Clinical and morphological features of large-cell neuroendocrine carcinomas and small-cell lung carcinomas expressing the DLL3 and ASCL1 oncoproteins.

Intratumoral similarities and differences between large-cell neuroendocrine carcinomas (LCNECs) and small-cell lung carcinomas (SCLCs) are determined partially by the Notch signaling pathway, which controls the switch from neuroendocrine to slight/non-neuroendocrine cell fate. LCNECs are divided into two subgroups according to genomic alterations: type I LCNECs exhibit a neuroendocrine profile characterized by achaete-scute homolog 1 (ASCL1)high/delta-like protein 3 (DLL3)high/NOTCHlow and type II LCNECs show the pattern ASCL1low/DLL3low/NOTCHhigh. Here, we used immunohistochemistry, transmission electron microscopy, and digital analysis to examine the role of the Notch ligand DLL3 as an immunomarker of the neuroendocrine state and ASCL1 as a regulator of cell-cell interactions in SCLCs and LCNECs. High DLL3 and ASCL1 expression was associated with atypical submicroscopic characteristics involving nuclear size, chromatin arrangement, Golgi apparatus, and endoplasmic reticulum, and was characteristic of type I LCNECs with similarity to SCLCs, whereas low DLL3 and ASCL1 expression was found in both SCLCs and type II LCNECs. In patients diagnosed at an early stage who did not have metastasis and who underwent chemotherapy, DLL3high and ASCL1high SCLCs and type I LCNECs were associated with a better prognosis and a lower risk of death. The present findings suggested that DLL3/ASCL1 are potential therapeutic targets and prognostic indicators in patients with SCLCs or LCNECs.

A semi-automated microscopic image analysis method for scoring Ki-67 nuclear immunostaining.

Nuclear proliferation marker MIB-1 (Ki-67) immunohistochemistry (IHC) is used to examine tumor cell proliferation. However, the diagnostic or prognostic value of the Ki-67 nuclear staining intensity and location, defined as nuclear gradient (NG), has not been assessed. This study examined the potential association between Ki-67 NG and cell cycle phases and its effect on the prognosis of pulmonary typical carcinoid (PTC) tumors. We propose a method for classifying the NG of Ki-67 during the cell cycle and compare the results between PTC, pulmonary adenocarcinoma (PAD), and breast ductal carcinoma (BDC). A literature review and objective analysis of IHC-stained paraffin sections were used to determine the Ki-67 labeling index and composed a stratification of the NG into NG1, NG2, and NG3/4 categories. A semi-automated image analysis protocol was established to determine the Ki-67 NG in PTC, PAD, and BDC. High intraobserver consistency and moderate interobserver agreement were achieved in the determination of Ki-67 NG in tumor specimens. NG1 and NG2 were lower in PTC than in PAD and BDC. Cox multivariate analysis of PTC after adjusting for age and number of metastatic lymph nodes showed that Ki-67 NG1 and NG2 significantly predicted clinical outcomes. The semi-automated method for quantification of Ki-67 nuclear immunostaining proposed in this study could become a valuable diagnostic and prognostic tool in PTC.

Concomitant TP53 mutation in early-stage resected EGFR-mutated non-small cell lung cancer: a narrative approach in a genetically admixed Brazilian cohort.

TP53 mutations are frequent in non-small cell lung cancer (NSCLC) and have been associated with poor outcome. The prognostic and predictive relevance of EGFR/TP53 co-mutations in NSCLC is controversial. We analyzed lung tissue specimens from 70 patients with NSCLC using next-generation sequencing to determine EGFR and TP53 status and the association between these status with baseline patient and tumor characteristics, adjuvant treatments, relapse, and progression-free (PFS) and overall survival (OS) after surgical resection. We found the EGFR mutation in 32.9% of patients (20% classical mutations and 12.9% uncommon mutations). TP53 missense mutations occurred in 25.7% and TP53/EGFR co-mutations occurred in 43.5% of patients. Stage after surgical resection was significantly associated with OS (P=0.028). We identified an association between progression-free survival and poor outcome in patients with distant metastases (P=0.007). We found a marginally significant difference in OS between genders (P=0.057) and between mutant and wild type TP53 (P=0.079). In univariate analysis, distant metastases (P=0.027), pathological stage (IIIA-IIIB vs I-II; P=0.028), and TP53 status (borderline significance between wild type and mutant; P=0.079) influenced OS. In multivariable analysis, a significant model for high risk of death and poor OS (P=0.029) selected patients in stage IIIA-IIIB, with relapse and distant metastases, non-responsive to platin-based chemotherapy and erlotinib, with tumors harboring EGFR uncommon mutations, with TP53 mutant, and with EGFR/TP53 co-mutations. Our study suggested that TP53 mutation tends to confer poor survival and a potentially negative predictive effect associated with a non-response to platinum-based chemotherapy and erlotinib in early-stage resected EGFR-mutated NSCLC.

