Rapid on-site evaluation (ROSE) during bronchoscopy provides the opportunity to assess the adequacy of the sampling site, thus enabling diagnosis. The rapid evaluation of smears is typically performed by cytopathologists, but this is not always feasible. At our institution, a university-trained clinical biochemist performs the evaluation of ROSE smears. Our aim was to compare ROSE evaluations conducted by a laboratory specialist and a cytopathologist. We assessed the adequacy of lymph node samples from 78 patients using ROSE: 37 samples were obtained via EBUS-TBNA and 41 via TBNA. All smears were examined by the laboratory specialist, and simultaneously prepared parallel smears from the same needle passes were sent to the Department of Pathology for evaluation. Of the 78 samples, both the laboratory specialist and the pathologist team deemed 63 samples adequate and 8 samples inadequate. In 7 cases, there was a disagreement between the pathologist team and laboratory specialist. This resulted in a 92.3% agreement. The Cohen’s kappa value was 0.71, indicating strong and Gwet’s AC1 value was 0.90 corresponding to almost perfect agreement. The diagnostic performance was also excellent. Our conclusion is that ROSE performed by a laboratory specialist is a suitable alternative to on-site evaluation by a pathologist. It may help to overcome the resource shortage of interventional pulmonologists and cytopathologists.

AIMS:Lymphangioleiomyomatosis (LAM) is a PEComa that primarily affects premenopausal women and is associated with the cystic destruction of the lung. Although smooth muscle and melanocytic markers can be helpful, histologic diagnosis of LAM remains challenging, especially in small biopsies. Glycoprotein non-metastatic melanoma protein B (GPNMB) expression has recently been identified as a highly specific and sensitive immunohistochemical marker for PEComas. Nevertheless, only a few LAM cases have been studied so far. The aim of this study was to assess the utility of GPNMB immunohistochemistry as an ancillary marker in a larger cohort of LAM samples. METHODS AND RESULTS:Immunohistochemistry for GPNMB was performed on cases of LAM (N = 15), potential differential diagnostic mimics (N = 30) and normal lung tissue (N = 2). Immunohistochemistry was assessed using the H-score method; an H-score above 100 was considered 'high'. All LAM cases showed strong cytoplasmic expression, with cell membrane accentuation in some areas. In contrast, differential diagnostic mimics were either negative or only weakly positive. Normal lung parenchyma was negative for GPNMB; however, weak-to-moderate expression was seen in alveolar macrophages in both the normal and diseased lung tissues in most cases. Considering only 'high' positivity, GPNMB immunohistochemistry showed 100% sensitivity and specificity for the diagnosis of LAM in our cohort. CONCLUSIONS:Based on its 100% sensitivity and specificity, GPNMB appears to be a highly valuable immunohistochemical marker for the diagnosis of pulmonary LAM. Besides its diagnostic value, the membrane positivity of GPNMB on LAM cells may predict a response to glembatumumab vedotin, an antibody-drug conjugate targeting GPNMB.

Small cell lung carcinoma (SCLC) is a highly malignant cancer with early metastatic dissemination and poor clinical outcomes. Mutations in the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) pathway, including the frequently occurring rapamycin-insensitive protein (RICTOR) amplification, have been described in these tumours. Moreover, the associated mTOR hyperactivity could be exploited for personalised treatment. Our aim was to study mTOR activity, RICTOR amplification, and their role during SCLC progression. In situ mTOR activity and Rictor expression were characterised by immunohistochemistry in 50 primary and 50 brain metastatic tumours, and 14 paired SCLC patient samples. RICTOR copy number changes were analysed by fluorescence in situ hybridisation of the paired SCLC patient samples and in vivo experiments. Additionally, in vitro sensitivity to cisplatin and mTOR inhibitors was evaluated in SCLC cell lines harbouring RICTOR amplification and other mTOR pathway mutations. High Rictor expression and mTOR complex 2 (mTORC2) hyperactivity were significantly associated with brain metastases and worse overall survival. An increase in RICTOR copy number was observed in paired samples during progression. The importance of these alterations was confirmed both by the sensitising effect of vistusertib in vitro and the RICTOR copy number/expression changes in xenografts. Our study highlights the role of mTORC2 in SCLC progression. Early detection of RICTOR amplification in primary tumours and targeting mTORC2 in these cases may represent a promising novel strategy to develop personalised therapy for metastasis prevention.

