Cytological features of stromal spindle cells and their prognostic significance in lung adenocarcinoma.

INTRODUCTION: Cancer-associated fibroblasts (CAFs) in the tumour microenvironment play a key role in tumour development, proliferation, invasion, and metastasis. The cytological features of spindle cells including CAFs-defined as stromal spindle cells (SSCs) adjacent to cancer cells-are frequently encountered in pulmonary adenocarcinomas. This study aimed to investigate the association between the presence of SSCs in cytological specimens and the clinicopathological features. METHODS: We evaluated 211 patients with pulmonary adenocarcinoma who underwent surgical resection. All participants had cytological specimens corresponding to the histological specimens available for review. RESULTS: Of the 211 cases examined, 89 were SSC-positive (SSC+ ) and 122 were SSC-negative (SSC- ). SSC+ cases were more frequently associated with higher pathological stage (P < 0.001), lymph node metastasis (P = 0.002), anaplastic lymphoma kinase (ALK) gene rearrangement (P = 0.04), high tumour grade (P < 0.001), solid and micropapillary predominant pattern (P = 0.02), and lymphatic vessel (P = 0.003), blood vessel (P < 0.001), and pleural invasion (P = 0.03) as compared to SSC- cases. Patients with SSC+ adenocarcinoma had a significantly shorter recurrence-free survival than those with SSC- adenocarcinoma (P = 0.009). Cytologically, necrotic background (P = 0.002), mucinous cancer cells (P = 0.02), pleomorphic cells (P < 0.001), and mutual cell inclusions (P = 0.01) were observed more frequently in SSC+ adenocarcinomas. CONCLUSIONS: The presence of SSCs could be an important cytological feature for predicting poor prognosis in lung adenocarcinomas.

CD73 expression defines immune, molecular, and clinicopathological subgroups of lung adenocarcinoma.

INTRODUCTION: CD73 is a membrane-bound enzyme crucial in adenosine generation. The adenosinergic pathway plays a critical role in immunosuppression and in anti-tumor effects of immune checkpoint inhibitors (ICI). Here, we interrogated CD73 expression in a richly annotated cohort of human lung adenocarcinoma (LUAD) and its association with clinicopathological, immune, and molecular features to better understand the role of this immune marker in LUAD pathobiology. MATERIALS AND METHODS: Protein expression of CD73 was evaluated by immunohistochemistry in 106 archived LUADs from patients that underwent surgical treatment without neoadjuvant therapy. Total CD73 (T +) was calculated as the average of luminal (L +) and basolateral (BL +) percentage membrane expression scores for each LUAD and was used to classify tumors into three groups based on the extent of T CD73 expression (high, low, and negative). RESULTS: CD73 expression was significantly and progressively increased across normal-appearing lung tissue, adenomatous atypical hyperplasia, adenocarcinoma in situ, minimally invasive adenocarcinoma, and LUAD. In LUAD, BL CD73 expression was associated with an increase in PD-L1 expression in tumor cells and increase of tumor-associated immune cells. Stratification of LUADs based on T CD73 extent also revealed that tumors with high expression of this enzyme overall exhibited significantly elevated immune infiltration and PD-L1 protein expression. Immune profiling demonstrated that T-cell inflammation and adenosine signatures were significantly higher in CD73-expressing lung adenocarcinomas relative to those lacking CD73. CONCLUSION: Our study suggests that higher CD73 expression is associated with an overall augmented host immune response, suggesting potential implications in the immune pathobiology of early stage lung adenocarcinoma. Our findings warrant further studies to explore the role of CD73 in immunotherapeutic response of LUAD.

Usefulness of Plasma Exosomal MicroRNA-451a as a Noninvasive Biomarker for Early Prediction of Recurrence and Prognosis of Non-Small Cell Lung Cancer.

