Zhaolin Xu

Loss of lkb1 Expression Reduces the Latency of ErbB2-Mediated Mammary Gland Tumorigenesis, Promoting Changes in Metabolic Pathways

The tumor suppressor kinase LKB1 is mutated in a broad range of cancers however, the role of LKB1 mammary gland tumorigenesis is not fully understood. Evaluation of human breast cancer tissue microarrays, indicate that 31% of HER2 positive samples lacked LKB1 expression. To expand on these observations, we crossed STK11fl/fl mice with mice genetically engineered to express activated Neu/HER2-MMTV-Cre (NIC) under the endogenous Erbb2 promoter, to generate STK11−/−/NIC mice. In these mice, the loss of lkb1 expression reduced the latency of ErbB2-mediated tumorigenesis compared to the latency of tumorigenesis in NIC mice alone. Analysis of STK11−/−/NIC mammary tumors revealed hyperactivation of mammalian target of rapamycin (mTOR) through both mTORC1 and mTORC2 pathways as determined by the phosphorylation status of ribosomal protein S6 and AKT. Furthermore, STK11−/−/NIC mammary tumors had elevated ATP levels along with changes in metabolic enzymes and metabolites. The treatment of primary mammary tumor cells with specific mTOR inhibitors AZD8055 and Torin1, that target both mTOR complexes, attenuated mTOR activity and decreased expression of glycolytic enzymes. Our findings underscore the existence of a molecular interplay between LKB1-AMPK-mTORC1 and ErbB2-AKT-mTORC2 pathways with mTOR at its epicenter, suggestive that loss of LKB1 expression may serve as a marker for hyperactivated mTOR in HER2 positive breast cancer and warranting further investigation into therapeutics that target LKB1-AMPK-mTOR and glycolytic pathways.

The role of atypical pathogens in community-acquired pneumonia.

The term atypical pneumonia was first used in 1938, and by the 1970s it was widely used to refer to pneumonia due to Mycoplasma pneumoniae, Legionella pneumophila (or other Legionella species), and Chlamydophila pneumoniae. However, in the purest sense all pneumonias other than the classic bacterial pneumonias are atypical. Currently many favor abolition of the term atypical pneumonia.This review categorizes atypical pneumonia pathogens as conventional ones; viral agents and emerging atypical pneumonia pathogens. We emphasize viral pneumonia because with the increasing availability of multiplex polymerase chain reaction we can identify the agent(s) responsible for viral pneumonia. By using a sensitive assay for procalcitonin one can distinguish between viral and bacterial pneumonia. This allows pneumonia to be categorized as bacterial or viral at the time of admission to hospital or at discharge from the emergency department and soon thereafter further classified as to the etiology, which should be stated as definite or probable.

MAPK Kinase 3 Is a Tumor Suppressor with Reduced Copy Number in Breast Cancer

Cancers are initiated as a result of changes that occur in the genome. Identification of gains and losses in the structure and expression of tumor-suppressor genes and oncogenes lies at the root of the understanding of cancer cell biology. Here, we show that the mitogen-activated protein kinase (MAPK) MKK3 suppresses the growth of breast cancer, in which it varies in copy number. A pervasive loss of MKK3 gene copy number in patients with breast cancer is associated with an impairment of MKK3 expression and protein level in malignant tissues. To assess the functional role of MKK3 in breast cancer, we showed in an animal model that MKK3 activity is required for suppression of tumor growth. Active MKK3 enhanced expression of the cyclin-dependent kinase inhibitors p21(Cip1/Waf1) and p27(Kip1), leading to increased cell-cycle arrest in G1 phase of the cell cycle. Our results reveal the functional significance of MKK3 as a tumor suppressor and improve understanding of the dynamic role of the MAPK pathway in tumor progression.

Physiologic and biologic characteristics of three experimental models of acute lung injury in rats.