RICTOR gene, which encodes the scaffold protein of mTORC2, can be amplified in various tumor types, including squamous cell carcinoma (SCC) of the lung. RICTOR amplification can lead to hyperactivation of mTORC2 and may serve as a targetable genetic alteration, including in lung SCC patients with no PD-L1 expression who are not expected to benefit from immune checkpoint inhibitor therapy. This study aimed to compare RICTOR amplification detected by fluorescence in situ hybridization (FISH) with Rictor and PD-L1 protein expression detected by immunohistochemistry (IHC) in SCC of the lung. The study was complemented by analysis of the publicly available Lung Squamous Cell Carcinoma (TCGA, Firehose legacy) dataset. RICTOR amplification was observed in 20% of our cases and 16% of the lung SCC cases of the TCGA dataset. Rictor and PD-L1 expression was seen in 74% and 44% of the cases, respectively. Rictor IHC showed two staining patterns: membrane staining (16% of the cases) and cytoplasmic staining (58% of the cases). Rictor membrane staining predicted RICTOR amplification as detected by FISH with high specificity (95%) and sensitivity (70%). We did not find any correlation between RICTOR amplification and PD-L1 expression; RICTOR amplification was detected in 18% and 26% of PD-L1 positive and negative cases, respectively. The TCGA dataset analysis showed similar results; RICTOR copy number correlated with Rictor mRNA and protein expression but showed no association with PD-L1 mRNA and protein expression. In conclusion, the correlation between RICTOR amplification and Rictor membrane staining suggests that the latter can potentially be used as a surrogate marker to identify lung SCC cases with RICTOR amplification. Since a significant proportion of PD-L1 negative SCC cases harbor RICTOR amplification, analyzing PD-L1 negative tumors by RICTOR FISH or Rictor IHC can help select patients who may benefit from mTORC2 inhibitor therapy.

The increasing knowledge of molecular alterations in malignancies, including mutations and regulatory failures in the mTOR (mechanistic target of rapamycin) signaling pathway, highlights the importance of mTOR hyperactivity as a validated target in common and rare malignancies. This review summarises recent findings on the characterization and prognostic role of mTOR kinase complexes (mTORC1 and mTORC2) activity regarding differences in their function, structure, regulatory mechanisms, and inhibitor sensitivity. We have recently identified new tumor types with RICTOR (rapamycin-insensitive companion of mTOR) amplification and associated mTORC2 hyperactivity as useful potential targets for developing targeted therapies in lung cancer and other newly described malignancies. The activity of mTOR complexes is recommended to be assessed and considered in cancers before mTOR inhibitor therapy, as current first-generation mTOR inhibitors (rapamycin and analogs) can be ineffective in the presence of mTORC2 hyperactivity. We have introduced and proposed a marker panel to determine tissue characteristics of mTOR activity in biopsy specimens, patient materials, and cell lines. Ongoing phase trials of new inhibitors and combination therapies are promising in advanced-stage patients selected by genetic alterations, molecular markers, and/or protein expression changes in the mTOR signaling pathway. Hopefully, the summarized results, our findings, and the suggested characterization of mTOR activity will support therapeutic decisions.

INTRODUCTION:Recent advances in the subclassification of small cell lung carcinomas (SCLCs) may help to overcome the unmet need for targeted therapies and improve survival. However, limited information is available on how the expression of the subtype markers changes during tumour progression. Our study aimed to compare the expression of these markers in primary and brain metastatic SCLCs. MATERIALS AND METHODS:Immunohistochemical analysis of the subtype markers was performed on 120 SCLCs (including 10 matched samples) and SCLC xenografts. RESULTS:Compared to primary SCLCs, there was a significant increase in the proportion of mixed subtypes in brain metastases, with a rate of ASCL1high/NeuroD1high and ASCL1high/NeuroD1high/YAP1high subtypes increasing to 48 % and 18 %, respectively. The subtype of the paired samples matched in only one-third of the cases. Although we did not observe a significant change after chemotherapy, a continuous decrease in ASCL1 expression coupled with an increase in the NeuroD1 expression was detected in the xenografts in a long-term experiment. DISCUSSION:Our results indicate that the expression of subtype markers frequently changes during disease progression, and subtype analysis of the primary SCLC may not provide accurate information about the characteristics of the recurrent or metastatic tumour. Therefore, repeated sampling and subtyping may be necessary for subtype-specific targeted therapy.