OBJECTIVES: The aim of this study was to clarify the usefulness of plasma exosomal microRNA-451a (miR-451a) as a novel biomarker for the early prediction of recurrence and prognosis in non-small cell lung cancer (NSCLC) patients after curative resection. METHODS: Before surgery, plasma samples were collected and exosomal microRNA (miRNA) levels were evaluated. We first profiled specific exosomal miRNAs related to recurrence in 6 NSCLC patients with stage IA cancer by miRNA microarray. We then validated the usefulness of selected miRNAs as biomarkers using the other 285 NSCLC patients. RESULTS: Plasma exosomal miR-451a showed the highest upregulation in the NSCLC patients with recurrence in the miRNA microarray analysis. A significant positive correlation was demonstrated between exosomal miR-451a levels and NSCLC tissue miR-451a levels. Exosomal miR-451a showed a significant association with lymph node metastasis, vascular invasion, and stage. In stage I, II, or III patients, the overall survival (OS) and disease-free survival (DFS) rates among the high-exosomal-miR-451a patients were significantly worse than those among the low-exosomal-miR-451a patients. In Cox multivariate analysis, exosomal miR-451a showed significance for OS and DFS. CONCLUSION: Plasma exosomal miR-451a might serve as a reliable biomarker for the prediction of recurrence and prognosis in NSCLC patients with stage I, II, or III cancer.

Nuclear factor-kappa B influences early phase of compensatory lung growth after pneumonectomy in mice.

BACKGROUND: Compensatory lung growth (CLG) is a well-established lung regeneration model. However, the sequential mechanisms, including unknown molecular triggers or regulators, remain unclear. Nuclear factor- kappa B (NF-κB) is known to be essential for inflammation and tissue regeneration; therefore, we investigated the role of NF-κB in CLG. METHODS: C57BL/6 J mice underwent either a left pneumonectomy or a thoracotomy (n = 77). Gene microarray analysis was performed to detect genes that were upregulated at 12 h after pneumonectomy. NF-κB protein expression was examined by immunohistochemistry and Western blot. To investigate the influence of NF-κB on CLG, either an NF-κB inhibitor SN50 or saline was administered following pneumonectomy and the degree of CLG was evaluated in each group by measuring the lung dry weight index (LDWI) and the mean linear intercept. RESULTS: Gene microarray analysis identified 11 genes that were significantly but transiently increased at 12 h after pneumonectomy. Among the 11 genes, NF-κB was selected based on its reported functions. Western blot analysis showed that NF-κB protein expression after pneumonectomy was significantly higher at 12 h compared to 48 h. Additionally, NF-κB protein expression at 12 h after pneumonectomy was significantly higher than at both 12 and 48 h after thoracotomy (p < 0.029 for all). NF-κB protein expression, evaluated through immunohistochemistry, was expressed mainly in type 2 alveolar epithelial cells and was significant increased 12 h after pneumonectomy compared to 48 h after pneumonectomy and both 12 and 48 h after thoracotomy (p < 0.001 for all). SN50 administration following pneumonectomy induced a significant decrease in NF-κB expression (p = 0.004) and LDWI compared to the vehicle administration (p = 0.009). CONCLUSIONS: This is the first report demonstrating that NF-κB signaling may play a key role in CLG. Given its pathway is crucial in tissue regeneration of various organs, NF-κB may shed light on identification of molecular triggers or clinically usable key regulators of CLG.

Predictors of long-term compensatory response of pulmonary function following major lung resection for non-small cell lung cancer.

