Publications by authors named "Teh-Ying Chou"

152 Publications

Seroprevalence of COVID-19 in Taiwan revealed by testing anti-SARS-CoV-2 serological antibodies on 14,765 hospital patients.

Lancet Reg Health West Pac 2020 Oct 10;3:100041. Epub 2020 Oct 10.

The Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, 201, Section 2, Shipai Road, Taipei 11217, Taiwan.

Background: Coronavirus Disease 2019 (COVID-19) has become a worldwide pandemic and affected more than 227 countries or territories, resulting in more than 25 million cases with over 0•85 million deaths, as of September 2, 2020. Taiwan has been successful in countering the COVID-19 outbreak, however, the potential risk for asymptomatic infections and the prevalence rates remain unknown. We aimed to estimate the seroprevalence of COVID-19 in Taiwan via serologically testing hospital patients with neither symptoms indicative of nor positive nucleic acid test for SARS-CoV-2 infection.

Methods: Residual specimens from laboratory blood tests for outpatient and emergency department patients visiting a medical centre in Taipei, Taiwan, within one week in May and another in July, 2020, were collected. We used Elecsys Anti-SARS-CoV-2 Assay to screen and further validated cases with high cutoff index by a confirmatory ELISA assay. We also analysed antibody responses against SARS-CoV-2 along disease progression in four nucleic acid test confirmed COVID-19 patients.

Findings: Blood samples from a total of 14,765 patients were tested. The unweighted seroprevalence of anti-SARS-CoV-2 antibodies was 0•07% [95% CI, 0•04%-0•13%]; after weighting with the population demographics of Taiwan, the estimated overall seroprevalence was 0•05% [95% CI, 0•02%-0•10%]. Furthermore, based on data of the four COVID-19 cases, the seroconversion dates for IgM were as early as 9 days and that for IgG 11 days after symptoms onset.

Interpretation: We screened the anti-SARS-CoV-2 antibodies in a small-scale population-based study and observed an approximately 0•05% seroprevalence of COVID-19, indicating that the current containment protocols emphasising mask wearing, hand washing, social distancing and mandatory quarantine for all incomers are effective in Taiwan.

Funding: Taipei Veterans General Hospital, Taipei, Taiwan.
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http://dx.doi.org/10.1016/j.lanwpc.2020.100041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7546957PMC
October 2020

Whole-exome Sequencing of Epstein-Barr Virus-associated Pulmonary Carcinoma With Low Lymphocytic Infiltration Shows Molecular Features Similar to Those of Classic Pulmonary Lymphoepithelioma-like Carcinoma: Evidence to Support Grouping Together as One Disease Entity.

Am J Surg Pathol 2021 11;45(11):1476-1486

Institute of Biomedical Informatics.

Pulmonary lymphoepithelioma-like carcinoma (LELC) is a distinct type of Epstein-Barr virus (EBV)-associated non-small cell carcinoma characterized by a syncytial growth pattern with heavy lymphocytic infiltration. We recently identified a group of non-small cell carcinomas, which are also associated with EBV but lack significant lymphocytic infiltration. These EBV-associated pulmonary carcinomas with low lymphocytic infiltration morphologically resemble nonkeratinizing squamous cell carcinoma, but their patient characteristics are more similar to those of LELC, including female sex and nonsmoking status. To clarify the relationships between these disease entities, in this study, we explored the molecular characteristics of the EBV-associated carcinomas with low lymphocytic infiltration using whole-exome sequencing and compared their molecular profiles with those of classic LELC and pulmonary squamous cell carcinoma. We demonstrate that the molecular characteristics of EBV-associated carcinomas with low lymphocytic infiltration are highly similar to those of classic LELC. Both show low tumor mutational burden, lack of commonly mutated driver genes in other types of non-small cell lung cancer, similar mutational signature involving APOBEC-related mutations, and enrichment of CD274 (programmed death-ligand 1) amplification. These molecular characteristics are very different from those of pulmonary squamous cell carcinoma. The unique patient demographics and molecular characteristics shared by EBV-associated carcinomas with low lymphocytic infiltration and classic LELC suggest that these tumors represent one single disease entity defined by EBV association. This study supports the proposal for the usage of the term "EBV-associated pulmonary carcinoma" to encompass the entire morphologic spectrum of this distinct EBV-associated disease entity.
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http://dx.doi.org/10.1097/PAS.0000000000001722DOI Listing
November 2021

Feedback Regulation of -GlcNAc Transferase through Translation Control to Maintain Intracellular -GlcNAc Homeostasis.

Int J Mol Sci 2021 Mar 27;22(7). Epub 2021 Mar 27.

Division of Molecular Pathology, Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taipei 11217, Taiwan.

Protein -GlcNAcylation is a dynamic post-translational modification involving the attachment of -acetylglucosamine (GlcNAc) to the hydroxyl groups of Ser/Thr residues on numerous nucleocytoplasmic proteins. Two enzymes are responsible for -GlcNAc cycling on substrate proteins: -GlcNAc transferase (OGT) catalyzes the addition while -GlcNAcase (OGA) helps the removal of GlcNAc. -GlcNAcylation modifies protein functions; therefore, dysregulation of -GlcNAcylation affects cell physiology and contributes to pathogenesis. To maintain homeostasis of cellular -GlcNAcylation, there exists feedback regulation of OGT and OGA expression responding to fluctuations of -GlcNAc levels; yet, little is known about the molecular mechanisms involved. In this study, we investigated the -GlcNAc-feedback regulation of OGT and OGA expression in lung cancer cells. Results suggest that, upon alterations in -GlcNAcylation, the regulation of OGA expression occurs at the mRNA level and likely involves epigenetic mechanisms, while modulation of OGT expression is through translation control. Further analyses revealed that the eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) contributes to the downregulation of OGT induced by hyper--GlcNAcylation; the S5A/S6A -GlcNAcylation-site mutant of 4E-BP1 cannot support this regulation, suggesting an important role of -GlcNAcylation. The results provide additional insight into the molecular mechanisms through which cells may fine-tune intracellular -GlcNAc levels to maintain homeostasis.
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http://dx.doi.org/10.3390/ijms22073463DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8037101PMC
March 2021

Lung Cancer in Republic of China.

J Thorac Oncol 2021 04;16(4):519-527

Department of Chest Medicine, Taipei Veterans General Hospital, Taipei, Republic of China; School of Medicine, National Yang-Ming University, Taipei, Republic of China. Electronic address:

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http://dx.doi.org/10.1016/j.jtho.2020.10.155DOI Listing
April 2021

Human Costars Family Protein ABRACL Modulates Actin Dynamics and Cell Migration and Associates with Tumorigenic Growth.

Int J Mol Sci 2021 Feb 18;22(4). Epub 2021 Feb 18.

Institute of Biochemistry and Molecular Biology, National Yang-Ming University, Taipei 11221, Taiwan.