Background Kidney transplant recipients (KTRs) face an increased risk of renal cell carcinoma (RCC), in which the immunosuppressive regimen plays an important role. This study aimed to identify intracellular signalling alterations associated with post-transplant (post-tx) tumour formation. Methods Expression of mTOR-related proteins were analysed in kidneys obtained from end-stage renal disease (ESRD) patients and RCCs developed in KTRs or non-transplant patients. The effects of tacrolimus (TAC) and rapamycin (RAPA) on mTOR activity, proliferation, and tumour growth were investigated through different in vitro and in vivo experiments. Results Elevated mTORC1/C2 activity was observed in post-tx RCCs and in kidneys of TAC-treated ESRD patients. In vitro experiments demonstrated that TAC increases mTOR activity in a normal tubular epithelial cell line and in the investigated RCC cell lines, moreover, promotes the proliferation of some RCC cell line. In vivo, TAC elevated mTORC1/C2 activity in ischaemic kidneys of mice and enhanced tumour growth in xenograft model. Conclusions We observed significantly increased mTOR activity in ischaemic kidneys and post-tx RCCs, which highlights involvement of mTOR pathway both in the healing or fibrotic processes of kidney and in tumorigenesis. TAC-treatment further augmented the already elevated mTOR activity of injured kidney, potentially contributing to tumorigenesis during immunosuppression.

Lung carcinoma is one of the most common cancer types for both men and women. Despite recent breakthroughs in targeted therapy and immunotherapy, it is characterized by a high metastatic rate, which can significantly affect quality of life and prognosis. Rictor (encoded by the RICTOR gene) is known as a scaffold protein for the multiprotein complex mTORC2. Among its diverse roles in regulating essential cellular functions, mTORC2 also facilitates epithelial–mesenchymal transition and metastasis formation. Amplification of the RICTOR gene and subsequent overexpression of the Rictor protein can result in the activation of mTORC2, which promotes cell survival and migration. Based on recent studies, RICTOR amplification or Rictor overexpression can serve as a marker for mTORC2 activation, which in turn provides a promising druggable target. Although selective inhibitors of Rictor and the Rictor-mTOR association are only in a preclinical phase, they seem to be potent novel approaches to reduce tumor cell migration and metastasis formation. Here, we summarize recent advances that support an important role for Rictor and mTORC2 as potential therapeutic targets in the treatment of lung cancer. This is a traditional (narrative) review based on Pubmed and Google Scholar searches for the following keywords: Rictor, RICTOR amplification, mTORC2, Rictor complexes, lung cancer, metastasis, progression, mTOR inhibitors.

Alterations in mTOR signalling molecules, including RICTOR amplification, have been previously described in many cancers, particularly associated with poor prognosis. In this study, RICTOR copy number variation (CNV) results of diagnostic next-generation sequencing (NGS) were analysed in 420 various human malignant tissues. RICTOR amplification was tested by Droplet Digital PCR (ddPCR) and validated using the “gold standard” fluorescence in situ hybridisation (FISH). Additionally, the consequences of Rictor protein expression were also studied by immunohistochemistry. RICTOR amplification was presumed in 37 cases with CNV ≥ 3 by NGS, among these, 16 cases (16/420; 3.8%) could be validated by FISH, however, ddPCR confirmed only 11 RICTOR -amplified cases with lower sensitivity. Based on these, neither NGS nor ddPCR could replace traditional FISH in proof of RICTOR amplification. However, NGS could be beneficial to highlight potential RICTOR -amplified cases. The obtained results of the 14 different tumour types with FISH-validated RICTOR amplification demonstrate the importance of RICTOR amplification in a broad spectrum of tumours. The newly described RICTOR -amplified entities could initiate further collaborative studies with larger cohorts to analyse the prevalence of RICTOR amplification in rare diseases. Finally, our and further work could help to improve and expand future therapeutic opportunities for mTOR-targeted therapies.

Rictor protein (encoded by the RICTOR gene) is the scaffold protein for mTORC2, which plays essential roles in cytoskeletal reorganization and cell survival. Amplification of RICTOR has been reported in various cancer types, including approximately 15% of lung SQCC. RICTOR amplification can lead to hyperactivation of mTORC2 and serve as a targetable genetic alteration. Selection of patients for targeted therapy is an important consideration. Fluorescence in situ hybridization (FISH) is considered the diagnostic standard for detection of RICTOR amplification.