BACKGROUND: Strategies to attenuate ventilator-associated lung injury have been tested in various experimental methods of acute lung injury (ALI). Conclusions are often drawn from physiologic and biologic effects, but the influence of the model on these results is not known. Our aim in this study was to characterize frequently used models of experimental ALI. METHODS: Twenty Sprague Dawley rats were anesthetized and their lungs mechanically ventilated for 5 hours. Three models of ALI (surfactant washout, acid aspiration, and high tidal volume ventilation) were investigated with regard to hemodynamics, respiratory mechanics, gas exchange, lung pathology, and inflammatory reactions. Animals without ALI served as controls. RESULTS: Five animals in each group were analyzed. Dynamic compliance and PaO2/fraction of inspired oxygen ratio decreased by at least 50% in all groups after 1 hour. Whereas compliance remained decreased in all models, oxygenation returned to baseline values in the lavage group after 5 hours. Diffuse alveolar damage was worse in the high tidal volume model and was not different between the control and lavage animals. Interleukin-6 was increased in bronchoalveolar lavage fluid in the aspiration and high tidal volume models. CONCLUSIONS: Although comparable physiologic effects meeting acute respiratory distress syndrome criteria were achieved in all models, the biologic responses varied among lung injury models. The acid aspiration model created both respiratory and inflammatory responses typically seen in ALI; these data suggest that it may be the most clinically applicable model to study the intermediate-term effects of ventilator-associated lung injury in rats.

Predicting EGFR mutation status in lung cancer:Proposal for a scoring model using imaging and demographic characteristics

ObjectiveTo determine if a combination of CT and demographic features can predict EGFR mutation status in bronchogenic carcinoma.MethodsWe reviewed demographic and CT features for patients with molecular profiling for resected non-small cell lung carcinoma. Using multivariate logistic regression, we identified features predictive of EGFR mutation. Prognostic factors identified from the logistic regression model were then used to build a more practical scoring system.ResultsA scoring system awarding 5 points for no or minimal smoking history, 3 points for tumours with ground glass component, 3 points for airbronchograms, 2 points for absence of preoperative evidence of nodal enlargement or metastases and 1 point for doubling time of more than a year, resulted in an AUROC of 0.861. A total score of at least 8 yielded a specificity of 95 %. On multivariate analysis sex was not found to be predictor of EGFR status.ConclusionsA weighted scoring system combining imaging and demographic data holds promise as a predictor of EGFR status. Further studies are necessary to determine reproducibility in other patient groups. A predictive score may help determine which patients would benefit from molecular profiling and may help inform treatment decisions when molecular profiling is not possible.Key points• EGFR mutation-targeted chemotherapy for bronchogenic carcinoma has a high success rate.• Mutation testing is not possible in all patients.• EGFR associations include subsolid density, slow tumour growth and minimal/no smoking history.• Demographic or imaging features alone are weak predictors of EGFR status.• A scoring system, using imaging and demographic features, is more predictive.

Cystic fibrosis transmembrane conductance regulator dysfunction in VIP knockout mice

Vasoactive intestinal peptide (VIP), a neuropeptide, controls multiple functions in exocrine tissues, including inflammation, and relaxation of airway and vascular smooth muscles, and regulates CFTR-dependent secretion, which contributes to mucus hydration and local innate defense of the lung. We had previously reported that VIP stimulates the VPAC1 receptor, PKCϵ signaling cascade, and increases CFTR stability and function at the apical membrane of airway epithelial cells by reducing its internalization rate. Moreover, prolonged VIP stimulation corrects the molecular defects associated with F508del, the most common CFTR mutation responsible for the genetic disease cystic fibrosis. In the present study, we have examined the impact of the absence of VIP on CFTR maturation, cellular localization, and function in vivo using VIP knockout mice. We have conducted pathological assessments and detected signs of lung and intestinal disease. Immunodetection methods have shown that the absence of VIP results in CFTR intracellular retention despite normal expression and maturation levels. A subsequent loss of CFTR-dependent chloride current was measured in functional assays with Ussing chamber analysis of the small intestine ex vivo, creating a cystic fibrosis-like condition. Interestingly, intraperitoneal administration of VIP corrected tissue abnormalities, close to the wild-type phenotype, as well as associated defects in the vital CFTR protein. The results show in vivo a primary role for VIP chronic exposure in CFTR membrane stability and function and confirm in vitro data.

