Small cell lung cancer (SCLC) exhibits a high incidence of perineural invasion (PNI), a clinical feature associated with poor prognosis. Here, we establish PNI as an independent adverse prognostic factor in a surgical SCLC cohort. We further show that the neural microenvironment upregulates stathmin-2 (STMN2) in SCLC cells. STMN2, in a concentration-dependent manner, activates the β-alanine metabolic pathway, leading to intracellular β-alanine accumulation, which enhances tumor cell migration and invasion. In vivo, STMN2 knockdown suppresses neural invasion, an effect reversible upon β-alanine supplementation. This work defines a neural-STMN2-β-alanine-invasion axis that drives PNI in SCLC, providing mechanistic insights and highlighting a promising metabolic vulnerability for therapeutic intervention.

Background: Although combinations of third-generation epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) with chemotherapy or amivantamab have significantly improved progression-free survival (PFS) and overall survival (OS) compared with EGFR-TKI monotherapy in EGFR -mutated advanced non-small cell lung cancer (NSCLC), the increased toxicities and requirement for intravenous administration of chemotherapy limit their practicality. Patients with EGFR -mutated advanced NSCLC who develop M1c2 disease or exhibit elevated programmed death-ligand 1 (PD-L1) expression represent a clinically refractory subgroup with a high unmet need for effective and convenient combination strategies. Objectives: This study is designed to evaluate the efficacy and safety of an all-oral regimen combining vinorelbine and firmonertinib (a third-generation EGFR-TKI) as first-line treatment in this refractory population. Design: This open-label, non-randomized, two-cohort study is conducted at two centers in China. Systematic treatment-naïve patients with histopathology confirmed EGFR -mutated (exon 19 deletion or exon 21 L858R mutation) NSCLC, who are not amenable to curative surgery or radiotherapy, will be recruited. Cohort 1 will comprise 54 metastatic NSCLC patients with M1c2 disease, and Cohort 2 will consist of 77 locally advanced or metastatic NSCLC patients with a PD-L1 tumor proportion score of ⩾25%. Recruitment began on December 11, 2025, and is currently in progress. Methods and analysis: During the one-cycle safety run-in period, six patients received oral vinorelbine (60 mg/m 2 on days 1 and 8) plus firmonertinib (80 mg once daily) as the starting dose. Dosing regimen for the subsequent expansion cohort will be determined based on the safety data from the safety run-in period. The treatment cycle is repeated every 3 weeks and will continue until disease progression or unacceptable toxicity. The primary endpoint is investigator-assessed PFS. Secondary endpoints are objective response rate, disease control rate, OS, and safety. Ethics: This study has received ethics approval from the ethical committee of the Shanghai Chest Hospital (approval number: IS25204). Discussion: To the best of our knowledge, this study represents the first evaluation of vinorelbine plus firmonertinib in EGFR -mutated locally advanced or metastatic NSCLC with M1c2 disease or elevated PD-L1 expression, aiming to provide an effective, tolerable, and convenient all-oral treatment option for this refractory population. Trial registration: The study protocol (version 1.2; July 18, 2025) was registered on October 24, 2025, at the Chinese Clinical Trial Registry (ChiCTR2500111096).

