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Cancer tissue is composed of both a cancer cell component and a stromal component. The aim of this study was to investigate if the component ratio predicts a prognosis for lung squamous cell carcinoma (SqCC) patients by using a machine learning method.A total of 135 peripheral SqCC cases (tumor size: 3-5 cm) were enrolled in this study. The areas of the cancer cell component, the necrotic component, and the stromal component were accurately measured via a machine learning method. Each case was divided into the following three subtypes: 1) predominant cancer cell, 2) predominant necrosis, and 3) predominant stroma. The study examined if a particular subtype had prognostic significance.The number of cases per subtype of predominant cancer cell, predominant necrosis, and predominant stroma was 59, 6, and 70, respectively. Patients with the predominant stroma subtype had a significantly shorter recurrence free survival (RFS) than did those with the predominant cancer cell subtype (5-yr RFS: 42.3 % vs. 84.3 %,p0.01). Also, in pathological stage I patients, the 5-year RFS rate for the predominant stroma subtype was significantly shorter (5-yr RFS: 64.3 % vs. 88.4 %, p0.01). In the multivariate analysis of p-stage I patients, the predominant stroma subtype was confirmed to be an independent prognostic factor for RFS (p0.01).Using machine learning, the study confirmed that the predominant stroma subtype was an independent factor for RFS, suggesting that the ratio of the stromal component correlates with the malignant potential of SqCC.

MS-assisted lipidomic tissue analysis is a valuable tool to assess sphingolipid metabolism dysfunction in disease. These analyses can reveal potential pharmacological targets or direct mechanistic studies to better understand the molecular underpinnings and influence of sphingolipid metabolism alterations on disease etiology. But procuring sufficient human tissues for adequately powered studies can be challenging. Therefore, biorepositories, which hold large collections of cryopreserved human tissues, are an ideal retrospective source of specimens. However, this resource has been vastly underutilized by lipid biologists, as the components of OCT compound used in cryopreservation are incompatible with MS analyses. Here, we report results indicating that OCT compound also interferes with protein quantification assays, and that the presence of OCT compound impacts the quantification of extracted sphingolipids by LC-ESI-MS/MS. We developed and validated a simple and inexpensive method that removes OCT compound from OCT compound-embedded tissues. Our results indicate that removal of OCT compound from cryopreserved tissues does not significantly affect the accuracy of sphingolipid measurements with LC-ESI-MS/MS. We used the validated method to analyze sphingolipid alterations in tumors compared with normal adjacent uninvolved lung tissues from individuals with lung cancer and to determine the long-term stability of sphingolipids in OCT compound-cryopreserved normal lung tissues. We show that lung cancer tumors have significantly altered sphingolipid profiles and that sphingolipids are stable for up to 16 years in OCT compound-cryopreserved normal lung tissues. This validated sphingolipidomic OCT compound-removal protocol should be a valuable addition to the lipid biologist's toolbox.

Background: Variations in fibroblast growth factor receptor 1 (FGFR1), which occur frequently, are common driver mutations of lung squamous cell carcinoma. Immune checkpoint inhibitors targeting programmed death-1 (PD-1) and programmed death ligand-1 (PD-L1) are powerful anticancer weapons. Activation of FGFR1 leads to tumorigenesis through multiple downstream molecules, including Yes-associated protein (YAP), but whether and how FGFR1 regulates tumor immune evasion remain largely unclear. Methods: H520 and HCC95 cells were treated with siRNA and plasmids to increase or decrease the expression of FGFR1, YAP and PD-L1, as assessed by molecular assays of protein and mRNA expression. The interaction between YAP and PD-L1 was verified by chromatin immunoprecipitation. After FGFR1 knockdown by shRNA, cancer cells were cocultured with Jurkat T cells, and then cell proliferation and activity were assessed. In C57BL/6 mice, the tumor immune microenvironment was analyzed by flow cytometry, immunofluorescence and immunohistochemistry after FGFR1 knockdown. The effect of the combination of FGFR1 knockdown and PD-1 blockade was explored both in vitro and in vivo. Results: In H520 and HCC95 cells, FGFR1 upregulated PD-L1 expression via YAP, and YAP initiated the transcription of PD-L1 after binding to its promoter region. Both in vitro and in vivo, FGFR1 knockdown decreased tumor growth and reduced immune escape and reactivation of T cells. The combination of FGFR1 knockdown and PD-1 blockade synergistically exerted antitumor effects. In human LSQCC, the expression of fibroblast growth factor 2 (FGF2), the activator of FGFR1, was positively correlated with that of PD-L1 at the mRNA level. Conclusions: The FGFR1/YAP/PD-L1 regulatory axis mediates tumor-associated immune suppression in lung squamous cell carcinoma, and FGFR1 knockdown reactivates T cells in the tumor microenvironment. Synergistic inhibition of both FGFR1 and PD-1/PD-L1 may be a possible treatment for lung cancer patients.

