers are imperfect. Due to the complex cancer-immune interactions among tumor cells, the tumor microenvironment and host immunity, collaborative efforts are needed to establish a multidimensional immunogram to integrate complementary predictive biomarkers for personalized immunotherapy. Furthermore, as a result of the wide use of ICIs, managing acquired resistance to ICI treatment remains an inevitable challenge. A deeper understanding of the underlying biological mechanisms of acquired resistance to ICIs is helpful to overcome these obstacles. In this review, we describe the cutting-edge progress made in patients with lung cancer, the optimal duration of ICI treatment, ICIs in some special populations, the unique response patterns during ICI treatment, the emerging predictive biomarkers, and our understanding of primary and acquired resistance mechanisms to ICI treatment.

Purpose of Review: Despite the significant advances in EGFR-mutant nonsmall cell lung cancer (NSCLC), some challenges remain. One of the permanent and inevitable issues is the emergence of acquired resistance. Therefore, blocking the activation of EGFR pathway and overcoming drug resistance with novel agents are still in high demand. Here, we review the development of novel drugs in EGFR-mutant, advanced NSCLC, including targeting EGFR exon 20 insertion (EGFR20ins), and novel role of epidermal growth factor receptor, tyrosine kinase inhibitor (EGFR-TKIs) in early-stage NSCLC., Recent Findings: EGFR-TKIs as adjuvant therapy or neoadjuvant therapy in patients with early-stage NSCLC with EGFR-sensitizing mutations have shown promising efficacy. The resistance mechanisms of third-generation EGFR-TKIs can be divided into two types: EGFR dependent and EGFR independent. Several clinical trials have demonstrated that the addition of MET inhibitors to EGFR-TKIs was an effective option for patients who had acquired resistance to EGFR-TKIs caused by hepatocyte growth factor receptor gene (MET) amplification or overexpression. Novel compounds that selectively and potently inhibit EGFR20ins are being investigated in phase III studies., Summary: A better characterization and understanding of resistance mechanisms to first-line osimertinib and adjuvant osimertinib is helpful to guide further treatment.

Non-small cell lung cancer (NSCLC) is a leading cause of cancer death in both men and women. microRNAs (miRs) can exert important functions in cancer development. However, the role of miR-877 in NSCLC as it relates to tartrate resistant acid phosphatase 5 (ACP5) is unknown. For this study, the gain-and-loss-of-function experiments were performed to explore the effects of miR-877 and ACP5 on NSCLC. miR-877 expression in LC and paracancerous tissues, lung epithelial cell line and NSCLC cell lines was detected, and the association between miR-877 expression and clinical features of LC patients was analyzed. The levels of ACP5, epithelial-mesenchymal transition (EMT) markers and apoptosis-related proteins were measured. In vivo experiments were conducted for further validation. Consequently, we found that miR-877 expression was lowered in LC tissues and cell lines, and correlated with clinical stage, differentiation, lymph node metastasis and prognosis of NSCLC patients. Additionally, miR-877 was determined to inhibit ACP5 activity, and miR-877 downregulated the PI3K/AKT pathway by silencing ACP5. Furthermore, overexpression of miR-877 inhibited the viability, migration, invasion and EMT of NSCLC cells, but promoted cell apoptosis. In conclusion, miR-877 overexpression inhibited malignant biological behaviors of NSCLC cells by downregulating ACP5 and inactivating the PI3K/AKT pathway.

Immunohistochemical (IHC) stain for PD-L1 as a biomarker for immunotherapy is recommended in non-small cell lung cancer (NSCLC). Under the FDA, the selection of patients for pembrolizumab requires companion diagnostic testing using the Dako Agilent PD-L1 IHC 22C3 pharmDx kit performed on the Dako Autostainer Link 48 platform. However, because it is not widely available, there is need for cross-platform validation. Existing studies provide incomplete protocol detail. In our study, 73 lung tumors were stained using the FDA-approved test ('gold standard'). The same blocks were stained using two different models of the Ventana DISCOVERY platform (ULTRA, n = 73 and XT, n = 70) using different parameters, and interpreted by three pathologists. The ULTRA group met College of American Pathologists (CAP) validation criteria (concordance 91.8%) while the XT group did not (concordance 67.1%). Using tumor proportion score (TPS) ≥1% and TPS ≥50% as cut-offs, the ULTRA protocol had higher sensitivity (97.8% and 91.7%) than XT (73.3% and 60.9%) and similar specificity (ULTRA 88.9% and 100%, XT 88% and 100%). Discordance between ULTRA and XT was 27%, and in all these cases ULTRA was concordant with gold standard. Interobserver reliability was substantial for ULTRA and almost perfect for XT, providing evidence that staining rather than observer variability accounts for XT's inferior performance. Cross-validation of the clinically used 22C3 anti PD-L1 antibody test with substantial interobserver agreement is possible on the commonly used the Ventana DISCOVERY ULTRA automated instrument, while the validation failed on the XT. Cautious attention to detail must be paid when choosing cross-validation parameters.

