Jeffrey A. Kern

Segregation of receptor and ligand regulates activation of epithelial growth factor receptor.

Interactions between ligands and receptors are central to communication between cells and tissues. Human airway epithelia constitutively produce both a ligand, the growth factor heregulin, and its receptors—erbB2, erbB3 and erbB4 (refs 1–3). Although heregulin binding initiates cellular proliferation and differentiation, airway epithelia have a low rate of cell division. This raises the question of how ligand–receptor interactions are controlled in epithelia. Here we show that in differentiated human airway epithelia, heregulin-α is present exclusively in the apical membrane and the overlying airway surface liquid, physically separated from erbB2–4, which segregate to the basolateral membrane. This physical arrangement creates a ligand–receptor pair poised for activation whenever epithelial integrity is disrupted. Indeed, immediately following a mechanical injury, heregulin-α activates erbB2 in cells at the edge of the wound, and this process hastens restoration of epithelial integrity. Likewise, when epithelial cells are not separated into apical and basolateral membranes (‘polarized’), or when tight junctions between adjacent cells are opened, heregulin-α activates its receptor. This mechanism of ligand–receptor segregation on either side of epithelial tight junctions may be vital for rapid restoration of integrity following injury, and hence critical for survival. This model also suggests a mechanism for abnormal receptor activation in diseases with increased epithelial permeability.

Dexamethasone inhibition of interleukin 1 beta production by human monocytes. Posttranscriptional mechanisms.

Dexamethasone is known to have an inhibitory effect on IL-1 production. To determine the mechanism(s) of this inhibition, adherent human blood monocytes were stimulated with Escherichia coli lipopolysaccharide (LPS) (10 micrograms/ml) in the presence of dexamethasone. Nuclear transcription run-off assays showed that LPS induced IL-1 beta gene transcription two- to fourfold and that this induction was unaffected by dexamethasone exposure (10(-5) M). The lack of dexamethasones transcriptional effects was further supported by the absence of any significant change in IL-1 beta mRNA accumulation between LPS-stimulated monocytes exposed or unexposed to dexamethasone, as determined by Northern blot analysis. Posttranscriptionally, dexamethasone was found to have multiple effects: slight prolongation of IL-1 beta mRNA half-life, moderate inhibition of translation of the IL-1 beta precursor, and profound inhibition of the release of IL-1 beta into the extracellular fluid. The data indicate that IL-1 beta is first translated as the 33,000-D pro-IL-1 beta protein, the predominant intracellular form, and the processed to a 17,500-D IL-1 beta protein before or during extracellular transport. The major inhibitory effects of dexamethasone appear to be directed at the translational and posttranslational steps involved in these events.

Proliferative Response of Bronchoalveolar Lymphocytes to Beryllium: A Test for Chronic Beryllium Disease

STUDY OBJECTIVE to ascertain whether measuring the proliferative response of bronchoalveolar lymphocytes to beryllium salts is useful for diagnosing chronic beryllium disease. DESIGN prospective case series compared to normal volunteers and patients with sarcoidosis. SETTING university referral center. PATIENTS twenty-three consecutive beryllium workers were evaluated. Fourteen had chronic beryllium disease diagnosed on the basis of histologic evidence of a progressive pulmonary granulomatosis. Four had biopsy evidence of non-beryllium disease. Three had probable chronic beryllium disease but did not have lung biopsies. Two did not have biopsies and had basilar fibrosis on chest roentgenogram suggestive of non-beryllium lung disease. These patients were compared with 6 normal volunteers and 16 patients with sarcoidosis. MEASUREMENTS AND MAIN RESULTS bronchoalveolar lavage was done and the proliferative response of the lung cells to two beryllium salts was tested. A positive proliferative test was defined as a stimulation index of more than five on two determinations. The sensitivity of this test was 100% in the 14 patients with definite chronic beryllium disease. The specificity of the test was also 100% among the normal volunteers and the 16 patients with sarcoidosis. The test was positive in none of the four patients with biopsy evidence of non-beryllium disease, none out of two patients with lower lobe fibrosis suggestive of non-beryllium disease, and all of three patients with probable chronic beryllium lung disease. CONCLUSIONS the proliferative response of bronchoalveolar lymphocytes to beryllium appears to be a useful test for chronic beryllium disease.

C-erbB-2 expression and codon 12 K-ras mutations both predict shortened survival for patients with pulmonary adenocarcinomas.

