Molecular profiling of lung adenosquamous carcinoma: hybrid or genuine type?

Lung adenosquamous carcinoma is a particular subtype of non-small cell lung carcinoma that is defined by the coexistence of adenocarcinoma and squamous cell carcinoma components. The aim of this study was to assess the mutational profile in each component of 16 adenosquamous carcinoma samples from a Caucasian population by a combination of next generation sequencing using the cancer hotspot panel as well as the colon and lung cancer panel and FISH. Identified mutations were confirmed by Sanger sequencing of DNA from cancer cells of each component collected by Laser Capture microdissection. Mutations typical for adenocarcinoma as well as squamous cell carcinoma were identified. Driver mutations were predominantly in the trunk suggesting a monoclonal origin of adenosquamous carcinoma. Most remarkably, EGFR mutations and mutations in the PI3K signaling pathway, which accounted for 30% and 25% of tumors respectively, were more prevalent while KRAS mutations were less prevalent than expected for a Caucasian population. Surprisingly, expression of classifier miR-205 was intermediate between that of classical adenocarcinoma and squamous cell carcinoma suggesting that adenosquamous carcinoma is a transitional stage between these tumor types. The high prevalence of therapy-relevant targets opens new options of therapeutic intervention for adenosquamous carcinoma patients.

Prophylactic antibiotics or G(M)-CSF for the prevention of infections and improvement of survival in cancer patients receiving myelotoxic chemotherapy.

BACKGROUND Febrile neutropenia (FN) and other infectious complications are some of the most serious treatment-related toxicities of chemotherapy for cancer, with a mortality rate of 2% to 21%. The two main types of prophylactic regimens are granulocyte (macrophage) colony-stimulating factors (G(M)-CSF) and antibiotics, frequently quinolones or cotrimoxazole. Current guidelines recommend the use of colony-stimulating factors when the risk of febrile neutropenia is above 20%, but they do not mention the use of antibiotics. However, both regimens have been shown to reduce the incidence of infections. Since no systematic review has compared the two regimens, a systematic review was undertaken. OBJECTIVES To compare the efficacy and safety of G(M)-CSF compared to antibiotics in cancer patients receiving myelotoxic chemotherapy. SEARCH METHODS We searched The Cochrane Library, MEDLINE, EMBASE, databases of ongoing trials, and conference proceedings of the American Society of Clinical Oncology and the American Society of Hematology (1980 to December 2015). We planned to include both full-text and abstract publications. Two review authors independently screened search results. SELECTION CRITERIA We included randomised controlled trials (RCTs) comparing prophylaxis with G(M)-CSF versus antibiotics for the prevention of infection in cancer patients of all ages receiving chemotherapy. All study arms had to receive identical chemotherapy regimes and other supportive care. We included full-text, abstracts, and unpublished data if sufficient information on study design, participant characteristics, interventions and outcomes was available. We excluded cross-over trials, quasi-randomised trials and post-hoc retrospective trials. DATA COLLECTION AND ANALYSIS Two review authors independently screened the results of the search strategies, extracted data, assessed risk of bias, and analysed data according to standard Cochrane methods. We did final interpretation together with an experienced clinician. MAIN RESULTS In this updated review, we included no new randomised controlled trials. We included two trials in the review, one with 40 breast cancer patients receiving high-dose chemotherapy and G-CSF compared to antibiotics, a second one evaluating 155 patients with small-cell lung cancer receiving GM-CSF or antibiotics.We judge the overall risk of bias as high in the G-CSF trial, as neither patients nor physicians were blinded and not all included patients were analysed as randomised (7 out of 40 patients). We considered the overall risk of bias in the GM-CSF to be moderate, because of the risk of performance bias (neither patients nor personnel were blinded), but low risk of selection and attrition bias.For the trial comparing G-CSF to antibiotics, all cause mortality was not reported. There was no evidence of a difference for infection-related mortality, with zero events in each arm. Microbiologically or clinically documented infections, severe infections, quality of life, and adverse events were not reported. There was no evidence of a difference in frequency of febrile neutropenia (risk ratio (RR) 1.22; 95% confidence interval (CI) 0.53 to 2.84). The quality of the evidence for the two reported outcomes, infection-related mortality and frequency of febrile neutropenia, was very low, due to the low number of patients evaluated (high imprecision) and the high risk of bias.There was no evidence of a difference in terms of median survival time in the trial comparing GM-CSF and antibiotics. Two-year survival times were 6% (0 to 12%) in both arms (high imprecision, low quality of evidence). There were four toxic deaths in the GM-CSF arm and three in the antibiotics arm (3.8%), without evidence of a difference (RR 1.32; 95% CI 0.30 to 5.69; P = 0.71; low quality of evidence). There were 28% grade III or IV infections in the GM-CSF arm and 18% in the antibiotics arm, without any evidence of a difference (RR 1.55; 95% CI 0.86 to 2.80; P = 0.15, low quality of evidence). There were 5 episodes out of 360 cycles of grade IV infections in the GM-CSF arm and 3 episodes out of 334 cycles in the cotrimoxazole arm (0.8%), with no evidence of a difference (RR 1.55; 95% CI 0.37 to 6.42; P = 0.55; low quality of evidence). There was no significant difference between the two arms for non-haematological toxicities like diarrhoea, stomatitis, infections, neurologic, respiratory, or cardiac adverse events. Grade III and IV thrombopenia occurred significantly more frequently in the GM-CSF arm (60.8%) compared to the antibiotics arm (28.9%); (RR 2.10; 95% CI 1.41 to 3.12; P = 0.0002; low quality of evidence). Neither infection-related mortality, incidence of febrile neutropenia, nor quality of life were reported in this trial. AUTHORS' CONCLUSIONS As we only found two small trials with 195 patients altogether, no conclusion for clinical practice is possible. More trials are necessary to assess the benefits and harms of G(M)-CSF compared to antibiotics for infection prevention in cancer patients receiving chemotherapy.