Clinical outcome of Brazilian patients with non-small cell lung cancer in early stage harboring rare mutations in epidermal growth factor receptor.

The common epidermal growth factor receptor (EGFR) mutations, such as the L858R point mutation in exon 21 and the in-frame deletional mutation in exon 19, have been definitively associated with response to EGFR-tyrosine kinase inhibitors (EGFR-TKI). However, the clinical outcome and response to treatment for many other rarer mutations are still unclear. In this study, we report the results of Brazilian patients in stage IB-IIIA non-small cell lung cancer (NSCLC) following complete resection with minimal residual disease and EGFR mutations treated with adjuvant chemotherapy and/or EGFR-TKIs. The frequency of EGFR mutations was investigated in 70 cases of early stage NSCLC. Mutations in exons 18 and 20, uncommon mutations in exons 19 and 21, as well as in exons 3, 7, 14, 16, 22, 27, and 28, and/or the presence of different mutations in a single tumor (complex mutations) are considered rare. EGFR mutations were detected in 23 tumors (32.9%). Fourteen cases carried rare mutations and were treated with platinum-based chemotherapy and two cases were treated with erlotinib. The clinical outcome is described case by case with references to the literature. Notably, we found two rare EGFR mutations and one of them with an unknown response to chemotherapy and/or EGFR-TKIs. We have provided complementary information concerning the clinical outcome and treatment of patients with early stage NSCLC for several rare EGFR mutations not previously or only rarely reported. Description of cases harboring rare mutations can support the decision-making process in this subset of patients.

A semi-automated microscopic image analysis method for scoring Ki-67 nuclear immunostaining

Nuclear proliferation marker MIB-1 (Ki-67) immunohistochemistry (IHC) is used to examine tumor cell proliferation. However, the diagnostic or prognostic value of the Ki-67 nuclear staining intensity and location, defined as nuclear gradient (NG), has not been assessed. This study examined the potential association between Ki-67 NG and cell cycle phases and its effect on the prognosis of pulmonary typical carcinoid (PTC) tumors. We propose a method for classifying the NG of Ki-67 during the cell cycle and compare the results between PTC, pulmonary adenocarcinoma (PAD), and breast ductal carcinoma (BDC). A literature review and objective analysis of IHC-stained paraffin sections were used to determine the Ki-67 labeling index and composed a stratification of the NG into NG1, NG2, and NG3/4 categories. A semi-automated image analysis protocol was established to determine the Ki-67 NG in PTC, PAD, and BDC. High intraobserver consistency and moderate interobserver agreement were achieved in the determination of Ki-67 NG in tumor specimens. NG1 and NG2 were lower in PTC than in PAD and BDC. Cox multivariate analysis of PTC after adjusting for age and number of metastatic lymph nodes showed that Ki-67 NG1 and NG2 significantly predicted clinical outcomes. The semi-automated method for quantification of Ki-67 nuclear immunostaining proposed in this study could become a valuable diagnostic and prognostic tool in PTC.