e20726 Background: At present, third-generation epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs) are the standard first-line treatment for advanced EGFR-mutant non-small cell lung cancer (NSCLC). Nevertheless, the combination of first-/second-generation EGFR-TKIs and chemotherapy or anti-angiogenic agents remains commonly used in real-world practice in China. This study aimed to compare the real-world effectiveness of these two first-line strategies. Methods: In this retrospective study, data were collected from Shanghai Chest Hospital from January 2017 to December 2023. Previously-untreated patients with stage III/IV EGFR-mutant NSCLC were enrolled and classified into two groups: those receiving first-line first-/second-generation EGFR-TKI combination therapy, and those receiving third-generation EGFR-TKI monotherapy. Propensity-score matching (PSM) (1:2 ratio) was performed to balance baseline characteristics. The primary endpoint was progression-free survival (PFS). Overall survival (OS) was a secondary endpoint. Results: A total of 512 eligible patients were enrolled and the median follow-up was 40.4 months. After PSM, 364 patients were included. The median PFS was significantly longer in the third-generation TKI monotherapy group (n = 130, PFS: 20.5 months; 95% confidence interval [CI], 16.45-24.55) compared to the first-/second-generation TKI combination group (n = 234, median PFS: 16.0 months; 95% CI, 14.16-17.91; P < 0.001). Subgroup analysis of PFS consistently favored the third-generation EGFR-TKI across almost all variables. No significant difference in OS was observed between the two groups (46.1 months, 95%CI 39.64-52.49 vs. 41.6 months, 95%CI 38.51-45.14; P = 0.721). Besides, 165 (70.5%) patients in the combination group received third-generation EGFR-TKIs in later lines, and this subgroup demonstrated significantly longer OS than those who did not (45.4 vs 33.5 months, P = 0.017). Conclusions: This real-world study demonstrates that first-line third-generation EGFR-TKI monotherapy is associated with a significantly improved PFS compared to first-/second-generation EGFR-TKI combination therapy in EGFR-mutant NSCLC. The absence of OS difference may be attributed to the high proportion of patients in the combination group who subsequently received third-generation TKIs as a later-line therapy. These findings support the superior efficacy of third-generation EGFR-TKIs as the recommended first-line treatment and also indicate that the first-/second-generation EGFR-TKI combination therapy is a viable alternative in real-world practice.

BACKGROUND:Lung cancer remains the primary cause of cancer-related mortality globally, despite significant advancements in therapeutic strategies. Overall survival rates remain unsatisfactory. Chronic inflammation and microRNAs both play pivotal roles in cancer development. METHODS:This study aimed to elucidate the roles of key microRNAs in inflammation-associated non-small cell lung cancer (NSCLC) development. RESULTS:Our findings reveal a significant reduction in miR-125b expression within NSCLC cell lines when stimulated by IL-10. Furthermore, when stimulated by IFN-γ, the expression levels of miR-125b markedly increase. Enforced expression of miR-125b markedly bolstered cell proliferation, migration, and invasion, while diminishing cell apoptosis. Conversely, inhibition of miR-125b produced opposing effects. Mechanistically, DAZAP2 was identified as a direct regulatory target of miR-125b However, because both miR-125b inhibition and DAZAP2 knockdown suppressed malignant phenotypes, DAZAP2 may represent one component of a broader miR-125b-associated regulatory network rather than the sole mediator of miR-125b function. Combined inhibition of miR-125b and DAZAP2 produced more pronounced tumor-suppressive effects both in vitro and in vivo. CONCLUSIONS:Our data suggest that miR-125b and DAZAP2 are involved in the cytokine-responsive regulatory network of inflammation-related NSCLC progression.

Although first-line immunotherapy plus chemotherapy has made substantial progress in extensive-stage small-cell lung cancer (ES-SCLC), the survival benefit remains limited. DURABLE was a prospective, multicenter, open-label, randomized, phase II trial (NCT04985851) that evaluated the efficacy and safety of durvalumab plus anlotinib as consolidative maintenance therapy following first-line durvalumab plus platinum-etoposide chemotherapy in ES-SCLC. The primary endpoint was blinded independent central review-assessed progression-free survival (PFS) from randomization, with a two-sided p-value of <0.20 considered statistically significant. 66 patients were randomly assigned to maintenance therapy with durvalumab plus anlotinib (n = 34) or durvalumab alone (n = 32). Durvalumab plus anlotinib significantly improved PFS compared with durvalumab alone, with median PFS from randomization of 5.4 months versus 1.9 months (HR = 0.64; 80% CI, 0.44-0.94; p = 0.12). The incidence of grade 3-4 treatment-related adverse events in the combination and monotherapy groups was 24.2% and 12.5%, respectively. Patients with impaired antigen presenting capacity or low bTMB tended to show improved outcomes with combined maintenance therapy. While similar efficacy between the two groups was observed in patients with high antigen presenting capacity or high bTMB. These findings suggest that durvalumab plus anlotinib might be an effective and well-tolerated maintenance treatment option in ES-SCLC.