Regulation of cellular actin dynamics is pivotal in driving cell motility. During cancer development, cells migrate to invade and spread; therefore, dysregulation of actin regulators is often associated with cancer progression. Here we report the role of ABRACL, a human homolog of the actin regulator Costars, in migration and tumorigenic growth of cancer cells. We found a correlation between ABRACL expression and the migratory ability of cancer cells. Cell staining revealed the colocalization of ABRACL and F-actin signals at the leading edge of migrating cells. Analysis of the relative F-/G-actin contents in cells lacking or overexpressing ABRACL suggested that ABRACL promotes cellular actin distribution to the polymerized fraction. Physical interaction between ABRACL and cofilin was supported by immunofluorescence staining and proximity ligation. Additionally, ABRACL hindered cofilin-simulated pyrene F-actin fluorescence decay in vitro, indicating a functional interplay. Lastly, analysis on a colorectal cancer cohort demonstrated that high ABRACL expression was associated with distant metastasis, and further exploration showed that depletion of ABRACL expression in colon cancer cells resulted in reduced cell proliferation and tumorigenic growth. Together, results suggest that ABRACL modulates actin dynamics through its interaction with cofilin and thereby regulates cancer cell migration and participates in cancer pathogenesis.
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http://dx.doi.org/10.3390/ijms22042037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7922284PMC
February 2021

The International Association for the Study of Lung Cancer Global Survey on Programmed Death-Ligand 1 Testing for NSCLC.

J Thorac Oncol 2021 04 2;16(4):686-696. Epub 2021 Mar 2.

Carolinas Pathology Group, Charlotte, North Carolina.

Introduction: Programmed death-ligand 1 (PD-L1) immunohistochemistry (IHC) is required to determine the eligibility for pembrolizumab monotherapy in advanced NSCLC worldwide and for several other indications depending on the country. Four assays have been approved/ Communauté Européene-In vitro Diagnostic (CV-IVD)-marked, but PD-L1 IHC seems diversely implemented across regions and laboratories with the application of laboratory-developed tests (LDTs).

Method: To assess the practice of PD-L1 IHC and identify issues and disparities, the International Association for the Study of Lung Cancer Pathology Committee conducted a global survey for pathologists from January to May 2019, comprising multiple questions on preanalytical, analytical, and postanalytical conditions.

Result: A total of 344 pathologists from 64 countries participated with 41% from Europe, 24% from North America, and 18% from Asia. Besides biopsies and resections, cellblocks were used by 75% of the participants and smears by 11%. The clone 22C3 was most often used (69%) followed by SP263 (51%). They were applied as an LDT by 40% and 30% of the users, respectively, and 76% of the participants developed at least one LDT. Half of the participants reported a turnaround time of less than or equal to 2 days, whereas 13% reported that of greater than or equal to 5 days. In addition, quality assurance (QA), formal training for scoring, and standardized reporting were not implemented by 18%, 16%, and 14% of the participants, respectively.

Conclusions: Heterogeneity in PD-L1 testing is marked across regions and laboratories in terms of antibody clones, IHC assays, samples, turnaround times, and QA measures. The lack of QA, formal training, and standardized reporting stated by a considerable minority identifies a need for additional QA measures and training opportunities.
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http://dx.doi.org/10.1016/j.jtho.2020.12.026DOI Listing
April 2021

Primary Signet Ring Cell/Histiocytoid Carcinoma of the Eyelid: Somatic Mutations in and Other Clinically Actionable Mutations Imply Early Use of Targeted Agents.

Curr Oncol 2021 02 16;28(1):918-927. Epub 2021 Feb 16.

Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taipei 11217, Taiwan.

Primary signet ring cell/histiocytoid carcinoma of the eyelid is a rare ocular malignancy and its diagnosis is often delayed. This neoplasm presents as an insidious, diffusely infiltrative mass in the periocular area that later infiltrates the orbit. An exenteration is usually indicated; however, nearly one-third of patients develop local recurrence or metastasis. Morphologically, it resembles signet ring cell carcinoma of the stomach and breast, raising the possibility of mutations in , the gene encoding E-cadherin. To determine whether primary signet ring cell/histiocytoid carcinoma harbors the mutation or other actionable mutations, we analyzed the tumor tissue via next-generation sequencing. We identified only one case of primary signet ring cell carcinoma of the eyelid with adequate DNA quality for sequencing from the pathological archive during the period 2000 to 2020. A comprehensive evaluation including histopathology, immunohistochemistry, and next-generation sequencing assay was performed on tumor tissue. Immunohistochemically, the tumor exhibited E-cadherin membranous staining with the aberrant cytoplasmic staining of β-catenin. Using next-generation sequencing, we demonstrated the mutation in the gene. In addition, other clinically actionable mutations including and were also detected. The alterations in other actionable genes indicate a need for larger studies to evaluate the pathogenesis and potential therapies for primary signet ring cell/histiocytoid carcinoma of the eyelid.
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http://dx.doi.org/10.3390/curroncol28010090DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7985776PMC
February 2021

Endosomal TLR3 co-receptor CLEC18A enhances host immune response to viral infection.

Commun Biol 2021 02 18;4(1):229. Epub 2021 Feb 18.

Genomics Research Center, Academia Sinica, Taipei, Taiwan.

Human C-type lectin member 18A (CLEC18A) is ubiquitously expressed in human, and highest expression levels are found in human myeloid cells and liver. In contrast, mouse CLEC18A (mCLEC18A) is only expressed in brain, kidney and heart. However, the biological functions of CLEC18A are still unclear. We have shown that a single amino acid change (S →R) in CTLD domain has profound effect in their binding to polysaccharides and house dust mite allergens. In this study, we further demonstrate that CLEC18A and its mutant CLEC18A(S339R) associate with TLR3 in endosome and bind poly (I:C) specifically. Compared to TLR3 alone, binding affinity to poly (I:C) is further increased in TLR3-CLEC18A and TLR3-CLEC18A(S339R) complexes. Moreover, CLEC18A and CLEC18A(S339R) enhance the production of type I and type III interferons (IFNs), but not proinflammatory cytokines, in response to poly (I:C) or H5N1 influenza A virus (IAV) infection. Compared to wild type (WT) mice, ROSA-CLEC18A and ROSA-CLEC18A(S339R) mice generate higher amounts of interferons and are more resistant to H5N1 IAV infection. Thus, CLEC18A is a TLR3 co-receptor, and may contribute to the differential immune responses to poly (I:C) and IAV infection between human and mouse.
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http://dx.doi.org/10.1038/s42003-021-01745-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7893028PMC
February 2021

Utility of Cerebrospinal Fluid Cell-Free DNA in Patients with EGFR-Mutant Non-Small-Cell Lung Cancer with Leptomeningeal Metastasis.

Target Oncol 2021 03 10;16(2):207-214. Epub 2021 Feb 10.

Department of Chest Medicine, Taipei Veterans General Hospital, 201, Section 2, Shih-Pai Road, Taipei, 112, Taiwan.

Background: Leptomeningeal metastasis (LM) is a fatal complication of advanced non-small-cell lung cancer (NSCLC).

Objective: The aim of this study was to evaluate the utility of cerebrospinal fluid (CSF) as a medium for epidermal growth factor receptor (EGFR) mutation testing in clinical practice.