Objectives: Immune checkpoint inhibitors (ICIs) have revolutionized the oncologic treatment landscape, but have been accompanied by immune-related adverse events (irAEs). ICI-related pneumonitis (ICI-pneumonitis) is a potentially fatal irAE. However, the risk factors associated with ICI-pneumonitis remain unclear. There is an urgent need to identify risk factors for ICI-pneumonitis using reliable and accessible parameters. Here, we aimed to identify baseline peripheral-blood biomarkers correlated with ICI-pneumonitis and clinical outcomes in patients with advanced non-small cell lung cancer (NSCLC) who were treated with ICIs., Materials and Methods: We conducted a retrospective analysis of eligible patients with advanced NSCLC who were treated with ICIs at our center. Receiver operating characteristic (ROC) curve was used to determine the optimal cutoff value for analyzing risk of ICI-pneumonitis. Multivariate logistic analysis was performed to identify risk factors of ICI-pneumonitis. Clinical characteristics and treatment outcomes were collected and compared according to the optimal cutoff value., Results: A total of 300 patients were included, in which 54 patients (18 %) experienced ICI-pneumonitis. Patients with ICI-pneumonitis had a high level of baseline peripheral-blood absolute eosinophil count (AEC) than those without ICI-pneumonitis (P = 0.013). The optimal threshold of baseline peripheral-blood AEC to predict ICI-pneumonitis was 0.125 × 10 9 cells/L. The incidence of ICI-pneumonitis was higher in the high-AEC group (AEC ≥ 0.125 × 10 9 cells/L; 27.7 %) than in the low-AEC group (AEC < 0.125 × 10 9 cells/L; 9.8 %, P < 0.001). Moreover, patients with high AEC (compared with those with low AEC) had a higher objective response rate (ORR) (40.9 % versus 28.8 %, P = 0.029) and longer median progression-free survival (PFS) (8.93 months versus 5.87 months, P = 0.038)., Conclusions: Among patients treated with ICIs, a baseline feature of high AEC (≥0.125 × 10 9 cells/L) was associated with an increasing risk of ICI-pneumonitis, and with a better clinical outcome., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Radiation resistance affects survival in non-small-cell lung cancer (NSCLC) patients. Further exploration of mechanisms and targets is urgently needed. Using bioinformatic analyses, we found that UBE2T is associated with survival, tumor size, lymph node metastasis and distant metastasis. Then, real-time PCR and immunohistochemistry were performed to explore the differentially expressed genes between normal and NSCLC tissues. Furthermore, we used colony formation, EdU incorporation, scratch, transwell assays, flow cytometry, immunofluorescence and western blot to assess the role of UBE2T in vitro and in vivo. RNA-Seq and coimmunoprecipitation were used to explore the mechanism. The results showed that UBE2T promotes proliferation, migration, invasion, and radiation resistance in vitro and in vivo by accelerating the G2/M transition and inhibiting apoptosis. Mechanistically, UBE2T promotes epithelial-mesenchymal transition (EMT) via ubiquitination-mediated FOXO1 degradation and Wnt/β-catenin signaling pathway activation. Moreover, FOXO1 reversed radiation resistance and EMT. Therefore, UBE2T may be a potential target for enhancing radiotherapy sensitivity and serve as a biomarker to predict prognosis., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: Bone metastasis (BoM) is common in patients with advanced non-small cell lung cancer (NSCLC) and considered as one of the negative prognostic factors. However, the impact of BoM on clinical outcomes of patients with advanced NSCLC treated with immune checkpoint inhibitors (ICIs) remains unclear., Methods: A total of 103 patients treated with ICI monotherapy and 101 patients treated with ICIs combined with chemotherapy or antiangiogenesis therapy were retrospectively analyzed. The differences in progression-free survival (PFS), overall survival (OS) and objective response rate (ORR) between BoM+ and BoM- were investigated., Results: Of those 101 patients who received combination therapy, no significant difference between BoM- and BoM+ in terms of both median PFS and median OS (median PFS, 10.1 vs. 12.1 months, P = 0.6; median OS, NR vs. 24.6 months, P = 0.713) was determined. In contrast, of the 103 patients who received ICI monotherapy, BoM+ patients had an inferior PFS (4.2 vs. 6.7 months, P = 0.0484) and OS (12.5 vs. 23.9 months, P = 0.0036) compared with BoM- patients. The univariate and multivariate analysis in the ICI monotherapy group also identified BoM as an independent factor attenuating the efficacy of ICI monotherapy. Of all BoM+ patients who received ICI monotherapy, neither palliative radiotherapy nor bisphosphonate drugs improved OS (palliative radiotherapy: 12.5 vs. 16.7 months, P = 0.487; bisphosphonate drugs: 12.5 vs. 9.7 months, P = 0.568)., Conclusions: BoM attenuated the efficacy of ICI monotherapy in patients with advanced NSCLC. Of BoM+ patients who received ICI monotherapy, neither palliative radiotherapy nor bisphosphonate drugs improved OS. Other therapeutic strategies are needed for patients with BoM., (© 2020 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.)