Clinical and morphological features of large-cell neuroendocrine carcinomas and small-cell lung carcinomas expressing the DLL3 and ASCL1 oncoproteins

Abstract Intratumoral similarities and differences between large-cell neuroendocrine carcinomas (LCNECs) and small-cell lung carcinomas (SCLCs) are determined partially by the Notch signaling pathway, which controls the switch from neuroendocrine to slight/non-neuroendocrine cell fate. LCNECs are divided into two subgroups according to genomic alterations: type I LCNECs exhibit a neuroendocrine profile characterized by achaete‐scute homolog 1 (ASCL1)high/delta-like protein 3 (DLL3)high/NOTCHlow and type II LCNECs show the pattern ASCL1low/DLL3low/NOTCHhigh. Here, we used immunohistochemistry, transmission electron microscopy, and digital analysis to examine the role of the Notch ligand DLL3 as an immunomarker of the neuroendocrine state and ASCL1 as a regulator of cell-cell interactions in SCLCs and LCNECs. High DLL3 and ASCL1 expression was associated with atypical submicroscopic characteristics involving nuclear size, chromatin arrangement, Golgi apparatus, and endoplasmic reticulum, and was characteristic of type I LCNECs with similarity to SCLCs, whereas low DLL3 and ASCL1 expression was found in both SCLCs and type II LCNECs. In patients diagnosed at an early stage who did not have metastasis and who underwent chemotherapy, DLL3high and ASCL1high SCLCs and type I LCNECs were associated with a better prognosis and a lower risk of death. The present findings suggested that DLL3/ASCL1 are potential therapeutic targets and prognostic indicators in patients with SCLCs or LCNECs.

Concomitant TP53 mutation in early-stage resected EGFR-mutated non-small cell lung cancer: a narrative approach in a genetically admixed Brazilian cohort

TP53 mutations are frequent in non-small cell lung cancer (NSCLC) and have been associated with poor outcome. The prognostic and predictive relevance of EGFR/TP53 co-mutations in NSCLC is controversial. We analyzed lung tissue specimens from 70 patients with NSCLC using next-generation sequencing to determine EGFR and TP53 status and the association between these status with baseline patient and tumor characteristics, adjuvant treatments, relapse, and progression-free (PFS) and overall survival (OS) after surgical resection. We found the EGFR mutation in 32.9% of patients (20% classical mutations and 12.9% uncommon mutations). TP53 missense mutations occurred in 25.7% and TP53/EGFR co-mutations occurred in 43.5% of patients. Stage after surgical resection was significantly associated with OS (P=0.028). We identified an association between progression-free survival and poor outcome in patients with distant metastases (P=0.007). We found a marginally significant difference in OS between genders (P=0.057) and between mutant and wild type TP53 (P=0.079). In univariate analysis, distant metastases (P=0.027), pathological stage (IIIA-IIIB vs I-II; P=0.028), and TP53 status (borderline significance between wild type and mutant; P=0.079) influenced OS. In multivariable analysis, a significant model for high risk of death and poor OS (P=0.029) selected patients in stage IIIA-IIIB, with relapse and distant metastases, non-responsive to platin-based chemotherapy and erlotinib, with tumors harboring EGFR uncommon mutations, with TP53 mutant, and with EGFR/TP53 co-mutations. Our study suggested that TP53 mutation tends to confer poor survival and a potentially negative predictive effect associated with a non-response to platinum-based chemotherapy and erlotinib in early-stage resected EGFR-mutated NSCLC.

Monitoring non-small cell lung cancer progression and treatment response through hyaluronic acid in sputum.

We evaluated whether hyaluronan (HA) levels in the sputum could be used as a noninvasive tool to predict progressive disease and treatment response, as detected in a computed tomography scan in non-small cell lung cancer (NSCLC) patients. Sputum samples were collected from 84 patients with histological confirmation of NSCLC, 33 of which were in early-stage and 51 in advanced-stage disease. Patients received systemic chemotherapy (CT) after surgery (n=36), combined CT and immunotherapy (IO) (n=15), or targeted therapy for driver mutation and disease relapse (N=4). The primary end-point was to compare sputum HA levels in two different concentrations of hypertonic saline solution with overall survival (OS) and the secondary and exploratory end-points were radiologic responses to treatment and patient outcome. Higher concentrations of HA in the sputum were significantly associated to factors related to tumor stage, phenotype, response to treatment, and outcome. In the early stage, patients with lower sputum HA levels before treatment achieved a complete tumor response after systemic CT with better progression-free survival (PFS) than those with high HA levels. We also examined the importance of the sputum HA concentration and tumor response in the 51 patients who developed metastatic disease and received CT+IO. Patients with low levels of sputum HA showed a complete tumor response in the computed tomography scan and stable disease after CT+IO treatment, as well as a better PFS than those receiving CT alone. HA levels in sputum of NSCLC patients may serve as a candidate biomarker to detect progressive disease and monitor treatment response in computed tomography scans.