Patients And Methods: We prospectively enrolled patients with EGFR-mutant NSCLC who underwent CSF sampling for suspected LM. The supernatant of CSF after routine cytology examination was collected. The diagnosis of LM was established according to EANO-ESMO criteria. CSF and plasma cell-free DNA (cfDNA) were retrieved for EGFR mutation testing.

Results: Fifty-one patients with a median age of 62.7 years were enrolled. The median duration from initial diagnosis to CSF sampling was 23.0 months and most patients (94.1%) had received at least one EGFR-tyrosine kinase inhibitor. Adenocarcinoma cells were found in 37 CSF samples (72.5%), and 48 (94.1%) patients had confirmed or probable LM. Thirty-five of these 48 patients (72.9%) had valid EGFR mutation-testing results using CSF cfDNA and tended to have higher white blood cell counts and positive cytology in their CSF compared to those with invalid mutation testing results. The overall detection rate of EGFR mutation in CSF cfDNA was 68.8%, and the T790M detection rate was 14.6%. In 37 patients with paired CSF and plasma samples, the concordance rate of the EGFR mutation results was 29.7%.

Conclusions: For patients with EGFR-mutant NSCLC with LM, CSF supernatant is a valuable source for EGFR mutation testing and may provide important information.
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http://dx.doi.org/10.1007/s11523-021-00791-9DOI Listing
March 2021

The clinical manifestations and interval changes of reverse-transcriptase quantitative polymerase chain reactions among different specimens of coronavirus disease 2019 patients.

J Chin Med Assoc 2021 02;84(2):151-157

Division of Infectious Diseases, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan, ROC.

Background: Since December 2019, a number of cases and deaths due to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic have been reported worldwide. In spite of clinical manifestations similar to the SARS-CoV epidemic in 2003, affected organs and severity are yet to be defined. Moreover, viral load alterations and viral shielding among different specimens remained scarce. Therefore, clarifying clinical presentations and correlations among viral loads, disease severity, and viral shielding of SARS-CoV-2 infection is crucial in the disease prevention.

Methods: The clinical courses of SARS-CoV-2 cases were presented through Gantt charts. Laboratory examinations and reverse-transcriptase quantitative polymerase chain reactions (RT-qPCR) among different specimens were tested periodically. Cycle thresholds (CT) were recorded and presented as viral loads.

Results: From March 2020 to April 2020, 4 SARS-CoV-2 cases were presented, of which, cases 1 and 2 manifested the symptoms severer than cases 3 and 4, along with higher serum lactate dehydrogenase levels and graded for lymphocytopenia. Case 4 initially exhibited anosmia but recovered within a short period. Curves of the CT of all the cases, except case 2, concaved upward after prescribing hydroxychloroquine (HCQ) and azithromycin. Except for case 4, the CT in most stool specimens remained undetectable; however, none of the cases presented gastrointestinal symptoms. Surprisingly, the CT values of the saliva specimens were inconsistent with those of the nasopharyngeal swabs and sputum.

Conclusion: SARS-CoV-2 manifests various symptoms. Sudden onset of central nervous system symptoms should be considered. The timing of HCQ and azithromycin administration might be a key factor in the viral load reduction. Positive prediction values of RT-qPCR of different specimens should be tested carefully to prevent false-negative results.
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http://dx.doi.org/10.1097/JCMA.0000000000000451DOI Listing
February 2021

Establishing diagnostic algorithms for SARS-CoV-2 nucleic acid testing in clinical practice.

J Chin Med Assoc 2020 Nov 9. Epub 2020 Nov 9.

Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taipei, Taiwan, ROC.

Background: Coronavirus disease 2019 (COVID-19), which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has become a global pandemic. Our laboratory initially used a two-step molecular assay, first reported by Corman et al., for SARS-CoV-2 identification (the T-CDC method). As rapid and accurate diagnosis of COVID-19 is required to control the spread of this infectious disease, the current study evaluated three commercially available assays, including the TaqPath COVID-19 Combo kit, the cobas® SARS-CoV-2 test and the Rendu 2019-nCoV Assay kit, to establish diagnostic algorithms for clinical laboratories.

Methods: A total of 790 clinical specimens, including nasopharyngeal swabs, throat swabs, sputum, saliva, stool, endotracheal aspirate and serum were obtained from patients who were suspected or already confirmed to have COVID-19 at the Taipei Veterans General Hospital from February to May 2020. These specimens were tested for SARS-CoV-2 using the different assays and the performance variance between the assays was analyzed.

Results: Of the assays we evaluated, the T-CDC method and the TaqPath COVID-19 Combo kit require lots of hands-on practical lab work, while the cobas® SARS-CoV-2 test and the Rendu 2019-nCoV Assay kit are fully automated detection systems. The T-CDC method and the TaqPath COVID-19 Combo kit showed similar detection sensitivity, however, the T-CDC method frequently delivered false positive signals for E and/or RdRP gene detection, thus increasing the risk of reporting false positive results. A manual test-based testing strategy combining the T-CDC method and the TaqPath COVID-19 Combo kit was developed, which demonstrated excellent concordance rates (>99%) with the cobas and Rendu automatic systems. There were a few cases showing discrepant results, which may be due to the varied detection sensitivities as well as targets among the different platforms. Moreover, the concordance rate between the cobas and Rendu assays was 100%.

Conclusion: Based on our evaluation, two SARS-CoV-2 diagnostic algorithms, one focusing on the manual assays and the other on the automatic platforms, were proposed. Our results provide valuable information that allow clinical laboratories to implement optimal diagnostic strategies for SARS-CoV-2 testing based on their clinical needs, such as test volume, turn-around time, and staff/resource limitations.
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http://dx.doi.org/10.1097/JCMA.0000000000000456DOI Listing
November 2020

TAZ negatively regulates the novel tumor suppressor ANKRD52 and promotes PAK1 dephosphorylation in lung adenocarcinomas.

Biochim Biophys Acta Mol Cell Res 2021 02 20;1868(2):118891. Epub 2020 Oct 20.

Institute of Clinical Medicine, National Yang-Ming University, Taipei, Taiwan; Institute of Microbiology and Immunology, National Yang-Ming University, Taipei, Taiwan; Institute of Biochemistry and Molecular Biology, National Yang-Ming University, Taipei, Taiwan; Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan; National Health Research Institute, Miaoli, Taiwan; National Chiao Tung University, Hsinchu, Taiwan. Electronic address:

Lung cancer is the leading cause of cancer death, and therefore the discovery of novel therapeutic targets is crucial. P21-activated kinase (PAK1) is an important oncogene involved in the signaling of actin cytoskeleton organization. Although PAK1 inhibition has been shown to suppress cancer progression, specific PAK1 inhibitors are not available due to the complex structure and insufficient understanding of this kinase. The Hippo signaling effector TAZ is known to be elevated in multiple human cancers and to promote cancer metastasis. This study aimed to explore the role of TAZ in regulating the tumor suppressor ankyrin repeat domain 52 (ANKRD52) and PAK1 activity. A negative correlation between TAZ and ANKRD52 was observed, with knockdown of TAZ leading to enhanced ANKRD52 promoter activity and increased mRNA levels. Moreover, reduced ANKRD52 levels were associated with late-stage lung cancer. Knockdowns of ANKRD52 resulted in elevated cell mobility, while forced ANKRD52 expression attenuated cell mobility. ANKRD52 is a subunit of the protein phosphatase 6 (PP6) holoenzyme. Mass spectrometry analysis revealed the interaction between PAK1 and the ANKRD52-PP6 complex. Knockdown of ANKRD52 or PP6c resulted in upregulated PAK1 phosphorylation. Our study demonstrates that the novel tumor suppressor protein ANKRD52 is transcriptionally inhibited by TAZ, regulating cell mobility through interactions with PP6c and dephosphorylation of PAK1.
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http://dx.doi.org/10.1016/j.bbamcr.2020.118891DOI Listing
February 2021

Multidisciplinary team discussion results in survival benefit for patients with stage III non-small-cell lung cancer.