Significant improvements in the survival rates of paediatric cancer have been achieved over the past decade owing to recent advances in therapeutic and diagnostic strategies. However, disease progression and relapse remain a major challenge for the clinical management of paediatric angiosarcoma. Comprehensive genomic profiling of these rare tumours using high-throughput sequencing technologies may improve patient stratification and identify actionable biomarkers for therapeutic intervention. Here, we describe the clinical, histopathological, immunohistochemical and molecular profile of a novel and precision medicine-informed case where a KHDRBS1-NTRK3 fusion determined by next-generation sequencing-based comprehensive genomic profiling led to complete and sustained remission (clinical and radiological response) in an otherwise incurable disease. Our patient represents the first paediatric angiosarcoma harbouring a targetable NTRK3 fusion in the literature and demonstrates the first example of targeting this alteration in angiosarcoma using larotrectinib, an NTRK inhibitor. Clinical and radiological remission was achieved in under two months of therapy, and the patient is currently in complete remission, 4 month after stopping larotrectinib therapy, which was given over 17 months with only mild side effects reported. Therefore, this remarkable case exemplifies the true essence of precision-based care by incorporating conventional pathology with the why, when, and how to test for rare oncogenic drivers and agnostic biomarkers in paediatric angiosarcoma.

Failures of anti-tumour therapies and drug resistance initiate difficulties in cancer treatments often caused by alterations in signalling network activity, including PI3K/Akt/mTOR hyperactivity due to oncogenic mutations. In this review, we summarise the relevance of mTOR (mechanistic target of rapamycin) dysregulation identified decades ago, which is now known to be characteristic of many tumours. In this context, we present differences in activity, function and testability of mTOR kinase complexes (mTORC1 and mTORC2) differing in structure, regulatory mechanisms and inhibitor sensitivity. We highlight that genetic alterations, including RICTOR amplification and associated mTOR hyperactivity, are relevant in targeted therapy development. It is recommended to investigate mTOR profile activity in patients for whom mTOR inhibitor therapies are considered since the current first-generation mTOR inhibitors (rapamycin and analogues) may be ineffective in case of mTORC2 hyperactivity. Ongoing phase trials of new inhibitors and combination therapies are promising in advanced stage patients selected by molecular markers.

Lung cancer is the leading cause of tumor-related mortality, therefore significant effort is directed towards understanding molecular alterations occurring at the origin of the disease to improve current treatment options. The aim of our pilot-scale study was to carry out a detailed proteomic analysis of formalin-fixed paraffin-embedded tissue sections from patients with small cell or non-small cell lung cancer (adenocarcinoma, squamous cell carcinoma, and large cell carcinoma). Tissue surface digestion was performed on relatively small cancerous and tumor-adjacent normal regions and differentially expressed proteins were identified using label-free quantitative mass spectrometry and subsequent statistical analysis. Principal component analysis clearly distinguished cancerous and cancer adjacent normal samples, while the four lung cancer types investigated had distinct molecular profiles and gene set enrichment analysis revealed specific dysregulated biological processes as well. Furthermore, proteins with altered expression unique to a specific lung cancer type were identified and could be the targets of future studies.

Metabolic characteristics of kidney cancers have mainly been obtained from the most frequent clear cell renal cell carcinoma (CCRCC) studies. Moreover, the bioenergetic perturbances that affect metabolic adaptation possibilities of papillary renal cell carcinoma (PRCC) have not yet been detailed. Therefore, our study aimed to analyze the in situ metabolic features of PRCC vs. CCRCC tissues and compared the metabolic characteristics of PRCC, CCRCC, and normal tubular epithelial cell lines. The protein and mRNA expressions of the molecular elements in mammalian target of rapamycin (mTOR) and additional metabolic pathways were analyzed in human PRCC cases compared to CCRCC. The metabolic protein expression pattern, metabolite content, mTOR, and metabolic inhibitor sensitivity of renal carcinoma cell lines were also studied and compared with tubular epithelial cells, as "normal" control. We observed higher protein expressions of the "alternative bioenergetic pathway" elements, in correlation with the possible higher glutamine and acetate consumption in PRCC cells instead of higher glycolytic and mTOR activity in CCRCCs. Increased expression of certain metabolic pathway markers correlates with the detected differences in metabolite ratios, as well. The lower lactate/pyruvate, lactate/malate, and higher pyruvate/citrate intracellular metabolite ratios in PRCC compared to CCRCC cell lines suggest that ACHN (PRCC) have lower Warburg glycolytic capacity, less pronounced pyruvate to lactate producing activity and shifted OXPHOS phenotype. However, both studied renal carcinoma cell lines showed higher mTOR activity than tubular epithelial cells cultured in vitro, the metabolite ratio, the enzyme expression profiles, and the higher mitochondrial content also suggest increased importance of mitochondrial functions, including mitochondrial OXPHOS in PRCCs. Additionally, PRCC cells showed significant mTOR inhibitor sensitivity and the used metabolic inhibitors increased the effect of rapamycin in combined treatments. Our study revealed in situ metabolic differences in mTOR and metabolic protein expression patterns of human PRCC and CCRCC tissues as well as in cell lines. These underline the importance in the development of specific new treatment strategies, new mTOR inhibitors, and other anti-metabolic drug combinations in PRCC therapy.

