Search

Lung adenocarcinoma (LUAD) is a heterogeneous disease. Our study aimed to understand the unique molecular features of preinvasive to invasive LUAD subtypes. We retrospectively analyzed the clinical, histopathological, and molecular data of 3,254 Chinese patients with preinvasive lesions (n = 252), minimally invasive adenocarcinomas (n = 479), and invasive LUAD (n = 2,523). Molecular data were elucidated using a targeted 68-gene next-generation sequencing panel. Our findings revealed four preinvasive lesion-predominant gene mutations, including MAP2K1 insertion-deletions (indels), BRAF non-V600E kinase mutations, and exon 20 insertions (20ins) in both EGFR and ERBB2, which we referred to as mutations enriched in AIS (MEA). The detection rate of MEA in invasive tumors was relatively lower. MAP2K1 missense mutations, which were likely passenger mutations, co-occurred with oncogenic driver mutations, while small indels were mutually exclusive from other genes regardless of the invasion level. BRAF non-V600E kinase-mutant invasive adenocarcinomas (IAC) had significantly higher mutation rates in tumor suppressor genes but lower frequency of co-occurring oncogenic driver mutations than non-kinase-mutant IAC, suggesting the potential oncogenic activity of BRAF non-V600E kinase mutations albeit weaker than BRAF V600E. Moreover, similar to the extremely low frequency of MAP2K1 indels in IAC, BRAF non-V600E kinase domain mutations co-occurring with TSC1 mutations were exclusively found in preinvasive lesions. Compared with EGFR L858R and exon 19 deletion, patients with preinvasive lesions harboring 20ins in either EGFR or ERBB2 were significantly younger, while those with IAC had similar age. Furthermore, our study demonstrated distinct mutational features for subtypes of oncogene mutations favored by different invasion patterns in adenocarcinomas. In conclusion, our data demonstrate distinct mutational features between preinvasive lesions and invasive tumors with MEA, suggesting the involvement of MEA in the early stages of tumorigenesis. Further pre-clinical studies are required to establish the role of these genes in the malignant transformation of LUAD.

The authors have requested that this preprint be removed from Research Square.

AimsAn increasing number of small pulmonary nodules are being screened by CT, and an intraoperative diagnosis is necessary for preventing excessive treatment. However, there is limited literature on the frozen diagnosis of small sclerosing pneumocytomas (SPs). In particular, tumours smaller than 1 cm are challenging for pathologists performing intraoperative frozen diagnosis.MethodsIn total, 230 cases of SP were surgically resected between January 2015 and March 2019 at Shanghai Chest Hospital, and of them, 76 cases were smaller than 1 cm. The histology and clinical information of these 76 cases (33.0%, 76/230) were reviewed retrospectively, 54 cases of which were diagnosed intraoperatively, and the pitfalls were summarised. All diagnoses were confirmed on permanent sections and immunohistochemical sections.ResultsHistologically, 78.9% (60/76) of the small SP was dominated by one growth pattern, and solid and papillary growth pattern were the most commonly misdiagnosed circumstances. The rate of intraoperative misdiagnosis of these SP smaller than 1 cm was 11.1% (6/54).ConclusionsThe main reason for misdiagnosis was failure to recognise the dual cell populations and the cellular atypia. Diagnostic clues include the gross morphology, the presence of dual-cell populations and a hypercellular papillary core, foam cell accumulation in glandular spaces and haemorrhage and haemosiderin on the periphery. In spite of awareness of pitfalls some cases may still be essentially impossible to diagnose on frozen section.

Background: Different pathological subtypes of lung adenocarcinoma lead to different treatment decisions and prognoses, and it is clinically important to distinguish invasive lung adenocarcinoma from preinvasive adenocarcinoma (adenocarcinoma in situ and minimally invasive adenocarcinoma). This study aims to investigate the performance of the deep learning approach on the classification of tumor invasiveness and compare it with the performances of currently available approaches.Methods: In this study, we propose a deep learning approach based on 3D conventional networks to automatically predict the invasiveness of pulmonary nodules. A total of 901 early-stage non-small cell lung cancer patients who underwent surgical treatment at Shanghai Chest Hospital between November 2015 and March 2017 were retrospectively included and randomly assigned to a training set (n=814) or testing set 1 (n=87). We subsequently included 116 patients who underwent surgical treatment and intraoperative frozen section between April 2019 and January 2020 to form testing set 2. We compared the performance of our deep learning approach in predicting tumor invasiveness with that of intraoperative frozen section analysis and human experts (radiologists and surgeons).Results: The deep learning approach yielded an area under the receiver operating characteristic curve (AUC) of 0.946 for distinguishing preinvasive adenocarcinoma from invasive lung adenocarcinoma in testing set 1, which is significantly higher than the AUCs of human experts (P0.05) and junior thoracic surgeons (0.768, P>0.05).Conclusions: We developed a deep learning approach that achieved comparable performance to intraoperative frozen section analysis in determining tumor invasiveness. The proposed method may contribute to clinical decisions related to the extent of surgical resection.