BACKGROUND AND OBJECTIVE: Long-term pulmonary function which might include compensatory response (CR) significantly influences quality of life of long-term survivor after major lung resection. We investigated long-term pulmonary function after major lung resection. METHODS: A total of 137 patients who had undergone lobar resection for non-small cell lung cancer (NSCLC) from May 2013 to June 2014 had spirometry at 10-14 months after surgery. Actual post-operative forced expiratory volume in 1 s (FEV1 ) (FEV1apo )/predicted post-operative FEV1 (FEV1ppo ), actual post-operative forced vital capacity (FVC) (FVCapo )/predicted post-operative FVC (FVCppo ), its relationship with clinicopathological factors and immunohistochemistry for pro-surfactant protein C (pro-SPC), thyroid transcription factor-1 (TTF-1) and vascular endothelial growth factor receptor 2 (VEGFR2) were investigated. RESULTS: FEV1apo /FEV1ppo showed strong correlation with FVCapo /FVCppo (r = 0.628; P < 0.001). We defined greater CR as both FEV1apo /FEV1ppo and FVCapo /FVCppo were >120%. Greater CR was significantly associated with decreased smoking index (P < 0.001) and greater resected subsegments (P = 0.037). The never-smoker group revealed significantly greater CR compared with the smoker group in both FEV1apo /FEV1ppo (119.9 ± 12.5% vs 107.5 ± 14.2%; P = 0.030) and FVCapo /FVCppo (117.9 ± 9.98% vs 107.2 ± 13.1%; P = 0.046) in case-matched comparison. The expression of pro-SPC, TTF-1 and VEGFR2 in the normal lung parenchyma of greater CR group was significantly higher than those of lesser CR group (P < 0.001 for each). In addition, pro-SPC, TTF-1 and VEGFR2 expressions showed a significant correlation to the degree of CR especially in the smoker group (r = 0.631, 0.705 and 0.732, respectively; P < 0.001 for each). CONCLUSION: Our data suggest that smokers may develop lesser long-term CR after major lung resection. Decreased expression of pro-SPC, TTF-1 and VEGFR2 may indicate decreased capacity of CR, especially in patients who smoke.

Optimized magnitude of cryosurgery facilitating anti-tumor immunoreaction in a mouse model of Lewis lung cancer.

BACKGROUND: Cryosurgery has reemerged as a less invasive local treatment with possible immune-regulatory effects. However, the optimal magnitude of cryosurgery for achieving immune-regulatory responses at abscopal tumor sites remains unclear. We aimed to investigate appropriate magnitude of cryosurgery for this goal using a mouse model. METHODS: C57BL/6J mice were inoculated with Lewis lung carcinoma cells or B16 melanoma cells in bilateral flanks. The left-sided tumor was cryoablated with repeated freeze/thaw cycles either once, twice, or thrice. The peritumoral injections of LPS were performed. Abscopal tumor volumes were measured, immunohistochemistry was performed for CD4, CD8, Foxp3, and Ki-67, and proinflammatory cytokines were measured in lavage fluid of cryoablated tumor. RESULTS: The growth rate of the abscopal tumor was slowest in the Cryosurgery ×2 group among the five experimental groups. The proportions of CD4(+) T cells and CD8(+) T cells in the abscopal tumor were also significantly higher in the Cryosurgery ×2 group. The levels of IL-1ß, IL-2, IL-6, IL-12ß, IFN-γ, and TNF-α in the peritumoral lavage fluid in Cryosurgery ×2 + LPS group were significantly increased compared with the other groups. CONCLUSIONS: This study suggested that achievement of approximately 73 % damaged area in the cryoablated tumor by two cycles of cryosurgery generates the most favorable immune-regulatory response for abscopal tumors via activation of anti-tumor immune cells as well as increased secretion of proinflammatory cytokines.

Efficacy and hemodynamic response of pleural carbon dioxide insufflation during thoracoscopic surgery in a swine vessel injury model.

PURPOSES: Thoracoscopic anatomical lung resection is a minimally invasive technique, but intraoperative massive bleeding is a critical complication. We investigated the hemostatic efficacy and safety of intrapleural carbon dioxide (CO2) insufflation in thoracoscopic surgery in a swine vessel injury model. METHODS: Swines were assigned to one of four groups subjected to thoracoscopic surgery under target intrathoracic pressures of 0, 5, 10, or 15 mmHg CO2 insufflation, respectively. A pin-hole injury of the right cranial lobe pulmonary vein was inflicted thoracoscopically and we compared the blood loss and hemodynamic changes in each group. RESULTS: There were no signs or echographic findings of air embolus. Both the blood loss per minute and total blood loss during the experiment were significantly lower in the 10 and 15 mmHg groups than in the 0 mmHg group (p > 0.05, respectively). The hemodynamic signs, including heart rate, mean arterial pressure, and peripheral oxygen saturation, were not significantly different in the 0 and 10 mmHg groups at most times, although they were significantly correlated with the insufflation pressure during the experiments (p < 0.05). CONCLUSIONS: CO2 insufflation in thoracoscopic major lung resection appears to be safe, even in the short term, and can help to control vessel injury.