Inhibition of epidermal growth factor receptor expression by RNA interference in A549 cells

AIM: To investigate the biological features of A549 cells in which epidermal growth factor (EGF) receptors expression were suppressed by RNA interference (RNAi). METHODS: A549 cells were transfected using short small interfering RNAs (siRNAs) formulated with Lipofectamine 2000. The EGF receptor numbers were determined by Western blotting and flowcytometry. The antiproliferative effects of sequence specific double stranded RNA (dsRNA) were assessed using cell count, colony assay and scratch assay. The chemosensitivity of transfected cells to cisplatin was measured by MTT. RESULTS: Sequence specific dsRNA-EGFR down-regulated EGF receptor expression dramatically. Compared with the control group, dsRNA-EGFR reduced the cell number by 85.0 %, decreased the colonies by 63.3 %, inhibited the migration by 87.2 %, and increased the sensitivity of A549 to cisplatin by four-fold. CONCLUSION: Sequence specific dsRNA-EGFR were capable of suppressing EGF receptor expression, hence significantly inhibiting cellular proliferation and motility, and enhancing chemosensitivity of A549 cells to cisplatin. The successful application of dsRNA-EGFR for inhibition of proliferation in EGF receptor overexpressing cells can help extend the list of available therapeutic modalities in the treatment of non-small-cell lung carcinoma (NSCLC).

Implementação da técnica Silver In Situ Hibridization para avaliação do status do gene EGFR em doentes com CPNPC

Avaliação do estado do gene Epidermal Growth Factor Receptor (EGFR), por Silver In Situ Hibridization (SISH), tem-se destacado como biomarcador preditivo na resposta à terapêutica. O principal objectivo foi optimizar a etapa de recuperação por calor da metodologia automatizada SISH Dual-Colour, em carcinomas pulmonares fixados em formol durante 24 e 72 horas. A optimização levou a um aumento da preservação do contorno nuclear e da intensidade e contraste dos sinais para os dois tempos de fixação, permitindo avaliar o estado do EGFR em 83,3% dos casos em estudo. A SISH Dual-Colour é uma alternativa para avaliar o estado do EGFR.