Background There is currently no definite clinical implication for the subtypes of lung squamous cell carcinoma according to the 2015 WHO classification. This study aimed to investigate postoperative recurrence of the two major subtypes of lung squamous cell carcinoma: keratinizing squamous cell carcinoma (KSCC) and nonkeratinizing squamous cell carcinoma (NKSCC). Methods We identified the patients with KSCC and NKSCC who had undergone complete resection in Shanghai Chest Hospital between April 2015 and June 2016. Disease-free survival (DFS) was compared using Kaplan-Meier statistical analysis. Variables selected by univariate analysis were evaluated in multivariate analysis using the Cox proportional hazard model. Results A total of 334 patients included 231 (69.2%) cases with KSCC and 103 (30.8%) cases with NKSCC. There were more smokers in keratinizing than nonkeratinizing subtype (84.8% versus 72.8%, p = 0.009). The percentage of stage Ⅲ was higher in NKSCC than that in KSCC (35% versus 22.9%, p = 0.012). The 2-year DFS rates of stage Ⅰ, stage Ⅱ and stage Ⅲ were 90.1%, 66.4% and 37.7% in KSCC, 83.3%, 67.7% and 52.8% in NKSCC, respectively. There were no significant differences of 2-year DFS rates between KSCC and NKSCC. Furthermore, KSCC and NKSCC had no significant differences in recurrence patterns and metastatic sites. Conclusion There were no significant differences of postoperative recurrence between KSCC and NKSCC.

In 2015, the World Health Organization classification of lung cancer proposed the concept of spread through air spaces (STAS) as a new pattern of invasion in lung adenocarcinoma. The definition of STAS included one or more pathologic micropapillary clusters, solid nests or single cells beyond the edge of the tumor into air spaces in the surrounding lung parenchyma, and separation from the main tumor other than tumor islands. The roles of STAS has been investigated in many studies. The results indicated that STAS is associated with key clinical variables and the prognosis of patients both in lung adenocarcinoma, lung squamous cell carcinoma, small-cell lung cancer, and lung pleomorphic carcinoma. This mini review will be focused on the developments and perspectives of STAS in lung cancer.

BackgroundThis study aims to profile integrative genomic spectra of Chinese patients with different subtypes of lung squamous cell carcinoma (LUSC) and explore potential molecular prognosis factors.MethodsWe retrospectively identified 204 surgically resected LUSC patients in Shanghai Chest Hospital who underwent capture-based targeted next-generation sequencing (NGS) with a panel of 68 lung cancer‐related genes from September 2017 to January 2019. NGS was used to profile comprehensive molecular characterizations.ResultsOf 204 cases, 114 (55.9%) were keratinizing squamous cell carcinoma (KSCC), 77 (37.7%) were non-keratinizing squamous cell carcinoma (NKSCC), 13 (6.4%) were basaloid squamous cell carcinoma (BSCC), respectively. All subtypes presented similarly high proportions of mutations, including TP53, CDKN2A, and NOTCH1. A comparable prevalence of FGFR1 amplifications was identified between KSCC and NKSCC (11.4 versus 26.9%, p = 0.007). Compared with NKSCC, IGF1R amplifications were more frequent in BSCC (0 versus 15.4%, p = 0.019). We found cases with TP53 alterations had less EGFR alterations in KSCC (P = 0.013, OR = 0.158). Compared with TCGA cohorts, our Chinese cohorts exhibited statistic differences in both somatic mutations and signaling pathways. We found that STK 11 alterations and TOP2A alterations were significantly associated with higher risk of recurrence in patients with LUSC.ConclusionsSignificant differences exist among three subtypes of LUSC in molecular characterizations.