PLoS One 2020 8;15(10):e0236503. Epub 2020 Oct 8.

Department of Chest Medicine, Taipei Veterans General Hospital, Taipei, Taiwan, Republic of China (R.O.C).

Background: The treatment for stage III non-small cell lung cancer (NSCLC) often involves multi-modality treatment. This retrospective study aimed to evaluate whether multidisciplinary team (MDT) discussion results in better patient survival.

Materials And Methods: MDT discussion was optional before February 2016 and was actively encouraged by the MDT committee beginning February 2016. We reviewed the medical charts and computer records of patients with stage III NSCLC between January 2013 and December 2018.

Results: A total of 515 patients were included. The median survival of all the patients was 33.9 months (M). The median survival of patients who were treated after MDT discussion was 41.2 M and that of patients treated without MDT discussion was 25.7 M (p = 0.018). The median survival of patients treated before February 2016 was 25.7 M and that of patients treated after February 2016 was 33.9 M (p = 0.003). The median survival of patients with stage IIIA tumors and those with stage IIIB tumors was 39.4 M and 25.7 M, respectively (p = 0.141). Multivariate analysis showed that MDT or not (p<0.001), T staging (p = 0.009), performance status (p<0.001), and surgery (p = 0.016) to be significant prognostic factors.

Conclusion: The results of the study show that MDT discussion results in survival benefit in patients with stage III NSCLC. The MDT discussion, performance status, and if surgery was performed were independent prognostic factors for patients with stage III NSCLC.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0236503PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7544080PMC
November 2020

Programmed death-ligand 1 (PD-L1)/thyroid transcription factor-1 double immunohistochemical staining facilitates scoring of tumor PD-L1 expression in cytopathology specimens from lung adenocarcinoma patients.

Cancer Cytopathol 2021 02 25;129(2):148-155. Epub 2020 Sep 25.

Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taipei, Taiwan.

Background: Immune checkpoint inhibitor therapy has revolutionized lung adenocarcinoma therapy. Treatment with antibodies against the immune checkpoint molecules programmed death-1 (PD-1) and programmed death-ligand 1 (PD-L1) can induce a durable response in a subset of patients. Immunohistochemistry characterization of tumor PD-L1 expression using either a histopathology specimen or a cytopathology specimen has been shown to correlate with treatment response. However, the current practice relies on pathologists' visual estimation of tumor PD-L1 staining, which can be variable in certain conditions. Highlighting tumor cells via double immunostaining with PD-L1 and thyroid transcription factor-1 (TTF-1) may improve estimation accuracy.

Methods: We performed PD-L1 single staining and PD-L1/TTF-1 double staining in 42 pairs of cytopathology and histopathology specimens from lung adenocarcinoma patients. An experienced pathologist visually estimated PD-L1 expression in each case and placed tumor PD-L1 expression into 1 of 3 categories: <1%, 1%-49%, or ≥50%. A medical technologist also performed estimations of the same cases based on a count of 200 tumor cells, and the results were compared.

Results: PD-L1/TTF-1 double immunohistochemistry could better identify the PD-L1-positive tumor cells in cytopathology specimens compared with PD-L1 single staining. The concordance of PD-L1 expression categorization between the pathologist's visual estimation and the medical technologist's counting was increased by double staining in cytopathology specimens (Cohen's weighted kappa: single stain, 0.784; double stain, 0.880). Double staining reduced possible error in the pathologist's visual estimation of PD-L1 expression from 9.5% to 4.8%. The benefit was not observed in histopathology specimens.

Conclusion: A simple PD-L1/TTF-1 double immunohistochemistry technique can be applied successfully to cytopathology specimens in better identifying patients who can potentially benefit from immune checkpoint blockade treatment.
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http://dx.doi.org/10.1002/cncy.22359DOI Listing
February 2021

Low-Dose Nicotine Activates EGFR Signaling via α5-nAChR and Promotes Lung Adenocarcinoma Progression.

Int J Mol Sci 2020 Sep 17;21(18). Epub 2020 Sep 17.

Institute of Clinical Medicine, National Yang Ming University, Taipei 112304, Taiwan.

Nicotine in tobacco smoke is considered carcinogenic in several malignancies including lung cancer. The high incidence of lung adenocarcinoma (LAC) in non-smokers, however, remains unexplained. Although LAC has long been less associated with smoking behavior based on previous epidemiological correlation studies, the effect of environmental smoke contributing to low-dose nicotine exposure in non-smoking population could be underestimated. Here we provide experimental evidence of how low-dose nicotine promotes LAC growth in vitro and in vivo. Screening of nicotinic acetylcholine receptor subunits in lung cancer cell lines demonstrated a particularly high expression level of nicotinic acetylcholine receptor subunit α5 (α 5-nAChR) in LAC cell lines. Clinical specimen analysis revealed up-regulation of α 5-nAChR in LAC tumor tissues compared to non-tumor counterparts. In LAC cell lines α 5-nAChR interacts with epidermal growth factor receptor (EGFR), positively regulates EGFR pathway, enhances the expression of epithelial-mesenchymal transition markers, and is essential for low-dose nicotine-induced EGFR phosphorylation. Functionally, low-dose nicotine requires α 5-nAChR to enhance cell migration, invasion, and proliferation. Knockdown of α 5-nAChR inhibits the xenograft tumor growth of LAC. Clinical analysis indicated that high level of tumor α 5-nAChR is correlated with poor survival rates of LAC patients, particularly in those expressing wild-type EGFR. Our data identified α 5-nAChR as an essential mediator for low-dose nicotine-dependent LAC progression possibly through signaling crosstalk with EGFR, supporting the involvement of environmental smoke in tumor progression in LAC patients.
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http://dx.doi.org/10.3390/ijms21186829DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7555382PMC
September 2020

Diffuse Intratumoral Stromal Inflammation in Ovarian Clear Cell Carcinoma is Associated With Loss of Mismatch Repair Protein and High PD-L1 Expression.