Monitoring non-small cell lung cancer progression and treatment response through hyaluronic acid in sputum

We evaluated whether hyaluronan (HA) levels in the sputum could be used as a noninvasive tool to predict progressive disease and treatment response, as detected in a computed tomography scan in non-small cell lung cancer (NSCLC) patients. Sputum samples were collected from 84 patients with histological confirmation of NSCLC, 33 of which were in early-stage and 51 in advanced-stage disease. Patients received systemic chemotherapy (CT) after surgery (n=36), combined CT and immunotherapy (IO) (n=15), or targeted therapy for driver mutation and disease relapse (N=4). The primary end-point was to compare sputum HA levels in two different concentrations of hypertonic saline solution with overall survival (OS) and the secondary and exploratory end-points were radiologic responses to treatment and patient outcome. Higher concentrations of HA in the sputum were significantly associated to factors related to tumor stage, phenotype, response to treatment, and outcome. In the early stage, patients with lower sputum HA levels before treatment achieved a complete tumor response after systemic CT with better progression-free survival (PFS) than those with high HA levels. We also examined the importance of the sputum HA concentration and tumor response in the 51 patients who developed metastatic disease and received CT+IO. Patients with low levels of sputum HA showed a complete tumor response in the computed tomography scan and stable disease after CT+IO treatment, as well as a better PFS than those receiving CT alone. HA levels in sputum of NSCLC patients may serve as a candidate biomarker to detect progressive disease and monitor treatment response in computed tomography scans.

Clinical outcome of Brazilian patients with non-small cell lung cancer in early stage harboring rare mutations in epidermal growth factor receptor

The common epidermal growth factor receptor (EGFR) mutations, such as the L858R point mutation in exon 21 and the in-frame deletional mutation in exon 19, have been definitively associated with response to EGFR-tyrosine kinase inhibitors (EGFR-TKI). However, the clinical outcome and response to treatment for many other rarer mutations are still unclear. In this study, we report the results of Brazilian patients in stage IB-IIIA non-small cell lung cancer (NSCLC) following complete resection with minimal residual disease and EGFR mutations treated with adjuvant chemotherapy and/or EGFR-TKIs. The frequency of EGFR mutations was investigated in 70 cases of early stage NSCLC. Mutations in exons 18 and 20, uncommon mutations in exons 19 and 21, as well as in exons 3, 7, 14, 16, 22, 27, and 28, and/or the presence of different mutations in a single tumor (complex mutations) are considered rare. EGFR mutations were detected in 23 tumors (32.9%). Fourteen cases carried rare mutations and were treated with platinum-based chemotherapy and two cases were treated with erlotinib. The clinical outcome is described case by case with references to the literature. Notably, we found two rare EGFR mutations and one of them with an unknown response to chemotherapy and/or EGFR-TKIs. We have provided complementary information concerning the clinical outcome and treatment of patients with early stage NSCLC for several rare EGFR mutations not previously or only rarely reported. Description of cases harboring rare mutations can support the decision-making process in this subset of patients.

In vivo evidence of angiogenesis inhibition by ß2-glycoprotein I subfractions in the chorioallantoic membrane of chicken embryos.