Lung cancer remains the leading cancer in incidence and mortality in both genders and most countries. Global cancer statistics show a declining trend of lung squamous cell carcinoma (LSCC) but an uprising trend of lung adenocarcinoma (LADC). The reasons behind their opposite trends are unclear. This study aims to analyze the global trends of LSCC and LADC during 1990–2012 in relation to tobacco consumption and air pollution. Results show a 1% decline of smoking prevalence of 7 years ago is associated with a 9% (95% confidence interval: 8%, 10%) drop in the LSCC incidence globally, whereas a 0.1 μg/m3 increment of BC of 7 years ago is associated with a 12% (9%, 16%) increase in LADC incidence globally. Association between BC and LSCC (or LADC) is more prominent in females, with a 14% (7%, 20%) increase in LSCC [or 14% (11%, 19%) increase in LADC] incidence for a 0.1 μg/m3 increment of BC of 8 (or 6) years ago. Associations vary with different genders across different continents. For instance, concentration of BC is positively associated with incidence of both LSCC and LADC in Europe and North America, whereas concentration of sulfate is positively associated with LSCC incidence in Europe and Oceania, and with LADC incidence in Asia, Oceania and South America. We conclude global decreasing LSCC incidence is associated with the reduced tobacco consumption, whereas the global increasing LADC incidence is likely associated with air pollution. Various particulate species have divergent effects on LADC incidence in different continents.

MicroRNA (miRNA) are attractive markers of lung cancer, due to their regulatory role in cell cycle. However, we know more about function of miRNA in cancer development, there is still little known about role of their precursors (primary miRNA; pri-miRNA) in tumorgenesis. In present study we investigated potential role of miRNA-944 and its precursor pri-miRNA-944 in development of squamous-cell lung cancer (SCC) and explored interdependence between miRNA precursor and its mature form. This is a first available literature report analyzing pri-miRNA as a cancer diagnostic marker.Expression of miRNA-944 and its precursor was analyzed in 58 fresh-frozen tissues of non-small cell lung cancer and corresponding adjacent non-cancerous tissues using qRT-PCR. Expression of pri-miRNA-944 was correlated with TP63 and miRNA-944. Using ROC analysis diagnostic accuracy of studied markers was evaluated.miRNA-944 and its precursor were significantly overexspressed in SCC compared to adenocarcinoma (AC) and non-cancerous tissue. pri-miRNA-944 strongly and positively correlated with TP63 (r = 0.739, p 0.001) and with mature miRNA-944 expression (r = 0.691, p 0.001). Also, TP63 expression significantly correlated with mature miRNA (r = 0.785, p 0.001). Combined analysis of pri-miRNA-944 and mature miRNA-944 allowed to distinguish SCC tissue form AC with sensitivity of 93.3% and specificity of 100% (AUC = 0.978), and SCC from non-cancerous tissue with 92.9% sensitivity and 100% specificity (AUC = 0.992).We assumed that pri-miRNA-944 and miRNA-944 may be involved in early squamous-type differentiation of lung tumors. Moreover, analysis of both markers provided high diagnostic accuracy for SCC detection.