Perioperative in-stent thrombosis after lung resection performed within 3 months of coronary stenting

BACKGROUND: Incidence of perioperative in-stent thrombosis associated with myocardial infarction in patients undergoing major lung resection within 3 months of coronary stenting. METHODS: Retrospective multi-institutional trial including all patients undergoing major lung resection (lobectomy or pneumonectomy) within 3 months of coronary stenting with non-drug-eluting stents between 1999 and 2004. RESULTS: There were 32 patients (29 men and 3 women), with age ranging from 46 to 82 years. One, two or four coronary stents were deployed in 72%, 22% and 6% of the patients, respectively. The time intervals between stenting and lung surgery were <30 days, 30-60 days and 61-90 days in 22%, 53% and 25% of the patients, respectively. All patients had dual antiplatelet therapy after stenting. Perioperative medication consisted of heparin alone or heparin plus aspirin in 34% and 66% of the patients, respectively. Perioperative in-stent thrombosis with myocardial infarction occurred in three patients (9%) with fatal outcome in one (3%). Twenty patients underwent lung resection after 4 weeks of dual antiplatelet therapy as recommended by the ACC/AHA Guideline Update; however, two out of three perioperative in-stent thrombosis occurred in this group of patients. CONCLUSIONS: Major lung resection performed within 3 months of coronary stenting may be complicated by perioperative in-stent thrombosis despite 4 weeks of dual antiplatelet therapy after stenting as recommended by the ACC/AHA Guideline Update.

Micropapillary pattern in lung adenocarcinoma: aspect on 18F-fluorodeoxyglucose positron emission tomography/computed tomography imaging.

We diagnosed a non-small cell lung carcinoma in a 49-year-old female patient with the histopathological diagnosis of stage IIIB mixed bronchioloalveolar and papillary adenocarcinoma with extensive micropapillary feature, which was not visualized on the preoperative multimodality imaging with positron emission tomography (PET) and computed tomography (CT). The micropapillary component characterized by a unique growth pattern with particular morphological features can be observed in all subtypes of lung adenocarcinoma. Micropapillary component is increasingly recognized as a distinct entity associated with higher aggressiveness. Even the most modern multimodality PET/CT imaging technology may fail to adequately visualize this important component with highly relevant prognostic implications. Thus, the pathologist needs to consciously look for a micropapillary component in the surgical specimen or in preoperative biopsies or cytology. This may have potential future treatment implications, as adjuvant or neoadjuvant chemotherapy may be of relevance, even in the early stages of the disease.

The cooperative induction of hypoxia-inducible factor-1 alpha and STAT3 during hypoxia induced an impairment of tumor susceptibility to CTL-mediated cell lysis.

Hypoxia is an essential component of tumor microenvironment. In this study, we investigated the influence of hypoxia (1% PO(2)) on CTL-mediated tumor cell lysis. We demonstrate that exposure of target tumor cells to hypoxia has an inhibitory effect on the CTL clone (Heu171)-induced autologous target cell lysis. Such inhibition correlates with hypoxia-inducible factor-1alpha (HIF-1alpha) induction but is not associated with an alteration of CTL reactivity as revealed by granzyme B polarization or morphological change. Western blot analysis indicates that although hypoxia had no effect on p53 accumulation, it induced the phosphorylation of STAT3 in tumor cells by a mechanism at least in part involving vascular endothelial growth factor secretion. We additionally show that a simultaneous nuclear translocation of HIF-1alpha and phospho-STAT3 was observed. Interestingly, gene silencing of STAT3 by small interfering RNA resulted in HIF-1alpha inhibition and a significant restoration of target cell susceptibility to CTL-induced killing under hypoxic conditions by a mechanism involving at least in part down-regulation of AKT phosphorylation. Moreover, knockdown of HIF-1alpha resulted in the restoration of target cell lysis under hypoxic conditions. This was further supported by DNA microarray analysis where STAT3 inhibition resulted in a partly reversal of the hypoxia-induced gene expression profile. The present study demonstrates that the concomitant hypoxic induction of phospho-STAT3 and HIF-1alpha are functionally linked to the alteration of non-small cell lung carcinoma target susceptibility to CTL-mediated killing. Considering the eminent functions of STAT3 and HIF-1alpha in the tumor microenvironment, their targeting may represent novel strategies for immunotherapeutic intervention.