Abstract A26: Identification of a novel therapeutic target in lung adenocarcinoma

The reactivation of biologic signaling events that occur throughout fetal development has been observed during malignant cell transformation and tumor progression. Transcription factors are typically at the hub of these signaling events, such as NKX2-1 and several ETS transcription factors. ELF3 is an uncharacterized ETS family member that is highly expressed during fetal lung development and could play a biologic role in lung cancer based on its location within the recurrently gained chromosome 1q. We hypothesize that ELF3 is a novel oncogenic transcription factor and a therapeutic target. Multiple independent datasets encompassing 1,685 clinical samples of lung adenocarcinoma (LUAD), lung squamous cell carcinoma (LUSC), small cell lung cancer, and nonmalignant lung tissues were analyzed to establish the frequency of ELF3 overexpression and underlying genetic mechanisms of selection. ELF3 overexpression was validated by immunohistochemistry. Associations with patient survival were tested using the log-rank method. Isogenic cell lines were established to assess oncogenic phenotypes including tumor growth in a xenograft model. Protein-protein interaction (PPI) networks were constructed around ELF3, and integrated pathway analysis was performed to decipher the signaling network disruptions resulting from ELF3 overexpression. ELF3 overexpression was frequently observed in LUAD (>2-fold: BCCA 73% TCGA 40%), but was not observed in other lung cancer subtypes. Similarly, high ELF3 expression was significantly associated with poor overall survival of LUAD patients (p We have deciphered the oncogenic role of ELF3 in LUAD. Its requirement for tumor growth in our model indicates that therapeutic targeting of ELF3 could benefit the 73% of patients who display ELF3 overexpression. Citation Format: Katey S.S. Enfield, Erin A. Marshall, Christine Anderson, Kevin W. Ng, Sara Rahmati, Zhaolin Xu, Calum E. MacAulay, Stephen Lam, William W. Lockwood, Raj Chari, Aly Karsan, Igor Jurisica, Wan L. Lam. Identification of a novel therapeutic target in lung adenocarcinoma [abstract]. In: Proceedings of the Fifth AACR-IASLC International Joint Conference: Lung Cancer Translational Science from the Bench to the Clinic; Jan 8-11, 2018; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(17_Suppl):Abstract nr A26.

Abstract B36: Novel miRNAs as tissue-of-origin markers for distinguishing malignant pleural mesothelioma from lung adenocarcinoma

The outcome of patients with malignant pleural mesothelioma (MPM) is poor, and diagnosis is complicated by a lack of biomarkers capable of distinguishing primary MPM from cancers that have metastasized to the pleura. Clinical diagnosis and tissue of origin is currently assessed through the use of a panel of positive and negative markers; however, there remains a subset of cases that are not identifiable by current clinical biomarkers. Recent studies suggest that the human genome encodes more miRNAs than are currently annotated, and that the novel miRNAs may display enhanced tissue and lineage specificity. We conducted a de novo search for novel miRNAs by applying a prediction algorithm to the small RNA-sequence data in a cohort of MPM tumors (n=87) from The Cancer Genome Atlas (TCGA). This analysis yielded 424 predicted novel miRNA-like sequences, which were subsequently filtered by RNA structure, abundance, and genomic location to identify 154 previously unannotated miRNA sequences. This represents a significant increase to the repertoire of 1,597 annotated miRNAs in MPM. Protein-coding genes predicted to be targeted by these novel miRNAs, using the miRanda algorithm, include genes involved in MPM biology. One of the most highly expressed novel miRNAs identified targets the Ataxia Telangiectasia Mutated (ATM) gene. Another target gene, BRCA1 Associated Protein 1 (BAP1), is also in the DNA damage response pathway. To investigate the ability of these 154 novel miRNAs to distinguish MPM from other thoracic cancers, we assessed their expression in 1,093 lung tumors from four independent cohorts from TCGA and the BC Cancer Agency (BCCA): two adenocarcinoma (LUAD) cohorts (TCGA n=497, BCCA n=94) and two squamous cell carcinoma (LUSC) cohorts (TCGA n=467, BCCA n=35). Principal component analyses revealed that novel miRNA expression was able to unambiguously distinguish MPM from LUAD and LUSC. Furthermore, we developed an miRNA-based classifier model using the weighted voting class prediction method. A 10 novel miRNAs classifier was deduced by comparing MPM and LUAD cases from TCGA and validated by comparing MPM against LUAD cases from the BCCA cohort. Remarkably, this classifier successfully identified 86 out of the 87 MPM cases (98.8%) and 100% of LUAD cases (true positive rate = 98.85%, false positive rate = 1.15%). The strikingly high sensitivity and specificity in distinguishing MPM from LUAD illustrates the potential of using novel miRNAs to supplement current clinical markers to define MPM. Citation Format: Erin A. Marshall, Christine Anderson, Kevin W. Ng, Brenda C. Minatel, Katey S.S. Enfield, Adam P. Sage, Zhaolin Xu, Wan L. Lam, Victor D. Martinez. Novel miRNAs as tissue-of-origin markers for distinguishing malignant pleural mesothelioma from lung adenocarcinoma [abstract]. In: Proceedings of the Fifth AACR-IASLC International Joint Conference: Lung Cancer Translational Science from the Bench to the Clinic; Jan 8-11, 2018; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(17_Suppl):Abstract nr B36.