Int J Gynecol Pathol 2021 Mar;40(2):148-155

Ovarian clear cell carcinoma (OCCC) is an aggressive chemotherapy-resistant cancer with limited treatment options, and some OCCCs have mismatch repair (MMR) deficiency (MMRD). Emerging evidence has revealed that various cancers with MMRD are susceptible to anti-programmed death-1/programmed death ligand-1 (anti-PD-1/PD-L1) immunotherapy, and certain histologic features are associated with MMRD. However, few studies have addressed this in OCCC. We reviewed 76 OCCCs for tumor-associated inflammation (intratumoral stromal inflammation and peritumoral lymphocytes) and performed immunohistochemistry for 4 MMR proteins and PD-L1. MMR-deficient OCCCs were analyzed for microsatellite instability (MSI), and those with MLH1 loss were tested for MLH1 promoter methylation. No patients fulfilled the Amsterdam II criteria for the diagnosis of Lynch syndrome. Four (5.3%) tumors showed diffuse intratumoral stromal inflammation obliterating the tumor-stroma interfaces, and none had peritumoral lymphoid aggregates. MMRD was found in 2 (2.6%) tumors; one had MLH1/PMS2 loss (MSI-high and MLH1 promoter methylation was detected) and the other had MSH2/MSH6 loss (MSI-low). Twenty (26.3%) tumors showed tumoral PD-L1 expression ≥1%. Both MMR-deficient tumors showed diffuse intratumoral stromal inflammation and tumoral PD-L1 expression ≥50%. Three of the 4 (75%) tumors with diffuse intratumoral stromal inflammation also showed tumoral PD-L1 expression ≥50%. None of the tumors without diffuse intratumoral stromal inflammation showed MMRD (P=0.021) or tumoral PD-L1 expression ≥50% (P=0.0001). We identified a strong correlation among diffuse intratumoral stromal inflammation, MMRD, and high tumoral PD-L1 expression in a small but significant subset of OCCCs. Histologic evaluation can facilitate patient selection for subsequent anti-PD-1/PD-L1 immunotherapy.
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http://dx.doi.org/10.1097/PGP.0000000000000682DOI Listing
March 2021

A Grading System for Invasive Pulmonary Adenocarcinoma: A Proposal From the International Association for the Study of Lung Cancer Pathology Committee.

J Thorac Oncol 2020 10 17;15(10):1599-1610. Epub 2020 Jun 17.

Department of Diagnostic Pathology, National Cancer Center Hospital, Tokyo, Japan.

Introduction: A grading system for pulmonary adenocarcinoma has not been established. The International Association for the Study of Lung Cancer pathology panel evaluated a set of histologic criteria associated with prognosis aimed at establishing a grading system for invasive pulmonary adenocarcinoma.

Methods: A multi-institutional study involving multiple cohorts of invasive pulmonary adenocarcinomas was conducted. A cohort of 284 stage I pulmonary adenocarcinomas was used as a training set to identify histologic features associated with patient outcomes (recurrence-free survival [RFS] and overall survival [OS]). Receiver operating characteristic curve analysis was used to select the best model, which was validated (n = 212) and tested (n = 300, including stage I-III) in independent cohorts. Reproducibility of the model was assessed using kappa statistics.

Results: The best model (area under the receiver operating characteristic curve [AUC] = 0.749 for RFS and 0.787 for OS) was composed of a combination of predominant plus high-grade histologic pattern with a cutoff of 20% for the latter. The model consists of the following: grade 1, lepidic predominant tumor; grade 2, acinar or papillary predominant tumor, both with no or less than 20% of high-grade patterns; and grade 3, any tumor with 20% or more of high-grade patterns (solid, micropapillary, or complex gland). Similar results were seen in the validation (AUC = 0.732 for RFS and 0.787 for OS) and test cohorts (AUC = 0.690 for RFS and 0.743 for OS), confirming the predictive value of the model. Interobserver reproducibility revealed good agreement (k = 0.617).

Conclusions: A grading system based on the predominant and high-grade patterns is practical and prognostic for invasive pulmonary adenocarcinoma.
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http://dx.doi.org/10.1016/j.jtho.2020.06.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8362286PMC
October 2020

The Promises and Challenges of Tumor Mutation Burden as an Immunotherapy Biomarker: A Perspective from the International Association for the Study of Lung Cancer Pathology Committee.

J Thorac Oncol 2020 09 6;15(9):1409-1424. Epub 2020 Jun 6.

Department of Pathology, Memorial Sloan Kettering Cancer Center, New York, New York.

Immune checkpoint inhibitor (ICI) therapies have revolutionized the management of patients with NSCLC and have led to unprecedented improvements in response rates and survival in a subset of patients with this fatal disease. However, the available therapies work only for a minority of patients, are associated with substantial societal cost, and may lead to considerable immune-related adverse events. Therefore, patient selection must be optimized through the use of relevant biomarkers. Programmed death-ligand 1 protein expression by immunohistochemistry is widely used today for the selection of programmed cell death protein 1 inhibitor therapy in patients with NSCLC; however, this approach lacks robust sensitivity and specificity for predicting response. Tumor mutation burden (TMB), or the number of somatic mutations derived from next-generation sequencing techniques, has been widely explored as an alternative or complementary biomarker for response to ICIs. In theory, a higher TMB increases the probability of tumor neoantigen production and therefore, the likelihood of immune recognition and tumor cell killing. Although TMB alone is a simplistic surrogate of this complex interplay, it is a quantitative variable that can be relatively readily measured using currently available sequencing techniques. A large number of clinical trials and retrospective analyses, employing both tumor and blood-based sequencing tools, have evaluated the performance of TMB as a predictive biomarker, and in many cases reveal a correlation between high TMB and ICI response rates and progression-free survival. Many challenges remain before the implementation of TMB as a biomarker in clinical practice. These include the following: (1) identification of therapies whose response is best informed by TMB status; (2) robust definition of a predictive TMB cut point; (3) acceptable sequencing panel size and design; and (4) the need for robust technical and informatic rigor to generate precise and accurate TMB measurements across different laboratories. Finally, effective prediction of response to ICI therapy will likely require integration of TMB with a host of other potential biomarkers, including tumor genomic driver alterations, tumor-immune milieu, and other features of the host immune system. This perspective piece will review the current clinical evidence for TMB as a biomarker and address the technical sequencing considerations and ongoing challenges in the use of TMB in routine practice.
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http://dx.doi.org/10.1016/j.jtho.2020.05.019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8363213PMC
September 2020

CT-Guided Core Biopsy for Peripheral Sub-solid Pulmonary Nodules to Predict Predominant Histological and Aggressive Subtypes of Lung Adenocarcinoma.

Ann Surg Oncol 2020 Oct 3;27(11):4405-4412. Epub 2020 May 3.

School of Medicine, National Yang-Ming University, Taipei, Taiwan.

Background: Adenocarcinoma is the most common type of lung cancer, and pre-operative biopsy plays an important role to determine its major subtypes. As proposed by the International Association for the Study of Lung Cancer (IASLC), American Thoracic Society (ATS), and European Respiratory Society (ERS) in 2011, the predominant histological subtype of adenocarcinoma is an indicator of outcomes and recurrence rate. However, the value of CT-guided core biopsy in predicting the predominant subtype and detecting the presence of an aggressive subtype of adenocarcinoma, peripheral sub-solid nodule, has less been discussed.