e20146 Background: Extensive-stage small cell lung cancer (ES-SCLC) is a highly aggressive malignancy with limited treatment options. While immunochemotherapy constitutes standard first-line therapy, predictive biomarkers for treatment response remain undefined, impeding personalized therapeutic strategies. Methods: We investigated circulating small extracellular vesicle (sEV)-derived microRNAs (miRNAs) as non-invasive predictive biomarkers in treatment-naïve ES-SCLC patients receiving immunochemotherapy. Plasma samples from a training cohort (n = 33) and an independent prospective cohort (n = 5) were collected pre-treatment. sEVs were isolated, characterized, and subjected to small RNA sequencing to quantify miRNA expression. Results: Small RNA sequencing revealed 23 differentially expressed sEV miRNAs between responders (n = 19) and non-responders (n = 14). Machine learning refined these candidates into a predictive model. Recursive feature elimination (RFE) yielded an 11-sEV-miRNA signature. The top-performing model (Extra Trees Gini) incorporated 5 sEV miRNAs and 1 clinical feature, demonstrating high predictive accuracy in the training set (AUC = 0.855, sensitivity = 95%, specificity = 80%). Preliminary validation in the prospective cohort achieved 80% accuracy (4/5 correct classifications). Conclusions: We established a novel sEV-miRNA biomarker panel that robustly predicts immunochemotherapy response in ES-SCLC. High discriminatory performance and initial prospective validation underscore its clinical utility for guiding treatment decisions and optimizing outcomes by avoiding ineffective therapy in non-responders.

Immunotherapy has revolutionized the treatment landscape of advanced non-small-cell lung cancer (NSCLC). However, its efficacy in patients with epidermal growth factor receptor (EGFR)-mutant NSCLC, particularly after failure of EGFR-tyrosine kinase inhibitors (TKIs), remains unclear and highly heterogeneous across individuals. This difference in treatment response is closely associated with the dynamic remodeling of the tumor microenvironment (TME) driven by the selective pressure of EGFR-TKIs. This review outlines the evolution of the TME during EGFR-TKI treatment and after the development of TKI resistance. Additionally, it summarizes current clinical evidence regarding immunotherapy after EGFR-TKI resistance, and discusses predictive biomarkers and novel therapeutic approaches. By integrating mechanistic insights with clinical translation, this review provides a comprehensive perspective on the immunotherapy landscape after TKI failure, aiming to identify patient subgroups most likely to benefit from immunotherapy and optimize treatment strategies for EGFR-mutant NSCLC.

Oncogenic KRAS mutations are frequently detected in NSCLC. It remains a major challenge to target all KRAS mutants. MEK inhibitors are considered candidates for treating KRAS-mutant NSCLC; however, their easy adaptive resistance precludes further application. Here, we found that MEK inhibitor-trametinib treatment results in the feedback activation of multiple receptor tyrosine kinases (RTKs) and that treatment with the pan-RTK inhibitor anlotinib effectively inhibits the progression of KRAS-mutant NSCLC. Furthermore, we evaluated this strategy in a clinical study (NCT04967079) involving 33 advanced non-G12C KRAS-mutant NSCLC patients. The phase Ia containing 13 patients showed that the recommended phase 2 dose (RP2D) is trametinib (2 mg) plus anlotinib (8 mg), the objective response rate (ORR) is 69.2% (95% CI: 38.6-90.9), the median progression-free survival (PFS) is 6.9 months (95% CI: 3.9 to could not be evaluated), disease control rate (DCR) is 92% (95% CI: 64.0–99.8) and the rate of adverse events (AEs) ≥grade 3 is 23%. The phase Ib containing 20 patients demonstrated the high efficacy of this combinational therapy with RP2D, with the ORR at 65% (95% CI: 40.8–84.6), the median PFS is 11.5 months (95% CI: 8.3–15.5), the median overall survival (OS) is 15.5 months (95% CI: 15.5 to could not be evaluated), the DCR at 100% (95% CI: 83.2–100.0), the median duration of overall response (DoR) is 9.3 months (95% CI: 2.5–12.1), and the rate of AEs ≥ grade 3 at 35%. Overall, this study provides a potential combinational therapeutic strategy for KRAS-mutant NSCLC through the cotargeting of MEK and RTKs.