Monolayer cultures, the less standard three-dimensional (3D) culturing systems, and xenografts are the main tools used in current basic and drug development studies of cancer research. The aim of biofabrication is to design and construct a more representative in vivo 3D environment, replacing two-dimensional (2D) cell cultures. Here, we aim to provide a complex comparative analysis of 2D and 3D spheroid culturing, and 3D bioprinted and xenografted breast cancer models. We established a protocol to produce alginate-based hydrogel bioink for 3D bioprinting and the long-term culturing of tumour cells in vitro. Cell proliferation and tumourigenicity were assessed with various tests. Additionally, the results of rapamycin, doxycycline and doxorubicin monotreatments and combinations were also compared. The sensitivity and protein expression profile of 3D bioprinted tissue-mimetic scaffolds showed the highest similarity to the less drug-sensitive xenograft models. Several metabolic protein expressions were examined, and the in situ tissue heterogeneity representing the characteristics of human breast cancers was also verified in 3D bioprinted and cultured tissue-mimetic structures. Our results provide additional steps in the direction of representing in vivo 3D situations in in vitro studies. Future use of these models could help to reduce the number of animal experiments and increase the success rate of clinical phase trials.

Background Hereditary angioedema (HAE) is a rare, life-threatening disease. The knowledge about the molecular pathogenesis of HAE has derived mainly from investigating blood samples. However, limited data are available on the role of the molecular mechanisms in the affected tissues during HAE attack. Objective The aim of our study was to explore the histological changes occurring in HAE attacks. Methods Post mortem macro-, microscopic and immunohistological assessment of upper airway tissues of a patient with HAE due to C1 inhibitor deficiency (C1-INH-HAE) type 2 who died from laryngeal HAE attack was compared with a non-HAE patient who died from other condition without any signs of angioedema. Results Compared to the control patient, we demonstrated stronger T cell/monocyte infiltration and a more intense C1-INH staining in the C1-INH-HAE patient. The expression of both bradykinin receptors (B1/B2) was observed with a slightly lower level in the C1-INH-HAE patient than in the control patient. PAR1 expression was strongly reduced in the C1-INH-HAE patient suggesting overactivation of this hyperpermeability inducing receptor. Conclusion Our unique case and novel results correspond to the knowledge about C1-INH and BDKRs observed in plasma; however, it revealed new information about the pathomechanism of HAE attack focusing on the potential involvement of PAR1 in edema formation. This observation, if it is verified by subcutaneous biopsy studies, may designate a new therapeutic target in HAE.

Soft tissue sarcomas are a heterogeneous group of malignant tumours, accounting for the fifth most common paediatric cancer. Next-generation sequencing-based (NGS) comprehensive genomic profiling (CGP) of paediatric soft tissue sarcomas tumours associated with a poor prognosis can identify actionable biomarkers and determine prognostic and therapeutic stratification of patient management. Diagnostic samples of 20 patients diagnosed with various paediatric sarcomas were investigated using a CGP assay (Illumina TruSight Oncology 500). The identified single nucleotide variants, small insertions or deletions, copy number alterations, gene fusions, microsatellite stability status, and the tumour mutation burden were analysed. Using Clinical insight (Qiagen) and PierianDx (Pierian) software, actionable variants were determined. Our study revealed potentially actionable alterations for therapeutic intervention in 75% of cases (15/20). Novel variants were detected in several tumour entities which have not previously been published. All cases were characterised by low tumour mutation burden, with none of the cases characterised by microsatellite instability. The targetable gene fusions included three NTRK fusions, TFG-ROS1, ROS1-GIT2 and EML4-ALK fusions. The most frequent copy number alterations affected the CDK4, ALK and FGFR1 genes, identified in 28.6% (4/20), 14.3% (2/14) and 14.3% (2/14) of the cases, respectively. Single nucleotide variants were detected in 7 genes, including ALK, CHEK2, ESR1, FANCL, MET, NBN and NRAS, across six cases. Based on the NGS results, targeted therapy was prescribed in 5 cases (tazemetostat in epithelioid sarcoma, entrectinib in inflammatory myofibroblastic tumour and infantile fibrosarcoma, larotrectinib in angiosarcoma and infantile fibrosarcoma). At least partial remission was achieved in all patients receiving targeted therapies. In our study, routine application of the TSO500 CPG assay led to administration of targeted therapies in 25% of the patients with at least partial clinical response in this patient cohort.