The vascular network expansion and functioning are important factors affecting normal intra-uterine fetal development. This study addressed the previously reported antiangiogenic potential of beta-2-glycoprotein I (ß2GPI) in vivo in the chick embryo model of angiogenesis. The effects of two naturally occurring ß2GPI forms on the development of the chorioallantoic membrane (CAM) vessels and the chicken embryo were investigated. ß2GPI monomers and dimers were obtained by fractioned purification and characterized using SDS-PAGE, immunoblot, and ELISA. The egg exposure was performed by injection of small volumes of 2.5 µg/mL solutions of the ß2GPI subfractions. Angiogenesis was evaluated through quantitative measurements of vascular architecture parameters in the captured CAM images, using computational analysis of texture contrasts and computer vision techniques. Quantitative information was assigned to the CAM vasculature modifications. In vivo, the ß2GPI dimer completely halted the formation of CAM vessels and led to embryo death after 48 h of exposure. The ß2GPI monomer allowed the embryo to develop up to the 10th day, despite early changes of CAM vessels. The impaired normal vessel growth proceeded as a self-limited effect. The ß2GPI monomer-exposed eggs showed reduced vascularization on the 6th day of incubation, but embryos were viable on the 10th day of incubation, with ingurgitated CAM vessels implying sequelae of the angiogenesis inhibition. Both subfractions impaired CAM vasculature development. The ß2GPI dimer proved to be largely more harmful than the ß2GPI monomer. ß2GPI modification by cleavage or dimerization may play a role in angiogenesis control in vivo.

In vivo evidence of angiogenesis inhibition by ß2-glycoprotein I subfractions in the chorioallantoic membrane of chicken embryos

The vascular network expansion and functioning are important factors affecting normal intra-uterine fetal development. This study addressed the previously reported antiangiogenic potential of beta-2-glycoprotein I (β2GPI) in vivo in the chick embryo model of angiogenesis. The effects of two naturally occurring β2GPI forms on the development of the chorioallantoic membrane (CAM) vessels and the chicken embryo were investigated. β2GPI monomers and dimers were obtained by fractioned purification and characterized using SDS-PAGE, immunoblot, and ELISA. The egg exposure was performed by injection of small volumes of 2.5 µg/mL solutions of the β2GPI subfractions. Angiogenesis was evaluated through quantitative measurements of vascular architecture parameters in the captured CAM images, using computational analysis of texture contrasts and computer vision techniques. Quantitative information was assigned to the CAM vasculature modifications. In vivo, the β2GPI dimer completely halted the formation of CAM vessels and led to embryo death after 48 h of exposure. The β2GPI monomer allowed the embryo to develop up to the 10th day, despite early changes of CAM vessels. The impaired normal vessel growth proceeded as a self-limited effect. The β2GPI monomer-exposed eggs showed reduced vascularization on the 6th day of incubation, but embryos were viable on the 10th day of incubation, with ingurgitated CAM vessels implying sequelae of the angiogenesis inhibition. Both subfractions impaired CAM vasculature development. The β2GPI dimer proved to be largely more harmful than the β2GPI monomer. β2GPI modification by cleavage or dimerization may play a role in angiogenesis control in vivo.

Detection of sputum cofilin-1 as indicator of malignancy.

Cofilin-1 (CFL1), a small protein of 18 kDa, has been studied as a biomarker due to its involvement in tumor cell migration and invasion. Our aim was to evaluate CFL1 as an indicator of malignancy and aggressiveness in sputum samples. CFL1 was analyzed by ELISA immunoassay in the sputum of 73 lung cancer patients, 13 cancer-free patients, and 6 healthy volunteers. Statistical analyses included ANOVA, ROC curves, Spearman correlation, and logistic regression. Sputum CFL1 levels were increased in cancer patients compared to cancer-free patients and volunteers (P<0.05). High expression of sputum CFL1 was correlated to T4 stage (P=0.01) and N stage (P=0.03), tobacco history (P=0.01), and squamous cell carcinoma histologic type (P=0.04). The accuracy of sputum CFL1 in discriminating cancer patients from cancer-free patients and healthy volunteers were 0.78 and 0.69, respectively. CFL1 at a cut-off value of 415.25 pg/mL showed sensitivity/specificity of 0.80/0.70 in differentiating between healthy volunteers and cancer patients. Sputum CFL1 was also able to identify cancer-free patients from patients with lung cancer. The AUC was 0.70 and, at a cut-off point ≥662.63 pg/mL, we obtained 60% sensitivity and 54% specificity. Logistic regression analysis controlled for tobacco history, histologic types, and N stage showed that cancer cell-associated CFL1 was an independent predictor of death. Smoker patients with squamous cell carcinoma, lymph node metastasis and sputum CFL1>1.475 pg/mL showed augmented chance of death, suggesting lung cancer aggressiveness. CFL1 presented diagnostic value in detecting lung cancer and was associated to tumor aggressiveness.