Background:Treatment of tyrosine kinase inhibitor (TKI)-resistant anaplastic lymphoma kinase (ALK) rearranged non-small cell lung cancer (NSCLC) remains an unmet need. Among these patients, the efficacy of immunotherapy has not been thoroughly investigated. The purpose of our study was to evaluate the efficacy of immunotherapy in patients with ALK-TKI-resistant NSCLC, stratified by programmed cell death ligand-1 (PD-L1) expression. Methods:We retrospectively collected the data of advanced NSCLC patients with ALK-rearrangement, who were treated with immunotherapy or chemotherapy after the development of ALK-TKI resistance at the Shanghai Chest Hospital. Progression-free survival (PFS) was used to evaluate the outcomes. Results:The final analysis included 89 patients between June 1, 2018, and December 31, 2022, who met the selection criteria. The entire cohort had a median follow-up time of 33.4 months. The patients who received immunotherapy had better PFS than those who received non-immunotherapy (median PFS: 5.3 vs. 2.5 months; P=0.009). The PD-L1-positive patients who received immunotherapy had a median PFS of 7.1 months, while those who received non-immunotherapy had a median PFS of 2.5 months (P=0.02). No such statistically significant difference was observed in the PD-L1-negative patients (median PFS for with immunotherapy vs. without immunotherapy: 1.5 vs. 2.9 months; P=0.68). The PD-L1-positive patients who underwent re-biopsy after the development of TKI resistance and who received immunotherapy had a PFS of 7.8 months, while those who received non-immunotherapy had a PFS of 2.7 months (P=0.002). Conclusions:This was the first real-world retrospective study to show that some patients with positive PD-L1 expression may benefit from immune-based therapy after the development of ALK-TKI resistance. However, we still recommend biopsy for patients who develop ALK-TKI resistance to provide further treatment guidance.

Background:As a rare tumor with poor prognosis, the first-line treatment strategy of advanced SMARCA4-deficient thoracic tumors is inconclusive. Although previous studies have shown immunotherapy to be effective, the efficacy and safety of different immune checkpoint inhibitors (ICIs) combination treatment strategies have not been explored in detail. This study aims to identify optimal immunotherapeutic combinations for this population. Methods:We collected the clinical and pathological information of 55 patients with SMARCA4-deficient non-small cell lung cancer (SMARCA4-deficient NSCLC) and SMARCA4-deficient undifferentiated tumor (SMARCA4-deficient UT), after which we evaluated and analyzed the survival status and clinicopathological characteristics of the patients. Results:Following statistical analysis, it was found that the patients were mainly male smokers with a mean age of 66 years (range, 46-81 years old). Histologically, NSCLC accounts for the majority (n=40, 72.7%). Survival analysis demonstrated that overall survival (OS) was significantly longer in patients who received first-line immunotherapy compared to those who did not receive immunotherapy for first line (21.67 vs. 8.80 months, P=0.003). A trend of prolonged OS was observed in patients who received immunotherapy in the first line compared with those who received immunotherapy in the latter line (21.67 vs. 15.30 months, P=0.14). Furthermore, the OS of patients who received anti-angiogenesis therapy plus immunotherapy was superior to that of patients who received three other first-line treatments [not reached vs. 21.67 months (chemotherapy plus immunotherapy) vs. 8.80 months (chemotherapy plus anti-angiogenesis therapy) vs. 7.83 months (chemotherapy), P=0.02]. Conclusions:The early use of ICI-based treatment may result in superior survival outcomes for these patients with SMARCA4-deficient thoracic tumors compared to other treatment modalities. Besides, ICIs combined with anti-angiogenesis therapy may be a potential first-line treatment.