We evaluated the prognostic significance of p185c-erbB-2 expression and ras gene mutations in all patients diagnosed with a pulmonary adenocarcinoma between 1982 and 1985 at the University of Iowa. p185c-erbB-2 expression was detected in 15 cases (34%). A ras gene mutation was found in 16 cases (36%) and all were in codon-12 of K-ras. No N-ras mutations were identified. Both p185c-erbB-2 expression and a K-ras mutation were found only in codon-12 and present in six cases (14%). By univariate analysis p185c-erbB-2 expression was associated with shortened survival (P = 0.02) while the presence of a K-ras mutation was not (P = 0.16). Multivariate analysis by the Cox proportional hazards model, controlling for patient age and tumor stage, also continued to identify p185c-erbB-2 expression as an independent unfavorable prognostic factor (P = 0.01). In this model a K-ras mutation also approached significance as a poor prognostic indicator (P = 0.06). The impact of both p185c-erbB-2 expression and a K-ras mutation on survival was additive and highly significant (P = 0.004). This additive nature suggests that together these two markers identify a high-risk population of lung adenocarcinoma patients that may benefit from aggressive therapy.

Dual MET–EGFR combinatorial inhibition against T790M-EGFR-mediated erlotinib-resistant lung cancer

Despite clinical approval of erlotinib, most advanced lung cancer patients are primary non-responders. Initial responders invariably develop secondary resistance, which can be accounted for by T790M-EGFR mutation in half of the relapses. We show that MET is highly expressed in lung cancer, often concomitantly with epidermal growth factor receptor (EGFR), including H1975 cell line. The erlotinib-resistant lung cancer cell line H1975, which expresses L858R/T790M-EGFR in-cis, was used to test for the effect of MET inhibition using the small molecule inhibitor SU11274. H1975 cells express wild-type MET, without genomic amplification (CNV=1.1). At 2 μM, SU11274 had significant in vitro pro-apoptotic effect in H1975 cells, 3.9-fold (P=0.0015) higher than erlotinib, but had no effect on the MET and EGFR-negative H520 cells. In vivo, SU11274 also induced significant tumour cytoreduction in H1975 murine xenografts in our bioluminescence molecular imaging assay. Using small-animal microPET/MRI, SU11274 treatment was found to induce an early tumour metabolic response in H1975 tumour xenografts. MET and EGFR pathways were found to exhibit collaborative signalling with receptor cross-activation, which had different patterns between wild type (A549) and L858R/T790M-EGFR (H1975). SU11274 plus erlotinib/CL-387,785 potentiated MET inhibition of downstream cell proliferative survival signalling. Knockdown studies in H1975 cells using siRNA against MET alone, EGFR alone, or both, confirmed the enhanced downstream inhibition with dual MET–EGFR signal path inhibition. Finally, in our time-lapse video-microscopy and in vivo multimodal molecular imaging studies, dual SU11274-erlotinib concurrent treatment effectively inhibited H1975 cells with enhanced abrogation of cytoskeletal functions and complete regression of the xenograft growth. Together, our results suggest that MET-based targeted inhibition using small-molecule MET inhibitor can be a potential treatment strategy for T790M-EGFR-mediated erlotinib-resistant non-small-cell lung cancer. Furthermore, optimised inhibition may be further achieved with MET inhibition in combination with erlotinib or an irreversible EGFR-TKI.

Autonomous decision-making: a data mining approach

The researchers and practitioners of today create models, algorithms, functions, and other constructs defined in abstract spaces. The research of the future will likely be data driven. Symbolic and numeric data that are becoming available in large volumes will define the need for new data analysis techniques and tools. Data mining is an emerging area of computational intelligence that offers new theories, techniques, and tools for analysis of large data sets. In this paper, a novel approach for autonomous decision-making is developed based on the rough set theory of data mining. The approach has been tested on a medical data set for patients with lung abnormalities referred to as solitary pulmonary nodules (SPNs). The two independent algorithms developed in this paper either generate an accurate diagnosis or make no decision. The methodology discussed in the paper depart from the developments in data mining as well as current medical literature, thus creating a variable approach for autonomous decision-making.

Multicenter clinical trial of ultrasmall superparamagnetic iron oxide in the evaluation of mediastinal lymph nodes in patients with primary lung carcinoma

The purpose of this study was to evaluate the clinical efficacy of ultrasmall superparamagnetic iron oxide particles as a magnetic resonance (MR) contrast agent in differentiating metastatic from benign lymph nodes. Eighteen patients with primary lung malignancy and suspected regional lymph node metastases underwent MR imaging before and after Combidex® infusion in a multi‐institutional study. All MR sequences were interpreted by one or more board‐certified radiologists experienced in imaging thoracic malignancy. Each patient was evaluated for the number and location of lymph nodes, homogeneity of nodal signal, and possible change of MR signal post contrast. All patients underwent resection or sampling of the MR‐identified lymph node(s) 1–35 day(s) post contrast MR imaging. In all, 27 lymph nodes or nodal groups were available for histopathologic correlation. Combidex had a sensitivity of 92% and a specificity of 80% in identifying pathologically confirmed metastatic mediastinal lymph nodes. Based on our preliminary data, Combidex MR imaging may provide additional functional information useful in the staging of mediastinal lymph nodes. J. Magn. Reson. Imaging 1999;10:468–473.