Pregabalin reduces post-surgical pain after thoracotomy: a prospective, randomized, controlled trial.

PURPOSE: A new perioperative management method was explored by assessing the safety and the efficacy of pregabalin for the treatment of intercostal neuralgia after thoracotomy. METHODS: The study was conducted on 68 adult patients scheduled to undergo thoracotomy. Patients were randomly divided into two groups; a NSAIDs group, where 34 patients were orally administered loxoprofen three times daily and a pregabalin group, where 34 patients were orally administered 75 mg of pregabalin twice daily, starting on the day of surgery and continuing for 2 weeks. The pain scores, sleep interference and the incidence of neuropathic pain were evaluated on days 1, 3 and 7, and at weeks 4, 8 and 12 after surgery. The frequency of pain medication use in the first week after surgery and the adverse effects in each group were also examined. RESULTS: The pain scores, sleep interference and incidence of neuropathic pain were significantly lower (p < 0.001) at all time points in the pregabalin group. The use of additional pain medication during the first week after surgery was not significantly different between the groups. The only significant adverse effect was a stomach ache in the NSAIDs group, while mild drowsiness was reported in the pregabalin group. CONCLUSION: Pregabalin is considered to be an effective and safe drug for the treatment of pain after thoracotomy.

Mediastinal hemorrhage due to ectopic parathyroid hyperplasia with long-term hemodialysis: report of a case.

Secondary hyperparathyroidism can sometimes occur among long-term hemodialysis patients. We herein present the case of a 48-year-old female who underwent surgical resection to treat a mediastinal hemorrhage from an ectopic parathyroid. She had been receiving dialysis for the past 16 years due to renal failure. She visited the hospital due to chest pain, and a CT scan revealed a tumorous lesion in the anterior mediastinum. An increase in size of the tumorous lesion, accompanied by bilateral pleural fluid, was observed. Emergency surgery was performed due to a diagnosis of a mediastinal hemorrhage from the tumorous lesion, accompanied by a decrease in the Hb value. The pathological findings of the hematoma revealed parathyroid hyperplasia. This is a rare case report of an ectopic parathyroid developing hyperplasia which caused a mediastinal hemorrhage due to secondary hyperparathyroidism.

[Castleman's Disease of the Chest Wall Successfully Resected by Thoracoscopic Surgery；Report of a Case].

A 61-year-old woman without a significant past medical history was pointed out the abnormal shadow on the annual medical checkup. Chest computed tomography (CT) revealed a well-defined paravertebral chest wall tumor of 20 mm in maxmum size. Furthermore, diffusion weighted image on magnetic resonance imaging (MRI) showed high intensity, and standardized uptake value (SUV) max on positron emission tomography/computed tomography (PET/CT) was 13.4. Schwanoma, solitary fibrous tumor (SFT) or malignant lymphoma was suggested. Complete resection was performed with thoracoscopic surgery. The histological diagnosis was Castleman's disease with hyalineized type.

A Comparison of the Efficacies of OK-432 and Talc Slurry for Pleurodesis in Patients with Prolonged Air Leak after Pulmonary Resection.

PURPOSE: A prolonged air leak (PAL) is one of the common postoperative complications of pulmonary resection. The aim of this study was to evaluate the efficacy and safety of pleurodesis with sterile talc or OK-432 for postoperative air leak. METHODS: Patients with postoperative air leak who received chemical pleurodesis using sterile talc or OK-432 were retrospectively identified from medical records data. For pleurodesis with either agent, prior assessment and approval by the hospital safety department were carried out for each case, in addition to individual consent. RESULTS: Between February 2016 and June 2022, 39 patients had PALs and underwent chemical pleurodesis. Among them, 24 patients received pleurodesis with talc (Talc group) and 15 with OK-432 (OK-432 group). The leak resolved after less than two pleurodesis treatments in 22 patients (91.7%) in the Talc group compared with 14 patients (93.3%) in the OK-432 group. Pleurodesis significantly increased white blood cell counts, C-reactive protein concentration, and body temperature in the OK-432 group compared with that in the Talc group (p <0.001, p = 0.003, and p <0.001, respectively). CONCLUSIONS: Pleurodesis with talc may be an effective treatment option for postoperative air leak. Our findings suggest that talc was as effective as OK-432 and resulted in a milder systemic inflammatory response.