T-cell immune checkpoint blockers (ICBs) and natural killer (NK) cell activation have emerged as promising strategies for cancer therapy in recent years. In this approach, ICBs target inhibitory receptors on cytotoxic immune cells, such as programmed death Protein 1 (PD-1)/programmed cell death-ligand 1 (PD-L1), to enhance immune cell cytotoxicity against cancer cells in a CD8+ T cell-dependent manner. Meanwhile, NK cells play a critical role in immunosurveillance through their direct cytotoxic effects, which do not require prior activation. NK cell activation is mediated by receptors such as NK Group 2 member D (NKG2D), which regulates NK cell function and cytotoxicity through the upregulation of cytokine production. Individually, these treatments target only a limited subset of cancer patients and often face great resistance rates after treatment. However, combining ICBs with NK cell activation may produce a synergistic therapeutic effect, potentially improving treatment outcomes. This perspective article discusses the mechanisms of action of T cell-related PD-1/PDL1 pathways and NK cell activation through NKG2D, examining current studies that provide a rationale for combined NK/T cell combination therapy. The potential of this dual-combination approach to enhance anti-tumor immunity is highlighted. Future perspectives suggest the potential development of chimeric antibodies targeting both T cells and NK cells as a novel therapeutic strategy for cancer treatment.

Background:Kirsten rat sarcoma homolog (KRAS) mutations are one of the key drivers in non-small cell lung cancer (NSCLC) and FDA-approved specific inhibitors of KRAS-G12C mutation are available clinically. However, inhibitors of certain KRAS mutation subtypes remain unavailable, especially rare KRAS mutations including G13C, G13D, and Q61H. In this study, we retrospectively investigated the outcomes of NSCLC patients with rare KRAS-mutation to determine if they may benefit from immune checkpoint inhibitors (ICIs). Methods:Our retrospective study involved 240 advanced NSCLC patients with KRAS mutations, who visited Shanghai Chest Hospital from July 2018 to July 2021. Complete clinical and pathological data were recorded and progression-free survival (PFS) and overall survival (OS) were adopted as primary endpoints. Results:The median follow-up time was 36.5 months (range, 30.8-42.1 months) and the median OS was 9.7 months (range, 7.6-11.8 months). Of the 240 patients evaluated, 130 (54.2%) received chemotherapy and 110 (45.8%) received ICI-based treatment. Among the patients who received chemotherapy, patients with rare KRAS-mutations presented worse survival outcomes (median PFS, 3.4 vs. 4.1 months, P=0.047; median OS, 5.2 vs. 7.1 months, P=0.02) than conventional KRAS-mutant patients. PFS and OS of rare KRAS-mutation patients were prolonged after immunotherapy (median PFS 7.3 vs. 3.4 months, P<0.001; median OS, 13.3 vs. 5.2 months, P<0.001) and had no significant difference compared with conventional KRAS-mutant patients, in part of them whose programmed death-ligand 1 (PD-L1) expression data before immunotherapy were available (n=72), patients with a higher rate of PD-L1 positive tumor cells (≥50%) presented elevated PFS and OS. Conclusions:Despite having potential survival disadvantage compared with other NSCLC patients, rare KRAS-mutant patients (other than G12A, C, D, V) could benefit specifically from ICI-based therapy and survival outcomes are correlated with PD-L1 expression.

This research explores the potential of multimodal fusion for the differential diagnosis of early-stage lung adenocarcinoma (LUAD) (tumor sizes < 2 cm). It combines liquid biopsy biomarkers, specifically extracellular vesicle long RNA (evlRNA) and the computed tomography (CT) attributes. The fusion model achieves an impressive area under receiver operating characteristic curve (AUC) of 91.9% for the four-classification of adenocarcinoma, along with a benign-malignant AUC of 94.8% (sensitivity: 89.1%, specificity: 94.3%). These outcomes outperform the diagnostic capabilities of the single-modal models and human experts. A comprehensive SHapley Additive exPlanations (SHAP) is provided to offer deep insights into model predictions. Our findings reveal the complementary interplay between evlRNA and image-based characteristics, underscoring the significance of integrating diverse modalities in diagnosing early-stage LUAD.