Methods: We retrospectively reviewed 318 consecutive peripheral sub-solid nodules that underwent percutaneous CT-guided lung biopsy and surgical resection, between October 2015 and December 2018 and were diagnosed as adenocarcinoma with histological subtype. The subtyping results from biopsy and surgical pathology were compared to evaluate the concordance rate.

Results: The overall concordance rate between biopsy and surgical pathology in determining the predominant histological subtype was 64%. Better concordance was found in small tumors (≤ 2 cm), in predicting either predominant histology (χ = 7.091, P = 0.008) or high grade adenocarcinoma, micropapillary and/or solid subtype, MIP-SOL (χ = 22.301, P < 0.001). The analysis of ground glass opacity (GGO) component (C/T ratio) obtained significantly higher accuracy in the pure GGO group than in the other two groups in predicting predominant histology or high grade adenocarcinoma (χ = 17.560, P < 0.001 and χ = 61.938, P < 0.001, respectively).

Conclusions: CT-guided core biopsies provide additional value in predicting the histological subtype of lung adenocarcinoma after surgical resection, especially in small tumors (≤ 2 cm) or an initially pure GGO group.
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http://dx.doi.org/10.1245/s10434-020-08511-9DOI Listing
October 2020

Squamous cell carcinoma transformation after acquired resistance to osimertinib in a patient with lung adenocarcinoma harboring uncommon EGFR mutation.

J Formos Med Assoc 2020 Sep 12;119(9):1439-1441. Epub 2020 Mar 12.

Division of Thoracic Oncology, Department of Chest Medicine, Taipei Veterans General Hospital, Taipei, Taiwan; Faculty of Medicine, School of Medicine, National Yang-Ming University, Taipei, Taiwan. Electronic address:

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http://dx.doi.org/10.1016/j.jfma.2019.12.017DOI Listing
September 2020

Epidermal growth factor receptor mutations in non-small cell lung cancer undetected by high-sensitivity allele-specific real-time polymerase chain reaction-based assays.

J Chin Med Assoc 2020 Apr;83(4):345-349

Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taipei, Taiwan, ROC.

Background: Identifying epidermal growth factor receptor (EGFR) mutation status is critical for planning lung cancer treatment. Sanger sequencing detects both known and novel mutations but shows poor sensitivity. High-sensitivity allele-specific real-time polymerase chain reaction (ASRP)-based assays offer quick and reliable results, but may overlook uncommon mutations. We aimed to define the rate at which high-sensitivity ASRP-based assays missed uncommon EGFR mutations.

Methods: Non-small cell lung cancer specimens that were diagnosed as EGFR wild-type (EGFR-WT) by high-sensitivity ASRP-based assays and had residual DNA samples were sent for Sanger sequencing. Patient characteristics and clinical features were evaluated by chart review, and outcomes of EGFR-tyrosine kinase inhibitor (EGFR-TKI) therapy were studied.

Results: Hundred DNA specimens diagnosed by high-sensitivity ASRP-based assays as EGFR-WT were rechecked by Sanger sequencing. Two samples which were re-biopsy specimens from patients with EGFR mutations were excluded from the analysis. Sanger sequencing was failed in 24 samples. Among the remaining 74 samples, 6 (8.1%) had EGFR mutations-one exhibited exon 19 deletion (delT751_I759insS), two exhibited substitution mutations (S768I+V769L and L861Q), and three exhibited exon 20 insertions (N771_P772insN, P772_H773insHP, and H773_V774insAH). Only the patient with the exon 19 deletion had received EGFR-TKI therapy. Although the best tumor response was only stable disease, this was maintained for >10 months.

Conclusion: High-sensitivity ASRP-based assays can overlook uncommon mutations. This detection failure rate is worth noting, especially when treating patients from regions known to have a high prevalence of EGFR mutation. Patients carrying uncommon mutations may still benefit from EGFR-TKI therapy.
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http://dx.doi.org/10.1097/JCMA.0000000000000277DOI Listing
April 2020

IASLC Multidisciplinary Recommendations for Pathologic Assessment of Lung Cancer Resection Specimens After Neoadjuvant Therapy.

J Thorac Oncol 2020 05 28;15(5):709-740. Epub 2020 Jan 28.

Department of Pathology, University of Turin, Torino, Italy.

Currently, there is no established guidance on how to process and evaluate resected lung cancer specimens after neoadjuvant therapy in the setting of clinical trials and clinical practice. There is also a lack of precise definitions on the degree of pathologic response, including major pathologic response or complete pathologic response. For other cancers such as osteosarcoma and colorectal, breast, and esophageal carcinomas, there have been multiple studies investigating pathologic assessment of the effects of neoadjuvant therapy, including some detailed recommendations on how to handle these specimens. A comprehensive mapping approach to gross and histologic processing of osteosarcomas after induction therapy has been used for over 40 years. The purpose of this article is to outline detailed recommendations on how to process lung cancer resection specimens and to define pathologic response, including major pathologic response or complete pathologic response after neoadjuvant therapy. A standardized approach is recommended to assess the percentages of (1) viable tumor, (2) necrosis, and (3) stroma (including inflammation and fibrosis) with a total adding up to 100%. This is recommended for all systemic therapies, including chemotherapy, chemoradiation, molecular-targeted therapy, immunotherapy, or any future novel therapies yet to be discovered, whether administered alone or in combination. Specific issues may differ for certain therapies such as immunotherapy, but the grossing process should be similar, and the histologic evaluation should contain these basic elements. Standard pathologic response assessment should allow for comparisons between different therapies and correlations with disease-free survival and overall survival in ongoing and future trials. The International Association for the Study of Lung Cancer has an effort to collect such data from existing and future clinical trials. These recommendations are intended as guidance for clinical trials, although it is hoped they can be viewed as suggestion for good clinical practice outside of clinical trials, to improve consistency of pathologic assessment of treatment response.
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http://dx.doi.org/10.1016/j.jtho.2020.01.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8173999PMC
May 2020

Preoperative biopsy and tumor recurrence of stage I adenocarcinoma of the lung.

Surg Today 2020 Jul 23;50(7):673-684. Epub 2019 Dec 23.

Division of Thoracic Surgery, Department of Surgery, Taipei Veterans General Hospital, 201, Section 2, Shih-Pai Road, Taipei, Taiwan.

Purpose: To evaluate whether preoperative biopsy affects the outcomes of patients undergoing at least lobectomy for stage I lung adenocarcinoma.

Methods: We reviewed the medical records of patients who underwent surgery for stage I lung adenocarcinoma between 2006 and 2013. Tumor recurrence and survival were compared between patients who underwent preoperative biopsy, including computed tomographic-guided needle biopsy and transbronchial biopsy, and those who underwent intraoperative frozen section.

Results: Among 509 patients, 229 patients (44.9%) underwent preoperative biopsy and 280 patients had lung adenocarcinoma diagnosed by intraoperative frozen section (reference group). Recurrence developed in 65 (12.8%) patients within a median follow-up period of 54.4 months. Multivariate analysis demonstrated that preoperative biopsy (OR 1.97, p = 0.045), radiological solid appearance (OR 5.43, p < 0.001), and angiolymphatic invasion (OR 2.48, p = 0.010) were independent predictors of recurrence. In the overall cohort, preoperative biopsy appeared to worsen 5-year disease-free and overall survival significantly (76.6% vs. 93.0%, p < 0.001; and 83.8% vs. 94.5%, p = 0.002, respectively) compared with the reference group. After propensity matching, multivariable logistic regression still identified preoperative biopsy as an independent predictor of overall recurrence (OR 2.21, p = 0.048) after adjusting for tumor characteristics.