Arterial Embolization and Cone-Beam Computed Tomography-Guided Lung Resection for Anomalous Systemic Arterial Blood Supply to Normal Lung: Two Case Reports.

Systemic arterial blood supply to a normal lung is a rare anatomical abnormality. Surgery is usually indicated because this abnormality leads to pulmonary hypertension. Herein, we report our experience and ideas for safe vessel dissection. Case 1 was a woman in her 50s. We performed a left lower lobectomy following percutaneous coil embolization. The aberrant artery with emboli was confirmed intraoperatively by cone-beam computed tomography (CBCT) to safely dissect under thoracoscopic surgery (TS). Case 2 was a man in his 40s. Following percutaneous endovascular plug occlusion, we performed a left partial resection using indocyanine green fluorescence navigation. Intraoperatively, CBCT imaging demonstrated the aberrant artery and exact position of the emboli. This combination technique of interventional radiology and TS with CBCT imaging was considered safe and more secure for the treatment of anomalous systemic arterial blood supply to a normal lung.

Solitary pulmonary capillary hemangioma mimicking a preinvasive malignant lesion in an asymptomatic middle-aged female patient.

Pulmonary capillary hemangiomatosis (PCH) is a rare disease characterized by a proliferation of capillaries in the alveolar septa, bronchial and venous walls, pleura, and regional lymph nodes. However, the etiology of the disease remains unknown due to its rarity. Therefore, we present a case of a solitary PCH lesion without symptoms in a 38-year-old female patient. According to computed tomography, she was diagnosed with lung carcinoma, indicated by a tiny nodule with ground-glass opacity detected in her right upper lung. However, no other lesions were detected on systemic examination. Consequently, partial lung resection was conducted, since the lesion was suspected of lung adenocarcinoma. Pathologic results showed that the thick alveolar septa were caused by capillary growth without cellular atypia and hardly any infiltration of inflammatory cells. Finally, we diagnosed the pulmonary lesion as PCH, although solitary PCH has previously been reported in a few case reports. Therefore, further case studies are essential to clarify the causes of PCH.

A pilot study of intraoperative localization of peripheral small pulmonary tumors by cone-beam computed tomography: sandwich marking technique.

Background: Intraoperative identification of small pulmonary nodules has been an important technical issue. We aimed to develop a new localization method which is much safer and simple procedure compared with conventional methods. Methods: This was a retrospective study including patients with resected peripheral pulmonary nodules between November 2017 and April 2021 at Teikyo University School of Medicine, and Saitama Cardiovascular and Respiratory Center. All surgical procedure was wedge resection, and the tumor size was equal to or less than 20 mm which were detected by cone-beam computed tomography (CBCT; Philips Allura Xper FD 20, Philips). Some metal clips were put on several places of visceral pleura, where the target lesion was sandwiched by marking clips (sandwich marking technique). CBCT detected both the target lesion and metal clips, and video-assisted thoracoscopic surgery (VATS) was performed. Radiological and pathological findings were analyzed, and the correlation coefficient of tumor size was examined among pre-, intra-, and post-operative tumor sizes. Results: The average age of 90 patients was 65.2 years, and 47 were male (52.2%). All procedure was wedge resection including twelve bi-wedge resections, and one hundred nine peripheral pulmonary lesions were obtained by sandwich marking technique. The detection rate was 100%, and there was no marking-related complication. Conclusions: All small peripheral pulmonary lesions were successfully detected and resected by using CBCT with no marking-related complication. Sandwich marking technique was demonstrated to provide safe, reliable, and simple localization procedure for small peripheral pulmonary lesions.

Immune evolution from preneoplasia to invasive lung adenocarcinomas and underlying molecular features.