Purpose To investigate the relationship between food insecurity and cognitive decline among elderly Americans.Methods Utilizing NHANES 2011-2014 data, we examined cognitive function via the Immediate Recall Test (IRT), Delayed Recall Test (DRT), Animal Fluency Test (AFT), Digit Symbol Substitution Test (DSST) and assessed food security through the US Food Security Survey Module. Multiple regression models were used to adjust for demographic and health variables.Results Food insecurity demonstrated a significant association with lower cognitive function scores. The effects of food insecurity on cognitive function were moderated by factors such as smoking and alcohol use, indicating a direct influence of food insecurity on cognitive decline.Conclusion This study underscores the importance of food security for cognitive health in the elderly and advocates for targeted interventions to address nutritional disparities and enhance cognitive functioning in aging populations.

Background:Nanocarrier platforms have been indicated to have great potential in clinical practice to treat non-small cell lung cancer (NSCLC). Our previous Phase III clinical study revealed that polymeric micellar paclitaxel (Pm-Pac) is safe and efficacious in advanced NSCLC patients. However, the histopathological-toxicological profile of Pm-Pac in mammals remains unclear.Methods:We examined the Pm-Pac-induced antitumour effect in both A549/H226 cells and A549/H226-derived xenograft tumour models.. And then, we evaluated the short-term and long-term toxicity induced by Pm-Pac in healthy Sprague‒Dawley (SD) rats. The changes in body weight, survival, peripheral neuropathy, haematology, and histopathology were studied in SD rats administered Pm-Pac at different dosages.Results:In the A549-derived xenograft tumour model, better therapeutic efficacy was observed in the Pm-Pac group than in the solvent-based paclitaxel (Sb-Pac) group when an equal dosage of paclitaxel was administered. Toxicity assessments in healthy SD rats indicated that Pm-Pac caused toxicity at an approximately 2- to 3-fold greater dose than Sb-Pac when examining animal body weight, survival, peripheral neuropathy, haematology, and histopathology. Interestingly, based on histopathological examinations, we found that Pm-Pac could significantly decrease the incidences of paclitaxel-induced brain and liver injury but could potentially increase the prevalence of paclitaxel-induced male genital system toxicity.Conclusion:This study introduces the toxicological profile of the engineered nanoparticle Pm-Pac and provides a novel perspective on the Pm-Pac-induced histopathological-toxicological profile in a rat model.

IntroductionPlatinum-based chemotherapy is still the standard of care for Epidermal growth factor receptor (EGFR) mutated non-small cell lung cancer (NSCLC) patients after developing EGFR-TKI resistance. However, no study focusing on the role of immuno checkpoint inhibitor (ICI) based treatments for EGFR mutated NSCLC patients who carried programmed death ligand 1 (PD-L1) tumor proportion score (TPS) greater than 50% progressed after EGFR-TKI therapy. In this study, we retrospectively investigated the outcomes of ICI-based treatments for EGFR mutated NSCLC patients carried PD-L1 TPS≥50% after developing EGFR-TKI resistance and to explore the population that may benefited from ICI-based treatment.MethodsWe retrospectively collected data of advanced NSCLC patients with EGFR mutations and PD-L1 TPS≥50% who have failed prior EGFR-TKI therapies without T790M mutation at Shanghai Chest Hospital between January 2018 and June 2021. Progression-free survival (PFS) and overall survival (OS) were utilized to evaluate the outcomes of this study.ResultsA total of 146 patients were included. Up to June 20th, 2022, median follow-up was 36.7 months (IQR, 12.5-44.2 months). Among the population, 66 patients (45.2%) received chemotherapy, the remaning (54.8%) received ICI-based treatment, including 56 patients(70.0%) received ICI combined with chemotherapy (IC) and 24 patients (30.0%) received ICI monotherapy (IM). In IC group,31 patients received ICI combined with chemotherapy,19 patients received ICI combined with antiangiogenic therapy and remaing received ICI combined with chemotherapy and antiangiogenic therapy. Survival analysis shown that patients who received ICI-based treatment had better progress-free survival (PFS) and overall survival (OS) compared with those treated with other therapy (median PFS, 10.0 vs. 4.0 months, P<0.001; median OS, 39.5 vs. 24.2 months, P<0.001). What’s more, patients who treated with IC treatment had a superior survival time than those received IM treatment (median PFS, 10.3 vs. 7.0 months, P<0.001; median OS, 41.6 vs. 32.4 months, P<0.001). Subgroup analysis found that the PFS and OS benefit of IC was evident in all subgroups.ConclusionsFor advanced NSCLC patients with EGFR mutations and PD-L1 TPS≥50% who have failed prior EGFR-TKI therapies without T790M mutation, ICI-based treatment could provide a more favorable survival than classical chemotherapy. What’ s more, compared with ICI monotherapy, ICI combined with chemotherapy seems to be the preferred treatment.