Lung transplant recipients are at risk for life-threatening infections including severe acute respiratory syndrome coronavirus 2-associated COVID-19. Several viral infections have been associated with the development of chronic lung allograft dysfunction. Long-term outcomes of COVID-19 on graft function are not known.A 53-year-old female patient, who underwent bilateral lung transplantation 3 years before because of stage IV sarcoidosis and secondary pulmonary hypertension was admitted in the second wave of the pandemic because of COVID-19 with symptoms including dry cough. Chest computed tomography showed ground glass opacities affecting 25% to 50% of the lung parenchyma. She was admitted to the COVID-19 Unit of our clinic. She received oxygen via nasal cannula, remdesivir, and low-dose methylprednisolone while mycofenolate acid administration was stopped. Her clinical condition improved. The first follow-up visit 1 month after the infection demonstrated deterioration in lung function. Computed tomography scan showed almost complete resolution; transbronchial biopsy was performed and proved acute allograft rejection. During the hospitalization a new onset atrial fibrillation was confirmed. In the background of atrial fibrillation and simultaneous neck pain, severe hyperthyroidism was proven. Because of thyroiditis and lung allograft rejection, high-dose steroid treatment was initiated and everolimus was added to the immunosuppressive therapy. Donor specific antibodies were also detected, hence plasmapheresis was indicated and continued with photoferesis. On the follow-up spirometry the values were stable; however, they did not reach pre-COVID levels.In lung transplant recipients COVID-19 might trigger allograft rejection in addition to virus-related thyroid disease.

BACKGROUND:Although lung adenocarcinoma (LADC) with sensitizing mutations of the epidermal growth factor receptor (EGFR) is highly sensitive to EGFR tyrosine kinase inhibitors (EGFR-TKIs), in most cases disease progression inevitably occurs. Our aim was to investigate the predictive and prognostic significance of adjusted tumoral EGFR variant allele frequency (EGFR-aVAF) in the above setting.METHODS:Eighty-nine Caucasian advanced-stage LADC patients with known exon-specific EGFR mutations undergoing EGFR-TKI treatment were included. The correlations of EGFR-aVAF with clinicopathological variables including progression-free and overall survival (PFS and OS, respectively) were retrospectively analyzed.RESULTS:Of 89 EGFR-mutant LADC patients, 46 (51.7%) had exon 19 deletion, while 41 (46.1%) and 2 (2.2%) patients had exon 21- and exon 18-point mutations, respectively. Tumoral EGFR-aVAF was significantly higher in patients harboring EGFR exon 19 mutations than in those with exon 21-mutant tumors (P<0.001). Notably, patients with EGFR exon 19 mutant tumors demonstrated significantly improved PFS (P=0.003) and OS (P=0.02) compared to patients with exon 21 mutations. Irrespective of specific exon mutations, a statistically significant positive linear correlation was found between EGFR-aVAF of tumoral tissue and PFS (r=0.319; P=0.002). High (≥70%) EGFR-aVAF was an independent predictor of longer PFS [vs. low (<70%) EGFR-aVAF; median PFSs were 52 vs. 26 weeks, respectively; P<0.001]. Additionally, patients with high EGFR-aVAF also had significantly improved OS than those with low EGFR-aVAF (P=0.011).CONCLUSIONS:Our study suggests that high (≥70%) EGFR-aVAF of tumoral tissue predicts benefit from EGFR-TKI treatment in advanced LADC and, moreover, that exon 19 EGFR mutation is associated with high EGFR-aVAF and improved survival outcomes.