Conclusion: Preoperative biopsy might be considered a prognosticator of recurrence of stage I adenocarcinoma of the lungs in patients who undergo at least anatomic lobectomy without postoperative adjuvant chemotherapy.
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http://dx.doi.org/10.1007/s00595-019-01941-3DOI Listing
July 2020

PD-L1 Testing for Lung Cancer in 2019: Perspective From the IASLC Pathology Committee.

J Thorac Oncol 2020 04 20;15(4):499-519. Epub 2019 Dec 20.

Royal Brompton and Harefield NHS Foundation Trust, London, and National Heart and Lung Institute, Imperial College, United Kingdom.

The recent development of immune checkpoint inhibitors (ICIs) has led to promising advances in the treatment of patients with NSCLC and SCLC with advanced or metastatic disease. Most ICIs target programmed cell death protein 1 (PD-1) or programmed death ligand 1 (PD-L1) axis with the aim of restoring antitumor immunity. Multiple clinical trials for ICIs have evaluated a predictive value of PD-L1 protein expression in tumor cells and tumor-infiltrating immune cells (ICs) by immunohistochemistry (IHC), for which different assays with specific IHC platforms were applied. Of those, some PD-L1 IHC assays have been validated for the prescription of the corresponding agent for first- or second-line treatment. However, not all laboratories are equipped with the dedicated platforms, and many laboratories have set up in-house or laboratory-developed tests that are more affordable than the generally expensive clinical trial-validated assays. Although PD-L1 IHC test is now deployed in most pathology laboratories, its appropriate implementation and interpretation are critical as a predictive biomarker and can be challenging owing to the multiple antibody clones and platforms or assays available and given the typically small size of samples provided. Because many articles have been published since the issue of the IASLC Atlas of PD-L1 Immunohistochemistry Testing in Lung Cancer, this review by the IASLC Pathology Committee provides updates on the indications of ICIs for lung cancer in 2019 and discusses important considerations on preanalytical, analytical, and postanalytical aspects of PD-L1 IHC testing, including specimen type, validation of assays, external quality assurance, and training.
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http://dx.doi.org/10.1016/j.jtho.2019.12.107DOI Listing
April 2020

Integrative analyses of noncoding RNAs reveal the potential mechanisms augmenting tumor malignancy in lung adenocarcinoma.

Nucleic Acids Res 2020 02;48(3):1175-1191

Genome and Systems Biology Program, National Taiwan University and Academia Sinica, Taipei 10617, Taiwan.

Precise noncoding RNA (ncRNA)-based network prediction is necessary to reveal ncRNA functions and pathological mechanisms. Here, we established a systemic pipeline to identify prognostic ncRNAs, predict their functions and explore their pathological mechanisms in lung adenocarcinoma (LUAD). After in silico and experimental validation based on evaluations of prognostic value in multiple LUAD cohorts, we selected the PTTG3P pseudogene from among other prognostic ncRNAs (MIR497HG, HSP078, TBX5-AS1, LOC100506990 and C14orf64) for mechanistic studies. PTTG3P upregulation in LUAD cells shortens the metaphase to anaphase transition in mitosis, increases cell viability after cisplatin or paclitaxel treatment, facilitates tumor growth that leads to poor survival in orthotopic lung models, and is associated with a poor survival rate in LUAD patients in the TCGA cohort who received chemotherapy. Mechanistically, PTTG3P acts as an ncRNA that interacts with the transcription factor FOXM1 to regulate the transcriptional activation of the mitotic checkpoint kinase BUB1B, which augments tumor growth and chemoresistance and leads to poor outcomes for LUAD patients. Overall, we established a systematic strategy to uncover prognostic ncRNAs with functional prediction methods suitable for pan-cancer studies. Moreover, we revealed that PTTG3P, due to its upregulation of the PTTG3P/FOXM1/BUB1B axis, could be a therapeutic target for LUAD patients.
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http://dx.doi.org/10.1093/nar/gkz1149DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7026595PMC
February 2020

A two-dimensional immunomagnetic nano-net for the efficient isolation of circulating tumor cells in whole blood.

Nanoscale 2019 Nov 20;11(44):21119-21127. Epub 2019 Sep 20.

Genomics Research Center, Academia Sinica, Taipei, Taiwan. and Department of Chemical Engineering, Stanford University, Stanford, California, USA.

An immunomagnetic "nano-net" was designed and synthesized for specifically capturing rare cells of interest from mixtures. The nano-net, [email protected], consists of conjugated antibody molecules on a lipid coated magnetic nanoparticle-graphene oxide sheet complex. The magnetism, chemical composition, and the morphology of the construct and its precursors were characterized by SQUID, FTIR, TGA, DLS and SEM, to confirm the feasibility of the synthetic steps and the resulting properties suitable for solution phase immuno-recognition for cell capture. When applied to capturing circulating tumor cells (CTCs) in oral, colon and lung cancer clinical patients' blood samples, the nano-net construct exhibited far superior ability whereas conventional immunomagnetic beads in some cases were unable to capture any CTCs, even by increasing the bead concentration. Confocal images showed that the nano-net wrapped around the CTCs while the immunomagnetic beads attached them with point contacts. A stable, patch-like multivalent matrix nano-net was demonstrated to tackle the shortcomings of single point contact of immunomagnetic beads to the target cell. This strategy is universal for any cell separation in complex fluids.
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http://dx.doi.org/10.1039/c9nr06256dDOI Listing
November 2019

AKT1 internal tandem duplications and point mutations are the genetic hallmarks of sclerosing pneumocytoma.

Mod Pathol 2020 03 16;33(3):391-403. Epub 2019 Sep 16.

Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taipei, Taiwan.

Sclerosing pneumocytoma is a unique benign neoplasm of the lungs. The molecular alterations in sclerosing pneumocytoma are not well understood. In a previous whole-exome sequencing study, recurrent AKT1 point mutation was observed in about half of the cases of sclerosing pneumocytoma. However, in the remaining half, cancer-related mutations have still not been identified. In this study, we first analyzed the raw sequence data from the previous whole-exome sequencing study (PRJNA297066 cohort). Using Genomon-ITDetector, a special software for detection of internal tandem duplications, we identified recurrent internal tandem duplications in the AKT1 gene in 22 of the 44 tumor samples (50%). All the cases positive for AKT1 internal tandem duplications lacked AKT1 point mutations. Next, we performed targeted next-generation sequencing in an independent cohort of sclerosing pneumocytoma from our hospital (VGH-TPE cohort), and again identified recurrent AKT1 internal tandem duplications in 20 of the 40 (50%) tumor samples analyzed. The internal tandem duplications resulted in duplications of 7 to 16 amino acids in a narrow region of the Pleckstrin homology domain of the AKT1 protein. This region contains the interaction interface between the Pleckstrin homology and kinase domains, which is known to play a critical role in the activation of the AKT1 protein. Moreover, we found that AKT1 internal tandem duplications were mutually exclusive of other forms of AKT1 mutations, including point mutations and short indels. Taking all forms of AKT1 mutations together, we detected AKT1 mutations in almost all the sclerosing pneumocytomas in our study (PRJNA297066 cohort: 41 out of 44 cases, 93%; VGH-TPE cohort: 40 out of 40 cases, 100%). Our results suggest that AKT1 mutation is the genetic hallmark of sclerosing pneumocytoma. These results would help in better understanding of the pathogenesis of sclerosing pneumocytoma.
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http://dx.doi.org/10.1038/s41379-019-0357-yDOI Listing
March 2020

Cellular prion protein transcriptionally regulated by NFIL3 enhances lung cancer cell lamellipodium formation and migration through JNK signaling.