Background:Severe mouth opening difficulty may increase the risk of airway management during anesthesia induction, and awake fiberoptic nasotracheal intubation (AFNI) is the first option for patients with orofacial anatomical changes. Case summary:A 54-year-old man was scheduled to undergo wedge resection of the right upper lung in August 2021. The patient had a history of enlarged right maxillary lesion resection and partial right maxillary resection surgery in April 2020, which led to orofacial anatomical changes and severe mouth opening difficulty. To avoid difficult airway-related emergency scenarios, the AFNI was successfully performed through intravenous injection of sufentanil and dexmedetomidine combined with lidocaine topical anesthesia under a conscious state without any uncomfortable feeling or complications. Conclusions:Intravenous injection of sufentanil and dexmedetomidine combined with lidocaine topical anesthesia can be used as an alternative medication scheme to relieve uncomfortable suffering for AFNI in patients with severe mouth opening difficulty.

Dear Editor, In our previous study (NCT03628521), the efficacy and safety of sintilimab plus anlotinib regime have been evaluated upon the patients with advanced non-small cell lung cancer (NSCLC).1 Liquid biopsy plays an important role in screening responders in clinical practice.2-5 Therefore, understanding the circulating tumour DNA (ctDNA) as biomarker for monitoring the therapeutic outcome potentially provides new insights into the combined regime-based stratification. In the present study, we performed an exploratory study to screen the potential ctDNA-based biomarker. To further understand the underlying biomarker for stratifying the responders, we performed the ctDNA analysis via customized-panel consists of 1021 genes6-8 (Geneplus, China; Table S1) at four timepoints, that was baseline (BL), before cycle 2 (C2), best response (BR), all the way to after progression disease (PD). Our data showed a ctDNA positive rate of 84.2% at BL, 60% at C2, 50% at BR, and 70% at PD (Figure 1A). The correlation analysis of blood tumour mutational burden (bTMB) and tissue tumour mutational burden (tTMB) at BL indicated that there is a favourable consistency between bTMB and tTMB, suggesting bTMB could serve as a long-term monitoring for evaluating immunotherapy outcomes (Figure S1). Consequently, we attempted to analyse the association between bTMB and progression-free survival (PFS)/overall survival (OS). Regrettably, results showed that bTMB was not associated with PFS/OS in the NSCLC patients who received sintilimab plus anlotinib therapy (Figure S2A–-D). Besides, we also performed the association analysis between the brain metastasis, tumour stage or PD-L1 expression with PFS/OS, and the results indicated that these clinical characteristics are not associated with prognosis in this cohort (Figure S3). Furthermore, our data indicated that the maximum somatic allele frequency (MSAF) corrected bTMB was also not correlated with PFS/OS (Figure S4). Therefore, we wondered that whether the MSAF could be used as biomarker for stratifying the responders who received sintilimab plus anlotinib therapy. Interestingly, our results indicated that the NSCLC patients with high MSAF received more PFS benefit than those with low MSAF at BL (p = .033; Figure 1B). Furthermore, Kaplan–Meier curve analysis showed that the NSCLC patients with high MSAF received a median PFS of 836 days, while the patients with low MSAF received a median PFS of 322 days at BL (p = .011; Figure 1C). Regrettably, we could not obtain a similar result when the MSAF was used for OS stratification (p = .085; Figure 1D). These results suggested that the MSAF potentially be used as a biomarker for PFS stratification but not OS stratification. Furthermore, we found that the MSAF levels of majority patients decreased significantly at C2 and then maintain a relative stability during the later therapy in this clinical trial (Figure 1E–H, Figure S5A–C). Therefore, we would like to know that whether the MSAF alterations from BL to C2 can be used as a valuable biomarker for PFS/OS stratification. Firstly, we defined the MSAF alterations from BL to C2 as δMSAF, which calculated via the MSAF of BL minus the MSAF of C2, then divided by MSAF of BL. Secondly, we performed Cox regression to investigate the association