Detection of sputum cofilin-1 as indicator of malignancy

Cofilin-1 (CFL1), a small protein of 18 kDa, has been studied as a biomarker due to its involvement in tumor cell migration and invasion. Our aim was to evaluate CFL1 as an indicator of malignancy and aggressiveness in sputum samples. CFL1 was analyzed by ELISA immunoassay in the sputum of 73 lung cancer patients, 13 cancer-free patients, and 6 healthy volunteers. Statistical analyses included ANOVA, ROC curves, Spearman correlation, and logistic regression. Sputum CFL1 levels were increased in cancer patients compared to cancer-free patients and volunteers (P<0.05). High expression of sputum CFL1 was correlated to T4 stage (P=0.01) and N stage (P=0.03), tobacco history (P=0.01), and squamous cell carcinoma histologic type (P=0.04). The accuracy of sputum CFL1 in discriminating cancer patients from cancer-free patients and healthy volunteers were 0.78 and 0.69, respectively. CFL1 at a cut-off value of 415.25 pg/mL showed sensitivity/specificity of 0.80/0.70 in differentiating between healthy volunteers and cancer patients. Sputum CFL1 was also able to identify cancer-free patients from patients with lung cancer. The AUC was 0.70 and, at a cut-off point ≥662.63 pg/mL, we obtained 60% sensitivity and 54% specificity. Logistic regression analysis controlled for tobacco history, histologic types, and N stage showed that cancer cell-associated CFL1 was an independent predictor of death. Smoker patients with squamous cell carcinoma, lymph node metastasis and sputum CFL1>1.475 pg/mL showed augmented chance of death, suggesting lung cancer aggressiveness. CFL1 presented diagnostic value in detecting lung cancer and was associated to tumor aggressiveness.

Comparison of different degrees of variability in tidal volume to prevent deterioration of respiratory system elastance in experimental acute lung inflammation.

BACKGROUND: Variable ventilation improves respiratory function, but it is not known whether the amount of variability in tidal volume (VT) can be reduced in recruited lungs without a deterioration of respiratory system elastance. METHODS: Acute lung inflammation was induced by intratracheal instillation of lipopolysaccharide in 35 Wistar rats. Twenty-eight animals were anaesthetized and ventilated in volume-controlled mode. Lungs were recruited by random variation of VT (mean 6 ml kg(-1), coefficient of variation 30%, normal distribution) for 30 min. Animals were randomly assigned to different amounts of VT variability (n=7 for 90 min per group): 30, 15, 7.5, or 0%. Lung function, diffuse alveolar damage, and gene expression of biological markers associated with cell mechanical stress, inflammation, and fibrogenesis were assessed. Seven animals were not ventilated and served as controls for post-mortem analyses. RESULTS: A VT variability of 30%, but not 15, 7.5, or 0%, prevented deterioration of respiratory system elastance [Mean (SD) -7.5 (8.7%), P<0.05; 21.1 (9.6%), P<0.05; 43.3 (25.9), P<0.05; and 41.2 (16.4), P<0.05, respectively]. Diffuse alveolar damage was lower with a VT variability of 30% than with 0% and without ventilation, because of reduced oedema and haemorrhage. A VT variability of 30, 15, or 7.5% reduced the gene expression of amphiregulin, cytokine-induced neutrophil chemoattractant-1, and tumour necrosis factor α compared with a VT variability of 0%. CONCLUSIONS: In this model of acute lung inflammation, a VT variability of 30%, compared with 15 and 7.5%, was necessary to avoid deterioration of respiratory system elastance and was not associated with lung histological damage.