Objectives Small cell lung cancer (SCLC) is the most malignant lung cancer. Some of them are mixed with non-small cell lung cancer(NSCLC, Non SCLC),which are called combined small cell lung cancer (C-SCLC).Due to the difficulty of pathological diagnosis and the complexity of treatment, studies of C-SCLC have just been rising in recent years. This study is to evaluate the clinical and pathologic characteristics of C-SCLC. Methods Stage Ⅰ-Ⅲa C-SCLC patients who received radical R0 surgery between 2009–2018 in Shanghai Chest Hospital were enrolled. Clinical characteristics and prognosis were analyzed. Results Totally 181 patients were included, most of them were small cell combined with large cell neuroendocrine components(SCLC/LCNEC,58.0 %,N = 105),then with adenocarcinoma(SCLC/ADC:13.8 %,N = 25),and finally with squamous cell carcinoma(SCLC/SCC:13.3 %,N = 24).Median DFS and OS of C-SCLC patients underwent radical surgery were 32.5 and 49.7 months.1,3 and 5 years DFS rates of the entire cohort were 68.5 %,32.6 % and 16.0 %,respectively. Patients with SCLC/LCNEC had longer DFS (44.1 m vs. 20.4 m, p = 0.040) and longer OS trend (62.1 m vs. 33.2 m, p = 0.122).Groups of whether tumor invaded the pleura(p = 0.028 and p = 0.050),lymph node stage(p = 0.029 and p = 0.010) and the courses of adjuvant chemotherapy(p = 0.011 and p = 0.001) had statistical differences on DFS and OS. Conclusions SCLC/LCNEC was the most common type of C-SCLC. Patients' DFS and OS were also longer than other combined types. Adjuvant chemotherapy for SCLC is still the main treatment for surgical C-SCLC. Further studies are needed to clarify the clinical characteristics and prognosis of C-SCLC.

Introduction It has been well established that EGFR Thr790Met is one of the major resistance mechanisms to first- and second-generation EGFR tyrosine kinase inhibitors (TKIs). Nevertheless, whether EGFR Thr790Leu (T790L), which shares the mutation site of Thr790 with EGFR Thr790Met, mediates resistance to EGFR TKIs remains elusive. The treatment options for patients harboring this rare mutation have not been reported. Methods Capture-based targeted ultradeep sequencing was performed on tumor and plasma samples collected at various treatment milestones from three patients with advanced lung adenocarcinoma undergoing targeted therapy. Results Needle biopsy of lymph node metastasis from patient 1 revealed EGFR T790L at disease progression on first-line treatment of gefitinib. Patient 2 had EGFR T790L identified from needle biopsy of lung tissue at disease progression on icotinib treatment. This patient was subsequently treated with osimertinib and achieved stable disease with a progression-free survival of 9 months. For patient 3, at disease recurrence after surgery, resected lung tumor tissue was retrieved for molecular profiling and revealed EGFR exon 19 deletion and EGFR T790L. The patient subsequently received osimertinib treatment and continued to benefit for 16 months and counting. She has maintained stable disease at the time of submission of this manuscript. Conclusions We revealed for the first time that EGFR T790L may serve as a potential resistance mechanism to first-generation EGFR TKIs. We also report the first clinical evidence of efficacy generated by osimertinib in patients with lung adenocarcinoma harboring primary or acquired EGFR T790L, shedding light on treatment options for this subset of patients.