Background Because the relationship between the timing of surgery following diagnosis of lung cancer and survival has not been precisely described, guidelines on what constitutes a clinically meaningful delay of resection of early-stage lung cancer do not exist. This study tested the hypothesis that increasing the time between diagnosis and lobectomy for stage IA squamous cell carcinoma (SCC) would be associated with worse survival. Methods The association between timing of lobectomy and survival for patients with clinical stage IA SCC in the National Cancer Data Base (2006-2011) was assessed using multivariable Cox proportional hazards analysis and restricted cubic spline (RCS) functions. Results The 5-year overall survival of 4,984 patients who met study inclusion criteria was 58.3% (95% CI, 56.3-60.2). Surgery was performed within 30 days of diagnosis in 1,811 (36%) patients, whereas the median time to surgery was 38 days (interquartile range, 23, 58). In multivariable analysis, patients who had surgery 38 days or more after diagnosis had significantly worse 5-year survival than patients who had surgery earlier (hazard ratio, 1.13 [95% CI, 1.02-1.25]; P = .022). Multivariable RCS analysis demonstrated the hazard ratio associated with time to surgery increased steadily the longer resection was delayed; the threshold time associated with statistically significant worse survival was ∼90 days or greater. Conclusions Longer intervals between diagnosis of early-stage lung SCC and surgery are associated with worse survival. Although factors other than the timing of treatment may contribute to this finding, these results suggest that efforts to minimize delays beyond those needed to perform a complete preoperative evaluation may improve survival.

PURPOSE/OBJECTIVE(S) We aimed to evaluate the safety of and maximum tolerated dose (MTD) for 5-fraction stereotactic ablative radiotherapy (SABR) for early-stage lung squamous cell carcinoma (SCC), for which local failure rates appears higher than that of early-stage adenocarcinomas. MATERIALS/METHODS A single institution phase I study was conducted in pathologically confirmed early-stage lung SCC. The 3+3 design involved 4 dose levels of 11, 12, 13, and 14 Gy x 5 fractions delivered every other day (dose levels DL1, DL2, DL3, and DL4, respectively). The primary objective was to determine the MTD of 5-fraction SABR, which was defined as the dose associated with a 33% rate of dose-limiting toxicities, defined as grade ≥3 respiratory/mediastinal or cardiac events (or grade ≥4 gastrointestinal or chest wall events), with toxicities reported until data lock in October 2020. Target volume delineation and organ-at-risk (OAR) constraints were based on RTOG 0813, using OAR constraints expressed in percent of prescription as fixed constraints from the 11 Gy per fraction prescription for all DL. Institutional constraints for the chest wall were used (V30 < 30cm3, up to 70 cm3 acceptable; maximum point dose 52.5 Gy). Target coverage objectives included ≥95% of the planning target volume (PTV) to be covered by 100% of the prescription dose, and 100% of the internal target volume (ITV) to receive ≥110% of the prescription dose. For technically challenging cases, it was permitted to prescribe as low as 50 Gy (BED of 100 Gy10) to a small portion of the GTV, ITV (if used) and corresponding PTV adjacent to a critical OAR (split target volume prescription). SABR was delivered using free breathing, deep inspiration breathhold, or expiratory gating. Thus, ITVs were not used for all patients. RESULTS From October 2017 to June 2020, 13 patients were enrolled: 3 in DL1, 3 in DL2, 4 in DL3, and 3 in DL4. Seven (IASLC definition) or 6 (RTOG) patients had central lesions, and the rest abutted the chest wall. All patients completed SABR per protocol without major deviations. At 18.4 months median follow-up (range, 2.4-25.6 months), no patient developed dose limiting toxicities. There were no grade 4-5 events and 1 grade 3 event (painful brachial plexopathy, which occurred in DL1 at 16 months post-SABR at the time of local progression). There were 7 instances of grade 2 toxicities. These included fatigue (n = 1, DL2), chest wall pain (n = 2, DL2 & DL4), cough (n = 1, DL3), dyspnea (n = 2, DL3 & DL4), nausea (n = 1, DL4), and wheezing (n = 1, DL4). Eight (62%) patients required utilization of "split target volume" prescriptions. The mean total dose to the GTV ranged from 60.9-62.2 Gy for DL1, 63.2-68.6 Gy for DL2, 67.3-78.8 Gy for DL3, and 75.5-82.2 Gy for DL4. The median GTV was 19.9 cc (range 5-57cc). CONCLUSION 5-fraction SABR for early-stage lung SCC up to 14 Gy x 5 fractions using a split target prescription for OAR adjacent tumor appears safe. Long term follow up will be needed to assess late toxicities and tumor control. (NCT03321747).