Hyaluronidases and hyaluronan synthases expression is inversely correlated with malignancy in lung/bronchial pre-neoplastic and neoplastic lesions, affecting prognosis

We collected a series of 136 lung/bronchial and 56 matched lung parenchyma tissue samples from patients who underwent lung/bronchial biopsies and presented invasive carcinoma after lung surgery. The lung/bronchial samples included basal cell hyperplasia, squamous metaplasia, moderate dysplasia, adenomatous hyperplasia, severe dysplasia, squamous cell carcinoma and adenocarcinoma. Matched lung parenchyma tissue samples included 25 squamous cell carcinomas and 31 adenocarcinomas. Immunohistochemistry was performed to analyze for the distribution of hyaluronidase (Hyal)-1 and −3, and hyaluronan synthases (HAS)-1, −2, and −3. Hyal-1 showed significantly higher expression in basal cell hyperplasia than in moderate dysplasia (P=0.01), atypical adenomatous hyperplasia (P=0.0001), or severe dysplasia (P=0.03). Lower expression of Hyal-3 was found in atypical adenomatous hyperplasia than in basal cell hyperplasia (P=0.01) or moderate dysplasia (P=0.02). HAS-2 was significantly higher in severe dysplasia (P=0.002) and in squamous metaplasia (P=0.04) compared with basal cell hyperplasia. HAS-3 was significantly expressed in basal cell hyperplasia compared with atypical adenomatous hyperplasia (P=0.05) and severe dysplasia (P=0.02). Lower expression of HAS-3 was found in severe dysplasia compared with squamous metaplasia (P=0.01) and moderate dysplasia (P=0.01). Epithelial Hyal-1 and −3 and HAS-1, −2, and −3 expressions were significantly higher in pre-neoplastic lesions than in neoplastic lesions. Comparative Cox multivariate analysis controlled by N stage and histologic tumor type showed that patients with high HAS-3 expression in pre-neoplastic cells obtained by lung/bronchial biopsy presented a significantly higher risk of death (HR=1.19; P=0.04). We concluded that localization of Hyal and HAS in lung/bronchial pre-neoplastic and neoplastic lesions was inversely related to malignancy, which implied that visualizing these factors could be a useful diagnostic procedure for suspected lung cancer. Finalizing this conclusion will require a wider study in a randomized and prospective trial.

Hyaluronidases and hyaluronan synthases expression is inversely correlated with malignancy in lung/bronchial pre-neoplastic and neoplastic lesions, affecting prognosis.

We collected a series of 136 lung/bronchial and 56 matched lung parenchyma tissue samples from patients who underwent lung/bronchial biopsies and presented invasive carcinoma after lung surgery. The lung/bronchial samples included basal cell hyperplasia, squamous metaplasia, moderate dysplasia, adenomatous hyperplasia, severe dysplasia, squamous cell carcinoma and adenocarcinoma. Matched lung parenchyma tissue samples included 25 squamous cell carcinomas and 31 adenocarcinomas. Immunohistochemistry was performed to analyze for the distribution of hyaluronidase (Hyal)-1 and -3, and hyaluronan synthases (HAS)-1, -2, and -3. Hyal-1 showed significantly higher expression in basal cell hyperplasia than in moderate dysplasia (P=0.01), atypical adenomatous hyperplasia (P=0.0001), or severe dysplasia (P=0.03). Lower expression of Hyal-3 was found in atypical adenomatous hyperplasia than in basal cell hyperplasia (P=0.01) or moderate dysplasia (P=0.02). HAS-2 was significantly higher in severe dysplasia (P=0.002) and in squamous metaplasia (P=0.04) compared with basal cell hyperplasia. HAS-3 was significantly expressed in basal cell hyperplasia compared with atypical adenomatous hyperplasia (P=0.05) and severe dysplasia (P=0.02). Lower expression of HAS-3 was found in severe dysplasia compared with squamous metaplasia (P=0.01) and moderate dysplasia (P=0.01). Epithelial Hyal-1 and -3 and HAS-1, -2, and -3 expressions were significantly higher in pre-neoplastic lesions than in neoplastic lesions. Comparative Cox multivariate analysis controlled by N stage and histologic tumor type showed that patients with high HAS-3 expression in pre-neoplastic cells obtained by lung/bronchial biopsy presented a significantly higher risk of death (HR=1.19; P=0.04). We concluded that localization of Hyal and HAS in lung/bronchial pre-neoplastic and neoplastic lesions was inversely related to malignancy, which implied that visualizing these factors could be a useful diagnostic procedure for suspected lung cancer. Finalizing this conclusion will require a wider study in a randomized and prospective trial.