between MSAF/δMSAF and PFS/OS. The results indicated that the hazard ratios derived from δMSAF stratification show better performance than those of MSAF (Figure 2A–F). Lastly, we performed PFS/OS analysis via δMSAF-based stratification. Results suggested that the NSCLC patients with high δMSAF received more PFS benefit than those with low δMSAF the patients (p = .029; Figure 2G). Furthermore, Kaplan–Meier curve analysis showed that the NSCLC patients with high δMSAF received more PFS and OS benefit than those patients with low δMSAF (PFS: p = .027, Figure 2H; OS: p = .032, Figure 2I). To further understand the association between δMSAF and tumour volume, we performed the analysis of the images of computed tomography (CT) as well as the δMSAF for each patient. Among the 18 patients who have δMSAF, the δMSAF values of majority patients are greater than zero (Figure 2J). The results of combined analysis of CT images and δMSAF indicated that the greater the tumour volume shrinked, the greater the δMSAF obtained, suggesting the more potential predictive value of the δMSAF for responsive stratification of the NSCLC patients who received sintilimab plus anlotinib as 1st line therapy (Figure 2K,L, Figures S6 and S7). Lastly, we evaluated the predictive value of MSAF, which derived from C2 and BR. Results suggested that the MSAF both from the C2 and BR also have predictive value (Figure S8). Based on the abovementioned results, we offered the first evidence that the dynamic alterations of ctDNA associate with the therapeutic outcome in the advanced NSCLC patients who received sintilimab plus anlotinib regime as 1st line therapy. Previously, we reported that the MSAF at BL potentially guided PFS stratification for the advanced NSCLC patients who received anlotinib as 3rd or more line therapy.9 Consistence with previous results, here, we also found that the MSAF at BL potentially guided PFS stratification for the advanced NSCLC patients who received sintilimab plus anlotinib as 1st line therapy. However, the MSAF at BL could not guide the OS stratification. Interestingly, here, we found the δMSAF not only has predictive value for guiding PFS stratification but also has predictive value for guiding OS stratification. Therefore, the δMSAF provided the potential value for clinical practice. The limitations of this study were summarized. Firstly, the present cohort consisted a limited sample size due to the biomarker screening based on the phase I clinical trial (NCT03628521). However, the phase II clinical trial (NCT04124731) consisted of 89 patients has been finished during November 2019 to July 2022. The corresponding validation will be performed in the extensive cohort, and the validated results will be opened in future. Secondly, the present study found the high MSAF associated with better PFS; however, the previous studies showed the low MSAF associated with better PFS.10 For understanding the phenomenon, the association between blood TMB and MSAF was interrogated, and the results demonstrated that MSAF and bTMB were positively correlated (Figure S9). This result provided evidence that MSAF could be used as the potential biomarker for the regime of sintilimab plus anlotinib. In any event, the contradictory results provided a novelty thought for developing the predictive biomarker-δMSAF. Collectively, this study provided a candidate biomarker-δMSAF that potentially be used for stratifying the responders for the NSCLC patients who received sintilimab and anlotinib as the 1st line therapy and offered a novelty perspective for screening liquid biopsy-based biomarker via ctDNA sequencing. This work was funded by Medical innovation project of Scientific and Technological innovation action plan of the Shanghai Committee of Science and Technology (grant number: 21Y11913500), Foundation of Shanghai Chest Hospital (project number: YJXT20190102), National Natural Science Foundation of China grants (project number: 82272913), Shanghai Jiao Tong University (grant number: YG2021QN121), Chinese Society of Clinical Oncology (project numbers: Y-2019AZZD-0355 and Y-QL2019-0125) and National Multi-disciplinary Treatment Project for Major Disease (project number: 2020NMDTP). JY, CZ, JL and XX are current employees of Geneplus Company. No other actual or potential conflict of interest is declared. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.