9051 Background: Lung adenocarcinoma (LUAD) is further classified into several histological subtypes with adenocarcinoma in situ (AIS), minimally invasive adenocarcinoma (MIA), and invasive adenocarcinoma (IAC) as the three major subtypes according to the extent of invasion. AIS has been considered as a precursor of IAC. Considering the significantly higher mutation burden among IAC tumors than AIS tumors, it seems likely that AIS tumors undergo a process of accumulating various somatic mutations to gain invasive ability. To understand the gene mutations involved in this transformation, we compared the mutational features of AIS and IAC tumors. Methods: This retrospective study included 2,769 Chinese patients diagnosed with stage 0-IIIA LUAD. Targeted sequencing was performed on tissue DNA isolated from 246 AIS tumors and 2,523 IAC tumors using 68 lung cancer-related genes (Lung Core, Burning Rock Biotech). Results: Analysis of mutation profiles revealed that mutation count was significantly lower for AIS ( P< 0.01) as compared to IAC tumors. Moreover, AIS tumors had significantly higher mutation detection rates for ERBB2 exon 20 insertion (20ins) ( P≤0.05), EGFR 20ins ( P≤0.05), non-V600E BRAF mutations ( P≤0.05), and MAP2K1 small insertion-deletion variants ( P≤0.05). These 4 gene mutations were grouped and referred to as AIS-like mutations for further analysis. Detection rates of AIS-like mutations were 54.9% for AIS tumors and 7.8% for IAC tumors. Patients with AIS-like mutation-positive AIS tumors were significantly younger than those with AIS tumors without AIS-like mutations ( P =0.018), while age were similar for IAC tumors with or without AIS-like mutations. Mutation count was similar between AIS tumors with or without AIS-like mutations. Interestingly, IAC tumors harboring AIS-like mutations had a significantly higher mutation count than those harboring known oncogenic drivers ( P= 0.045). Further investigation of the molecular profiles of IAC tumors harboring AIS-like mutations (n = 198) revealed the presence of various concurrent mutations in 8 genes including TP53 (39.4%), EGFR (non-20ins) (16.7%), RB1 (7.1%), PIK3CA (6.6%), MET (5.1%), ROS1 (4.0%), FLT3 (4.0%), and PTEN (3.5%), which were absent among AIS tumors, particularly those that harbor AIS-like mutations. In addition to TP53 (35.8%), PIK3CA (4.5%), and RB1 (4.0%), IAC tumors without AIS-like mutations (n = 2,324) had additional concurrent mutations in 2 other genes CDK4 (5.7%) and STK11 (3.9%) as compared to AIS tumors. Conclusions: Our data suggest that AIS-like mutations could be involved in the early stages of tumorigenesis by initiating the accumulation of other gene mutations that are required for the transformation of AIS tumors into IAC tumors. Our study contributes to a deeper understanding of the distinct gene mutations between AIS and IAC tumors among Chinese LUAD patients.

Background. The risk of atherosclerosis is elevated in abnormal lipid metabolism and circadian rhythm disorder. We investigated whether abnormal lighting condition would have influenced the circadian expression of clock genes and clock-controlled lipid metabolism-related genes in ApoE-KO mice.Methods. A mouse model of atherosclerosis with circadian clock genes expression disorder was established using ApoE-KO mice (ApoE-KO LD/DL mice) by altering exposure to light. C57 BL/6J mice (C57 mice) and ApoE-KO mice (ApoE-KO mice) exposed to normal day and night and normal diet served as control mice. According to zeitgeber time samples were acquired, to test atheromatous plaque formation, serum lipids levels and rhythmicity, clock genes, and lipid metabolism-related genes along with Sirtuin 1 (Sirt1) levels and rhythmicity.Results. Atherosclerosis plaques were formed in the aortic arch of ApoE-KO LD/DL mice. The serum lipids levels and oscillations in ApoE-KO LD/DL mice were altered, along with the levels and diurnal oscillations of circadian genes, lipid metabolism-associated genes, and Sirt1 compared with the control mice.Conclusions. Abnormal exposure to light aggravated plaque formation and exacerbated disorders of serum lipids and clock genes, lipid metabolism genes and Sirt1 levels, and circadian oscillation.

Background Dysregulation of circadian rhythms can contribute to diseases of lipid metabolism. NAD-dependent deacetylase sirtuin-1(SIRT1) is an important hub which links lipid metabolism with circadian clock by its deacetylation activity depends on intracellular NAD+/NADH content ratio. Hydrogen sulfide (H2S) is an endogenous reductant which can affect the intracellular redox state. Therefore, we hypothesized that exogenous H2S can affect the expression of circadian clock genes mediated by sirt1 thereby affecting body's lipid metabolism. And also because the liver is a typical peripheral circadian clock oscillator that is intimately linked to lipid metabolism. Thus the effect of H2S were observed on 24-hour dynamic expression of 4 central circadian clock genes and sirt1gene in primary cultured hepatocytes. Results We established a hepatocyte model that showed a circadian rhythm by serum shock method. And detected that the expression level and the peak of circadian clock genes decreased gradually and H2S could maintain the expression and amplitude of circadian clock genes such as Clock, Per2, Bmal1 and Rev-erbαwithin a certain period time. Accordingly the expression level of sirt1 in H2S group was significantly higher than that in the control group. Conclusion Exogenous reductant H2S maintain the circadian rhythm of clock gene in isolated liver cells. We speculated that H2S has changed NAD+/NADH content ratio in hepatocytes and enhanced the activity of SIRT1 protein directly or indirectly, so as to maintain the rhythm of expression of circadian clock genes, they play a role in the prevention and treatment of lipid metabolism-related disease caused by the biological clock disorders.