Oncogene 2020 01 2;39(2):385-398. Epub 2019 Sep 2.

Program in Molecular Medicine, National Yang-Ming University and Academia Sinica, Taipei, 11221, Taiwan.

Tumor invasion and metastasis are the major causes of treatment failure and mortality in lung cancer patients. In this study, we identified a group of genes with differential expression in in situ and invasive lung adenocarcinoma tissues by expression profiling; among these genes we further characterized the association of the upregulation of PRNP, the gene encoding cellular Prion protein (PrPc), with lung adenocarcinoma invasiveness. Immunohistochemistry on clinical specimens showed an association of PrPc expression with invasive but not in situ lung adenocarcinoma. Consistently, the expression of PrPc was higher in the highly invasive than in the lowly invasive lung adenocarcinoma cell lines. Knockdown of PrPc expression in cultured lung adenocarcinoma cells decreased their lamellipodium formation, in vitro migration and invasion, and in vivo experimental lung metastasis. Phosphorylation of JNKs was found to correlate with PrPc expression and the inhibition of JNKs suppressed the PrPc-induced up-regulation of lamellipodium formation, cell migration, and invasion. Moreover, we identified the nuclear factor, interleukin 3 regulated (NFIL3) protein as a transcriptional activator of the PRNP promoter. Accordingly, NFIL3 promoted lung cancer cell migration and invasion in a PrPc-dependent manner. High NFIL3 expression in clinical specimens of lung adenocarcinoma was also associated with tumor invasiveness. Overall, our observations suggest that the NFIL3/PrPc axis, through regulating lamellipodium formation and cell mobility via JNK signaling, plays a critical role in lung cancer invasiveness and metastasis.
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http://dx.doi.org/10.1038/s41388-019-0994-0DOI Listing
January 2020

The importance of EGFR mutation testing in squamous cell carcinoma or non-small cell carcinoma favor squamous cell carcinoma diagnosed from small lung biopsies.

Diagn Pathol 2019 Jun 21;14(1):59. Epub 2019 Jun 21.

Division of Molecular Pathology, Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taipei, Taiwan.

Background: Adenosquamous carcinoma (ADSC) of the lung, a rare but aggressive subtype of non-small cell lung cancer (NSCLC), is defined as a carcinoma containing components of adenocarcinoma (ADC) and squamous cell carcinoma (SqCC). Mutations of epidermal growth factor receptor (EGFR) are found at a frequency of 15 to 44% in Asian ADSC, and EGFR tyrosine kinase inhibitors (EGFR-TKIs) are a more effective treatment for EGFR-mutated ADSC compared to chemotherapy. However, ADSC in small lung biopsies could be misdiagnosed as SqCC or non-small cell carcinoma (NSCC) favor SqCC due to undersampling, which may result in neglecting of EGFR mutation testing and affecting patients' clinical management, particularly in Asian patients that relatively have high prevalence of EGFR mutation.

Methods: A total of 148 small lung biopsy cases with pathological diagnosis of SqCC or NSCC favor SqCC were retrospectively enrolled. The frequency of EGFR mutations and the correlation between patients' EGFR mutation status and clinicopathological characteristics were evaluated.

Results: EGFR mutations were found in 8.8% (13 /148) of all cases with 5.2% (7/135) in SqCC and 46.2% (6/13) in NSCC favor SqCC. There were 7 (53.8%) L858R mutation, 4 (30.8%) exon 19 deletions, and 2 (15.4%) cases with coexistent L858R and T790 M mutations. Multivariate analysis showed that EGFR mutations were more prevalent in never-smokers (83.3% versus 16.7%, p = 0.006) and patients diagnosed as NSCC favor SqCC (46.2% versus 5.2%, p = 0.001). Moreover, 75% (3/4) of EGFR mutation-positive cases with subsequent surgical resection or rebiopsy were further diagnosed as ADSC.

Conclusions: EGFR mutation testing should be performed in Asian patients with SqCC diagnosed from small lung biopsies, especially in never-smokers and patients with diagnosis of NSCC favor SqCC, which have a high probability of being ADSC.
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http://dx.doi.org/10.1186/s13000-019-0840-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6587243PMC
June 2019

PD-L1 is a double-edged sword in colorectal cancer: the prognostic value of PD-L1 depends on the cell type expressing PD-L1.

J Cancer Res Clin Oncol 2019 Jul 25;145(7):1785-1794. Epub 2019 May 25.

Institute of Clinical Medicine, National Yang-Ming University, Taipei, Taiwan.

Purpose: To investigate the associations between programmed cell death ligand-1 (PD-L1) on tumor cells (TCs) or PD-L1 on tumor-infiltrating immune cells (TIICs) and the microsatellite instability (MSI) status in colorectal cancer (CRC).

Methods: In total, 238 CRC patients were enrolled. PD-L1 expression and MSI status were studied by immunohistochemical staining and polymerase chain reaction. The χ test was used to compare characteristics. The Kaplan-Meier method was used for survival analysis. Cox proportional hazards models were used to determine the prognostic influence of clinicopathological factors.

Results: Eighteen patients (7.6%) were had MSI-high (MSI-H) CRC. The number of patients with PD-L1 expression on TCs, stromal TIICs and intraepithelial TIICs was 13 (5.5%), 64 (26.9%) and 45 (18.9%), respectively. The MSI-H phenotype was significantly associated with younger age, right sidedness, mucinous component, high grade, stromal TIICs expressing PD-L1 (P = 0.042) and intraepithelial TIICs expressing PD-L1 (P < 0.001), but not TCs expressing PD-L1. PD-L1-expressing TCs were an independent marker of poor prognosis [hazard ratio (HR) = 3.387, P = 0.003], and PD-L1-expressing stromal TIICs were an independent marker of good prognosis (HR = 0.551, P < 0.001).

Conclusions: PD-L1-expressing TCs were a marker of poor prognosis; in contrast, PD-L1-expressing TIICs were a marker of good prognosis. The MSI-H phenotype was associated with the presence of PD-L1-expressing TIICs, but not of PD-L1-expressing TCs.
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http://dx.doi.org/10.1007/s00432-019-02942-yDOI Listing
July 2019
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