Novos produtos naturais capazes de atuar na estabilização de microtúbulos, um importante alvo no combate ao câncer

Microtubules are involved in many aspects of cellular biology and represent an important target of anticancer chemotherapeutics. In the past five years, novel natural products such as epothilones, discodermolide, sarcodictyin, eleutherobin, and laulimalide, all of which have biological activities similar to those of paclitaxel (Taxolâ), have been discovered. Taxolâ is an important antitumor drug approved by the FDA for the treatment of ovarian, breast and non-small-cell lung carcinomas and became the first natural product described that stabilized microtubules avoiding the cellular replication. The present article reports new natural products that are able to act on the stabilization of microtubules.

Eficacia del diagnóstico prenatal no invasivo por células fetales libres en sangre materna 1990-2014. Revisión sistemática

ANTECEDENTES: El aislamiento de células fetales libres o ADN fetal en sangre materna abre una ventana de posibilidades diagnósticas no invasivas para patologías monogénicas y cromosómicas, además de permitir la identificación del sexo y del RH fetal. Actualmente existen múltiples estudios que evalúan la eficacia de estos métodos, mostrando resultados costo-efectivos y de menor riesgo que el estándar de oro. Este trabajo describe la evidencia encontrada acerca del diagnóstico prenatal no invasivo luego de realizar una revisión sistemática de la literatura. OBJETIVOS: El objetivo de este estudio fue reunir la evidencia que cumpla con los criterios de búsqueda, en el tema del diagnóstico fetal no invasivo por células fetales libres en sangre materna para determinar su utilidad diagnóstica.  MÉTODOS: Se realizó una revisión sistemática de la literatura con el fin de determinar si el diagnóstico prenatal no invasivo por células fetales libres en sangre materna es efectivo como método de diagnóstico.  RESULTADOS: Se encontraron 5,893 artículos que cumplían con los criterios de búsqueda; 67 cumplieron los criterios de inclusión: 49.3% (33/67) correspondieron a estudios de corte transversal, 38,8% (26/67) a estudios de cohortes y el 11.9% (8/67) a estudios casos y controles. Se obtuvieron resultados de sensibilidad, especificidad y tipo de prueba. CONCLUSIÓN: En la presente revisión sistemática, se evidencia como el diagnóstico prenatal no invasivo es una técnica feasible, reproducible y sensible para el diagnóstico fetal, evitando el riesgo de un diagnóstico invasivo.

Increased numbers of spontaneous SCLC metastasis in absence of NK cells after subcutaneous inoculation of different SCLC cell lines into pfp/rag2 double knock out mice

Spontaneous metastases in small cell lung cancer (SCLC) occur regularly in patients but seldom if any in conventional xenograft mouse models. To overcome this problem, SCLC cells were grafted subcutaneously onto pore forming protein and recombination activating gene 2 double knock out (pfp/rag2) mice and in severe combined immunodeficient (scid) mice. Primary tumours grew well in both mouse strains, while metastases occurred frequently in the pfp/rag2 mice and infrequently in scid mice. Hence NK cells, which are inactive in pfp/rag2 mice, play an important role in SCLC metastasis formation in xenograft models. This observation is in agreement with clinical studies, where a high NK cell number in the blood is correlated with a better prognosis of the patient.

Regulation of {\it neu\/} gene expression by the simian virus 40 large T antigen and tumor suppressors Rb and p53

The neu gene (also c-erbB-2 or HER2) encodes a 185 kilodalton protein that is frequently overexpressed in breast, ovarian and non-small cell lung cancers. Study of the regulation of neu indicates that neu gene expression can be modulated by c-myc or by the adenovirus 5 E1a gene product. This study demonstrates that the transforming protein, large T antigen, of the simian virus 40 represses neu promoter activity. Repression of neu by large T antigen is mediated through the region $-$172 to $-$79 (relative to first ATG) of the neu promoter--unlike through $-$312 to $-$172 for c-myc or E1a. This suggests a different pathway for repression of neu by large T antigen. The 10 amino acid region of large T required for binding the tumor suppressor, retinoblastoma gene product, Rb, is not necessary for repression of neu. Moreover, the tumor suppressors, Rb and p53 can independently inhibit neu promoter activity. Rb inhibits neu through a 10 base pair G-rich enhancer (GTG element) ($-$243 to $-$234) and also through regions close to transcription initiation sites ($-$172 to $-$79). Mutant Rb unable to complex large T is able to repress the region close to transcription initiation but not the GTG enhancer. Thus, Rb inhibits the two regulatory domains of the neu gene by different mechanisms. Both Rb and p53 can repress the transforming activity of activated neu in focus forming assays. These data provide evidence that tumor suppressors regulate expression of growth stimulatory genes such as neu. Therefore, one reason for the overexpression of neu that is frequently seen in breast cancer cells may be due to functional inactivation of Rb and p53 which is also a common occurrence in breast cancer cells. ^