Abstract B21: Investigating targeted driver mutations and PD-L1 expression for improved therapy of non-small cell lung cancer

Most lung cancer patients are diagnosed at an advanced stage, limiting their treatment options to chemotherapies that have very low response rate. New therapies that target driver gene mutations (e.g., EGFR, ALK, ROS1, BRAF) are being used to treat patients who have tumors with these mutations. In addition, a type of immunotherapy called immune checkpoint inhibitor is being used to treat lung cancer patients. For instance, patients with tumors that express PD-L1 are responsive to anti-PD-1/PD-L1 therapy. Thus, being able to identify the presence of driver mutations and PD-L1 will help patients to benefit from different therapies. A total of 844 cases of non-small cell lung cancer samples have been profiled for the presence of EGFR, KRAS, BRAF, PIK3CA, and HER2 mutations by SNaPshot/sizing genotyping. Immunohistochemistry (IHC) was used to identify the protein expression of ALK and PD-L1. Histologic examination was performed to determine the pathologic type, grade, and lymphatic/vascular invasion. Statistical analysis revealed a number of correlations between the presence of the mutations, PD-L1 expression and the patient pathologic data. Specifically, it was determined that women had lung tumors with a significantly greater number of EGFR mutations than men (p value = 0.001). Examining the presence of ALK, EGFR, KRAS, BRAF, PIK3CA, and HER2 mutations against the presence of pleural invasions yielded a p-value of 0.071, which implies evidence that different mutations are not associated with pleural invasion. However, EGFR mutations were associated with the absence of vascular and lymphatic invasions in lung cancer patients (p value = 0.001, 0.002 respectively). In addition, while the expression of PD-L1 does not associate with the patients who express KRAS mutation, it is associated with lung cancer patients who express EGFR mutation (p value = 0.002). Knowing the mutational and PD-L1 status in lung cancer patients will help patients benefit from targeted therapies and/or checkpoint inhibitors. Citation Format: Akram Alwithenani, Marika Forsythe, Mathieu Castonguay, Wenda Greer, Gorden Flowerdew, Drew Bethune, Harry Henteleff, Madelaine Plourde, Aneil Mujoomdar, Daniel French, Micheal Johnston, Paola Marcato, Zhaolin Xu. Investigating targeted driver mutations and PD-L1 expression for improved therapy of non-small cell lung cancer [abstract]. In: Proceedings of the Fifth AACR-IASLC International Joint Conference: Lung Cancer Translational Science from the Bench to the Clinic; Jan 8-11, 2018; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(17_Suppl):Abstract nr B21.