Objectives: Recent investigations have shown strong correlations between cytology and surgical non-small cell lung carcinoma (NSCLC) specimens in programmed death-ligand 1 (PD-L1) immunohistochemical (IHC) evaluations. Our study aims to evaluate the reproducibility of PD-L1 IHC scoring in NSCLC cytology cell blocks (CBs) and to assess the impact of CB cellularity, method of sample collection, and observer subspecialty on scoring agreement., Methods: PD-L1 IHC was performed on 54 NSCLC cytology CBs and was scored independently by seven cytopathologists (three of seven with expertise in pulmonary pathology). Three-tier scoring of negative (<1%), low positive (1%-49%), and high positive (≥50%) and interrater agreement were assessed., Results: Total and majority agreement among cytopathologists was achieved in 48% and 98% of cases, respectively, with κ = 0.608 (substantial agreement; 95% confidence interval, 0.50-0.72). Cytopathologists with pulmonary pathology expertise agreed in 67% of cases (κ = 0.633, substantial agreement), whereas the remaining cytopathologists agreed in 56% of cases (κ = 0.62, substantial agreement). CB cellularity (P = .36) and sample collection type (P = .59) had no statistically significant difference between raters., Conclusions: There is substantial agreement in PD-L1 IHC scoring in cytology CB specimens among cytopathologists. Additional expertise in pulmonary pathology, sample collection type, and CB cellularity have no statistically significant impact on interobserver agreement., (© American Society for Clinical Pathology, 2020. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

BACKGROUND Metastasis contributes to the high mortality rate of non-small cell lung cancer (NSCLC), and gaining a better understanding of its metastatic mechanisms would aid in initiating effective clinical treatment. MATERIAL AND METHODS In this study, bioinformatics analyses of the GEO database and TCGA-LUAD were first used to identify the key node gene regulating NSCLC malignant progression. Further in vitro experiments, including wound healing assay, invasion assay, Western blot assay, and luciferase report assay, were used to clarify the functions and mechanism of TPX2 in NSCLC. RESULTS Results of the TCGA analysis showed that TPX2 was significantly positively correlated with tumor metastasis and growth and the clinical stage of NSCLC. In addition, high levels of TPX2 significantly indicated a poor survival rate. In vitro experimental results also revealed that the upregulation of TPX2 significantly promoted NSCLC cell migration and invasion and could affect cell replasticity. Further results indicated that TPX2 significantly activated the epithelial-mesenchymal transition process and promoted the expression and activities of matrix metalloproteinase (MMP)2 and MMP9. CONCLUSIONS This study demonstrated that TPX2 promotes the metastasis and malignant progression of NSCLC and could thus serve as a marker of poor prognosis in NSCLC.

Nonsmall cell lung carcinoma (NSCLC) contributes to the highest number of cancer deaths globally. Metastases and chemoresistance are two major confounders to the treatment efficacy in NSCLC. Transducin (β)-like 1 X-linked receptor 1 (TBL1XR1) has been associated with high rates of metastases in breast, gastric, and stomach cancers. However, the role of TBL1XR1 in lung cancers remains underexplored. We selected matched and cancerous lung tissues to establish the upregulation of TBL1XR1. Using in vitro assays, we assessed the influence of TBL1XR1 on various cancer phenotypes, namely cell proliferation, chemoresistance, invasion, and metastases in a CRISPR-Cas9-mediated knock out model (A549 cells), and H460 cell lines overexpressing TBL1XR1. We found that TBL1XR1 is overexpressed in NSCLC tissue and patient sera in comparison to paired adjacent normal tissue. Overexpression of TBL1XR1 in NSCLC cell lines mediates cell survival, proliferation, and metastases. TBL1XR1 was found to regulate MEK and Akt pathways through their master regulator c-Met. We observed that activation of c-Met is downregulated in the absence of TBL1XR1. Our study strengthens the contention that TBL1XR1 is a biomarker for prognosis of NSCLC. It may also be considered as an adjunct or core therapeutic target to overcome cisplatin resistance in lung cancers.

Methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) is a bifunctional enzyme located in the mitochondria. It has been reported to be overexpressed in several malignancies. However, the relationship between the expression of MTHFD2 and non-small cell lung cancer (NSCLC) remains largely unknown. In this study, we found that MTHFD2 was significantly overexpressed in NSCLC tissues and cell lines. Knockdown of MTHFD2 resulted in reduced cell growth and tumorigenicity in vitro and in vivo. Besides, the mRNA and protein expression level of cell cycle genes, such as CCNA2, MCM7 and SKP2, was decreased in MTHFD2 knockdown H1299 cells. Our results indicate that the inhibitory effect of MTHFD2 knockdown on NSCLC may be mediated via suppressing cell cycle-related genes. These findings delineate the role of MTHFD2 in the development of NSCLC and may have potential applications in the treatment of NSCLC., (© 2019 The Authors. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)