Anlotinib is a novel molecular targeted drug that has been approved for the treatment of lung adenocarcinoma. Currently these agents are rarely used in the treatment of lung squamous cell carcinoma (LSCC). Bronchoscope-guided radiofrequency ablation (RFA) is a new strategy proposed for the treatment of LSCC that is able to alleviate the obstruction of the respiratory tract caused by LSCC by direct destruction of the tumor tissues. The presence work aims to reveal whether Anlotinib could enhance the antitumor activity of RFA on LSCC cells. The results from real-time PCR (qPCR) confirmed overexpression of targets of anlotinib activity, including receptor tyrosine kinase or the MPAK/PI3K-AKT pathway kinases, in LSCC tissues. Treatment with anlotinib inhibited the survival, in vitro invasion, and migration of LSCC cells. Moreover, the antitumor effects of RFA were investigated using a rodent model of LSCC. The combination of RFA and anlotinib treatment enhanced the antitumor effect of RFA treatment. We propose a combinative strategy of RFA and anlotinib as a novel approach for successful management of LSCC.

Objective Programmed death-ligand 1 (PD-L1) is known to be over-expressed in non-small cell lung cancer (NSCLC). However, the impact of chemotherapy on the altered status of PD-L1 expression has not been examined for NSCLC. The present study was intended to examine the impact of neoadjuvant chemotherapy on PD-L1 expression and its prognostic significance in lung squamous cell carcinoma (SCC). Patients and methods Matched tumor samples were obtained from 76 SCC patients prior to and after neoadjuvant chemotherapy. The expression of PD-L1 was evaluated by immunohistochemistry. Survival analysis was performed by the Kaplan-Meier method. Results Prior to neoadjuvant chemotherapy, PD-L1 expression was identified in 52.6% (40/76) of SCC patients while 61.8% (47/76) were positive for PD-L1 expression after neoadjuvant chemotherapy. Nine patients switched from negative to positive while another two patients’ samples showed the reverse of the above result. Multivariate analysis demonstrated that postoperative expression of PD-L1 was an independent prognostic factor for overall survival (HR = 0.50, P = 0.003), but not for PD-L1 expression prior to neoadjuvant chemotherapy. Conclusion Neoadjuvant chemotherapy may up-regulate the expression of PD-L1. As compared with the status of PD-L1 expression prior to chemotherapy, the postoperative expression of PD-L1 is a better prognostic factor for overall survival in SCC.

Few effective therapies have been developed for the treatment of lung squamous cell carcinoma (SQCC), in part due to a lack of understanding regarding the mechanisms underlying the initiation and development of this disease. Whole transcriptome sequencing not only provides insight into the expression of all transcribed genes, but offers an efficient approach for identifying genetic variations, including gene fusions, mutations and alternative splicing. In this study, we performed whole transcriptome sequencing of 10 patients with stage IIIA lung SQCC, and discovered a large number of single nucleotide variants (SNVs; mean of 12.2 SNVs/Mb), with C>T/G>A and A>G/T>C transitions being the most frequently observed. Additionally, a total of 132 gene fusions were identified based upon TopHat alignments, 70.5% (93/132) of which occurred as a result of intra-chromosomal rearrangements. Based on the number of supporting reads for each fusion, we further validated 20 of the 26 top gene fusions by RT-PCR and Sanger sequencing. Taken together, these data provide an in-depth view of transcriptional alterations in lung SQCC patients, and may be useful for identification of new therapeutic targets.