Differential gene expression in p53-mediated apoptosis-resistant vs. apoptosis-sensitive tumor cell lines

Induction of wild-type p53 in the ECV-304 bladder carcinoma cell line by infection with a p53 recombinant adenovirus (Ad5CMV-p53) resulted in extensive apoptosis and eventual death of nearly all of the cells. As a strategy to determine the molecular events important to p53-mediated apoptosis in these transformed cells, ECV-304 cells were selected for resistance to p53 by repeated infections with Ad5CMV-p53. We compared the expression of 5,730 genes in p53-resistant (DECV) and p53-sensitive ECV-304 cells by reverse transcription–PCR, Northern blotting, and DNA microarray analysis. The expression of 480 genes differed by 2-fold or more between the two p53-infected cell lines. A number of potential targets for p53 were identified that play roles in cell cycle regulation, DNA repair, redox control, cell adhesion, apoptosis, and differentiation. Proline oxidase, a mitochondrial enzyme involved in the proline/pyrroline-5-carboxylate redox cycle, was up-regulated by p53 in ECV but not in DECV cells. Pyrroline-5-carboxylate (P5C), a proline-derived metabolite generated by proline oxidase, inhibited the proliferation and survival of ECV-304 and DECV cells and induced apoptosis in both cell lines. A recombinant proline oxidase protein tagged with a green fluorescent protein at the amino terminus localized to mitochondria and induced apoptosis in p53-null H1299 non-small cell lung carcinoma cells. The results directly implicate proline oxidase and the proline/P5C pathway in p53-induced growth suppression and apoptosis.

Closure of large intrathoracic airway defects using extrathoracic muscle flaps.

BACKGROUND: Prospective assessment of pedicled extrathoracic muscle flaps for the closure of large intrathoracic airway defects after noncircumferential resection in situations where an end-to-end reconstruction seemed risky (defects of > 4-cm length, desmoplastic reactions after previous infection or radiochemotherapy). METHODS: From 1996 to 2001, 13 intrathoracic muscle transpositions (6 latissimus dorsi and 7 serratus anterior muscle flaps) were performed to close defects of the intrathoracic airways after noncircumferential resection for tumor (n = 5), large tracheoesophageal fistula (n = 2), delayed tracheal injury (n = 1) and bronchopleural fistula (n = 5). In 2 patients, the extent of the tracheal defect required reinforcement of the reconstruction by use of a rib segment embedded into the muscle flap followed by temporary tracheal stenting. Patient follow-up was by clinical examination bronchoscopy and biopsy, pulmonary function tests, and dynamic virtual bronchoscopy by computed tomographic (CT) scan during inspiration and expiration. RESULTS: The airway defects ranged from 2 x 1 cm to 8 x 4 cm and involved up to 50% of the airway circumference. They were all successfully closed using muscle flaps with no mortality and all patients were extubated within 24 hours. Bronchoscopy revealed epithelialization of the reconstructions without dehiscence, stenosis, or recurrence of fistulas. The flow-volume loop was preserved in all patients and dynamic virtual bronchoscopy revealed no significant difference in the endoluminal cross surface areas of the airway between inspiration and expiration above (45 +/- 21 mm(2)), at the site (76 +/- 23 mm(2)) and below the reconstruction (65 +/- 40 mm(2)). CONCLUSIONS: Intrathoracic airway defects of up to 50% of the circumference may be repaired using extrathoracic muscle flaps when an end-to-end reconstruction is not feasible.