The mechanism by which anti-cancer immunity shapes early carcinogenesis of lung adenocarcinoma (ADC) is unknown. In this study, we characterize the immune contexture of invasive lung ADC and its precursors by transcriptomic immune profiling, T cell receptor (TCR) sequencing and multiplex immunofluorescence (mIF). Our results demonstrate that anti-tumor immunity evolved as a continuum from lung preneoplasia, to preinvasive ADC, minimally-invasive ADC and frankly invasive lung ADC with a gradually less effective and more intensively regulated immune response including down-regulation of immune-activation pathways, up-regulation of immunosuppressive pathways, lower infiltration of cytotoxic T cells (CTLs) and anti-tumor helper T cells (Th), higher infiltration of regulatory T cells (Tregs), decreased T cell clonality, and lower frequencies of top T cell clones in later-stages. Driver mutations, chromosomal copy number aberrations (CNAs) and aberrant DNA methylation may collectively impinge host immune responses and facilitate immune evasion, promoting the outgrowth of fit subclones in preneoplasia into dominant clones in invasive ADC.

Resolving the Spatial and Cellular Architecture of Lung Adenocarcinoma by Multiregion Single-Cell Sequencing.

Little is known of the geospatial architecture of individual cell populations in lung adenocarcinoma (LUAD) evolution. Here, we perform single-cell RNA sequencing of 186,916 cells from five early-stage LUADs and 14 multiregion normal lung tissues of defined spatial proximities from the tumors. We show that cellular lineages, states, and transcriptomic features geospatially evolve across normal regions to LUADs. LUADs also exhibit pronounced intratumor cell heterogeneity within single sites and transcriptional lineage-plasticity programs. T regulatory cell phenotypes are increased in normal tissues with proximity to LUAD, in contrast to diminished signatures and fractions of cytotoxic CD8+ T cells, antigen-presenting macrophages, and inflammatory dendritic cells. We further find that the LUAD ligand-receptor interactome harbors increased expression of epithelial CD24, which mediates protumor phenotypes. These data provide a spatial atlas of LUAD evolution, and a resource for identification of targets for its treatment. SIGNIFICANCE: The geospatial ecosystem of the peripheral lung and early-stage LUAD is not known. Our multiregion single-cell sequencing analyses unravel cell populations, states, and phenotypes in the spatial and ecologic evolution of LUAD from the lung that comprise high-potential targets for early interception.This article is highlighted in the In This Issue feature, p. 2355.

Neoadjuvant nivolumab or nivolumab plus ipilimumab in operable non-small cell lung cancer: the phase 2 randomized NEOSTAR trial.

Ipilimumab improves clinical outcomes when combined with nivolumab in metastatic non-small cell lung cancer (NSCLC), but its efficacy and impact on the immune microenvironment in operable NSCLC remain unclear. We report the results of the phase 2 randomized NEOSTAR trial (NCT03158129) of neoadjuvant nivolumab or nivolumab + ipilimumab followed by surgery in 44 patients with operable NSCLC, using major pathologic response (MPR) as the primary endpoint. The MPR rate for each treatment arm was tested against historical controls of neoadjuvant chemotherapy. The nivolumab + ipilimumab arm met the prespecified primary endpoint threshold of 6 MPRs in 21 patients, achieving a 38% MPR rate (8/21). We observed a 22% MPR rate (5/23) in the nivolumab arm. In 37 patients resected on trial, nivolumab and nivolumab + ipilimumab produced MPR rates of 24% (5/21) and 50% (8/16), respectively. Compared with nivolumab, nivolumab + ipilimumab resulted in higher pathologic complete response rates (10% versus 38%), less viable tumor (median 50% versus 9%), and greater frequencies of effector, tissue-resident memory and effector memory T cells. Increased abundance of gut Ruminococcus and Akkermansia spp. was associated with MPR to dual therapy. Our data indicate that neoadjuvant nivolumab + ipilimumab-based therapy enhances pathologic responses, tumor immune infiltrates and immunologic memory, and merits further investigation in operable NSCLC.

Lymphovascular Invasion Is Associated With Mutational Burden and PD-L1 in Resected Lung Cancer.