FDG-PET imaging and the diagnosis of non-small cell lung cancer in a region of high histoplasmosis prevalence

STUDY OBJECTIVE Determine the sensitivity and specificity of [F-18]-fluorine-2-deoxy-D-glucose positron emission tomography (FDG-PET) in differentiating non-small cell lung cancer (NSCLC) from benign solitary pulmonary nodules (SPNs) in a region with a high endemic rate of histoplamosis. DESIGN Prospective, clinical study. SETTING University, tertiary referral hospital in the upper Mississippi River valley. PATIENTS Ninety patients with SPNs. INTERVENTIONS Independent interpretation of FDG-PET imaging, computed tomography and pathologic evaluation of the SPNs. MEASUREMENTS AND RESULTS To detect malignant SPNs, FDG-PET imaging had a sensitivity of 93%, a specificity of 40%, a positive predictive value (PPV) of 88% and a negative predictive value (NPV) of 55%. CONCLUSIONS In a region with a high prevalence of pulmonary fungal infection, FDG-PET is sensitive but has a low specificity and NPV for identifying NSCLC. In our study cohort, FDG-PET does not appear to reduce the need for SPN biopsies.

The interactive effect of Ras, HER2, P53 and Bcl-2 expression in predicting the survival of non-small cell lung cancer patients

In patients with non-small cell lung cancer (NSCLC), tumor expression of P21-Ras, HER2, P53, and Bcl-2 has been reported as independent predictors of prognosis. However, the prognostic information carried by these proteins has usually been determined separately, and their potential interaction has not been taken into account. We conducted immunostaining for P21-Ras, HER2, P53 and Bcl-2 on 238 cases of NSCLC in a Korean population with 203 squamous cell carcinomas, and 35 adenocarcinomas. P21-Ras, HER2, P53 or Bcl-2 was expressed at high levels in 54.6, 42.0, 18.1 and 71.8% of the NSCLC studied, respectively. A total of 59 tumors (24.8%) expressed only one protein, while 70 (29.4%) expressed two, 59 (24.8) expressed three, and 17 tumors (7.1%) expressed all four proteins. Univariate analysis testing the association of marker expression with survival found Bcl-2 expression to be significantly associated with a poor prognosis, as well as the co-expression of Bcl-2 + HER2, Bcl-2 + HER2 + P53, and Bcl-2 + HER2 + P53 + P21-ras with an increasing hazard ratio. By multivariate analysis controlling for age, tumor stage and tumor type, only the combination of Bcl-2 + HER2 expression was an independent marker of poor prognosis (hazard ratio = 1.91, P = 0.003). Thus, a prospective analysis of the co-expression of Bcl-2 + HER2 in NSCLC patients may identify patients with a poor prognosis who may benefit from more aggressive therapy.

Role of Smad2/3 and p38 MAP kinase in TGF-β1-induced epithelial-mesenchymal transition of pulmonary epithelial cells.

Idiopathic pulmonary fibrosis is characterized by myofibroblast accumulation, extracellular matrix (ECM) remodeling, and excessive collagen deposition. ECM‐producing myofibroblasts may originate from epithelial cells through epithelial to mesenchymal transition (EMT). TGF‐β1 is an inducer of EMT in pulmonary epithelial cells in vitro and in vivo, though the mechanisms are unclear. We hypothesized that TGF‐β1 induced EMT through Smad‐dependent and ‐independent processes. To test this hypothesis, we studied the roles and mechanisms of TGF‐β1‐induced Smad and p38 mitogen‐activated protein kinase (MAPK) signaling in EMT‐related changes in pulmonary epithelial cells. Exposure of pulmonary epithelial 1HAEo− cells to TGF‐β1 resulted in morphological and molecular changes of EMT over a 96‐h period; loss of cell–cell contact, cell elongation, down‐regulation of E‐cadherin, up‐regulation of fibronectin, and up‐regulation of collagen I. Both Smad2/3 and p38 MAPK signaling pathways were activated by TGF‐β1. However, neither Smad2/3 nor p38 MAPK were required for the down‐regulation of E‐cadherin, yet p38 MAPK was associated with fibronectin up‐regulation. Both Smad2/3 and p38 MAPK had a role in regulation of TGF‐β1‐induced collagen expression. Furthermore, these data demonstrate that Smads and p38 MAPK differentially regulate EMT‐related changes in pulmonary epithelial cells. J. Cell. Physiol. 226: 1248–1254, 2011.