Caryocar brasiliense camb protects against genomic and oxidative damage in urethane-induced lung carcinogenesis.

The antioxidant effects of Caryocar brasiliense Camb, commonly known as the pequi fruit, have not been evaluated to determine their protective effects against oxidative damage in lung carcinogenesis. In the present study, we evaluated the role of pequi fruit against urethane-induced DNA damage and oxidative stress in forty 8-12 week old male BALB/C mice. An in vivo comet assay was performed to assess DNA damage in lung tissues and changes in lipid peroxidation and redox cycle antioxidants were monitored for oxidative stress. Prior supplementation with pequi oil or its extract (15 µL, 60 days) significantly reduced urethane-induced oxidative stress. A protective effect against DNA damage was associated with the modulation of lipid peroxidation and low protein and gene expression of nitric oxide synthase. These findings suggest that the intake of pequi fruit might protect against in vivo genotoxicity and oxidative stress.

Tissue hyaluronan expression, as reflected in the sputum of lung cancer patients, is an indicator of malignancy

Hyaluronan (HA) shows promise for detecting cancerous change in pleural effusion and urine. However, there is uncertainty about the localization of HA in tumor tissue and its relationship with different histological types and other components of the extracellular matrix, such as angiogenesis. We evaluated the association between HA and degree of malignancy through expression in lung tumor tissue and sputum. Tumoral tissue had significantly increased HA compared to normal tissue. Strong HA staining intensity associated with cancer cells was significant in squamous cell carcinoma compared to adenocarcinoma and large cell carcinoma. A significant direct association was found between tumors with a high percentage of HA and MVD (microvessel density) in tumoral stroma. Similarly significant was the direct association between N1 tumors and high levels of HA in cancer cells. Cox multivariate analysis showed significant association between better survival and low HA. HA increased in sputum from lung cancer patients compared to cancer-free and healthy volunteers and a significant correlation was found between HA in sputum and HA in cancer tissue. Localization of HA in tumor tissue was related to malignancy and reflected in sputum, making this an emerging factor for an important diagnostic procedure in patients suspected to have lung cancer. Further study in additional patients in a randomized prospective trial is required to finalize these results and to validate our quantitative assessment of HA, as well as to couple it to gold standard sputum cytology.

Tissue hyaluronan expression, as reflected in the sputum of lung cancer patients, is an indicator of malignancy.

Hyaluronan (HA) shows promise for detecting cancerous change in pleural effusion and urine. However, there is uncertainty about the localization of HA in tumor tissue and its relationship with different histological types and other components of the extracellular matrix, such as angiogenesis. We evaluated the association between HA and degree of malignancy through expression in lung tumor tissue and sputum. Tumoral tissue had significantly increased HA compared to normal tissue. Strong HA staining intensity associated with cancer cells was significant in squamous cell carcinoma compared to adenocarcinoma and large cell carcinoma. A significant direct association was found between tumors with a high percentage of HA and MVD (microvessel density) in tumoral stroma. Similarly significant was the direct association between N1 tumors and high levels of HA in cancer cells. Cox multivariate analysis showed significant association between better survival and low HA. HA increased in sputum from lung cancer patients compared to cancer-free and healthy volunteers and a significant correlation was found between HA in sputum and HA in cancer tissue. Localization of HA in tumor tissue was related to malignancy and reflected in sputum, making this an emerging factor for an important diagnostic procedure in patients suspected to have lung cancer. Further study in additional patients in a randomized prospective trial is required to finalize these results and to validate our quantitative assessment of HA, as well as to couple it to gold standard sputum cytology.