BACKGROUND: High tumor mutational burden (TMB) and programmed death ligand 1 (PD-L1) expression are leading biomarkers in metastatic non-small cell lung cancer (NSCLC) and predict favorable response to checkpoint inhibitors. We sought to identify clinicopathologic characteristics associated with elevated TMB and PD-L1 expression among patients who underwent resection for NSCLC. METHODS: NSCLC patients undergoing primary resection (2016-2018) were prospectively enrolled in an immunogenomic profiling project. Multiplex immunofluorescence quantified densities (cells/mm2) of CD3+, CD3+CD8+, CD3+CD8+PD-1+, malignant cells (MCs), MCsPD-L1+, CD68+, CD68+PD-L1+, and CD20+ cells. Whole-exome sequencing quantified TMB (mutations/megabase). TMB and MCsPD-L1+ were dichotomized according to the median of each. RESULTS: A total of 55 patients completed multiplex immunofluorescence and whole-exome sequencing profiling. In this sample, 41.8% (23 of 55) had pathologic stage I disease. Median TMB and MCsPD-L1+ were 3.91 and 0.62 cells/mm2, respectively. TMB was higher among smokers (P = .001) and tumors with lymphovascular invasion (LVI) (P = .051). TMB was positively correlated with densities of MCsPD-L1+ (r = 0.293, P = .030), CD68+PD-L1+ (r = 0.289, P = .033), and CD20+ (r = 0.310, P = .043) cells. The density of MCsPD-L1+ was associated with increased CD3+CD8+ (r = 0.319, P = .018) and CD68+PD-L1+ (r = 0.371, P = .005) cells. Patients with PD-L1HighTMBHigh tumors (30.9%, 17 of 55) had higher intratumoral densities of CD3+, CD3+CD8+, CD68+, CD68+PD-L1+, and CD20+ cells. On multivariable analysis LVI was associated with synchronous elevated TMB and PD-L1 expression (odds ratio 3.53, P = .039). CONCLUSIONS: NSCLC tumors with elevated TMB and PD-L1 expression are associated with LVI and increased intratumoral immune cell infiltration. These findings may potentially improve patient selection for checkpoint inhibitor therapy trials in the adjuvant setting.

Neutrophil expansion defines an immunoinhibitory peripheral and intratumoral inflammatory milieu in resected non-small cell lung cancer: a descriptive analysis of a prospectively immunoprofiled cohort.

BACKGROUND: The biological underpinnings of the prognostic and predictive significance of a relative neutrophilia in patients with non-small lung cancer (NSCLC) are undefined. We sought to comprehensively examine the relationships between circulating and intratumoral neutrophil populations and features of the immune contexture in patients undergoing NSCLC resection. METHODS: Preoperative soluble cytokine and angiogenic factors; tumor multiplex immunofluorescence; RNA, whole exome, and T-cell receptor sequencing; and flow cytometry were analyzed for relationships with populations of circulating (from complete blood counts) and intratumoral neutrophils (transcriptional signatures) in a prospectively enrolled resected NSCLC cohort (n=66). In a historical cohort (n=1524), preoperative circulating neutrophil and lymphocyte counts were analyzed for associations with overall survival (OS). RESULTS: Circulating neutrophil populations were positively correlated with increased tumor burden, and surgical tumor resection was followed by a subsequent reduction in peripheral neutrophil counts. Expansion of the circulating neutrophil compartment was associated with increased levels of pro-granulopoietic (IL-1ß, IL-17A, TNFα, IL-6) and TH2-associated (IL-5, IL-13) cytokines. Tumors with high intratumoral neutrophil burden were marked by a blunted T-cell response characterized by reduced expression of cytotoxic T-cell genes (CD8A, CD8B, GZMA, GZMB), decreased CD3+CD8+ cell infiltration, and diminished expression of IFNγ-related genes. The associations between increased intratumoral neutrophil burden and reduced CD3+CD8+ infiltration persisted after adjustment for tumor size, histology, mutational burden, and PD-L1 expression. In 1524 patients, elevated preoperative circulating neutrophil count was independently associated with worse OS (main effect HR 1.82, 95% CI 1.24 to 2.68, p=0.002). CONCLUSIONS: Our findings demonstrate that neutrophil expansion reflects protumorigenic and immunosuppressive processes that manifest as worse OS in patients undergoing NSCLC resection. These results justify further investigation of therapeutic strategies targeting neutrophil-associated immune evasion.