Current targeted therapy proves no effective outcomes in lung squamous cell carcinoma (SQCC). Recent studies suggested that FGFR1 would be promising. This systematic review elaborated FGFR1 amplification in lung SQCC.An electronic search was conducted on PubMed, EMBASE, Web of SCI, Google Scholar and Cochrane Library. Eligible studies regarding incidence of FGFR1 amplification in lung SQCC and correlation between FGFR1 amplification and clinicopathological features or survival were extracted and analyzed.We identified 13 eligible studies with a total of 1798 patients. The results showed about 19% of FGFR1 amplification (95% CI: 0.15-0.24; I(2)=84.5%; p=0.000). Using the same test method: FISH, definition and ethnicity, the rates were 17% (95% CI: 0.14-0.20; I(2)=53.1%; p=0.037), 21% (95% CI: 0.18-0.24; I(2)=0; p=0.615), and 16% (95% CI: 0.13-0.19; I(2)=72.1%; p=0.028), respectively. Pearson's correlation analysis suggested that smoking status was highly correlated with FGFR1 amplification (coefficient=0.961, p0.001). FGFR1 amplification was significantly correlated with lymph node metastasis (OR: 2.27; 95% CI: 1.62-3.20; p=0.000), but not correlated with gender (OR: 1.12; 95% CI: 0.90-1.38; p=0.91), differentiation (OR: 1.02; 95% CI: 0.76-1.38; p=0.959) and stage (OR: 0.93; 95% CI: 0.73-1.19; p=0.877) in lung SQCC patients. With respect to survival, FGFR1 amplification had no influence on PFS (HR: 1.57; 95% CI: 0.85-2.30; p=0.259) and OS (HR: 1.40; 95% CI: 0.90-1.89; p=0.416) for SQCC patients.FGFR1 amplification is about 19%. Gender, stage, differentiation, ethnicities and test methods have no influence on FGFR1 amplification. FGFR1 amplification trends to correlate with lymph node metastasis and smoking. Whether FGFR1 amplification has effect on survival remains controversial.

Background:Human chromosome 9q33.3 is one of the most important loci for pathophysiological stresses with complex genetic traits. We hypothesized that the common single-nucleotide polymorphisms on this region may affect non–small-cell lung cancer risk.Methods:We genotyped 43 single-nucleotide polymorphisms that span 13 genes on 9q33.3 in two independent cohorts: the discovery study including 485 cases and 532 controls (North China) and the replicative study including 1063 cases and 1247 controls (South China). Both of the discovery cohort and the replicative cohort were included in the combined study.Results:In the discovery study, we identified a potential protective locus rs10118570 in mitogen-activated protein kinase associated protein 1 with a lower population attributable risk under logistical regression adjusted by age, gender, smoking, and drinking status (adjusted odds ratio [OR] 0.26, 95% confidence interval [CI] 0.10–0.71, p = 1.138 × 10–3 for genotype GG in lung squamous cell carcinoma). This protective quality increased in a dose-dependent manner as genotype GG decreased (ptrend = 9.675 × 10–4). Replicative and combined studies showed consistent association for this genotype (replicative: adjusted OR 0.36, 95% CI 0.20–0.66, p = 6.109 × 10–4, ptrend = 7.386 × 10–4; combined: adjusted OR 0.33, 95% CI 0.18–0.55, p = 1.259 × 10–6, ptrend = 7.725 × 10–6).Conclusion:We concluded that mitogen-activated protein kinase associated protein 1 rs10118570 may be an important protective factor for developing better management strategies in lung squamous cell carcinoma.

Secreted protein acidic and rich in cysteine (SPARC) plays a crucial role in the malignant progression of a number of human cancers. However, the roles of SPARC in lung squamous cell carcinoma (LSCC) remain elusive. In this present study, we first detected SPARC expression and investigated the relationship between SPARC expression and the clinicopathological attributes of LSCC patients. Then we constructed SPARC-overexpression model in LSCC cell line to explore the characteristics of SPARC in LSCC development both in vitro and in vivo. The data demonstrated a remarkably higher level of SPARC in LSCC tissues than in corresponding non-cancerous tissues and elevated SPARC expression was significantly correlated with poor outcome in LSCC patients. Moreover, a serial of phenotypic experiments indicated that SPARC overexpression substantially facilitated the growth and inhibited the apoptosis in LSCC cells and xenografts. Taken together, our results suggest that SPARC is a novel prognostic marker for LSCC prognosis and SPARC significantly promotes LSCC tumorigenesis. Targeting SPARC may provide a novel therapeutic strategy for LSCC management.